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Evaluation of the amino acid methionine for biorational control of selected insect pests of economic and medical importance

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Evaluation of the amino acid methionine for biorational control of selected insect pests of economic and medical importance
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Long, Lewis Scotty
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xii, 115 leaves : ill. ; 29 cm.

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Amino acids ( jstor )
Beetles ( jstor )
Insects ( jstor )
Larvae ( jstor )
Mortality ( jstor )
Pesticides ( jstor )
Pests ( jstor )
Sugars ( jstor )
Sweeteners ( jstor )
Table sugars ( jstor )
Biological insecticides ( lcsh )
Dissertations, Academic -- Entomology and Nematology -- UF
Entomology and Nematology thesis, Ph. D
Insect pests -- Control ( lcsh )
Methionine ( lcsh )
Pests -- Integrated control ( lcsh )
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bibliography ( marcgt )
theses ( marcgt )
non-fiction ( marcgt )

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Thesis:
Thesis (Ph. D.)--University of Florida, 2004.
Bibliography:
Includes bibliographical references.
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Printout.
General Note:
Vita.
Statement of Responsibility:
by Lewis Scotty Long.

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EVALUATION OF THE AMINO ACID METHIONINE FOR BIORATIONAL CONTROL OF SELECTED INSECT PESTS OF ECONOMIC AND MEDICAL IMPORTANCE
















By

LEWIS SCOTTY LONG












A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT
OF THE REQUIREMENTS FOR THE DEGREE OF
DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2004
































Copyright 2004 by

Lewis Scotty Long














ACKNOWLEDGMENTS

I thank Jim Cuda and Bruce Stevens for giving me the financial and intellectual freedom that made this work possible. I want to thank Jim for housing me in his lab and providing the facilities to perform this work, and Bruce for allowing me to take his initial work and elaborate on it as well as including me as a co-inventor of the research presented. Most of all, I would like to express my sincere appreciation to Judy Gillmore. Without her support and help this research would not have been completed. Judy was integral in every aspect of this endeavor and put up with more than her fair share of my "research". I extend heartfelt thanks to George Gerencser, James Maruniak, Simon Yu, and Susan Webb for serving as members of my supervisory committee. I would like to also thank Jim Lloyd, Jerry Butler, and Carl Barfield for all the experiences and knowledge shared. Finally, I want to express my deepest, eternal gratitude to my fellow graduate students Jim Dunford and Heather Smith, for providing support and guidance that only colleagues, intellectual equals, and close friends can give. I can only hope to repay them for their help by providing the same amount of support for their endeavors as they did mine.












Uli































I dedicate this work to Karen, my wife and best friend. I thank her for putting up with living as a "graduate student" for the last 5 years in fulfillment of my childhood dream of being a "Doctor". She has been my pillar of support, and I would not have made it this far without her love and understanding.















TABLE OF CONTENTS

1age

ACKNOW LEDGM ENTS ................................................................................................. iii

LIST O F FIG U RES ......................................................................................................... viii

A B STR A CT ....................................................................................................................... xi

CHAPTERS

1 THE INTEGRATED PEST MANAGEMENT DILEMMA: ARE
CONVENTIONAL PESTICIDES THE ONLY ANSWER? ...............................I...

Introduction ...................................................................................................... 1
Importance of IPM in Florida and Surrounding States .................................. 2
Problems Associated with Pesticide Misuse ........................................ 4
Biorational Compounds- An Alternative to Chemical Pesticides ................ 5


2 HISTORY OF THE USE OF AMINO ACIDS AS A MEANS TO CONTROL
IN SECT PESTS ................................................................... .............................. 7

Non-Protein Amino Acids .................................................... 7
Essential Amino Acids...................................................................... 10
The Cation-Anion Modulated Amino Acid Transporter with Channel
Properties (CAATCH I) System ...................................................... 9
M ethionine .............................................................................................. 13
Research Objectives................................................. 16

3 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT
OF THE TOBACCO HORNWORM, Manduca sexta, UNDER
LABORATORY CONDITIONS ....................... ............ 17

Introduction.............................. .......................... ....................................... 17
M aterials and M ethods................................................................................ 18
Diets and Survivorship........................................................................... 18
Feeding and Development ..... ........................ 20
Preference Tests .................................................................................. 22
D ata Analysis .......................................................................................... ... 24
R esults ................................................................................................. ... 24


v









Diets and Survivorship ................... ...................................................... 24
Feeding and Development ..................................... 31
Choice Tests.................................................. ....................................... 31
Discussion ............................................................................................ 36


4 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF
THE COLORADO POTATO BEETLE, Leptinotarsa decemlineata, UNDER
LABORATORY CONDITIONS........................................................................ 39

Introduction..................................................................................................... 39
Materials and Methods......................................................................... 40
Survivorship .................................................................................. 40
Feeding and Development ................................................................ 41
Preference Tests ............................................................ 41
Data Analysis .................................................................................................. 42
Results ....................................................................................... ... 43
Survivorship ........................... ............................................... 43
Feeding and Development ................... ........... 43
Preference Tests ............................................................................. 47
Discussion ....................................................................................................... 47


5 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE
YELLOW FEVER MOSQUITO, Aedes aegypti, UNDER LABORATORY
CONDITIONS ....................................................... ..........................................52

Introduction..................................................................................................... 52
Materials and Methods.................................................................................... 53
Bioassay .................................................................................................... 53
Growth and Development ..................................... ..... ............ 54
Data Analysis .................................................................................................. 56
Results............................................................................................................. 56
Bioassay ............................................................................................... 56
Growth and Development ................................. .... ................ 59
Discussion .................................................................... ............................. 66


6 EVALUATION OF L-METHIONINE UNDER NATURAL FIELD CONDITIONS69

Introduction............................................................................................... 69
Materials and Methods............................................... 70
Preliminary Investigation of Silwet L-77 and L-methionine .................. 70
Plot Design........................................................................................... ....70
Fruit Yield .......................................................................................... 71
Pest Introduction ................................................................................. 71
Data Analysis............................................................................................ 74


vi









Results ............................................................................................................ 74
Effects of L-methionine and Silwett L-77 on Colorado Potato Beetle Adults
Under Laboratory Conditions ............................................................... 74
Effects of L-methionine and Silwett L-77 on yield ...................................... 74
Survival of CPB larvae ......................................................................... 74
Discussion .............................................................................. ....... 78


7 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE
NON-TARGET SPECIES ............................. ...................................................... 82

Introduction............................................................................................... 82
Materials and Methods................................................................................. 84
Coleomegilla maculata ........................... ....... 84
Neochetina eichhorniae ................................................................... 85
Lysiphlebus testaceipes............................................................................ 86
Data Analysis .................................................................................................. 86
Results ....................................................................................................... 87
Coleomegilla maculata ................................................................... 87
Neochetina eichhorniae .................................................................... 87
Lysiphlebus testaceipes ............................................................. 87
Discussion ....................................................................................................... 87


8 SUMMARY AND DISCUSSION........................................................... .... 96

LIST OF REFERENCES ..............................................................................................102

BIOGRAPHICAL SKETCH ...........................................................................................14






















vii








LIST OF FIGURES



Figure Page

2-1. The CAATCHI system identified from the midgut of the tobacco hornworm..............15

3-1. Rearing chamber for tobacco hornworm and Colorado potato beetle larvae used in
the artificial and excised leaf diet tests ............................................. ............. 19

3-2. Setup for whole plant studies involving tobacco hornworm ..................... 21

3-3. Chambers used for tobacco hornworm and Colorado potato beetle preference tests.....23 3-4. Amount of L-methionine present on leaf surface after treatment .............................25

3-5. Mortality of tobacco hornworm larvae exposed to various concentrations of
L-methionine (nTow=480) in artificial diet................................. .......... 26

3-6. Survivorship of THW larvae exposed to various concentrations of L-methionine
(nToaw= 1,540) on excised eggplant leaves ........................................28

3-7. Mortality of tobacco homworm larvae exposed to various concentrations of
L-methionine (nTow=256) on whole plants ..................................... ....29

3-8. Concentrations (%) of L-methionine required for the mortality of 50% of test
population of tobacco hornworm after 9 days exposure (nTow=1,540; n=180
for 3.0% L-methionine 10.0% L-methionine, n=200 for remainder) ...............30

3-9. Mortality of tobacco hornworm larvae exposed to various concentrations of Lmethionine (nTow= 160) on excised eggplant leaves for feeding and
development trials ................................................................................................32

3-10. Mean head capsule widths of tobacco hornworm larvae exposed to excised eggplant
leaves treated with various concentrations of L-methionine (nTow=320)......... 33 3-11. Total leaf area consumed by tobacco hornworm larvae exposed to excised eggplant
leaves treated with various concentrations of L-methionine (nToW=320) .....34 3-12. Mean leaf consumption by tobacco hornworm in the preference tests ........................35

4-1. Mortality of Colorado potato beetle larvae exposed to excised eggplant leaves treated
with various concentrations of L-methionine (nTrro=560) .................................44

4-2. Concentrations (%) of L-methionine concentrations required for the mortality of
50% of the test population of Colorado potato beetle after 8 days exposure
(nTo =220) ....................................................................................................... ..45


viii








4-3. Mean head capsule widths of Colorado potato beetle larvae exposed to excised
eggplant leaves treated with various concentrations of L-methionine
(nT ta=320) ......................................................... ........................................... 46

4-4. Total leaf area consumed by Colorado potato beetle larvae exposed to excised
eggplant leaves treated with various concentrations of L-methionine
(nro 320) ......................................................... ...........................................48

4-5. Mean leaf consumption by Colorado potato beetle in the preference tests ...............49

5-1. Bioassay setup for yellow fever mosquito larvae exposed to various concentrations
of am ino acids and Bti............................................... ....................................55

5-2. Mortality of yellow fever mosquito larvae exposed to various concentrations of
L-methionine (nTowt=240) ......................................57

5-3. Mortality of yellow fever mosquito larvae exposed to various concentrations of
D-methionine (nTot =240)........................................................................... ..58

5-4. Mortality of yellow fever mosquito larvae exposed to various concentrations of Trisbuffered L-methionine (nrow=240) ...................................................................60

5-5. Mortality of YFM larvae exposed to various concentrations of Proline (nTw=240).....61

5-6. Mortality of yellow fever mosquito larvae exposed to various concentrations of
L-leucine (nTow=240) .....................................................................................62

5-7. Mortality of YFM larvae exposed to various concentrations of Beta-alanine
(nTo =240) ...........................................................................................................63

5-8. Mean head capsule widths of yellow fever mosquito larvae exposed to various Tris
buffered (7.0 pH) concentrations of L-methionine (nToaw=320). ...................... 64

5-9. Concentrations (%) resulting in 50% mortality (LCso) of yellow fever mosquito
larvae exposed to various amino acids after 10 days (nrot=240 for each amino
acid) ......... .......................................................................................... 65

6-1. Overview of the design layout used to study the effects of L-methionine and Silwett
L-77@ solutions on yield of eggplant ...............................................................72

6-2. Weed Systems, Inc. KQ 3L CO2 backpack back sprayer used for application of
L-methionine and Silwett L-77@ solutions... .................................... ...73

6-3. Mortality of Colorado potato beetle adults exposed to excised eggplant leaves treated
with L-methionine and the adjuvant Silwett L-77@ (nToW=120) .....................75

6-4. Effects of L-methionine and Silwett L-77@ on eggplant yield (A) and mean weight
in grams of fruit (B) from 09 June to 31 August 2001 .....................................76


ix









6-5. Mortality of Colorado potato beetle larvae on eggplants treated with L-methionine
and Silwett L-77 ................ .......................................87

7-1. Mortality of Coleomegilla maculata adults after exposure to L-methionine treated
artificial diet ..........................................................................88

7-2. Mortality of Coleomegilla maculata adults after exposure to L-methionine treated
cotton plant leaves infested with aphids.... ...................................... ....89

7-3. Feeding scars on water hyacinth (Eichhornia crassipes) leaf after exposure to
Neochetina eichhorniae adults. .......................................................................90

7-4. Mortality of Neochetina eichhorniae on treated water hyacinth leaves ......................... 91

7-5. Feeding rate of Neochetina eichhorniae on water hyacinth leaves treated with
L-methionine and Proline .................................................................................92

7-6. Lysephlebius testiceipes parasitized aphids on cotton plants treated with
L-m ethionine ........................................................ ..........................................93

































x















Abstract of Dissertation Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy

EVALUATION OF THE AMINO ACID METHIONINE FOR BIORATIONAL CONTROL OF SELECTED INSECT PESTS OF ECONOMIC AND MEDICAL IMPORTANCE

By

Lewis Scotty Long

May, 2004


Chair: James P. Cuda
Cochair: Bruce R. Stevens
Major Department: Department of Entomology and Nematology

Integrated pest management (IPM) strategies were developed in an effort to

control pests with fewer pesticides. However, because of the misuse of pesticides and the failure to adopt IPM practices pesticide use is higher than ever. An alternative to conventional broad-spectrum pesticides is the use of biorational compounds; those that pose minimal risk to the environment and are specific to the target pests.

The recent discovery of the CAATCHI system in the midgut of the tobacco

hornworm (THW), Manduca sexta, has revealed a novel means to control certain insect pests. This membrane protein works in alkaline conditions as both an amino acid transporter and also independently as a cation channel. However, the amino acid L-methionine blocks amino acid transport thus altering the ion flow.





xi









Bioassays involving the tobacco hornworm, Colorado potato beetle (CPB),

Leptinotarsa decemlineata, and the yellow fever mosquito (YFM), Aedes aegypti, were conducted to determine the insecticidal properties of this compound. L-methionine in artificial and natural diets resulted in the mortality of 50 to 100% in concentrations of 0.3% and higher for THW and CPB. Feeding rates and larval development also were affected with reduced levels (>0.1%) of L-methionine. Bioassay trials involving YFM larvae were similar, concentrations greater than 0.1% L-methionine produced mortality rates of 70 to 100%. All three species responded better to higher concentrations of Lmethionine than to Bacillus thuringiensis, the most commonly used and commercially available biorational pesticide.

Field trials and non-target tests also were performed to determine L-methionine effectiveness under natural settings and safety to other organisms. Eggplant yield was not reduced by the application of L-methionine, which effectively controlled CPB larvae on the plants. Furthermore, several beneficial insects that were tested (a predator, a herbivore, and a parasitoid) were not affected by the addition of L-methionine to their diets.

Based on these results, L-methionine was found to be effective in controlling

selective agriculturally and medically important insect pest species, yet posed little threat to the crop plants applied to or to non-target organisms. The use of L-methionine as a pesticide, its relationship with insects and its possible use in delaying insecticide resistance were also examined.







xii













CHAPTER 1
THE INTEGRATED PEST MANAGEMENT DILEMMA: ARE CONVENTIONAL PESTICIDES THE ONLY ANSWER?

Introduction

Integrated Pest Management (IPM), the sustainable approach to the management of pest species using a combination of biological, chemical and cultural methods to reduce economic, environmental, and public health risk, was a result of economic losses associated with years of overuse of chemical control leading to resistance problems. The use of IPM strategies have certainly decreased pesticide usage and encouraged the use of methods that ensure a safer environment but many feel that it is not enough. After three decades of research efforts in the United States, IPM as it was envisioned in the 1970s was practiced on less than 8% of U.S. crop acreage based on Consumers Union estimates-well short of the national commitment to implement IPM on 75% of the total U.S. acreage by the end of the 1990s (Ehler and Bottrell 2000). This means that farm practices have changed little since the national IPM initiative was established in 1994 to implement biologically based alternatives to pesticides for controlling arthropod pests. It should be noted that the low percentage of IPM practices on commercial U.S. farmland may possibly be related to the lack of sufficient reporting means and actually may be higher than believed when the local growers and homeowners are included. However, the United States is considered the worlds' largest user of chemical pesticides, accounting for nearly 50% of total worldwide production and shows no sign of slowing (Deedat 1994). Pesticides remain the primary tool of pest consultants and farmers, because of the lack of economic incentives to adopt alternative strategies that require more effort to
1





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implement, produce unpredictable results, and require new knowledge (Barfield and Swisher 1994; Ehler and Bottrell 2000).

Importance of IPM in Florida and Surrounding States

Considerable effort has been devoted to developing IPM programs in Florida

because of its unique pest problems and crop production systems, sensitivity to chemical pollutants, and increased urbanization (Capinera et al. 1994; Rosen et al. 1996). The necessity for developing IPM protocols for Florida's major plant and animal pests was underscored in a new statewide initiative. In November 1999, the Institute of Food and Agricultural Sciences (IFAS) at the University of Florida launched Putting Florida FIRST

-Focusing IFAS Resources on Solutions for Tomorrow (Florida FIRST 1999). The Florida FIRST initiative was created (with input from stakeholders) to define the role of IFAS in shaping Florida's future in the 21st century. Increasing concerns (expressed repeatedly by Florida's scientific community and the general public) about environmental contamination, food safety issues, and human and animal health problems resulting from the indiscriminate use (and often misuse) of pesticides are making existing methods for pest management obsolete. Successful implementation of "true" IPM, as it was envisioned by those who envisioned the original concept, will have the added benefit of helping Florida ". .. enhance natural resources, provide consumers with a wide variety of safe and affordable foods, .. provide enhanced environments for homes, work places and vacations, maintain a sustainable food and fiber system, and improve the quality of life. ." (Florida FIRST 1999).

This effort to promote IPM programs in the state of Florida also benefits the

surrounding states. For example, solanaceous crops produced in the southeastern U.S. (such as tomato, tobacco, eggplant, peppers and potato) are subjected to the same





3


defoliation and fruit damage from various lepidopteran and coleopteran pests that also threaten Florida. The tomato pinworm [Keiferia lycopersicella (Walshingham) (Lepidoptera: Gelechidae)], armyworms [Spodoptera spp. (Lepidoptera: Noctuidae)], the Colorado potato beetle [Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae)], and hornworms [Manduca spp. (Lepidoptera: Sphingidae)] are some examples of pests that threaten both conventional producers and homeowners alike. For example, the estimated loss from and the cost of control of the tobacco hornworm, the number-one pest in tobacco crops in Georgia, reached $1.5 (and $2.3 million), respectively, for the years 1996-1997 (Jones and McPherson 1997). From 1992-1998, tomato, eggplant, and pepper producing areas in the Southeast had a total of 1,247,000 pounds of endosulfan applied over 270,000 acres (Aerts and Neshiem 1999; Neshiem and Vulinec 2001). The cost of insecticides applied in Florida tomato production alone for 1993-1994 amounted to approximately $1,052/hectare for a total of $2.1 million; and rose to $2550/acre, totaling $103M for the 1996-1997 season (Aerts and Neshiem 1999; Schuster et al. 1996). The use of pesticides in Florida tomato production is high because tomatoes account for 30% of the total vegetable-crop value and 13% of the total vegetable acreage for the state, with 99% of production aimed toward the fresh market (Schuster et al. 1996). For Florida potato producers, the cost of applying pesticides from 1995-1996 was $11.5M, and 96% of total Florida eggplant-crop acreage was treated with chemical insecticides (mainly methomyl and endosulfan) (Neshiem and Vulinec 2001). In addition to the monetary cost of pesticide use, commonly used insecticides such as endosulfan and fenvalerate show a high degree of toxicity to parasitoids of the tomato pinworm, thus negating the benefits of predation by natural enemies (Schuster et al. 1996). These figures may be the result of the "more is better" attitude of producers who want to avoid





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all risk of insect damage by using more applications and stronger pesticides (Schuster et al. 1996).

Problems Associated with Pesticide Misuse

The use of pesticides is not completely ruled out under IPM strategies, but rather IPM encourages responsible use to minimize environmental harm and to protect human safety and health (Deedat, 1994). However, the misuse (both intentional, in terms of "more is better;" and unintentional, as in agricultural runoff) also has resulted in resistance in some of the target pests. For example, surveys in North Carolina have shown that the Colorado potato beetle has become resistance to fenvalerate, carbofuran, and azinphosmethyl as a result of control failures in the field (Heim et al. 1990). Resistance to insecticides has also been observed in more than 450 arthropod pests (Romoser and Stoffolano 1998). Bills et al. (2004) found a 38% increase in the number of registered compounds used as pesticides from 1989-2000, and also a 16% increase in pesticide resistance of arthropod species worldwide.

Losses are not limited to agricultural systems alone. Across Africa for example, populations of insecticide-resistant mosquitoes are the result of a variety of mechanisms, including exposure to pesticide residues in agricultural runoff, mutation of target sites, and migration of resistant populations into areas where there were no previous problem (FIC-NIH 2003). Parts of southwest Asia have seen a resurgence of malaria in some areas where it was considered eradicated (due to a combination of resistance and the economics associated with control of mosquito vectors) (Deedat 1994). The importance of this example becomes even more relevant when one considers that one million individuals die every year as a result of malaria, with upwards of 500 million cases per year (Centers for Disease Control 2003). The existence of other mosquito-borne diseases





5


such as Dengue fever, yellow fever, and West Nile virus to name just a few, put countless millions more at risk. It would be dangerous to think that these diseases only occur in underdeveloped countries and not the United States. Integrated Pest Management practices also should be adopted for controlling the medical and veterinarian important insect vectors of these and other diseases.

Biorational Compounds: An Alternative to Traditional Chemical Insecticides

One way to reduce this reliance on traditional chemical pesticides and delay resistance is by increasing the variety and use biorational compounds. Biorational compounds are effective against selected pest species but are innocuous to nontarget or beneficial organisms; and have limited affect (if any) on biological control agents (Stansly et al. 1996). Biorational compounds include detergents, oils, pheromones, botanical products, microbes, and systemic and insect growth regulators (Perfect 1992; Wienzierl et al. 1998). Their safety lies in the low toxicity of the compound to nontarget organisms and the compound's short residual activity in the field. For example, Bacillus thuringiensis isrealensis (Bti) currently is one of the most widely used microbial pesticides for controlling aquatic dipteran pests (i.e., mosquitoes and black flies) because of its selectivity to this group and minimal nontarget effects (Glare and O'Callaghan 1998). However, resistance to Bt products has occurred in many species of lepidoptera from overuse of Bacillus thuringiensis kurstaki, and in some mosquito species to Bti, thus showing the need for alternatives to these compounds that are still effective (Brogdon and McAllister 1998; Marrone and Macintosh 1993). In addition to resistance, other problems are associated with the use of microbial control agents. Cook et al. (1996) discussed potential hazards, not properly identified in the planning stages, of displacement of native microorganisms, allergic responses in susceptible humans and





6


animals, and eventual toxicity to nontarget organisms. Because of these problems, alternatives are needed to prevent another crisis like the one from which IPM originally arose.













CHAPTER 2
HISTORY OF THE USE OF AMINO ACIDS AS A MEANS TO CONTROL INSECT PESTS

Non-Protein Amino Acids

One avenue of pest management explored in the field of biorational pesticides is the use nonprotein amino acids. Secondary plant materials such as these serve many functions in insect-plant relationships from attractants and repellents to crude insecticides (Dahlman 1980). Only a few nonprotein amino acids have been examined as a potential means to control insect pests. L-canavanine and its by-product of detoxification, Lcanaline, have been studied extensively, with a variety of effects ranging from developmental deformities to aberrant adult behavior (Dahlman and Rosenthal 1975; 1976; Rosenthal et al. 1995). L-canavanine is found mainly in leguminous plants, including several economic species (Bell 1978; Felton and Dahlman 1984). It is believed that plants produce this allelochemical for protection against feeding by phytophagous insects and herbivores (Rosenthal 1977). The mode of action for canavanine can be traced to several metabolic processes, including disruption of DNA/RNA and protein synthesis, arginine metabolism, uptake, anomalous canavanyl protein formation, and the reduction of active transport of K+ in the midgut epithelium (Kammer et al. 1978; Racioppi and Dahlman 1980; Rosenthal 1977; Rosenthal et al. 1977; Rosenthal and Dahlman 1991). In contrast, canaline possesses neurotoxic characteristics with an unknown mode of action (Kammer et al. 1978). The species of choice for studies involving nonprotein amino acids has been the tobacco hornworm (THW), Manduca sexta (L.) (Lepidoptera: Sphingidae).
7








L-canavanine exhibits a range of insecticidal effects in artificial diets when exposed to the THW. Dahlman (1977) demonstrated a reduction in consumption of artificial diet containing less than 1% canavanine (w/v) which resulted in a lower body mass and increased developmental time to the adult stage. Fecundity and fertility also was affected by L-canavanine. Rosenthal and Dahlman (1975) showed that concentrations as low as 0.5 mM L-canavanine in the diets of the THW resulted in the reduction of ovarial mass of adults, while Palumbo and Dahlman (1978) showed that concentrations of L-canavanine in agar-based diets resulted in the reduction of chorionated oocyte production in concentrations between 1.0 and 2.0 mM.

Under natural conditions, L-canavanine was found to retard development, and increased the susceptibility of exposed larvae to biotic and abiotic mortality factors (Dahlman 1980). However, field applications of L-canavanine were shown to be impractical because of the expense involved in synthesizing L-canavanine from its source, the jack bean (Canavalia ensiformis (L.) DC. (Family: Fabaceae)).

Other sources of L-canavanine (i.e., analogues and homologues) were sought in an attempt to find a more practical source of the amino acid. Structural homologues of canavanine were examined and found to contribute to pupal deformities (and to a lesser degree, to mortality) (Rosenthal et al. 1998). Long-chain esters of L-canavanine were found to be more toxic than the parent compound when injected or added to an artificial diet exposed to last instar of THW specimens (Rosenthal et al. 1998). Adding amino acids other than arginine (the parent compound to L-canavanine) to diets containing Lcanavanine increased deformities and mortality of THW larvae and was attributed to the structure and position of the functional groups on the added compounds (Dahlman and Rosenthal 1982). Although the THW has an effective means of degrading aberrant








proteins (produced by the assimilation of L-canavanine) into newly synthesized proteins; the proteases involved do not efficiently degrade enough to prevent some damage from occurring in the insect (Rosenthal and Dahlman 1986; 1988).

Surprisingly, L-canavanine also was shown to increase the effectiveness of

Bacillus thuringiensis in vivo by altering membrane properties, mainly gut permeability, and active transport in the midgut of the THW (Felton and Dahlman 1984). However, despite the possible synergistic relationship between the relatively safe Bt product and this amino acid, no further research has been conducted to evaluate the combination for future commercial use.

Other species of insects have also been tested for susceptibility to canavanine with a variety of results. Larvae of Drosophilia melanogaster Meigen (Diptera: Drosophilidae) showed no deleterious response to lower concentrations of canavanine, but showed mortality increased at concentrations over 1,000 ppm (Harrison and Holiday 1967). Lower concentrations also were ineffective against adult Pseudosarcophaga affinis (Fallen) (Diptera: Calliphoridae), with no effect on oocyte development (Hegdekar 1970). Dahlman et al. (1979) examined four species of muscoid flies and found greater than 70% mortality at the higher concentration (800 ppm) and decreased pupal weights as concentrations of canavanine increased.

Despite the toxicity of canavanine to some insects, others have evolved

detoxifying mechanisms to deal with high concentrations of this compound. Rosenthal et al. (1978) attributed the detoxification of canavanine in the bruchid Caryedes brasiliensis Thunberg (Coleoptera: Bruchidae) to the beetle's ability to convert canavanine to canaline, another toxic amino acid. Canaline is metabolized through reductive deamination to homoserine and ammonia, with the overall result being the detoxification





10


of the two antimetabolites. This process actually increases the nitrogen intake from the foodstuff (from the increase of ammonia) (Rosenthal et al. 1976; Rosenthal et al. 1977). Another insect, the tobacco budworm (Heliothis virescens (Fab.) (Lepidoptera: Noctuidae)) was able to metabolize far more canavanine then the bruchid beetle larva ever takes in during its development, suggesting that generalists may have more than a single detoxification mechanism for compounds they may encounter (Berge et al. 1986). Metabolism of L-canavanine by the tobacco budworm was attributed to the gut enzyme canavanine hydrolase, and may have been the result of feeding on canavanine-containing plants of the Fabaceae (Melangeli et al. 1997).

Essential Amino Acids

In despite of the extensive toxicological research conducted on nonprotein amino acids, another group of amino acids, the essential ones, has been overlooked. One reason this avenue for research has not been pursued is that we do not want to give pests convenient access to an integral part of their diet. The fear of creating a "super" insect (that has been provided with compounds that actually aid in its development) is a rational one. Mittler (1967a; 1967b) found an increase in gustation in Myzus persicae (Sulzer) (Hemiptera: Aphididae), with amino acid levels as low as 0.2% concentration in a sucrose solution. Likewise, Sugarman and Jakinovich (1986) found increased gustatory response to both D-and L-methionine by Periplaneta americana (L) (Blattodea: Blattidae) adults. Another reason that essential amino acids have not been examined for use as a pesticide is the knowledge regarding the limited mode of action these compounds could be involved with (i.e., an active site or systemic response). Recent studies on the membrane proteins of insects show the possibility of a biophysiological system that can be exploited for insect control with certain essential amino acids.













The Cation-Anion Modulated Amino Acid Transporter With Channel Properties (CAATCH 1) System

Cation-Anion modulated Aminoacid Transporter with Channel properties (CAATCH1) is a recently cloned insect-membrane protein isolated from larval midgut/hindgut nutritive absorptive epithelium. This membrane protein exhibits a unique polypeptide and nucleotide sequence related to, but different from, mammalian Na+-, C1coupled neurotransmitter transporters (Feldman et al. 2000). Using a unique PCR-based strategy, the gene encoding CAATCH1 was cloned from the digestive midgut of THW larvae. The unique biochemical, physiological, and molecular properties of CAATCHI indicate that it is a multifunction protein that mediates thermodynamically uncoupled amino acid uptake, functions as an amino acid-modulated gated alkali cation channel, and is likely a key protein in electrolyte and organic-solute homeostasis of pest insects (Quick and Stevens 2001). In the presence of no amino acids, the cations K+ and Na' are transported through the membrane via the channel (Figure 2A). When exposed to proline, the amino acid is transported through the membrane with an increase in cation flow, especially Na+ (Figure 2B). However, when exposed to methionine, the amino acid transport is stopped and cation flow is altered, mainly the increased flow of K+ and the decreased flow ofNa+ (Figure 2C). The CAATCH1 system works in alkaline conditions, at a pH optimum 9.5. This alkaline condition is found in the midgut of several species (Nation 2001) and has been attributed to a variety of causes, from the detoxification of plant allelochemicals to amino acid uptake (Giordana et al., 2002; Leonardi et al. 2001).





12




No Amino Acid Proline Methionine

g Na ++ K Na N









SNa Na+ f Na+

A B C

Figure 2. The CAATCH1 system identified from the midgut of the tobacco
hornworm (modified from Quick and Stevens 2001). In the presence of no amino acids, ion flow is similar for both K+ and Na+ (A). With the addition of an amino acid, flows are changed.
When proline is added (B), the transport occurs but the binding of the amino acid increases the ion flow, notably Nat. However, when methionine is added (C) transport occurs and the binding of the amino acid greatly decreases the flow of Na+ while K+ is
increased





13

Several amino acids were found to initiate the blocking action of ion flow through the CAATCH 1 protein, including threonine, leucine, and methionine, with the latter producing the greater response, based on CAATCH1 research (Feldman et al. 2000; Stevens et al. 2002; Quick and Stevens 2001).

Methionine

The amino acid methionine is considered essential in the diets of many organisms. Methionine is considered an indispensable amino acid in humans. Because the body does not synthesize it, uptake of methionine must occur in the diet. The recommended daily allowance of methionine for a healthy lifestyle ranges from 13 to 27 mg/kg/day for infants to full-grown adults (Young and El-Khoury 1996). This amino acid is linked to a decrease in histamine levels, increased brain function, and is found in a variety of sources; with the highest concentration in various seeds, greens, beef, eggs, chicken, and fish (Dietary Supplement Information Bureau 2000). Recently, research has centered on the genetic modification of crop plants to overproduce methionine to increase its nutritional quality (Zeh et al. 2001). Wadsworth (1995) discussed using methionine as a feed supplement, as an aid in the therapy of ketosis in livestock, and as a treatment for urinary infections in domestic pets. Onifade et al. (2001) examined the use of housefly larvae as protein foodstuffs, and found an increase in body weight gain and erythrocyte counts in rats whose diets were supplemented with fly larvae and methionine. Likewise, Koo et al. (1980) suggested dry face fly pupae could be used as a dietary supplement and foodstuff extender for poultry because of the high concentration of methionine. The environmental safety of methionine is well known, as it poses no risk to vertebrates due to a rather high oral LDs0 of 36g/kg"' observed in rats (Mallinckrodt Baker 2001) and also





14

in its use as a feed supplement for livestock under the trade name of Alimet (Novus, Inc., St. Louis, MO).

In addition to vertebrates, methionine also is considered an essential amino acid for insects (Nation 2001). Based on research on nutritional requirements for insects, the amount of methionine needed in a diet for survival ranged from as little as 0.0007 mg/mL (for Aedes aegypti (L.) (Diptera: Culicidae) to as high as 100 mg/mL (for Heliothis zea (Broddie) (Lepidoptera: Noctuidae)) (Dadd and Krieger 1968; Eymann and Friend 1985; Friend et al. 1957; Kaldy and Harper 1979; Kasting et al. 1962; Koyama 1985; Koyama and Mitsuhashi 1975; Rock and Hodgson 1971; Singh and Brown 1957). Methionine occurs naturally as the L-isomer while the D-isomer (an optical enantiomer) is toxic to many insects and is not found in nature (Anand and Anand 1990). A few exceptions are known, (mainly Diptera and Lepidoptera) that actually are capable of metabolizing the normally unusable D-isomer (Dimond et al. 1958; Geer 1966; Rock 1971; Rock et al. 1973; Rock et al. 1975). The requirement for small amounts of this amino acid (as compared to other amino acids) may be a result of the ability for some insects to synthesize methionine from cysteine (a common sulfur containing amino acid) thus reducing the need to take in exogenous sources of methionine. Jaffe and Chrin (1979) found that A. aegypti adults were able to synthesize methionine from homocysteine with the aid of a methionine synthetase. They found this enzyme similar to those common in other metazoans, and found that the levels of methionine synthetase increased with the presence of filarial parasites. They hypothesized that this increase in methionine synthetase was a result of the parasite depleting the host of methionine.

Fertility and fecundity also have been associated with methionine in some insects (mainly D. melanogaster,) with the possibility if it being a limiting factor during egg





15


production (Sang and King 1961). Lack of methionine in the diet of the female may also explain the transfer of methionine in the ejaculate of the male during fertilization (Bownes and Partridge 1987). Methionine plays another role in insect biochemistry, especially in juvenile hormone biosynthesis, inhibitory allatostatins, and storage proteins known as hexamerins. Audsley et al. (1999) found that in vitro rates of juvenile hormone synthesis in females of the tomato moth (Mamestra oleracea (L.) (Lepidoptera: Noctuidae)) were dependent on the concentration of methionine present in the incubation medium. Tobe and Clarke (1985) found a direct relationship between methionine concentration and juvenile hormone biosynthesis in the cockroach, Diplopterapunctata (Eschscholtz) (Blattodea: Blaberidae), further supporting the idea that methionine plays an important role in insect biochemistry.

Storage proteins, or hexamerins, act as a storehouse for amino acids that can be sequestered for later use in the developmental cycle (Pan and Telfer 1996). Many Lepidoptera have been identified with hexamerins containing high concentrations of methionine and are metabolized during the last larval stage, and presumably used for egg production (Wheeler et al. 2000).

Methionine as a potential pesticide has not been overlooked entirely. Tzeng

(1988) tested a methionine and riboflavin mixture and found it successful in controlling various pests, including the larvae of Culex spp. (Diptera: Culicidae). The mode of action for this mixture was attributed to a photodynamic reaction and the production of oxygen rich radicals (Tzeng et al. 1990). Their research led to the use of this methionine compound as a control agent for sooty mold of strawberry (Tzeng and Devay 1989; Tzeng et al. 1990) but not as an insecticide.





16

Discovery of novel means for controlling various insect pests is one tenant of IPM. The amino acid methionine, an environmentally safe organic compound, appears to be a candidate for further study. Before it can be considered for use in controlling insects pests, several issues must be addressed, including the determination of concentrations needed to provide effective control, compatibility with current application systems, safety to nontarget organisms (i.e., beneficial or biological-control agents), and to phytotoxicity.

Research Objectives

Our overall goal was to evaluate the effects of L-methionine, and its amino acid analogues, on the CAATCH1 system putatively in the midgut/hindgut as a means to control different insect pests. The working hypothesis is that the L-methionine only affects the CAATCH1 system and no other system, especially those involving Na+ channels or pumps (i.e., nervous tissue). The L-isomer of methionine was chosen because of the inability of most insect species to utilize the D-isomer. Ideal targets for this research are those pests that cause severe damage to agricultural systems and to human health. Specific objectives were to

Examine the effects of L-methionine as an insecticide on the larvae of M sexta
(Tobacco hornworm), L. decemlineata (Colorado potato beetle) and A. aegypti
(Yellow-fever mosquito) under various conditions

Determine any adverse effects of L-methionine on plant health to ensure its safe
use in a cropping system

Examine the effects of L-methionine on various nontarget insect species to ensure
the environmental safety of L-methionine and thus its compatibility with natural
enemies in the context of IPM.













CHAPTER 3
EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE TOBACCO HORNWORM, Manduca sexta, UNDER LABORATORY CONDITIONS Introduction


Manduca sexta (L.) (Lepidoptera: Sphingidae), the tobacco homworm (THW), is a widespread species considered an economic pest throughout North and South America. The caterpillar is known for its voracious appetite. In Georgia, the THW was responsible for between approximately $1.2 to $1.5 million in losses and costs for control annually in tobacco from 1997 to 2001 (Jones and McPherson 1997; McPherson and Jones 2002). In addition to its well-earned reputation as an agricultural pest of solanaceous crops, the THW has shown to be resistant to common pesticides (such as endrin and endosulfan), with the possibility of cross-resistance (Bills et al. 2004).

The THW also is very important to scientific research outside the arena of

economic entomology, with studies ranging from molecular-based research to ecological and physiological research, mainly because of its availability and ease in culturing (Dwyer 1999). One research area of interest to scientists involves the chemistry and physiology of the midgut. Insect control (or the development of new insecticides) was probably not the main purpose of the research that resulted in identifying the CAATCH1 protein, yet it became the basis of our research project.

Because little information is available on the insecticidal properties of

methionine, several baseline experiments were necessary to determine that concentrations of this amino acid to test. It also was necessary to test L-methionine and THW


17





i8


interaction in a variety of ways, including artificial diet, natural diet (excised leaves, whole plant, and choice tests. The purpose of this portion of this study was to determine whether L-methionine was detrimental to the survival and development of the THW and to determine if L-methionine could be used to control this species.

Materials and Methods

Eggs of THW were obtained from the insectary of North Carolina State

University, and were held in 26.4L x 19.2W x 9.5H (cm) clear plastic rearing chambers with a hardware cloth (to facilitate cleaning) (Figure 3-1). Florida Reach-In Units (FRIUs) were used to control the environment for the rearing containers (Walker et al. 1993) Containers were held at 27* C, 60% relative humidity, and a 16L:8D photoperiod in FRIUs with either artificial or natural diet (excised eggplant leaves or whole plants) depending on the pending experiment. Neonates were allowed to feed for 2 days after eclosion before being transferred to treatment groups. A camel hair brush was used for transferring larvae, to minimize the risk of damage. Diets and Survivorship

The artificial diet was prepared using the procedures outlined in Baumhover et al. (1977) with the inclusion of L-methionine for the treatment concentrations of 0.1%,

0.3%, 0.5%, 1.0%, 3.0%, 5.0% and 10.0% (wt/wt). The artificial diet was changed on a regular basis to prevent desiccation and fungal growth. Larvae were exposed to the artificial diet in the clear plastic rearing chambers with a hardware cloth, and kept in the FRIUs programmed with the aforementioned environmental constants.

Natural diets consisted of excised eggplant leaves (Solanum melongena

L.,"Classic" variety) of potted plants grown and maintained at the University of Florida,






19





















Figure 3-1. Rearing chamber for tobacco hornworm and Colorado potato
beetle larvae used in the artificial and excised leaf diet tests.
Hardware cloth stage supporting the leaf allowed for easy clean up and minimized disease problems by preventing larvae from coming in contact with fecal material (paper liner
not shown).






20

Department of Entomology and Nematology green and shade houses. Excised leaves were dipped in solutions of deionized H20 containing different concentrations of methionine; depending on the experiment and exposed to larvae in the same rearing chambers as the artificial diet trials under the same conditions. Survivorship data were pooled from several different trials for data analysis.

In total, 64 potted eggplants were used for the whole-plant portion of the study. Plants were held in FRIUs under the same conditions as the artificial and excised leaf trials, in 38H x 15D (cm) plexiglas cylinders (Figure 3-2). Four THW neonates were placed on each plant for a total of 64 larvae (16 replicates) per treatment (nTow=256 larvae). The treatment of L-methionine was applied to the test plants (using a hand-held sprayer calibrated to deliver approximately 10 mL of solution to each plant) before the addition of larvae.

Feeding and Development

To test L-methionine on the developmental rates of THW, larvae were exposed to excised eggplant leaves dipped in solutions containing the same concentrations of Lmethionine used in the artificial diet trials. Additional treatments of proline (1.0%) and Bt-kurstaki (Dipel 86% WP@ at 3.5 grams/liter; Bonide, Oriskany, NY) were included as positive and negative controls, respectively. Leaves were scanned photometrically using the CI 203 Area Meter with conveyor attachment (CID, Inc.; Camas, WA) to measure leaf consumption before and after exposure to larvae. The difference in leaf areas resulting from the missing leaf tissue was assumed to be the amount eaten by the developing larvae. Larval head capsule widths were measured at the time of death or the





21

























Figure 3-2. Setup for whole plant studies involving tobacco hornworm. Top and
portions of the sides were replaced with fine mesh to allow for
airflow and to reduce condensation.





22


end of the trial (using an Olympus Tokyo Model 213598 stereomicroscope with a optical micrometer) to monitor larval development.

Trials to determine the total amount of L-methionine applied to excised leaves also were included to quantify how much of the amino acid was physically present on leaves at the different concentration levels. Leaves were weighed before dipping into the control (0%) and L-methionine solutions (0.1%-10%), allowed to air dry for 30 min and weighed again. The difference was assumed to be the actual amount of L-methionine residue on the leaf. This value then was used to determine the total amount of L-methionine on the leaf surface of the excised leaves and the amount of L-methionine consumed per gram of leaf material, based on calculations of the physical amount of the compound for each % concentration.

Preference Tests

It was unknown if the additional methionine acted to attract or repel larvae. Neonate larvae were used in the choice tests to determine if there was a preference between the control (deionized H20) and the Treatments (1.0% L-methionine). Leaves were obtained from potted plants maintained in the outdoor shade house. The tests consisted of 4 leaf disks (30 mm diameter) dipped into the control solution and placed into the chamber alternately with four leaf disks (30 mm diameter) dipped into the treatment solution and replicated with a total of 10 chambers. Each chamber consisted of a large petri dish (25.0 cm diameter x 9.0 cm depth) lined with a Seitz@ filter disk. The filter disk was moistened routinely with deionized H20 to prevent the leaf disks from desiccation (Figure 3-3). Chambers were held in FRIUs at the same environmental constants described previously. The leaf disks also were scanned photometrically and





23























Figure 3-3. Chambers used for tobacco hornworm and Colorado potato beetle
preference tests. Two treatments (control and 1.0% Lmethionine) were used to determine if any larvae exhibited any preference or avoidance to L-methionine. Treatments were alternated in the chamber and neonates were released in the center of the dish and allowed to search for food. The filter paper in the bottom of the dish was moistened to prevent desiccation of the
leaf disks and the test specimens.






24


larval head capsule measurements made using the same procedures described in the Feeding and Development section.

Data Analysis

Sample sizes of all experiments were chosen according to the guidelines recommended by Robertson and Preisler (1991) for optimal sample size and data analysis. Probit analysis and determination of mean Lethal Concentration (LCso) were performed using PROBIT Version 1.5 (Ecological Monitoring Research Division, USEPA) after Abbott's correction for control mortality (Abbott 1925). Survival data were normalized to the previous value when control mortality was greater than the treatment mortality, to produce a smoother trend line. Statistical analysis was performed on the data using Minitab Version 14 (Minitab, Inc.; State College, PA). Analysis of the data included One-way ANOVA and separation of significant means using Tukey's Multiple Comparison and Pearson Correlation was performed on the choice trial data to examine possible relationships between development and consumption of treated leaf material (Zar 1999). Regression analysis using lest squares were performed on the leaf weights before and after the L-methionine treatment for the equation used to convert % concentration to mg/g plant material (Figure 3-4).

Results

Diets and Survivorship

The artificial diet resulted in 100% mortality of THW larvae for the 3.0%

L-methionine to 10.0% L-methionine treatment after only one day of exposure (Figure 3-5). Approximately 80% mortality was observed in the 1.0% L-methionine treatment after 4 days, and 50% mortality for both the 0.3% L-methionine and 0.5% L-methionine














*~ 0.06





0.04

0
0.03

S. Y = 8.65E-04 + 4.76E-03X S0.02 R-Sq = 98.6 %
0
0.01
95% Confidence Intewl +/- 2SE (SE=.001997)

O

0 1 2 3 4 5 6 7 8 9 10
L-methionine Concentration (%)



Figure 3-4. Amount of L-methionine present on leaf surface after treatment.
Excised leaves were weighed, dipped into various concentrations of L-methionine, allowed to dry, and then re-weighed. Difference assumed to be the amount of L-methionine remaining on leaf
surface (T=22.43, and P<0.001).





26


100 ML


i l /OO%


60 30.30%10









Day I and the 0.3% L-methionine and 0.5% L-methionine treatments from
Day 1 toDay 1000


10/0

0 2 3 4 5 6 7 8 9 10 Days of Exposure


Figure 3-5. Mortality of tobacco hornworm larvae exposed to various concentrations of
L-methionine (nTotw=480) in artificial diet. Data were adjusted using Abbott's formula to account for control mortality. Note the overlap in trend lines for the 3.0% L-methionine-10.0% L-methionine concentrations after Day I and the 0.3% L-methionine and 0.5% L-methionine treatments from
Day 1 to Day 10.





27


treatment after 10 days of exposure. The 0.1% L-methionine concentration had lowest larval mortality with approximately 30% observed for the triaL

The excised leaf trials exhibited higher mortality rates associated with the

treatments than did the artificial diet trials. Again, complete mortality was observed with the 3.0% L-methionine thru 10.0% L-methionine concentrations after 1 day of exposure (Figure 3-6). Greater than 90% in the 0.5% L-methionine and 1.0% L-methionine treatments, followed by 80% mortality in the 0.3% L-methionine treatment, and greater than 60% mortality occurred in the 0.1% L-methionine treatment after 8 days.

Whole plant trials produced results similar to the excised leaf trials with greater than 90% larval mortality observed with the 1.0% L-methionine treated plants after 14 days (Figure 3-7). Mortalities exceeding 20% and 60% were observed for the 0.1% L-methionine and 0.5% L-methionine treatments, respectively, during the same time interval.

PROBIT analysis of a sample size of nTou= 1,540 for 7 treatments (0.1% L-methionine, 0.3% L-methionine, 0.5% L-methionine, 1.0% L-methionine, 3.0% L-methionine, 5.0% L-methionine and 10.0% L-methionine) revealed an overall LC50 of

0.66% (323 mg/g leaf material) concentration for the artificial diet and 0.074% (4.39 mg/g leaf material) concentration for the natural diet after 9 days of exposure (Figure 3-8). The LC50 for the THW exposed to artificial diet was approximately half the value of that for the natural diet for the 24 to 72 hour exposure period. The LC50 for the artificial diet of 1.08% (52.3 mg/g leaf material) for 24 h dropped to 1.0% (48.5 mg/g leaf material) after 48 h and to 0.57% (28.0 mg/g leaf material) after 72 h. As for the natural diet, the LC50 of 0.53% (26.1 mg/g leaf material) was found to be lower than the artificial






28



100


80


: 60 -- -Corol 0.30%
40 .1


20 --'-3.00%

0
"1P5.00%0


0 1 2 3 4 5 6 7 8 9 10 Days of Exposure



Figure 3-6. Mortality of tobacco hornworm larvae exposed to various concentrations of
L-methionine (nTos= 1,540) on excised eggplant leaves. Data were adjusted using Abbott's formula for control mortality. Note the overlap in trend lines for the 3.0% L-methionine-10.0% L-methionine concentrations
after Day 1.






29



100 -0Control
--60.10%
80 -.--- 40.50%
1.00%
--J
60

S40
X 40 -...-.-..-.--.-.--.--20

0
0 2 4 6 8 10 12 14 Days of Exposure



Figure 3-7. Survivorship of tobacco hornworm larvae exposed to various
concentrations of L-methionine (nTotal=256) on whole plants. Lmethionine was applied using a hand-held sprayer in the amount of 10 mL/treatment. Data were adjusted using Abbott's formula for
control mortality.





30



1.2
1.1 (0.94- 1.24)
1.0 (0.85- 1.17)
.o -IIArtificial Diet
S4 0-Natural Diet

S0.8
9 0.66 (0.02 1.21)
.57 (0.47 0.69
0
' 0.6
. 0.53 (0.35 0.69)

S0.4 0.40 (0.33 0.47)

S0.25 (0.18- 0.31)

0.07 (0.03 0.13)

0.0
24h 48h 72h Overall (216h)


Figure 3-8. Concentrations (%) of L-methionine required for the mortality of 50%
of test population of tobacco hornworm after 9 days exposure (nTota=1,540; n=180 for 3.0% L-methionine 10.0% L-methionine, n=200 for remainder). Number range following value is the 95% confidence limits. Determination of LC50 was performed using PROBIT Version 1.5 (Ecological Monitoring Research Division,





31


diet at 24 h and dropped to 0.4% (19.9 mg/g leaf material) at 48 h and 0.25% (12.8 mg/g leaf material) after 72 h exposure. Overall, the LC50 at the end of the experiment for the natural diet was well below the value for the artificial diet, with close to a 90% reduction. Feeding and Development

Mortality of THW for the developmental tests ranged from approximately 30%

for the 0.1% L-methionine treatment and over 40% for the proline treatment (Figure 3-9). Complete mortality for the 0.3% L-methionine occurred after 7 days while the 0.5% L-methionine treatment took only 5 days. The Btk treatment mortality was similar to the 0.7% L-methionine and 1.0-%L-methionine treatment, resulting in 100% mortality after 1 day of exposure to the amino acid. Both the mean head capsule width and amount of leaf material consumed showed significant differences between treatments, with the control, 0.1% L-methionine and proline treatments being different that the remaining treatments (Figures 3-10 and 3-11).

Preference Tests

The amount of control and 1.0% L-methionine leaf tissue consumed during the preference tests were found not to be statistically different (Figure 3-12). In addition to the amount of leaf material consumed between treatments not being different, the mean head capsule width (i.e., development) showed a correlation with the amount of control diet consumed (Pearson Correlation Coefficient 0.885, P<0.001) while no correlation to the Treatment diet consumed (Pearson Correlation Coefficient 0.630, P=0.051) (Figure 3-11).






32





100
100- O Control

80o --0.10% A AA 0.30% 60- -- 0.50% 0.70%
N40 --1.00% 20 -*-Proline
I-Btk


0 1 2 3 4 5 6 7 8 9 10 11 Days of Exposure

Figure 3-9. Mortality of tobacco hornworm larvae exposed to various concentrations of
L-methionine (nTot= 160) on excised eggplant leaves for feeding and development trials. Proline (1.0%) and Btk were included for comparison as positive and negative controls. Data were adjusted using Abbott's formula for control mortality. Note the overlap in the 0.7% L-methionine,
1.0% L-methionine and Btk treatments at Day 1.






33


(Error Bars @ 95%; F (o.05)7,1s2=2.37, F=18.2; P <0.001)
6
A

I A A
4 4



2 B B B
1

0
Control 0.1% 0.3% 0.5% 0.7% 1.0% Proline Btt


Figure 3-10. Mean head capsule widths of tobacco homworm larvae exposed
to excised eggplant leaves treated with various concentrations of L-methionine (nTotw=320). Proline (1.0%) and Btk were included for comparison as positive and negative controls. Error bars denote 2 SE. Bars within treatments having the same letter are
not statistically different (Tukey's MST, P<0.001).






34





(Error Bars @95%; F(o.os)7sz2-2.37,F=18.2; P<0.001) 300

A
250

A A 200


150


10oo


50
B B B B B
0
Control 0.1% 0.3% 0.5% 0.7% 1.0% Proline Btk



Figure 3-11. Total leaf area consumed by tobacco hornworm larvae exposed to
excised eggplant leaves treated with various concentrations of Lmethionine (nTota=320). Proline (1.0%) and Btk were included for comparison as positive and negative controls. Error bars denote 2 SE.
Bars within treatments having the same letter are not statistically
different (Tukey's MSTP, P<0.001).





35


(Error Bars @ 95%; F(o.os)las5.98, F=1.64; P=0.217)

30
A
25

20 A



15




0o
Control 1.0% Treatment Figure 3-12. Mean leaf consumption by tobacco hornworm in the preference tests. Error bars denote 95% SE, and treatments were found not to be statistically different. However, there was correlation between the control diet consumed and mean head capsule width (Pearson Correlation Coefficient 0.885, P-0.001) while no correlation was found between the Treatment diet consumed and mean head
capsule width (Pearson Correlation Coefficient 0.630, P-0.05).





36

Discussion

The initial studies involving the high concentrations of L-methionine (i.e., 3.0-10.0%, which are outside the range normally encountered in nature) showed that a concentration of 1.0% L-methionine was sufficient enough to provide good control of THW larvae reared on both artificial and natural diets. The 0.1%L-methionine concentration remained similar to that of the control for developmental and feeding trials (Figure 3-9), indicating a level of methionine that can be tolerated to some extent, as seen in the low mortality of this treatment. This is in stark contrast to the mortality seen in the excised leaf trials in which the same concentration had over 60% mortality (Figure 3-6). One possible explanation could be the amount of L-methionine present on the leaf disk being low enough and ingested at a slower rate than that of the whole leaf, which was left in the chamber with the larvae until the leaf was either completely consumed or too wilted for the larvae to ingest.

The preference tests did show some preference towards control leaf disks over the

1.0%L-methionine treated disks as seen in the correlation analysis of the diet consumed and the mean head capsule width of the larvae. Despite the lack of a statistical difference between the amount consumed, the larvae could have fed on the treated disks and then switched to the control disks based on a physiological cue. It is unclear if THW larvae possess specialized sensory structures to detect amino acids like those found in other Lepidoptera (Beck and Henec 1958; Dethier and Kuch 1971; Schoonhoven 1972), but the possible switch from the methionine rich treatment to the control leaf disks does indicate some sort of mechanism for detection. Del Campo and Renwick (2000) found THW larvae were induced to feeding on plants outside of their normal diet when the plants





37

were treated with an extract from potato foliage suggesting induced host preference, attraction, and dependence on this compound in the extent of sustained feeding and development. A combination of sensory structures may be involved for the detection of specific amino acids and host plant compounds, which may explain the selection of methionine depleted host plants to avoid problems with the CAATCH I system present in the midgut of the THW.

The difference in the LCso for the artificial and natural diets was striking considering the concentrations were the same. One possible explanation is the L-methionine on the natural diet was more readily available than that found in the artificial diet. With the artificial diet, the L-methionine is presumably spread throughout the diet and would therefore take longer for the THW to ingest enough to adversely affect the CAATCH1 system. In contrast, the L-methionine was found on the surface of the leaf in higher concentrations than that of the artificial diet and was also freely available once ingested. Thus, larvae were exposed to a higher concentration of L-methionine with less work to digest, resulting in lower survivorship in the same period of time.

The 1.0%L-methionine concentration had the same mortality, feeding and developmental rates for THW, as did the Btk treatments (Figure 3-9). The 0.3% L-methionine, 0.5% L-methionine and 0.7% L-methionine treatments were virtually the same for mortality (Figure 3-9), developmental rate (Figure 3-10) and total leaf material consumed (Figure 3-11) and statistically the same as the 1.0% L-methionine concentration and the Btk treatment. The similar mortality rate observed for the higher concentrations of L-methionine and Btk is encouraging considering the resistance to Bt seen in many insect species because of reduced receptor activity and binding (Bills et al.





38


2004; Nester et al. 2002). Resistance in insects involves a variety of mechanisms and many are the result of a combination of different pesticide classes. The CAATCHI system is one that could be used in cases where the only alternative is by adding more pesticides or at higher rates to break resistance. Further research is needed to determine compatibility of the different Bt insecticides and L-methionine with each other for cases in which Bt resistance is observed in natural populations. Given the safety of L-methionine and the shorter time required for 100% mortality (when compared to Btk results of this study), this compound could represent a viable alternative for pesticides currently used in the management of the THW.













CHAPTER 4
EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE COLORADO POTATO BEETLE, Leptinotarsa decemlineata, UNDER LABORATORY CONDITIONS

Introduction

Leptinotarsa decemlineta (Say) (Coleoptera: Chrysomelidae), the Colorado potato beetle (CPB), is considered an economic pest throughout North America. The larvae and adults of the CPB feed on a wide variety of solanaceous crop plants and are responsible for $150 million in losses and control related costs (Durham 2000). To further complicate matters, the CPB is resistant to numerous pesticides, including various pyrethroids and carbamates (Bills et al. 2004). Historically, CPB management relied heavily on chemical control methods that led to the development of resistance to different pesticides in several areas of the eastern United States (Forgash 1985; Gauthier et al. 1981). Control of CPB without the use of chemicals is further complicated given the species ability to develop resistance and the limitations on the use of resistant varieties of potato (Ragsdale and Radcliffe 1999). The use of plant varieties that are resistant to CPB and other pests also run the risk of developing tolerance to chemical pesticides in other pest species (Sorenson et al. 1989). Despite the success of Bacillus thuringiensistenebrionis (Btt) and the biocontrol agents Podisus maculiventris Say (Hemiptera: Pentatomidae) and Edovum puttleri Grissel (Hymenoptera: Eulophidae), more biorational alternatives are necessary for controlling CPB to prevent yet another devastating threat to the potato industry because of this insect's ability develop resistance and overcome control methods (Boucher 1999; Ferro 1985; Tipping et al. 1999). This makes the CPB


39





40

an excellent candidate for the evaluation of L-methionine as a possible means of controlling this devastating pest.

Because little information is available on the insecticidal properties of L-methionine, several baseline experiments were necessary to determine what concentrations of this amino acid to test. Therefore, it was necessary to test L-methionine and CPB interaction in a variety of ways including survivorship of both larvae and adults, development of larvae when exposed to different concentrations of the amino acid, and preference tests. The purpose of this portion of this study was to conduct bioassays to determine if exposure to L-methionine was detrimental to the survival and development of the CPB and to determine if L-methionine could be used to control this species.

Materials and Methods

Eggs of CPB were obtained under UDSA permit from the insectary of the New Jersey Department of Agriculture and held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a hardware cloth (to facilitate cleaning) and held at 270 C, 60% relative humidity and 16L/8D photoperiod in FRIUs (Figure 3-1). Excised eggplant leafs were placed in the chambers with the neonates and they were allowed to feed for 2 days after eclosion before being transferred to experiments. A camel hair brush was used for transferring the neonates to minimize the risk of damaging the larvae. Survivorship

Larvae and adults of the CPB were tested in preliminary experiments with the

highest concentration (1.0% L-methionine (wt/wt)) observed in tests done on the THW in the previous chapter. The diet for the larvae and adults consisted of excised eggplant leaves (Solanum melongena L.,"Classic" variety (Family: Solanaceae)) from plants grown and maintained at the University of Florida, Department of Entomology and





41

Nematology green and shade houses. Excised leaves were dipped in solutions of deionized H20 containing different concentrations of methionine and held in the clear plastic boxes and held at the aforementioned environmental conditions (Figure 3-1). Additional treatments of proline (1.0%) and Bt-tenebrionis (Novodor@ FC @12.4 mL/L; Valent Biosciences, Libertyville, IL) were included as positive and negative controls, respectively. Survivorship data were pooled from several different trials for data analysis.

Feeding and Development

To test L-methionine on the developmental rates of CPB, larvae were exposed to excised eggplant leaves dipped in different concentrations of L-methionine under the same conditions as the survivorship trials. Additional treatments of proline (1,0%) and Btt were included as positive and negative controls, respectively. Leaves were scanned photometrically using the CI 203 Area Meter with conveyor attachment (CID, Inc., Camas, WA) before exposure to the larvae and measuring after leaf consumption. The difference in leaf areas resulting from the missing leaf tissue was assumed to be the amount eaten by the developing larvae. Larval head capsule widths were measured at the time of death or the end of the trial (using an Olympus Tokyo Model 213598 stereomicroscope with an ocular micrometer) as an evaluation of larval development. Preference Tests

It was unknown if the additional methionine acted to attract or repel larvae. Neonate larvae were used in the choice tests to determine if there was a preference between the Control (deionized H20) and the treatments (1.0% L-methionine). Leaves were obtained from potted plants maintained in the outdoor shade house. The tests





42

consisted of 4 leaf disks (30 mm diameter) dipped into the Control solution and placed into the chamber alternately with four leaf disks (30 mm diameter) dipped into the treatment solution and replicated with a total of 10 chambers. Each chamber consisted of a large petri dish (25.0 cm diameter x 9.0 cm depth) lined with a Seitz@ filter disk. The filter disk was moistened routinely with deionized H20 to prevent the leaf disks from desiccation (Figure 3-3). Chambers were held in FRIUs at the same environmental constants described previously. The leaf disks also were scanned photometrically and larval head capsule measurements made using the same procedures described in the Feeding and Development section.

Data Analysis

Sample sizes of all experiments were chosen according to the guidelines recommended by Robertson and Preisler (1991) for optimal sample size and data analysis. Probit analysis and determination of mean Lethal Concentration (LC5o) were performed using PROBIT Version 1.5 (Ecological Monitoring Research Division, USEPA) after Abbott's correction for control mortality (Abbott 1925). Survival data were normalized to the previous value when control mortality was greater than the treatment mortality, to produce a smoother trend line. Statistical analysis was performed on the data using Minitab Version 14 (Minitab, Inc.; State College, PA). Analysis of the data included One-way ANOVA and separation of significant means using Tukey's Multiple Comparison and Pearson Correlation was performed on the choice trial data to examine possible relationships between development and consumption of treated leaf material (Zar 1999).





43


Results

Survivorship

Mortality of CPB larvae on treated excised eggplant leaves ranged from

approximately 20% for the 0.1% L-methionine treatment after 4 days, 80% mortality for the 0.3% L-methionine treatment after 8 days of exposure and 100% for the remaining concentrations with the highest dose of 1.0% L-methionine exhibiting complete control of CPB in 3 days post treatment (Figure 4-1). Some mortality (50%) was observed for the proline (1.0/o) treatment while the Btt larval treatment mortality was similar to the

1.0% L-methionine treatment, resulting in 100% mortality after 5 days.

PROBIT analysis of a sample size of ntot=1,320 for 6 treatments (Control), 0.1% L-methionine, 0.3% L-methionine, 0.5% L-methionine, 0.7% L-methionine and 1.0% L-methionine) revealed an overall LCso of 0.218% concentration for the CPB after 8 days of exposure (Figure 4-2). The LC5o of 2.9% for 24 hours dropped to 1.1% after 48 hours and to 0.22% after 72 hours.

Feeding and Development

Mean head capsule widths between treatments were found to be statistically different (Figure 4-3). Four distinct groups were observed, with the Control, 0.1% L-methionine and proline treatments forming the first group. The second group of proline and 0.5% L-methionine were statistically the same and likewise the third group of the 0.3% L-methionine, 0.5% L-methionine, and 0.7% L-methionine treatments. The final group of Btt and 1.0% L-methionine treatments was statistically different from all other treatments.





44




0 Control
100 0.10%o A 0.30%
80 4 0.50% A *1 0.70%

60 _A -- .00 S-+-Proline
E -Btt
40


20



0 1 2 3 4 5 6 7 8 Days of Exposure


Figure 4-1. Mortality of Colorado potato beetle larvae exposed to excised eggplant
leaves treated with various concentrations of L-methionine (nTowt=560).
Proline (1.0%) and Btt were included for comparison as positive and negative controls. Data were adjusted using Abbott's formula for
control mortality.






45


3.5
2.91
3.0 (1.83 11.0)


o 2.5


2.0


1.5
1.11


0.330
0.22
0o.s (0.04 0.51)
.(0.16 0.26)

0.0
24h 48h 72h Overall (184h)


Figure 4-2. Concentrations (%) of L-methionine concentrations required for
the mortality of 50% of the test population of Colorado potato beetle after 8 days exposure (nTota=220). Number range following value is the 95% confidence limits. Determination of LC50 was performed using PROBIT Version 1.5 (Ecological Monitoring Research Division, USEPA), including Abbott's





46

(Error Bars @ 95%; F(o.os)7,312=1.14;F=576.71; P<0.001)
2.5
A AB

52

1.5




S0.5

0
Control 0.1% 0.3% 0.5% 0.7% 1.0% Proline Btt


Figure 4-3. Mean head capsule widths of Colorado potato beetle larvae exposed to
excised eggplant leaves treated with various concentrations of Lmethionine (nTotw=320). Proline (1.0%) and Bt were included for comparison as positive and negative controls. Error bars denote 2 SE.
Bars within treatments having the same letter are not statistically
different (Tukey's MST, P<0.001).





47


Feeding rates of CPB also were found to be statistically different among treatments (Figure 4-4). Three distinct groups were observed with the first group containing the Control and 0.1% L-methionine treatments while the second group of the 0.1% Lmethionine and 0.3% L-methionine, treatments were found to be statistically the same. The 0.5% L-methionine, 0.7% L-methionine, 1.0% L-methionine and Btt treatments were statistically different from the other groups. Overlap occurred with the proline treatment across all groups indicating no statistical difference with the rest of the treatments. Preference Tests

The amount of Control and 1.0% L-methionine leaf tissue consumed during the preference tests was found not to be statistically different (Figure 4-5). In addition, the mean head capsule width (i.e., development) showed no relationship with either treatment based upon the low correlation coefficients.

Discussion

The 1.0% L-methionine concentration produced the same larval mortality, feeding and developmental rates for CPB, as did the Btt treatments (Figures 4-1, 4-3, and 4-4). The 0.3% L-methionine, 0.5% L-methionine and 0.7% L-methionine treatments took 4 days longer for complete control (Figure 4-1), but were statistically different for the developmental rates for the same treatments (Figure 4-3). As was the case with the THW survivorship, the 0.1% L-methionine concentration was not different from that of the Control. This may indicate a threshold of methionine that can be tolerated by the THW, and CPB to some extent, evidenced by the low mortality observed for this treatment.

The Preference tests did not indicate any preference of leaf disks with or without L-methionine. The high mortality (90%) of the CPB larvae could be explained by a






48



(Error Bars @ 95%; F(o0.o)7,312=1.14;F=40.1; P<0.O01)

400
" 350 A S300
SAB
S250
ABC
o 200
SBC a 150


50 C C


Control 0.1% 0.3% 0.5% 0.7% 1.0% Proline Bit


Figure 4-4. Total leaf area consumed by Colorado potato beetle larvae exposed
to excised eggplant leaves treated with various concentrations of Lmethionine (nTow=320). Proline (1.0%) and Btt were included for comparison as positive and negative controls. Error bars denote 2 SE. Bars within treatments having the same letter are not statistically
different (Tukey's MST, P<0.001).






49



(Error Bars @ 95%; F(0.0s)l,1s=5.98, F=1.64; P=0.217)

6
A
S5

0 4
5 A

3







Control 1.0% L-methionme


Figure 4-5. Mean leaf consumption by Colorado potato beetle in the preference
tests. Error bars denote 95% SE, and treatments were found not to be statistically different. No correlation between either Control or Treatment Diet consumed and mean head capsule width was found (Pearson Correlation Coefficient 0.466, P=0.175 and 0.665,
P=0.036, respectively).





50


combination of the early consumption of the treated disks and mortality occurring after 48 hours, when a lower concentration is required for mortality. The larvae could have fed on the treated disks and then switched to the Control based on a physiological cue. Mitchell (1974) and Mitchell and Schoonhoven (1974) examined the taste receptors of CPB and found physiological and behavioral responses to some amino acids, mainly gamma aminobutyric acid (GABA) and alanine. They discussed the possibility that host selection in solanaceous plants may have been the result of these chemosensory structures and the concentration of amino acids in the leaves. It should be noted that both studies excluded methionine and no electrophysiological data were collected on the response of CPB to this amino acid. This is not surprising considering the fact that the diet of the CPB is low in methionine and therefore would not be a candidate for the inclusion in feeding stimulatory studies (Cibula et al. 1967). It is unknown if these sensory structures can detect methionine and possibly act as a means to avoid plant material high in this amino acid. This appears to be contradicted by the data in Figure 4-5, in which there was no difference between the treatments. The larvae feeding on the Control treatment, consuming the majority and then moving to the 1.0% L-methionine treatment, could explain the lack of difference.

There are some differences between some of the Feeding and Development treatments should be noted. The mean head capsule of the larvae in the 0.5% L-methionine treatment was higher than the 0.3% L-methionine treatment while the amount of leaf material consumed for the same treatment were the same indicating another factor involved with the greater head capsule width. The differences could be the result of the larger size of females and possibly could have included more females.





51

The higher concentrations of L-methionine that produced mortality similar to the Btt is encouraging considering the occurrence of resistance to this compound seen in many pest insect species because of reduced receptor activity and binding (Bills et al. 2004; Nester et al. 2002). Resistance in insects involves a variety of mechanisms and many are the result of exposure to a combination of different pesticide classes. The Methionine-CAATCH I1 system could be exploited in cases where the only alternative is applying different pesticides or using higher rates to break resistance. Further research is needed to determine compatibility with Bt and L-methionine for cases in which resistance is observed in natural populations. Given the safety of L-methionine and the shorter time required for 100% mortality (when compared to Btt), this compound could represent a new biorational tool for the management of the CPB.













CHAPTER 5
EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE
YELLOW FEVER MOSQUITO, Aedes aegypti, UNDER LABORATORY CONDITIONS

Introduction

Integrated Pest Management practices are not restricted to agricultural pests. Medically important insect pests are responsible for epidemics that have changed the course of human existence, from bubonic plague spread by the Oriental rat flea (Xenopsylla cheops Rothschild (Siphonaptera: Pulicidae)), to malaria carried by anopheline mosquitoes. One medically important species that has had a significant impact on human existence is the yellow fever mosquito (YFM), Aedes aegypti (L.) (Diptera: Culicide). This cosmopolitan species is found worldwide and is the primary vector for human dengue and yellow fever despite concerted efforts at eradication in the United States (Womack, 1993). In the United States alone, upwards of 150,000 lives were lost to yellow fever in the period starting in the late 18" century and into the early 20"h century (Patterson, 1992). However, because of the development of a vaccine, yellow fever has been replaced by Dengue which is now second only to malaria as a worldwide threat (Gubler, 1998). Because Dengue fever is also vectored by the YFM, it poses a risk by affecting tens of millions of people worldwide (Gubler and Clark, 1995).

The inclusion of the YFM in this study was an effort borne of curiosity because of the lack of knowledge of the CAATCH1 system in other insects and the availability of specimens for study. Mosquito larvae are particulate feeders and have dietary



52






53

requirements of methionine in the amounts of 0.0007mg/ml for the YFM. This amino acid also is considered essential for other species of mosquito in untraceable (in those studies) amounts (Chen, 1958; Singh and Brown, 1957). Given the high alkalinity found in the midgut of the YFM as well as other mosquito species, this physiological condition indicates the possibility of the presence of the CAATCH1 system in larval mosquitoes (Dadd, 1975).

The purpose of this portion of the study was to examine the survival and

development of YFM larvae exposed to water treated with excess L-methionine (adults were not tested given the feeding nature). In addition to L-methionine, other amino acids were tested in an effort to see if their response (i.e., survivorship) was similar CAATCH1 responses to methionine found by Feldman et al. (2000).

Materials and Methods

Bioassay

The bioassay experiments consisted of six treatments (control, 0.1%, 0.3%, 0.5%,

0.7% and 1.0%) each with four replicates. Both L-methionine and D-methionine were tested along with proline, Beta-alanine and L-leucine to examine the other amino acids that were found to be reactive to the CAATCH-1 system (Feldman et al., 2000). Bt-isrealiensis (Aquabac@ @ a rate of 2.3 mL/m2; Biocontrol Network, Brentwood, TN) and proline also were included in some trials of L-methionine to allow for comparison of both positive and negative effects. Amino acids were weighed using a Denver Instruments Co. XD2-2KD digital scale and added to glass quart jars containing 500m of deionized H20. Concentrations were based on the proportion of Ig/100ml for a 1% solution and for corresponding concentrations. Solutions were allowed to sit at room






54


temperature (23*C) to permit the amino acid to fully dissolve before the addition of the larvae. An additional trial of L-methionine buffered with Tris to a pH of 7.0 using a Fisher Scientific Accumet pH 900 was conducted to determine if mortality was attributed to a change in pH or exposure to the L-methionine.

Larvae of YFM (third instar) were obtained from the mosquito colony maintained at the Department of Entomology and Nematology, University of Florida. Larvae were transferred to the treatment jars using a camel hair, with 10 larvae per replicate for a total of 40 larvae/treatment and nTotw=240 for each amino acid bioassay experiment (Figure 5-1). Approximately 0.5g of finely ground fish food was added to serve as a larval food source and nylon window screen was used to cover the tops of the jar to prevent the escape of any emerged adults. Jars were held at 23*C on a dedicated laboratory bench top for approximately one week. The numbers of larvae surviving were recorded each day.

Growth and Development

This experiment used the same Materials and Methods as the bioassay portion

with the exception of neonate larvae instead of 3rd instars. Eggs were placed in a tray of water and held at 23*C for 2 days after eclosion. Neonates were removed using a camel hair paintbrush and placed into each jar, with 10 larvae per replicate for a total of 40 larvae/treatment (nrota=240). Larval exuviae or dead larvae were removed and used to examine growth rates by measuring the head capsules. Larvae head capsule widths were measured (using an Olympus Tokyo Model 213598 stereomicroscope with an ocular micrometer) as an evaluation of larval development.





55























Figure 5-1. Bioassay setup for yellow fever mosquito larvae exposed to various
concentrations of amino acids and Bti. Jars contained 500mL of solution and were covered with screen to prevent the escape of
emerging adults.





56


Data Analysis

Sample sizes of all experiments were selected according to the guidelines of

Robertson and Preisler (1991) for optimal sample size and data analysis. Probit analysis and determination of mean Lethal Concentration (LC5o) were performed using PROBIT Version 1.5 (Ecological Monitoring Research Division, USEPA) after Abbott's correction for control mortality (Abbott 1925). Probit analysis was performed on different concentrations (0.1% / 0.3/o, 0.5%, 0.7% and 1.0%) of L-methionine, Trisbuffered L-methionine, D-methionine, Beta-alanine, proline and L-leucine for 24, 48, 72 and 168 hours (the end of the trials). Survival data were normalized to the previous value when control mortality was greater than the treatment mortality, to produce a smoother trend line. Statistical analyses were performed on the data using Minitab Version 12. Analysis (Minitab, Inc; State College, PA) of the data included One-way ANOVA and separation of means using Tukey's Multiple Comparison test (Zar 1999).

Results

Bioassay

Mortality of YFM larvae in both the unbuffered L-and D-methionine trials was similar with low or no mortality, at the 0.1% concentrations (Figures 5-2 and 5-3). The

0.3% concentration had lower mortality with D-methionine (45%) than L-methionine (75%) and greater than 80% mortality for the 0.5% concentration for both isomers. Higher concentrations of both D-and L-methionine forms produced 100% mortality of the larvae within 2 days after treatment.

Greater than 40% mortality was observed for the buffered 0.1% L-methionine concentration with complete mortality for the remaining treatments within 5 days of






57





100


so


60
80

S030 4 5 6 7 Exposure -- 0.50%
-0- 1.00%/0 20



1 2 3 4 5 6 7 Exposure (Days)


Figure 5-2. Mortality of yellow fever mosquito larvae exposed to various
concentrations of L-methionine (nTot=240). Data were
adjusted using Abbott's formula for control mortality.






58




100


80


60- -00.70% 40 A _A 1.000


20 0
0 1 2 3 4 5 6 7 Days of Exposure


Figure 5-3. Mortality of yellow fever mosquito larvae exposed to various concentrations of D-methionine (nrot=240). Data were adjusted using Abbott's formula for control mortality.





59


exposure (Figures 5-4). The 1.0/%L.mehioione treatment caused 100% mortality after 2 days while the Bti treatment took 3 days to reach the same level of control. The proline treatment caused less than 10% mortality.

In contrast to methionine, survival of YFM larvae exposed to proline and

L-leucine was higher, with only approximately 20% mortality for the higher 0.7% proline and 1.0% proline concentrations (Figure 5-5) and less than 3% mortality with the highest L-leucine concentration (Figure 5-6). Beta-alanine mortality was similar to the L-methionine treatments with between 75% and 83% mortality for the 0.5% Beta-alanine thru 1.0% Beta-alanine concentrations, respectively, greater than 40% mortality with the 0.3% Beta-alanine, and less than 5% mortality for the 0.1% Beta-alanine concentrations (Figure 5-7).

Growth and Development

Developmental rates of YFM larvae resulted in three distinct groups, with the control and proline treatments, producing virtually identical results; both were statistically different from the 0.1%L-methionine treatment and the remaining L-methionine treatments (Figure 5-8). The Bti treatment was statistically the same as the

0.3% L-methionine to 1.0% L-methionine treatments, with very little growth taking place.

Probit analysis for unbuffered L-methionine (nTotI=40 for 5 treatments; 0.1%, 0.3%, 0.5%, 0.7% and 1.0%) revealed an overall LC50 of 0.19% concentration for the YFM after 7 days of exposure (Figure 5-9). The LC50 of 1.2% for 24 hours dropped to 0.41% after 48 hours and to 0.24% after 72 hours. When the L-methionine treatments (same concentrations) were buffered to a pH Of 7.0, the values dropped to 0.64% for 24






60




100
0-Control
--0.10%
so A 0.30% 80--0.50%
-*0.70%
1.00%
60 -Proline



40


20




0 1 2 3 4 5 6 7 8 9 10 Days of Exposure


Figure 5-4. Mortality of yellow fever mosquito larvae exposed to various
concentrations of Tris-buffered L-methionine (nTota=240). Data were adjusted using Abbott's formula for control mortality.
Note the longer exposure because of the bioassay involving neonates instead of 3rd instars. Note the overlap in some of the trend lines on Day 1 with the 0.3% L-methionine and 0.5% Lmethionine treatments.






61




100
S--Control
80 +---__--O__OA 0.30%

--0.70%



20
__ __ M,000/ 40 IM M


20



0 1 2 3 4 5 6 7 Days of Exposure


Figure 5-5. Mortality of yellow fever mosquito larvae exposed to various
concentrations of Proline (nTota=240). Data were adjusted using Abbott's formula for control mortality. Note the overlap of trend lines for all treatments except the 0.7% L-methionine and
1.0% L-methionine treatments.






62



100


80 -_ --Control 8-01-0.10%
A 0.30%
60
t 1-00.50%
4- --* 0.70%
40 -
*-0 1.00%

20

0
0 1 2 3 4 5 6 7 Days of Exposure

Figure 5-6. Mortality of yellow fever mosquito larvae exposed to various concentrations of L-leucine (nrt,)=240). Data were adjusted using Abbott's formula for control mortality. Note the overlap in trend lines for all treatments.






63



100


--*Contn

0.600

Sa A a a 0.5 40
*****0.70W 20


0
0 1 2 3 4 5 6 7 Days of Exposure


Figure 5-7. Mortality of yellow fever mosquito larvae exposed to various
concentrations of Beta-alanine (nTw=240). Data were adjusted
using Abbott's formula for control mortality.






64



(Error Bars @ 95%; F(.s)7,312=1.14, F=40.1; P<4.001)

2.5
A A
2


1.5





0.5r C C C CC


0
Control 0.1% 0.3% 0.5% 0.7% 1.0% Proline Bti

Figure 5-8. Mean head capsule widths of yellow fever mosquito larvae
exposed to various Tris buffered (7.0 pH) concentrations of Lmethionine (nTotw=320). Proline (1.0%) and Bti were included for comparison as positive and negative controls. Error bars denote 2 SE. Bars within treatments having the same letter are
not statistically different (Tukey's MST, P<0.001).






65






1.4 a


1.2 1.20
(0.95- 2.1) -"L-methionine
*1 1.*6*L-methionine (Buffered)
(0.92- 1.4) iD-methionine (0.92- 1) Beta-alanine S0.8
i 0.41(0.21 0.74)
0.6 64
0.6- )4 0.44 (0.39 0.48)
(0.33 -1.2)
(3 -0.50 (0.43 0.59) 0.33 (0.29 -0.38) 0.32 (0.27 0.36)
S 0.44 0.35 (0.28- 0.42) 0.34 (0.27 0.41)
S1.02 2.09)

0.24 (0.19 -0.28) 0.19 (0.16- 0.22)
0.2 0.11 (0.07 0.15)

S0.11 (0.08- 0.13) 0.11 (0.08 0.13)
oiT--T- ---I~
24h 48h 72h Overall (240h)


Figure 5-9. Concentrations (%) resulting in 50% mortality (LCso) of yellow fever
mosquito larvae exposed to various amino acids after 10 days (n =T240 for each amino acid). Number range following value is the 95% confidence limits. Proline and L-leucine were also tested but did
not exhibit sufficient mortality to allow for Probit Analysis.





66


hours, and to 0.11% for 48-168 hours and remained constant since the trial lasted longer because of the use of neonates instead of 3rd instars. The D-methionine treatments were similar with 0.44% for 24 and 48 hours, 0.33% for 72 hours and 0.32% after 168 hours. While not as striking as the others, Beta-alanine had a LC5o concentration of 1.1% after 24 hours, dropped to 0.5% after 48 hours and leveled off around at 0.35% after 72 and 168 hours. Probit analysis of the Proline and L-leucine treatments was not performed, as the mortality associated with those treatments was too low (Figures 5-5 and 5-6).

Discussion

Although not commonly encountered, the D- form of methionine had virtually the same effect as the L- form on larval mosquito mortality. The D-and L-methionine trials showed that the D- form had lower mortality associated with it than the more reactive L-counterpart. Insects do not commonly use the D- form of amino acids, although D-methionine is metabolized by some orders to a limited extent (Ito and Inokuchi, 1981). The YFM could be an example of this phenomenon.

Because of the nature of the CAATCH1 system in the alkaline midgut, buffering may have acted to increase the effectiveness of the system. Buffering the solutions did result in an increase in mortality, with even the lowest concentration of 0.1% L-methionine exhibiting a two-fold increase with the buffered form (Figure 5-4). Complete mortality was reached sooner with the buffered forms even for concentrations that did not reach 100% in the unbuffered form. In a field setting, the addition of L-methionine would be buffered naturally by the chemical properties of the bodies of water to which it was applied and similar results would be expected.





67

Jaffe and Chrin (1979) found the adults of YFM females infected with Brugia, a filiaral parasite, were depleted of free form methionine because of the infection and were able to make up the difference by converting homocysteine to methionine with a special synthetase. The ability of YFM adults to synthesize methionine from homocysteine may be present in the larvae as well. This could be the result of the lack of methionine in the diet and possible evidence of the CAATCHI system being present in at least the adult stage. The susceptibility of the larvae to L-methionine also could be the result of overexposure to a compound that is normally not encountered in high concentrations (>0.1%). However, the alkalinity of the particulate feeding larvae and the high mortality to L-methionine suggests that the CAATCH 1 system is present and could be exploited in other species with similar midgut characteristics (Dadd, 1975).

The survival of YFM larvae exposed to both Beta-alanine and L-leucine was unusual in that they each had the opposite effect on the YFM larvae. L-leucine was expected to have similar blocking properties as L-methionine based on CAATCHI research (Feldman et al., 2000). Instead, almost no mortality was observed indicating the possibility of another system involved with the transport of this amino acid. Conversely, beta-alanine was not found to be reactive with the CAATCHI system based on the work of Feldman et al. (2000). The unusually high larval mortality associated with this amino acid may be the result of a yet to be discovered midgut property.

The similar mortalities observed for the higher concentrations of L-methionine and Bti is encouraging considering the resistance to this compound that has been documented in many insect species because of reduced receptor activity and binding (Bills et al., 2004; Nester et al., 2002). Resistance in insects involves a variety of





68

mechanisms and many are the result of a combination of different pesticide classes. The CAATCH1 system is one that could be exploited in cases where the only alternative is applying different or higher rates of pesticides to break resistance. Further research is needed to determine compatibility of Bti and L-methionine for cases in which resistance is observed in natural populations. Given the safety of L-methionine and the similar time required for 100% mortality (when compared to Bti), this compound could represent a viable alternative to traditional biorational compounds used in the management of the YFM or other susceptible pest mosquito species.













CHAPTER 6
FIELD EVALUATION OF L-METHIONINE AS AN INSECTICIDE Introduction

The role of methionine in animal systems is well known and only recently

understood in plants. Methionine is required for protein synthesis; it is a precursor to several important biochemical compounds including ethylene and polyamines, sulfate uptake and assimilation, and also acts as an activator of threonine-synthase (Giovanelli et al. 1980; Droux et al. 2000; Bourgis et al. 2000; Zeh el al. 2001). Recently, research has focused on the transgenic modification of crop plants to overproduce methionine in order to increase their nutritional quality without affecting other biochemical processes (Zeh et al. 2001). However, little work has been conducted on the effects of exogenous methionine and it became important to understand the role of externally applied methionine on plant health.

Furthermore, the application of L-methionine to plants exposed to natural

conditions presents additional problems in terms of how long the residue remains on the plant. Observations of other experiments using L-methionine revealed the tendency of this compound to crystallize after the aqueous portion evaporated forming a brittle, crusty coating that is easily removed. This coating does not appear to interfere with respiration and transpiration at the concentrations studied (1% and lower). To prevent the loss of L-methionine from the plants in a natural setting, the adjuvant Silwett L-77@ (Helena Chemical; Collierville, TN) was included in this portion of the study in an effort to increase residual activity on the plant. Silwet L-77@ is a nonionic organosilicate


69





70


surfactant that has wetting and spreading properties (Helena Chemicals 2002) and was found to be compatible with solutions of L-methionine.

The objectives for this portion of the study were to examine the effects of a

methionine and Silwet L-77 mixture on a crop plant (eggplant) in terms of yield (both fruit weight and total yield) and to evaluate this mixture as an insecticide under natural conditions.

Materials and Methods

Preliminary Investigation of Silwet L-77@ and L-methionine

Adult CPBs were obtained from the University of Florida Horticultural Unit,

Gainesville and held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a hardware cloth (to facilitate cleaning) and held at 270C, 60% relative humidity and 16L/8D photoperiod in FRIUs. Twenty-four adults were exposed used in each of the 5 treatments, with 4 replicates per treatment (nTow=120). Adults were used because of the lack of sufficient numbers of larvae to test. Excised leaves were dipped in solutions of deionized H20 containing different concentrations of methionine and Silwett L-77@ (0.5% concentration), 0.1% L-methionine, 0.5% L-methionine, 1.0% L-methionine and controls of deionized H20 and deionized H20 +Silwet L-77. The additional control was to determine the possible insecticidal properties of Silwet L-77 alone and to make sure the addition of this adjuvant did not affect mortality or deter feeding. Plot Design

Eggplants (Solanum melongena L.,"Classic" variety) were grown and maintained at the University of Florida Horticultural Unit, Gainesville, from 18 June to 04 November 2001. Eight, one hundred ft. rows of plants were used for this study, with two rows on each side consisting of buffer rows and four rows in the middle used for the experiments.





71

Each row contained the 4 treatment plots of 10 plants (control (0% L-methionine), 0.1% L-methionine, 0.5% L-methionine and 1.0% L-methionine in deionized water solutions) in a Latin square design. Plants within treatment plots were spaced 3 feet apart while treatment plots were 9 feet apart. Figure 6-1 shows the diagrammatic representation of the field plot.

Plant Yield

Before beginning the experiment, all developing eggplants were removed from the plants in an effort to standardize the treatments and ensure all eggplant development occurred after the exposure of methionine. Treatments were administered using a KQ 3L CO2 (Weed Systems, Inc.; Hawthorne, FL) backpack sprayer charged to 30 lbs PSI and a 3-nozzle boom to ensure complete coverage of the plant (Figure 6-2). Each treatment consisted of a 3L application over the 4 representative groups. The adjuvant Silwett L-77@ (0.5% concentration) was included to improve the residual effect of the methionine under the field conditions. Plants were sprayed a total of nine times at approximately two-week intervals. Fruits were harvested at various times during the study and were weighed in the field using a Tokyo Electronics hand-held digital scale. Pest Introduction

Neonate CPB larvae were reared on excised eggplant leaves for two days at 270C, 60% relative humidity and 16L/8D photoperiod in FRIUs to ensure healthy individuals for the test. Larvae were transferred to the field plants using a camel hairbrush and the branch marked with flagging tape. Introduction was made after the last spray treatment in November. Ten larvae were placed on each plant for a total sample size of 1,600 individuals. Plants were inspected for the next 5 days and larvae encountered noted.





72



Barrier Barrier
Rows Rows






















Control (A), 0.1% (C), 0.5% (B) and 1.0% (D)

Figure 6-1. Overview of the design layout used to study the effects of
L-methionine and Silwett L-77 solutions on yield of eggplant. Rows were four feet apart with individual plants three feet apart and treatments nine feet apart.
Each letter represents a group of ten eggplants.






73






















Figure 6-2. Weed Systems, Inc. KQ 3L CO2 backpack back sprayer used for application
of L-methionine and Silwett L-77@ solutions. Boom consisted of three nozzles (middle top and end of each arm). In total, 3L were applied per
treatment every two weeks from 09 July to 31 August 2001.






74


Data Analysis

Data from the fruit and the CPB experiments were analyzed with ANOVA using Minitab Version 12. Survivorship of CPB was corrected using Abbott's formula (Abbott 1925) to account for control mortality, mean separation was performed using Tukey's multiple comparison procedure (Zar 1999). Data for both the eggplant weight mean per treatment and also mean number of eggplants per treatment were analyzed using paired t-test.

Results

Effects of L-methionine and Silwett L-77@ on CPB Adults Under Laboratory Conditions

Little mortality was observed with the adult CPB at the 1.0% L-methionine concentration (Figure 6-3). The 0.5% L-methionine concentration had the highest mortality of all the treatments at approximately 20% with the other treatments showing no adverse effects after correction for control mortality. Effects of L-methionine and Silwett L-77@ on yield

In total, 735 eggplants were collected during the course of this study from 09 June to 31 August 2001. Mean weight and yield of eggplants between the treatments were not statistically different from each other (Figures 6-4). Control plants produced 195 fruits with a mean weight of 276.9 grams, followed by the 0.1% treatment with 191 fruits at 281.2 grams. The 0.5% and 1.0%/o treatments yielded 175 and 174 fruits with mean weights of 295.7 grams and 283.6 grams, respectively. Survival of CPB larvae

No statistical difference in survivorship of CPB larvae was observed between the three treatments for the first day after exposure (Figure 6-5) but treatment differences






75




100

--*-Control Water o A 0.10%
-0-0.50%
: 60 --0 1.00%
-- Control Silwet 40


20


0
0 1 2 3 4 5 6 7 8 9 Days of Exposure


Figure 6-3. Mortality of Colorado potato beetle adults exposed to excised eggplant
leaves treated with L-methionine and the adjuvant Silwett L-77@ (nTo.=120). Data corrected for control mortality using Abbott's formula.
Note the overlap in trend lines for the Control treatments and O.l%L-m/ahionne
treatment.





76




(Error Bars @ 95%; F(.156 =2.6626, F=0.37137; P= 0.77377) A 6


4- -



2-2




Control 0.1% 0.5% 1.0%
(n=195) (n=191) (n=-175) (n= 174)


(Error Bars @ 95%; F(O3,156 =2.6626, F-030963; P= 0.81840) B 320
300
280

~260 240

220 200
Control 0.1% 0.5% 1.0%
(n=195) (n=191) (n=175) (n=174)

Figure 6-4. Effects of L-methionine and Silwett L-77@ on eggplant yield
(A) and mean weight in grams of fruit (B) from 09 June to 31 August 2001. Error bars denote 2 SE. There was no statistical difference for either eggplant yield or mean
eggplant weight (Tukey's MST, P--0.05).





77


80



S-Control
so -U--0.10%
-+- 0.50% B
-4-1.00%

t 40

B B


20- AB


AB


0 2 3 4A Days after treatment




Figure 6-5. Mortality of Colorado potato beetle larvae on eggplants treated with
L-methionine and Silwett L-77@. Mortality of larvae corrected using Abbott's formula (Abbott, 1925). Analysis performed on arcsin transformed data. Error bars denote 2 SE. Data points having by the
same letter are not statistically different (Tukey's MST, P=0.05)





78

were observed thereafter. By Day 4 the 1.0% and 0.5% treatment were the only treatments that were statistically different from the control. There was substantial unexplained attrition of CPB larvae in the field for all treatments, which leveled off by Day 3. Data from day 5 was discounted because of the onset of a severe cold front that made it difficult to separate the effects of the weather from the treatments affects.

Discussion

The results of the field studies show that, using conventional application

techniques, a mixture of methionine and Silwett L-77@ did not appear to affect eggplant yield. Furthermore, the same combination produced substantial control of CPB larvae under natural field conditions after four days. Dahlman (1980) found that L-canavanine, a non-protein amino acid, could be used in the same manner for control of THW on tobacco, but the widespread use of this compound was limited by the cost ($107.85 for Ig L-canavanine versus $3.35 for Ig of L-methionine (Fisher Scientific International 2004)), adverse effect on plant development (Nakajima et al. 2001), and toxicity to vertebrates (Rosenthal 1977). Although complete coverage of the plant was not feasible, approximately 2.5 grams to 7.5 grams of L-methionine was applied to the plants in each of the treatment plots. Each plant, based on the amount applied, received approximately 7.5x106 jig for the 1.0% L-methionine treatment, 3.8x105 jig for the 0.5% L-methionine treatment and 2.5x104 pg for the 0.1% L-methionine treatment. This compares to only 4pig of L-canavanine, which resulted in decreased size, fecundity, and mortality of THW under field conditions (Dahlman 1980). It should be noted that the toxicity of L-canavanine is well documented and has a different mode of action than L-methionine and cannot be





79

compared directly. However, the cost for the amount of L-canavanine required for largescale application far exceeds that of the largest amount of L-methionine needed.

Despite the lack of a statistical difference between treatments for both mean weight and mean yield of eggplant, there were some interesting disparities within the data. First, there was an observable difference in mean weight of the eggplants between the treatments and the control. All eggplant weights were greater for the treatments than the control, with the 0.5% L-methionine concentration treatment producing the highest mean eggplant weight. It would appear that excess methionine decreases the number of fruit produced, but those fewer eggplants weighed more. Further research is needed to better understand the differences observed during this study.

The addition of Silwet L-77@ did not appear to adversely affect survival of CPB as seen in the preliminary tests on the adults and on the larvae during the field release (Figures 6-3 and 6-5). The low adult mortality observed could be attributed to the ability of this species to stop feeding and fly to a more suitable food source. Because the adults were unable to move to an untreated leaf, they were observed sitting motionless on the underside of the leaves. This was not observed in either of the controls as they were seen actively feeding the majority of the time.

One aspect of this research that was not examined is that of fertility and fecundity of adults exposed to excess amounts of L-methionine. Despite the fact that methionine is used for egg production in many insect species, excess concentrations may act as a deterrent to feeding causing the adults to stop feeding and to seek other food sources. The lag time from the cessation in feeding to finding another food source may be long





go

enough to significantly lower the fecundity of the females and possibly interfere with other behaviors such as mating.

During the course of this portion of the study, some anecdotal data were collected based on personal observations. Predators (mainly arachnids) were observed on the plants until the end of the experiment. Other insects also were observed feeding on plants after treatments including piercing-sucking insects (i.e., aphids, coreids and cicadellids) with foliage feeders such as caterpillars rarely encountered except found only on control plants. Attempts to control predators via manual removal were unsuccessful, and predation may have contributed to the observed decrease in CPB. Because predators were present on all treatments, loss from predation was corrected with the use of Abbott's formula. The presence of natural enemies indicates the selectivity of the L-methionine in the field. The amount of methionine ingested by the predators was probably very small because they fed on other insects not plant material.

Another set of observations on the safety of L-methionine was the exposure of

potted eggplants to high (1.0% methionine in distilled H20 solution). In total, five plants were sprayed daily with the methionine solution and compared to five plants sprayed with water alone for 14 days. The only difference in the plants was the browning of the leaf tips and edges of the methionine sprayed plants. This also was seen in the excised leaf experiments with THW and CPB. A possible reason for this occurrence was the excess sulfur in the methionine might have burned the leaves. As mentioned earlier, the concentration was very high and also applied daily. Applications of the same concentration did not affect the plants in the field plots, indicating that treatments conducted at 2-week intervals would be safe for the plant.








Overall, it appears that L-methionine can be used in a natural setting to control

CPB larvae without affecting crop production. The adjuvant Silwett L-77 worked well with L-methionine in controlling CPB larvae but not the adults. The lack of effectiveness on the adults may be attributed to their ability to stop feeding and living off of reserves acquired during the larval stage until suitable food sources can be found. It is unknown if L-methionine, alone or in combination with Silwett L-77@ adversely affects fecundity of the adults.













CHAPTER 7
EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF SELECTED NONTARGET SPECIES

Introduction

A biorational pesticide is defined as one that is effective against pest species but innocuous to non-target organisms and not disruptive to biological control agents and beneficial species (Stansly et al. 1996). To test L-methionine as a potential pesticide and determine if it could be considered biorational, it was necessary to examine the effects of this compound on selected nontarget species that could possibly come into contact with it, either directly while on the plant or indirectly via incidental contact or as a host that has come into direct contact with this compound. The species chosen reflect a variety of non-target organisms, mainly those that were shown to be important in controlling some pest species. The pink spotted ladybird beetle, Coleomegilla maculata (DeGeer), the mottled water hyacinth weevil, Neochetina eichhorniae Warner, and the greenbug parasitoid, Lysiphlebus testaceipes (Cresson) all are beneficial insects that have been effective against pests in the state of Florida and also are common and readily available. Each species also represents a different feeding guild (predator, herbivore and parasitoid, respectively) to ensure a thorough examination of the possible effects of methionine as it might be encountered in under natural conditions.

The pink spotted ladybird beetle (PSLB) is an abundant polyphagus species that is known to feed on many lepidopteran and coleopteran pests, including the Colorado potato beetle, in which it was responsible for over 50% of the predation on eggs and early



82





83

instars (Andow and Risch 1985; Giroux et al. 1995; Griffin and Yeargan 2002; Groden et al. 1990; Hazzard et al. 1991; Hilbeck and Kennedy 1996; Munyaneza and Obrycki 1998). This species is widespread throughout North America, and has been shown to provide effective biological control in several crop species, including corn, crucifers, tomato and potato (Hoffman and Frodsham 1993). However, the PSLB was found to be susceptible to carbaryl and menthamidophos, the same pesticides used for the control of many aphid species (Hoffman and Frodsham 1993).

Since its introduction into the United States in 1884, water hyacinth (Eichhornia crassipes (Mart.) Solms-Laubach) has infested waterways of the southeast that has cost upwards of $2 million to control in Florida alone (Schardt 1987). The mottled water hyacinth weevil (MWHW), native to Argentina, was first released in Florida in 1972 and subsequently to other states and countries in an effort to control water hyacinth (Center 1994). The genus is restricted to feeding on members of Pontederiaceae, with the MWHW feeding mainly on the introduced water hyacinth; it can be found virtually everywhere the host plant is present (Haag and Habeck 1991; Center et al. 1998).

The greenbug parasitoid (GBP) is an important natural enemy of many cereal aphids. This species is known for the production of "mummies", the bodies of parasitized aphids that act as a protective case for the developing wasp pupa, and is considered by many to be tolerant to cold temperatures (Elliott et al. 1999; Knutson et al. 1993; Wright 1995). However, this greenbug parasitoid is an insect and is just as susceptible to pesticides despite the protective case of the immature form (Knutson et al. 1993).





84

The purpose of this portion of the study was to examine the effects of Lmethionine on selected nontarget species that are both important in terms of being beneficial in controlling other pest species and also represent different feeding guilds that would come into contact with this compound in different ways (e.g., on prey items, on plant surfaces, hosts of parasitoids).

Materials and Methods

Coleomegilla maculata

Adults were obtained from ENTOMOS, LLC (Gainesville, Florida), and were held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a hardware cloth stage inserted (to facilitate cleaning) at 270C, 60% relative humidity and 16L/8D photoperiod in FRIUs. Natural diet consisted of excised cotton leafs infested with aphids (Aphis gossypii Glover (Hemiptera: Aphididae)). Leaves were then dipped into either a 1.0% L-methionine solution or 0% L-methionine (control) mixed with deionized H20. Five adults were used in each replicate for a total n=30 for each treatment. Leaves were replaced every other day from 27 October 2002 to 07 November 2002. Artificial diet was obtained from ENTOMOS and prepared according to their guidelines with the exception of the inclusion of methionine for the 1.0% L-methionine treatment (wt/wt). Diets were replaced every other day from 27 October 2002 to 07 November 2002. Ten adults were used for each replicate for a total n=60 for each treatment. Data was normalized to 0% mortality when the treatments were corrected for control mortality (i.e., when the control mortality was greater than that of the treatment).






85

Neochetina eichhorniae

Adults of the MWHW were used in this study since the larvae and pupae are

buried deep in plant tissue and therefore not likely to come into contact with methionine that could be present in a body of water. Specimens were supplied by Hydromentia, Inc. (Ocala, FL), from areas around South Florida. Weevils were maintained following the procedures outlined by Haag and Boucias (1991), with small petri dishes fitted with moistened filter paper and freshly cut water hyacinth leaves. Water hyacinth plants were collected from Lake Alice on the campus of the University of Florida and maintained in the University of Florida, Department of Entomology and Nematology greenhouse. Treatments consisted of cut leaves dipped in deionized H20 (control) or solutions containing 0.1% L-methionine, 0.5% L-methionine, 1.0% L-methionine or 1.0% proline.

Prior to weevil exposures, each leaf was inspected for feeding scars or damage and noted to ensure the counts were based on current feeding. Each treatment consisted of 4 replicates with n=5 per replicate (n=20 per treatment and total n=100). Weevils and hyacinth leaves were held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a hardware cloth (to facilitate cleaning) and maintained at 270 C, 60% relative humidity and 16L/8D photoperiod in FRIUs. Fresh leaves were provided every 4 days; exposed leaves were preserved in sealed plastic bags and placed in a refrigerator until scars could be counted. Feeding damage was determined (with the use of an Olympus Tokyo Model 213598 stereo microscope) by the total number of scars present with each counted scar marked with a fine tipped permanent marker (Figure 7-3).

Statistical analyses of the weevil data were performed using Minitab Version 12 (Minitab, Inc.; State College, PA). Feeding scars on control and treatment leafs were





86

compared with a One-way ANOVA and mean separation was performed using Tukey's Multiple Comparison test (Zar, 1999).

Lysiphlebus testaceipes

To test the effects of methionine on the GBP, cotton plants (Gossypium sp.;

Family: Malvacae) were grown and maintained at the University of Florida, Department of Entomology and Nematology green and shade houses from 07 October 2002 to 25 November 2002. Aphids (A. gossypii Glover) were supplied from other experiments using this organism and kept on plants within a sealed greenhouse to prevent unwanted parasitism. Plants were maintained in the sealed greenhouse, infested with aphids and then placed in the open shadehouse area to encourage parasitation. In total, 20 plants were used for 2 treatments, 1.0% L-methionine and 0% L-methionine (Control) mixed with deionized H20. Plants were sprayed weekly (12 October 2002 through 17 November 2002) with approximately 10 ml of solution using a hand-held spray bottle. Counts of parasitized aphids began approximately two weeks after placing plants outside to ensure adequate time for parasitism (Royer et al. 2001). Counts were made using a hand lens and counter; "mummies" with exit holes were enumerated and removed. A few parasitized aphids were removed and held in glass vials to ensure correct identification of the parasitoid.

Data Analysis

Data from the parasitoid experiments were analyzed using Minitab Version 12 (Minitab, Inc.; State College, PA). Control and experimental plants were compared against one another with a One-way ANOVA and separation of significant means was performed with Tukey's Multiple Comparison test (Zar, 1999).





87


Results

Coleomegilla maculata

There was virtually no difference between the control and treatment groups for either the artificial or natural diet tests after correction for control mortality. Mortality was slightly higher for the control groups than the 1.0% L-methionine treatment (Figures 7-1 and 7-2). Further analysis was not necessary because of the identical numbers. Neochetina eichhorniae

Total mortality for the treatments was less than 20% for all treatments, with the individual treatments having similar results (Figure 7-4). Feeding damage ranged between 2,000 and 4,000 scars per treatment and an average of 10.7 to 16.9 scars per survivor during the course of the experiment (Figure 7-5). No statistical differences were observed between the treatment and control groups Lysiphlebus testaceipes

In total, 188 and 232 aphid mummies with exit holes were found on treatment and control plants, respectively. Means for each treatment were not statistically different for each collection period or overall based on One-way ANOVA (Figure 7-6) with the only exception being the second and last collection period.

Discussion

In general, L-methionine did not have the same toxic effect on the non-target organisms tested when compared to the pest species exposed to the compound in previous chapters. The pink spotted ladybird beetle adults actually showed the least amount of susceptibility to L-methionine. Survival of the adult beetles was higher in the

1.0% L-methionine treatments than the control for both the artificial and natural diet






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100

80 -- Control- AD

60- 4W 1.0% L-methionine- AD


40

20 Survivorship of 1.00/L- thionine Grp> Control Grp

0
0 1 2 3 4 5 6 7 8 9 10 11 12 Days After Exposure Figure 7-1. Mortality of Coleomegilla maculata adults after exposure to L-methionine
treated artificial diet. Data corrected for control mortality using Abbott's
formula.




Full Text
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evaluation. Food and Nutrition Bulletin 17(3): 191-203.
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61
Figure 5-5. Mortality of yellow fever mosquito larvae exposed to various
concentrations of Proline (nTOtai=240). Data were adjusted using
Abbotts formula for control mortality. Note the overlap of
trend lines for all treatments except the 0.7% L-methionine and
1.0% L-methionine treatments.


83
supporting price directly to buying up excess sugar in the market, the cost of the program
is expected to diminish, resulting in welfare improvement. Yet, if the U.S. government
were to switch policies from the price support to buying up excess sugar, the timing to do
so will be important so as to minimize the cost incurred by the government; the cost of
buying up excess sugar will rise immediately after policies are switched while the cost of
the price support will not because the U.S. sugar price will be maintained relatively high
in the early stage of the forecast horizon. In practice, storage costs need to be considered.
When the United States controls sugar production with Mexico, welfare improves by
increased consumer surplus and tariff revenue combined with zero program cost incurred
by the government. This scenario also demonstrates that U.S. welfare becomes better off
at the expense of the sugar industries in both countries and Mexicos welfare. When the
policy of buying up excess sugar and the production control policy are compared, the
U.S. government could pursue the latter in light of the nations welfare; however, the
gain is very small compared to the loss bom by industries and Mexico. Both alternative
policies do not satisfy pareto optimality and the overall loss outweighs the gain by the
United States as a nation.
The impact of changes in U.S. price stabilization policy is illustrated in Table 5-5
with the assumption that the U.S. government maintains the minimum quotas for the rest
of the world no matter how much Mexico exports. When the U.S. government introduces
alternative policies to price support, U.S. welfare gains but both the U.S. and Mexican
sugar industries as well as Mexicos welfare lose to a larger degree. An extreme result
comes from Scenario 13. In this scenario, the U.S. sugar price support becomes
extremely costly if the U.S. government reserves the minimum quota for the rest of the


24
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100,000
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Figure 2-1. Distribution of Individual Farm Size of Mexican Sugarcane Growers
Source: COAAZUCAR, 2003b
Jalisco
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Morelos
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Chiapas
Indicates a processing facility (mill)
Figure 2-2. Map of Mexicos Sugar Producing and Processing States.


3
including NAFTA, devaluation, privatization of the sugar cane processing industry in the
1990s, and several changes in the policy regime, sugar production has shown steady
expansion over the past 10 years: Mexican sugar production expanded from 3.2 million
MT in 1990 to 4.7 million MT in 2000 (COAAZUCAR, 2003a). A significant amount of
surplus sugar destined to export has been generated since 1995, ranging from 200,000
MT in 1995 to over 1.1 million MT in 1998 (COAAZUCAR, 2003a). These records may
appear favorable; however, Mexico stood to benefit little from NAFTA. From 1996
through 1999 Mexico successfully received a 25,000 MT import quota as a result of
attaining net surplus producer status, yet it did not enjoy the expanded quota (250,000
MT) from 2000 through 2002 (USDA, 2003a), the amount equivalent to 20 percent of the
U.S. minimum sugar import requirement under GATT, because Mexicos production fell
short relative to its sweetener consumption. This indicates that Mexico missed the
opportunity to export sugar under-quota even though it generated a significant surplus.
Combined with a slump in production that occurred in 1999 and 2000, the Mexican
sugar industry underwent an economic crisis. In September 2001, the Mexican
government expropriated 27 of 60 of Mexicos functioning sugar mills in order to
maintain the industry (USDA, 2002b). Today, the circumstances surrounding the sugar
industry remain unfavorable. At the industry level, many mills are financially vulnerable
and suffer from low efficiency of production due to old technology or poor infrastructure.
Foreign investment has not been successfully encouraged to provide capital for needed
investments in new capital equipment. At the farm level, production efficiency is low due
to fragmented farmland, which is a result of the Ejido system (Mexicos agrarian law)
and social security program specifically tailored to sugar cane growers. Lack of credit


118
difficulty in allowing faster HFCS adoption to happen and thus will continue to struggle
to suppress HFCS adoption in its market.
The impact from these three factors on the United States is also significant;
however, U.S. domestic sugar production and consumption will remain relatively
unchanged, although various simulations showed fluctuating U.S. import from Mexico
and the rest of the world caused by changes in the Mexican sweetener situation. One
notable result is that when Mexicos high production and high HFCS adoption is
combined, Mexicos sugar export takes up the entire U.S. minimum import requirement,
leaving sugar export from the rest of the world marginalized. This scenario results in not
only reduced revenue to the U.S. sugar industry as well as reduced U.S. welfare, but also
triggers political pressures from the other sugar exporters whose shares in the U.S.
market are at stake.
Mexicos HFCS adoption turns out to be beneficial only to the U.S. HFCS industry,
but Mexicos tax on HFCS benefits all except for the U.S. HFCS industry. In the latter
case, Mexico will not be able to generate sugar surplus and thus export either under
quota, over-quota or quota-free; however, gains in producer surplus outweigh the losses
in consumer surplus in both countries due to high prices of domestic sugar. This policy
lever may not be acceptable in todays trade environment and in fact the decision by the
Mexican government has changed back and forth in the past.
The Impact of Changes in U.S. Sugar Policy
Quota allocation and price stabilization policies are examined as possible changes
in the U.S. sugar policy. The impact of a quota allocation policy poses a large impact on
both Mexico and the U.S. markets. For Mexico, the aforementioned large scale of sugar
export is possible only if the U.S. government allocates quotas among exporters in a


35
(Bror Bars @ 95%; /r(0.05)i, 18=5.98, F-1.64; P =0.217)
Control 1.0% Treatment
Figure 3-12. Mean leaf consumption by tobacco homworm in the preference
tests. Error bars denote 95% SE, and treatments were found not to
be statistically different. However, there was correlation between
the control diet consumed and mean head capsule width (Pearson
Correlation Coefficient 0.885, P=0.001) while no correlation was
found between the Treatment diet consumed and mean head
capsule width (Pearson Correlation Coefficient 0.630, P=0.05).


15
whereas white sugar is as purely refined as ordinary refined sugar found elsewhere. Both
kinds of refined sugar are indistinctly consumed by households as well as domestic bulk
sugar users such as soft-drink manufacturers and confectionaries. This unique aspect is
different from the U.S., where a purer form of sugar, equivalent to white sugar in Mexico,
is what most households purchase. Molasses, the heavy dark viscous liquid residue
discharged by the centrifugal from which no more sugar can be obtained by simple means
(Polopolus and Alvarez, 1991) is utilized in rum making in Mexico.
Mexicos national total sugar consumption was approximately 4,500,000 MT, raw
equivalent, in 2001 (FAO, 2003), making it the seventh largest sugar consumption
country/ region in the world (Table 2-2). Per capita sugar consumption was 44.6 kg (98.5
lbs.), raw equivalent, in 2000; relatively high among other major sugar-producing
countries (Table 2-2). When other kinds of sweeteners such as HFCS are included, the
largest per capita consumer is the United States, followed by Cuba, Brazil, Australia, and
Mexico (Table 2-2).
Mexico as a Sugar Exporter
Surplus raw sugar is either exported to the world market, primarily to the U.S.
market due to higher price, or stored as stock. The magnitude of sugar exports from
Mexico depends on the size of surplus determined by domestic production-consumption
balance and the quota limitation imposed on all sugar imports entering the U.S. market.
Geographically Mexico holds a sugar export advantage to the United States. One of
the main shipping ports in Mexico is Veracruz, located facing the Gulf of Mexico, only
830 miles from New Orleans and 1130 miles from South Florida where sugar refinery
facilities are located. Furthermore, the state of Veracruz produces approximately 40
percent of Mexicos domestic production of sugar.


31
diet at 24 h and dropped to 0.4% (19.9 mg/g leaf material) at 48 h and 0.25% (12.8 mg/g
leaf material) after 72 h exposure. Overall, the LC50 at the end of the experiment for the
natural diet was well below the value for the artificial diet, with close to a 90% reduction.
Feeding and Development
Mortality of THW for the developmental tests ranged from approximately 30%
for the 0.1% L-methionine treatment and over 40% for the proline treatment (Figure 3-9).
Complete mortality for the 0.3% L-methionine occurred after 7 days while the 0.5%
L-methionine treatment took only 5 days. The Btk treatment mortality was similar to the
0.7% L-methionine and 1.0-%L-methionine treatment, resulting in 100% mortality after 1
day of exposure to the amino acid. Both the mean head capsule width and amount of leaf
material consumed showed significant differences between treatments, with the control,
0.1% L-methionine and proline treatments being different that the remaining treatments
(Figures 3-10 and 3-11).
Preference Tests
The amount of control and 1.0% L-methionine leaf tissue consumed during the
preference tests were found not to be statistically different (Figure 3-12). In addition to
the amount of leaf material consumed between treatments not being different, the mean
head capsule width (i.e., development) showed a correlation with the amount of control
diet consumed (Pearson Correlation Coefficient 0.885, P<0.001) while no correlation to
the Treatment diet consumed (Pearson Correlation Coefficient 0.630, P=0.051) (Figure
3-11).


BIOGRAPHICAL SKETCH
Daisuke Sano was bom on January 27, 1967, in Japan. He received the Bachelor of
Science in 1989 and the Master of Science in 1991 in applied biochemistry from the
University of Tsukuba in Japan. He served the Ministry of Agriculture, Forestry and
Fisheries of Japan as Technical Officer from 1992 to 2000; meanwhile he received the
Master of Science in food and resource economics from the University of Florida in 1999
funded by a scholarship from Japan international Cooperation Agency (JICA). He
received the Doctor of Philosophy in food and resource economics from the University of
Florida in 2004.
136


80
enough to significantly lower the fecundity of the females and possibly interfere with
other behaviors such as mating.
During the course of this portion of the study, some anecdotal data were collected
based on personal observations. Predators (mainly arachnids) were observed on the
plants until the end of the experiment. Other insects also were observed feeding on plants
after treatments including piercing-sucking insects (i.e., aphids, coreids and cicadellids)
with foliage feeders such as caterpillars rarely encountered except found only on control
plants. Attempts to control predators via manual removal were unsuccessful, and
predation may have contributed to the observed decrease in CPB. Because predators
were present on all treatments, loss from predation was corrected with the use of Abbotts
formula. The presence of natural enemies indicates the selectivity of the L-methionine in
the field. The amount of methionine ingested by the predators was probably very small
because they fed on other insects not plant material.
Another set of observations on the safety of L-methionine was the exposure of
potted eggplants to high (1.0% methionine in distilled H2O solution). In total, five plants
were sprayed daily with the methionine solution and compared to five plants sprayed
with water alone for 14 days. The only difference in the plants was the browning of the
leaf tips and edges of the methionine sprayed plants. This also was seen in the excised
leaf experiments with THW and CPB. A possible reason for this occurrence was the
excess sulfur in the methionine might have burned the leaves. As mentioned earlier, the
concentration was very high and also applied daily. Applications of the same
concentration did not affect the plants in the field plots, indicating that treatments
conducted at 2-week intervals would be safe for the plant.


33
flow of the goods in each market and factors that influence markets are illustrated in
Figure 3-1.
In Mexico, sugar distributed in the market is supplied entirely by domestic sugar
production, which is also provided entirely by domestic sugar cane growers. Sugarcane
production depends on inputs such as labor from Mexican farmers, land, and agricultural
chemicals, technology to grow and harvest sugarcane, and other factors such as weather
and government support. Growers behavior is also influenced by their relationship with
mills. Sugar processing depends on inputs such as labor from mill workers, harvested
sugarcane, and energy such as petroleum to run the facilities, technology to produce
sugar from sugarcane, infrastructure, and government support. Note that government
plays an important role to support both growers and mills activities. HFCS is primarily
supplied by domestic production and the remained is imported from the United States.
Sugar is consumed by households and bulk users (such as soft-drink manufacturers),
while HFCS is consumed only by bulk users. Determinants of sweetener demand are
income, tastes and preferences, price and other factors such as population growth.
In the United States, domestic sugar supply is derived from domestic production
and supplemented by import from various origins. Unlike Mexico, domestic sugar is
derived from both sugarcane and sugar beet production. Sugar mills produce sugar not
only from domestically produced sugarcane and sugar beets, but also from imported raw
sugar. HFCS is supplied only from domestic com sweetener manufacturers and supplies
roughly half of caloric sweetener consumption in the United States. HFCS is also
exported, and Mexico is one of the main destinations.


94
trials. One possible explanation for this observation could be that the excess
L-methionine increased the dietary quality of the artificial and natural diets for the PSLB
in the treatments. However, because only adults were available, further tests are needed
to determine if the larvae, also predaceous on the same pests as the adults, are sensitive to
this compound. It should be noted that the midgut properties (i.e., alkalinity) for this
species are not well known and may not even have the CAATCH1 proteins present in the
midgut.
The mottled water hyacinth weevil also appears not to be adversely affected by
exposure to excess amounts of L-methionine despite its herbivorous habit like the THW
and CPB. Another weevil within the same family {Anthonomus granis Boheman
(Coleptera: Curculionidae)) is known to have an acidic midgut and the same could apply
to the MWHW based on these results (Nation 2001). Therefore, this species and possibly
other weevils may not be affected by compounds like L-methionine because of the lack
of an alkaline midgut needed for the CAATCH1 protein to operate (Feldman et al. 2000;
Quick and Stevens 2001). Again, further research is necessary to determine if
CAATCH1 proteins are present in this weevil species.
The greenbug parasitoid also was unaffected by exposure to the excess
L-methionine found on treated leaves infested with aphids. Dadd and Krieger (1968)
found higher methionine requirements for the greenbug Myzus persicae Sulzer
(Hemiptera: Aphididae) when cysteine is scarce because of its ability to transform excess
methionine to much needed sulfur and could possibly explain the parasitoids tolerance to
high methionine concentrations. Because of the life cycle of the GBP, and many other
parasitoids, direct contact with compounds such as L-methionine would occur inside the


98
to the safety of this compound and residual found on the plant does not pose the same
risk to the human population.
It is difficult to understand how a compound such as methionine can be
considered essential and deadly within the same organism. To understand this
dichotomy, an examination of the role of this compound and how it relates to
metabolism, development and reproduction is necessary.
Although the diet of the THW is lacking high concentrations of methionine, the
use of hexamerins may account for the levels needed for the biosynthesis of JH. The
larvae take in methionine, metabolizing what is needed and storing the rest for later on
during metamorphosis. In contrast, the larvae of the diamondback moth (Plutella
xylostella (Lepidoptera: Plutellidae)), feeding mainly on methionine-rich crucifers, lack
hexamerins with high methionine concentrations (Wheeler et al. 2000). The levels of
methionine encountered in a normal diet are below what the CAATCH1 proteins are
capable of processing and may also be affected by the presence of symbiotic bacteria that
is responsible for methionine oxidation in some insects (Gasnier-Fauchet and Nardon
1986a; 1986b). It is when the concentration exceeds the handling capacity of the midgut
that problems occur. The time it takes to digest material containing natural amounts of
methionine could be long enough for the CAATCH1 system to recover from exposure.
The difference between the artificial and natural diet LC50 for the THW (Figure 3-8)
appears to support the idea that bound methionine (i.e., incorporated into the diet and not
applied topically) takes longer to cause problems for the organism (if any) versus the
relatively quick kill associated with the free methionine present on the leaf surface. The


LC50 (% Concentration)
65
1.4
1.06
(0.92-1.4)
0.8
0.6
0.4
0.2
0
1.20
(0.95 2
24h
48h
72h
Overall (240h)
L-methionine
L-methionine (Buffered)
D-methionine
Beta-alanine
0.41(0.21-0.74)
0.44 (0.39 0.48)
0.50 (0.43 0.59)
0.33 (0.29 -0.38)
0.35 (0.28- 0.42)
0.32 (0.27 0.36)
0.34 (0.27-0.41)
(0.16-0.22)
Figure 5-9. Concentrations (%) resulting in 50% mortality (LC50) of yellow fever
mosquito larvae exposed to various amino acids after 10 days
(nT<#ai=240 for each amino acid). Number range following value is the
95% confidence limits. Proline and L-leucine were also tested but did
not exhibit sufficient mortality to allow for Probit Analysis.


76
(Error Bars @ 95%; F(1)3fl56 =2.6626, F =0.37137; P= 0.77377)
Control 0.1% 0.5% 1.0%
(n=195) (n= 191) (n=175) (n=174)
Control 0.1% 0.5% 1.0%
(n=T95) (n=191) (n=175) (n=174)
Figure 6-4. Effects of L-methionine and Silwett L-77 on eggplant yield
(A) and mean weight in grams of fruit (B) from 09 June to
31 August 2001. Error bars denote 2 SE. There was no
statistical difference for either eggplant yield or mean
eggplant weight (Tukeys MST, Z^O.05).


3
defoliation and fruit damage from various lepidopteran and coleopteran pests that also
threaten Florida. The tomato pinworm [Keiferia lycopersicella (Walshingham)
(Lepidoptera: Gelechidae)], armyworms [Spodoptera spp. (Lepidoptera: Noctuidae)], the
Colorado potato beetle [Leptinotarsa decemlineata (Say) (Coleptera: Chrysomelidae)],
and homworms [Manduca spp. (Lepidoptera: Sphingidae)] are some examples of pests
that threaten both conventional producers and homeowners alike. For example, the
estimated loss from and the cost of control of the tobacco homworm, the number-one
pest in tobacco crops in Georgia, reached $1.5 (and $2.3 million), respectively, for the
years 1996-1997 (Jones and McPherson 1997). From 1992-1998, tomato, eggplant, and
pepper producing areas in the Southeast had a total of 1,247,000 pounds of endosulfan
applied over 270,000 acres (Aerts and Neshiem 1999; Neshiem and Vulinec 2001). The
cost of insecticides applied in Florida tomato production alone for 1993-1994 amounted
to approximately $ 1,052/hectare for a total of $2.1 million; and rose to $2550/acre,
totaling $103M for the 1996-1997 season (Aerts and Neshiem 1999; Schuster et al.
1996). The use of pesticides in Florida tomato production is high because tomatoes
account for 30% of the total vegetable-crop value and 13% of the total vegetable acreage
for the state, with 99% of production aimed toward the fresh market (Schuster et al.
1996). For Florida potato producers, the cost of applying pesticides from 1995-1996
was $11.5M, and 96% of total Florida eggplant-crop acreage was treated with chemical
insecticides (mainly methomyl and endosulfan) (Neshiem and Vulinec 2001). In addition
to the monetary cost of pesticide use, commonly used insecticides such as endosulfan and
fenvalerate show a high degree of toxicity to parasitoids of the tomato pinworm, thus
negating the benefits of predation by natural enemies (Schuster et al. 1996). These
figures may be the result of the more is better attitude of producers who want to avoid


29
Figure 2-8. U.S. HFCS Supply
Source: USDA, 2002a
Figure 2-9. U.S. HFCS Export
Source: USDA, 2002a


96
o
o
o
E
J3
0
>
1,800
1,600
1,400
1,200
1,000
800
600
400
200
0
I
es m t-
OOO
000
O
O
(N
o o
o o
CN CS
T
ON O
o
OOO
es es es
o
es
o
es
o
es
o
es
Year
El Mexico under-quota export
Mexico over-quota export
Mexico quota-free export
El The rest of the world export to the U.S.
Figure 5-7. U.S. Sugar Import Forecast (Scenario 12 PA-C-F)
Figure 5-8. U.S. Sugar Import Forecast (Scenario 16 T-S-F)


6
animals, and eventual toxicity to nontarget organisms. Because of these problems,
alternatives are needed to prevent another crisis like the one from which IPM originally
arose.


133
Haley, S. and N.R. Suarez. U.S. Sugar Policy and Prospects for the U.S. Sugar
Industry. In Schmitz, Andrew, Thomas H. Spreen, William A. Messina, Jr., and Charles
B. Moss, Sugar and Related Sweetener Markets: International Perspectives (53-64). New
York: GAB I Publishing, 2002.
Institute Nacional de Estadstica, Geografa e Informtica (INEGI). Social and
demographic statistics. Internet site:
http://www.inegi.gob.mx/difusion/ingles/fiesoc.html (Accessed July 2003).
Koo, Won W. and Richard D. Taylor. 2000 Outlook of the U.S. and World Sugar
Markets. Agricultural Economics Report No.444, Northern Plains Trade Research
Center, North Dakota State University, Fargo, ND, July 2000.
Lopez, Rigoberto A. Economic Surplus in the U.S. Sugar Market. Northeastern
Journal of Agricultural Economics. 19(1): 28-36, April 1990.
Mas-Colell, Andreu, Michael D. Whinston and Jerry R. Green. Microeconomic
Theory, New York: Oxford University Press, 1995.
McCoy, Terry L. Latin American Sweetener Markets: Economic Reform and
Regional Integration. In Schmitz, Andrew, Thomas H. Spreen, William A. Messina, Jr.,
and Charles B. Moss, Sugar and Related Sweetener Markets: International Perspectives
(81-100). New York: GABI Publishing, 2002.
Morris, Peter. Introduction to Game Theory, New York: Springer-Verlag, 1994.
Moss, Charles B. and Andrew Schmitz. Coalition Structures and U.S. Sugar
Policy. In Schmitz, Andrew, Thomas H. Spreen, William A. Messina, Jr., and Charles
B. Moss, Sugar and Related Sweetener Markets: International Perspectives (53-64). New
York: GABI Publishing, 2002.
Offenbach, Lisa A. Effects of Sugar and Ethanol Related Policies on the Market for
High Fructose Com Syrup. Ph.D. Thesis. Department of Agricultural Economics. Kansas
State University, Manhattan, KS, 1995.
Organisation for Economic Co-operation and Development (OECD). Statistics
Portal. Internet site: http://www.oecd.org/statsportal/ (Accessed November 2003).
Petrolia, Daniel R. and P. Lynn Kennedy. A Partial-Equilibrium Simulation of
Increasing the U.S. Tariff-Rate Sugar Quota for Cuba and Mexico. Selected paper
presented at the American Agriculture Economics Association meeting, Long Beach, CA,
July 2002.
Polopolus, L and J. Alvarez. Marketing Sugar and Other Sweeteners. New York:
Elsevier Science Publishing Company Inc., 1991.


Head Capsule Width (mm
46
(Error Bars @ 95%; F(o.o5)7,3i2=1.14;F=576.71; P<0.001)
Figure 4-3. Mean head capsule widths of Colorado potato beetle larvae exposed to
excised eggplant leaves treated with various concentrations of L-
methionine (nTOtai=320). Proline (1.0%) and Bt were included for
comparison as positive and negative controls. Error bars denote 2 SE.
Bars within treatments having the same letter are not statistically
different (Tukeys MST, PO.OOl).


Mean Head Capsule Width (mm)
64
(Error Bars @ 95%; F 2.5
2
1.5
1
0.5
0
Figure 5-8. Mean head capsule widths of yellow fever mosquito larvae
exposed to various Tris buffered (7.0 pH) concentrations of L-
methionine (niotai-320). Proline (1.0%) and Bti were included
for comparison as positive and negative controls. Error bars
denote 2 SE. Bars within treatments having the same letter are
not statistically different (Tukeys MST, PO.OOl).


Sugar Production [MT]
26
Year
Figure 2-4. Mexican Sugar Production, 1988-2002
Source: COAAZUCAR 2003a
Sugarcane
Cane juice
[Raw sugar + International market or stored
Refined sugar
Standar d sugar
t
White sugar (purer)
Other grades
Molasses
Alcohol (rum)
Other by-products
Domestic market
t
Figure 2-5. Main Use of Sugarcane Derivatives in Mexico
Source: Adapted from Polopolus and Alvarez, 1991


62
Figure 5-6. Mortality of yellow fever mosquito larvae exposed to various
concentrations of L-leucine (nTotai=240). Data were adjusted
using Abbotts formula for control mortality. Note the
overlap in trend lines for all treatments.


79
compared directly. However, the cost for the amount of L-canavanine required for large-
scale application far exceeds that of the largest amount of L-methionine needed.
Despite the lack of a statistical difference between treatments for both mean
weight and mean yield of eggplant, there were some interesting disparities within the
data. First, there was an observable difference in mean weight of the eggplants between
the treatments and the control. All eggplant weights were greater for the treatments than
the control, with the 0.5% L-methionine concentration treatment producing the highest
mean eggplant weight. It would appear that excess methionine decreases the number of
fruit produced, but those fewer eggplants weighed more. Further research is needed to
better understand the differences observed during this study.
The addition of Silwet L-77 did not appear to adversely affect survival of CPB
as seen in the preliminary tests on the adults and on the larvae during the field release
(Figures 6-3 and 6-5). The low adult mortality observed could be attributed to the ability
of this species to stop feeding and fly to a more suitable food source. Because the adults
were unable to move to an untreated leaf, they were observed sitting motionless on the
underside of the leaves. This was not observed in either of the controls as they were seen
actively feeding the majority of the time.
One aspect of this research that was not examined is that of fertility and fecundity
of adults exposed to excess amounts of L-methionine. Despite the fact that methionine is
used for egg production in many insect species, excess concentrations may act as a
deterrent to feeding causing the adults to stop feeding and to seek other food sources.
The lag time from the cessation in feeding to finding another food source may be long


121
protecting the sugar industry consisting of a large number of growers and related
workers.
When the U.S. HFCS industry is included in a coalition, a conflict of interest
between the governments and industries becomes intensified. When pay-off to the U.S.
HFCS industry is pooled into a coalition, the industry coalition (the U.S. HFCS industry,
the U.S. sugar and Mexican sugar industries) will lobby for the strategy that involves
Mexicos HFCS adoption. Although the game has reached a different solution through
governments decision making, this is true because the industries recognize a better pay
off to the coalition. This contrast implies that the U.S. HFCS and the U.S. sugar
industries have a strong incentive to influence Mexicos choice of strategy. Furthermore,
since the Mexican sugar industry theoretically becomes better off from being
compensated by the industry coalition, it may be tempted to allow HFCS adoption rather
than expecting protection from its own government.
The results from games with a grand coalition indicate that there is no solution that
satisfies pareto optimality. If redistribution of pooled revenues from industries and
welfare is feasible, there are solutions that minimize the total losses for all entities.
In summary, in no case did the game reach a solution that included the current U.S.
policy of price support as the best policy; alternative policies examined in the study are
better sugar policies to pursue. Also, the stance of the U.S. HFCS industry on sugar
policy can be influential. If the U.S. HFCS industry is included in the decision-making
process, a sugar policy that differs from that chosen by the governments can be lobbied
for with enticing side agreements within the coalition.


6
1. What was the impact of changes in the trade regime in the U.S. and Mexican
sweetener market since NAFTA was implemented in 1994?
2. How much surplus sugar can Mexico generate and how much sugar will cross
the border both under- and over-quota? What will happen after 2008 when all
the restrictions are eliminated on Mexican sugar?
3. What will be the impact of changes in Mexicos market on both the United
States and Mexico? How much influence will HFCS adoption cause in both the
U.S. and Mexican sweetener markets?
4. What will be the impact of changes in U.S. sugar policy on both the United
States and Mexico?
5. Is there alternative sugar policy for the United States to current price support?
Objectives
The primary objective of the study is to develop a bilateral trade model of the U.S.-
Mexico sugar industry that reflects provisions of NAFTA, as well as related market
conditions in order to forecast the outlook of the sweetener market through various
simulations, encompassing hypothetical changes in Mexican sweetener situations and
U.S. sugar policy.
Secondly, the study aims to provide policy recommendations by examining
aggregated results from these simulations, paying attention to identify gainers and losers
under different scenarios. By doing so, the study hopes to illustrate conflicts of interest
among the various players in the U.S.-Mexico sweetener market.
Organization of the Study
The remainder of the dissertation is organized as follows. In chapter 2, the sugar
industries in both the United States and Mexico are introduced in the context of the
sweetener market in each country as well as the integrated market, paying close attention
to historical and political perspectives. In chapter 3, the conceptual and theoretical


Copyright 2004
by
Daisuke Sano


94
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£
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1,400
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CN
O
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CN
O
CN
E3 Mexico under-quota export
Mexico over-quota export
Mexico quota-free export
E3The rest of the world export to the U.S.
Figure 5-3. U.S. Sugar Import Forecast (Scenario 3 A-S-F)
o
o
o
QJ
£
a
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1,400 -
1,200
1,000
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es
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es
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es
Year
H Mexico under-quota export
Mexico over-quota export
Mexico quota-free export
O The rest of the world export to the U.S.
Figure 5-4. U.S. Sugar Import Forecast (Scenario 4 PA-S-F)


Barrier Barrier
Rows Rows
Figure 6-1. Overview of the design layout used to study the effects of
L-methionine and Silwett L-77 solutions on yield of
eggplant. Rows were four feet apart with individual
plants three feet apart and treatments nine feet apart.
Each letter represents a group of ten eggplants.


57
to the United States in order to iterate for solution from 2002 through 2015 using the
mathematical programming software package GAMS.
Intercepts of inverse linear demand and supply equations in quantity-price space
(IUi, IUUi, IM¡, and IMM¡ in equations [4.14] through [4.17], respectively) are calibrated
with the actual values realized in 2001 (base year) from equations [C.7b], [C8b], [C9b]
and [C.lOb] presented in Appendix C:
IU¡ = P us, 2001 (IU2*QDus, 2001 + Shifter us, 2001) (4.14)
IUUi = PSus, 2001 (IUU2*QSus, 2001 + Shifter s us, 2001 ) (4.15)
IMi = Pmx, 2001 (IM2*QTCmx, 2001 + Shifter0mx, 2001) (4.16)
IMM i = P5 mx, 2001 ~ (1MM2*QS mx, 2001 + Shifter^mx, 2001)- (4.17)
By calibrating intercepts, both excess supply in Mexico (quantity presented as A on
ESmx curve in Figure 4-1) and excess demand in the United States (quantity presented as
B on EDus curve in Figure 4-1) are set to correspond to the actual volume realized in
2001. Slopes for both inverse demand and inverse supply curves in both countries are
held constant, yet these curves shift over the forecast horizon according to the scenarios
proposed in the following section.
In order to represent Mexican sugar supply and export capacity adequately, the
model is further calibrated by adjusting the average transportation cost from the rest of
the world to the United States, based on the relationship expressed in equation [3.50] in
chapter 3. To do so, the transportation cost is calibrated so that Mexico exports sugar
over-quota at the minimum amount. This calibration procedure resulted in a rather high
transportation cost from the rest of the world to the United States; however, it insinuates
the irrational behavior of the Mexican sugar industry which has been suffering from


22
end of the trial (using an Olympus Tokyo Model 213598 stereomicroscope with a optical
micrometer) to monitor larval development
Trials to determine the total amount of L-methionine applied to excised leaves
also were included to quantify how much of the amino acid was physically present on
leaves at the different concentration levels. Leaves were weighed before dipping into the
control (0%) and L-methionine solutions (0.1%-10%), allowed to air dry for 30 min and
weighed again. The difference was assumed to be the actual amount of L-methionine
residue on the leaf. This value then was used to determine the total amount of
L-methionine on the leaf surface of the excised leaves and the amount of L-methionine
consumed per gram of leaf material, based on calculations of the physical amount of the
compound for each % concentration.
Preference Tests
It was unknown if the additional methionine acted to attract or repel larvae.
Neonate larvae were used in the choice tests to determine if there was a preference
between the control (deionized H2O) and the Treatments (1.0% L-methionine). Leaves
were obtained from potted plants maintained in the outdoor shade house. The tests
consisted of 4 leaf disks (30 mm diameter) dipped into the control solution and placed
into the chamber alternately with four leaf disks (30 mm diameter) dipped into the
treatment solution and replicated with a total of 10 chambers. Each chamber consisted of
a large petri dish (25.0 cm diameter x 9.0 cm depth) lined with a Seitz filter disk. The
filter disk was moistened routinely with deionized H2O to prevent the leaf disks from
desiccation (Figure 3-3). Chambers were held in FRIUs at the same environmental
constants described previously. The leaf disks also were scanned photometrically and


CHAPTER 3
EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE
TOBACCO HORNWORM, Manduca sexta, UNDER LABORATORY CONDITIONS
Introduction
Manduca sexta (L.) (Lepidoptera: Sphingidae), the tobacco homworm (THW), is
a widespread species considered an economic pest throughout North and South America.
The caterpillar is known for its voracious appetite. In Georgia, the THW was responsible
for between approximately $1.2 to $1.5 million in losses and costs for control annually in
tobacco from 1997 to 2001 (Jones and McPherson 1997; McPherson and Jones 2002). In
addition to its well-earned reputation as an agricultural pest of solanaceous crops, the
THW has shown to be resistant to common pesticides (such as endrin and endosulfan),
with the possibility of cross-resistance (Bills et al. 2004).
The THW also is very important to scientific research outside the arena of
economic entomology, with studies ranging from molecular-based research to ecological
and physiological research, mainly because of its availability and ease in culturing
(Dwyer 1999). One research area of interest to scientists involves the chemistry and
physiology of the midgut. Insect control (or the development of new insecticides) was
probably not the main purpose of the research that resulted in identifying the CAATCH1
protein, yet it became the basis of our research project.
Because little information is available on the insecticidal properties of
methionine, several baseline experiments were necessary to determine that concentrations
of this amino acid to test It also was necessary to test I .-methionine and THW
17


41
Xm, us Quota < 0 (3.29)
Xrow, us + XXmx, us USMin > 0 (3.30)
where Quotais the quota Mexico is assigned according to the NAFTA provisions and
USMin is the quota allocated to the rest of the world (the U.S. minimum sugar import
quota less the quota assigned to Mexico). Equation [3.25] and [3.26] satisfy domestic
demand; equation [3.27] and [3.28] regulate the quantities shipped from supply regions;
and equations [3.29] and [3.30] define quotas imposed on Mexican sugar and from the
rest of the world.
The implications of the model can be examined by writing the first-order
conditions obtained through using Kuhn-Tucker theorem. From equations from [3.24] to
[3.30] the Lagrangian form (L) is given as:
L= (1/2 )* IU2*(QDus)2 +m + Shifted us, )*Q us
+ (1/2)* IM2*(QTCmx)2 + (IM, + Shifter0mx)*QTCmx
- (1/2)* IUU2*(QSusf + (IUU,+ Shifter3us)*Qsus
- (1/2)* IMM2*(QSmx)2 + (IMM, + Shifter3mx)*QSmx
(Tmx, US)*XMX, US
(Tus, MX )*X US, MX
(Tmx, us + OQTarMx, us)*XXmx, us
- (Trow, us + Prow)*Xrow, us
+ PROW *XmX, row
+ VUS (X US, US + x MX, US + XX MX, US + X ROW, us QUS )
+ yMX (XMX, MX + X US, MX Q'^ Mx)
+ Wus (Q3us Xus, us X US, MX )


79
the U.S. quota. Thus, allocating such a large portion of the quota to one country may not
be a feasible policy option in the United States. In Scenario 13, it is conjectured that the
U.S. government maintains minimum quotas (the remainder of the minimum import
requirement less allocated to Mexico) for the rest of the world no matter how much
Mexico exports. As shown in Figure 5-5, Mexicos over-quota and quota-free export will
be dampened because of Mexicos comparative disadvantage to the rest of the world,
while the export from the rest of the world remains over 1.2 million MT over the entire
forecast horizon.
By contrast, a change in the U.S. governments policy on stabilization of the
domestic price does not pose much impact on sugar exports to the U.S. market. With the
same Mexican sweetener market situations (high production-high HFCS adoption), the
U.S. policy options of price support, buying up excess sugar in the market, and
production controls are compared (Figures 5-4, 5-6 and 5-7). The results indicate that all
three scenarios bring about the similar trends in Mexican sugar export. Yet, U.S.
production control with Mexico causes an overall increase in quota-free export after
2008. This is because the Mexican sugar price is maintained lower relative to the price
support or buying up excess sugar scenarios (Figures 5-9, 5-10, and 5-11). The prices of
sugar in both countries will converge in the integrated market without a U.S. price
support (Figure 5-10) while the difference in prices are kept after 2008 with production
control (Figure 5-11): the price difference corresponds to the transportation cost from
Mexico to the United States.
Mexicos tax on HFCS brings about very different results. In this scenario,
Mexico is unable not only to export either under-quota, over-quota or quota-free but also


90
Figure 7-3. Feeding scars on water hyacinth (Eichhornia crassipes) leaf after exposure
to Neochetina eichhorniae adults. Black marks represent feeding scars
marked with a fine tip marker to aid in counting (other side counted but not
shown).


73
Figure 6-2. Weed Systems, Inc. KQ 3L CO2 backpack back sprayer used for application
of L-methionine and Silwett L-77 solutions. Boom consisted of three
nozzles (middle top and end of each arm). In total, 3L were applied per
treatment every two weeks from 09 July to 31 August 2001.


71
Each row contained the 4 treatment plots of 10 plants (control (0% L-methionine), 0.1%
L-methionine, 0.5% L-methionine and 1.0% L-methionine in deionized water solutions)
in a Latin square design. Plants within treatment plots were spaced 3 feet apart while
treatment plots were 9 feet apart. Figure 6-1 shows the diagrammatic representation of
the field plot.
Plant Yield
Before beginning the experiment, all developing eggplants were removed from
the plants in an effort to standardize the treatments and ensure all eggplant development
occurred after the exposure of methionine. Treatments were administered using a KQ 3L
CO2 (Weed Systems, Inc.; Hawthorne, FL) backpack sprayer charged to 30 lbs PSI and a
3-nozzle boom to ensure complete coverage of the plant (Figure 6-2). Each treatment
consisted of a 3L application over the 4 representative groups. The adjuvant Silwett
L-77 (0.5% concentration) was included to improve the residual effect of the
methionine under the field conditions. Plants were sprayed a total of nine times at
approximately two-week intervals. Fruits were harvested at various times during the
study and were weighed in the field using a Tokyo Electronics hand-held digital scale.
Pest Introduction
Neonate CPB larvae were reared on excised eggplant leaves for two days at 27C,
60% relative humidity and 16L/8D photoperiod in FRJUs to ensure healthy individuals
for the test. Larvae were transferred to the field plants using a camel hairbrush and the
branch marked with flagging tape. Introduction was made after the last spray treatment
in November. Ten larvae were placed on each plant for a total sample size of 1,600
individuals. Plants were inspected for the next 5 days and larvae encountered noted.


59
and supply are held at Baseline level. The duration of harvest is held constant because
it is partly affected by the weather. Also, the assumption where Mexicos HFCS adoption
and tax on HFCS occur at the same time is not considered since the scenario becomes
incompatible to simulate.
The scenarios associated with HFCS adoption are consistent with an increasing
trend since 1994 when NAFTA went into effect and speculation of expansion of HFCS
production in Mexico. If Mexico continues to increase HFCS consumption, it may follow
a similar path as was seen in the United States in early 1980s when soft-drink
manufacturers decided to switch to HFCS from sugar. HFCS manufacturers, who are
mostly the U.S. corporations, see this phenomenon as a business opportunity in Mexico.
With HFCS capital-intensive facilities, existing HFCS plants in Mexico are operated by
the firms based in the United States. The HFCS Adoption case assumes that HFCS will
be adopted in a linear fashion until HFCS consumption is 50 percent of total indirect
sweetener consumption in 2008 and its share remains constant for the rest of the forecast
horizon. The 50 percent share of indirect consumption of sweetener is equivalent to about
27 percent of total consumption of sweeteners. In 2001 (base year), share of HFCS in
indirect sweetener consumption and total sweetener consumption were 25.3 percent and
11.6 percent, respectively. Comparisons of HFCS and indirect sugar consumption
forecast for Baseline scenario, HFCS adoption, and Tax on HFCS situations are
illustrated in Figure 4.2.
Assumptions related to U.S. sugar policy levers are summarized in Table 4-3. Two
kinds of policy levers are considered: one is to stabilize the demand price and the other is
to allocate quotas to exporters. For the former, two possible sugar policies are used as


CHAPTER 3
CONCEPTUAL AND THEORETICAL FRAMEWORK
In this chapter the conceptual and theoretical framework employed to analyze trade
between the United States and Mexico in the sugar market is discussed. In the conceptual
framework, factors that influence the market and the trade system are specified and
linked. Based on the conceptual framework, theoretical foundations are established in
two parts: an analysis of the sugar market for each country and an analysis of the market
balance in the U.S.-Mexico bilateral sugar trade system.
Conceptual Framework
The sugar market, one of the oldest and most common agricultural commodity
markets, is built upon sugarcane and sugar beet production and the resulting production
of processed sugar from these raw materials. Being an essential commodity for a daily
diet, sugar has been traded across borders for a long period of time. More recently, there
have been major changes to the trading pattern due to the emergence of an alternative in
the market. High fructose com syrup (HFCS), now the most widely adopted sugar
substitute, expanded the sugar market into a sweetener market. This is particularly true in
the case of the United States, where HFCS now occupies nearly half of the sweetener
market, and is becoming the case in Mexico as a result of the two markets becoming
more closely linked by the North American Trade Agreement (NAFTA). Understanding
this linkage between the United States and Mexico holds important clues to analyze
economic and political impacts in the sweetener markets. To illustrate this linkage, the
32


57
Figure 5-2. Mortality of yellow fever mosquito larvae exposed to various
concentrations of L-methionine (nTOtai=240). Data were
adjusted using Abbotts formula for control mortality.


CHAPTER 7
EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF
SELECTED NONTARGET SPECIES
Introduction
A biorational pesticide is defined as one that is effective against pest species but
innocuous to non-target organisms and not disruptive to biological control agents and
beneficial species (Stansly et al. 1996). To test L-methionine as a potential pesticide and
determine if it could be considered biorational, it was necessary to examine the effects of
this compound on selected nontarget species that could possibly come into contact with
it, either directly while on the plant or indirectly via incidental contact or as a host that
has come into direct contact with this compound. The species chosen reflect a variety of
non-target organisms, mainly those that were shown to be important in controlling some
pest species. The pink spotted ladybird beetle, Coleomegilla maculata (DeGeer), the
mottled water hyacinth weevil, Neochetina eichhorniae Warner, and the greenbug
parasitoid, Lysiphlebus testaceipes (Cresson) all are beneficial insects that have been
effective against pests in the state of Florida and also are common and readily available.
Each species also represents a different feeding guild (predator, herbivore and parasitoid,
respectively) to ensure a thorough examination of the possible effects of methionine as it
might be encountered in under natural conditions.
The pink spotted ladybird beetle (PSLB) is an abundant polyphagus species that is
known to feed on many lepidopteran and coleopteran pests, including the Colorado
potato beetle, in which it was responsible for over 50% of the predation on eggs and early
82


LCS0 (% L-methionine Concentration)
45
Figure 4-2. Concentrations (%) of L-methionine concentrations required for
the mortality of 50% of the test population of Colorado potato
beetle after 8 days exposure (nrotai=220). Number range
following value is the 95% confidence limits. Determination of
LC50 was performed using PROBIT Version 1.5 (Ecological
Monitoring Research Division, USEPA), including Abbotts


31
e World raw sugar price U.S rawsigir pnce, city fee paid
-k U.S spot price for HFCS42, Midwest markets A Lhit value of HFCSexpoted to Nfexico
Figure 2-12. Prices of Sugar and HFCS, 1994-2000
Source: USD A, 2001a


Table 5-6. Pay-off Matrix for the Trade Policy Game [Billion US$]
Mexico's strategies
Maintain the current policy
(status quo)
U.S.
HFCS
U.S.
sugar
U.S.
adjusted
welfare
Mexican
sugar
Mexico
welfare
High production
U.S.
HFCS
U.S.
sugar
U.S.
adjusted
welfare
Mexican
sugar
Mexico
welfare
High HFCS adoption
U.S.
HFCS
U.S.
sugar
U.S.
adjusted
welfare
Mexican
sugar
Mexico
welfare
High production -
High HFCS adoption
U.S.
HFCS
U.S.
sugar
U.S.
adjusted
welfare
Mexican
sugar
Mexico
welfare
Maintain price
support
(status quo)
5.68
35.71
354.37
21.74
74.44
5.68
35.56
353.90
21.87
76.24
10.13
35.31
353.97
21.322
65.726
10.13
35.37
354.08
21.31
72.29
u.s.
strategies
Buying up
excess sugar
in the market
5.68
35.40
354.79
22.15
69.77
5.68
35.40
354.73
21.87
76.24
10.13
35.09
354.89
21.355
65.739
10.13
35.21
354.85
21.31
72.29
Production
control with
Mexico
5.68
35.40
354.92
22.15
69.77
5.68
35.39
354.97
23.14
71.49
10.13
35.09
355.12
21.355
65.739
10.13
33.93
355.44
20.52
66.76
Mexico's strategies
Maintain the current policy
(status quo)
Tax on HFCS
U.S.
HFCS
U.S.
sugar
u.s.
adjusted
welfare
Mexican
sugar
Mexicos
welfare
U.S.
HFCS
U.S.
sugar
u.s.
adjusted
welfare
Mexican
sugar
Mexico's
welfare
U.S.
strategy
Maintain
price support
(status quo)
5.68
35.71
354.37
21.74
74.44
1.59
36.50
354.48
25.85
79.10
I11


I dedicate this work to Karen, my wife and best friend. I thank her for putting up with
living as a graduate student for the last 5 years in fulfillment of my childhood dream of
being a Doctor. She has been my pillar of support, and I would not have made it this
far without her love and understanding.


132
Comit de la Agroindustria Azucarera (COAAZUCAR). Acciones de Cosecha de
Caa que Reflejan Calidad Zafra 2000/2001. Internet site:
http://www.sagarpa.gob.mx/Coaazucar/menu4/ult_estim_prod.htm (Accessed May
2003e).
Comit de la Agroindustria Azucarera (COAAZUCAR). Resultados Econmicos
del Campo Caeros del Campo de las Zafras 1987/2002. Internet site:
http://www.sagarpa.gob.mx/Coaazucar/menu3/indexl.htm (Accessed May 2003f).
Comit de la Agroindustria Azucarera (COAAZUCAR). Relacin Histrica de los
Precios de la Caa de Azcar como Promedio Nacional. Internet site:
http://www.sagarpa.gob.mx/Coaazucar/menu7/com_precio.htm (Accessed May 2003g).
Congressional Research Service, Library of Congress, Agriculture: A Glossary of
Terms, Programs, and Laws, 2nd Edition, Washington, 1999.
Com Refiners Association. Food and Industrial Com Use 1980 to Present.
Internet site: http://www.com.org/web/foodseed.htm (accessed May 27, 2004).
Diario Oficial de la Federacin. Mexico, DF. Secretara de Medio Ambiente y
Recursos Naturales. March 26, 1997.
Farm Foundation. Trade Dispute in an Unsettled Industry: Mexican Sugar.
Internet site: http://farmfoundation.org/flags/schwedel.pdf (Accessed August 2003).
Food and Agriculture Organization of the United Nations Statistics (FAO). Internet
site: http://apps.fao.org/page/collections?subset=agriculture (Accessed February 2003).
Garcia Chaves, Luis R., Thomas H. Spreen and Gretchen Greene. Structural
Reform and Implications for Mexicos Sweetener Market. In Schmitz, Andrew, Thomas
H. Spreen, William A. Messina, Jr., and Charles B. Moss, Sugar and Related Sweetener
Markets: International Perspectives (81-100). New York: GABI Publishing, 2002.
Garcia Chaves, Luis R., Gretchen Greene, Thomas H. Spreen, Daisuke Sano and
Chris O. Andrew. Transitions in the Mexican Sugar Industry: An Analysis of the
Production and Marketing System. Lake Alfred: Florida Science Source, 2004.
General Accounting Office (GAO). Sugar Program: Supporting Prices Has
Increased Users Costs While Benefiting Producers. RCED-00-126, Washington DC,
2000.
Greene, Gretchen. Transitions in the Mexican Sugar Industry. Ph.D. dissertation.
University of Florida, 1998.
Haley, S. and N.R. Suarez. Sugar and Sweetener Outlook. Washington, DC:
USDA, January 2003.


CHAPTER 8
SUMMARY AND DISCUSSION
The creation and implementation of Integrated Pest Management (IPM) strategies
to combat pest species were developed as a response to the economic losses associated
with the overuse of chemical control. However. IPM strategies are not widely used
because of the lack of alternatives and the ease of use of pesticides. This has resulted in
the resistance to pesticides in many insect species, including economic and medical pests.
In an effort to provide alternatives to traditional chemical control, biorational methods
have been investigated and one such avenue is the use of non-protein amino acids.
Chapter 2 covered the history of the use of non-protein amino acids as a pesticide,
and discussed the CAATCH1 system and the safety of L-methionine. Only a handful of
these amino acids have been investigated as a means of controlling insect pests but still
lack the practicality and cost effectiveness as current chemical control methods. Recent
discovery of a new midgut membrane protein, CAATCH1, has revealed a new possibility
in insect control. The CAATCH1 system works in alkaline conditions and responds to
different amino acids, mainly the reduction in ion flow after exposure to methionine, an
essential amino acid required for normal development and metabolism of many species
including humans. The use of a compound such as methionine would be an excellent
addition to the IPM arsenal because of its relative safety to vertebrates and warrants
further study as a pesticide.
Chapters 3,4, and 5 were dedicated to examining the effects of L-methionine, a
common analog of methionine, on three different economic and medically important
96


106
Fogarty International Center and the U.S. National Institutes of Health (FIC-NIH). 2003.
Multilateral Initiative on Malaria. U.S. National Institutes of Health. Internet
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Forgash, AJ. 1985. Insecticide resistance in the Colorado potato beetle, pp. 33-52. IN
D.N. Ferro and R.H. Voss (eds.) Proceedings of the Symposium on Colorado
Potato Beetle. XVII International Congress of Entomology, Massachusetts
Agricultural Experiment Station Bulletin 704. Amherst Massachusetts.
Friend, W.G., R.H. Backs and L.M. Cass. 1957. Studies on amino acid requirements of
larvae of the onion maggot, Hylema antiqua (MG.), under aseptic conditions.
Can. J. Zool. 35: 535-543.
Gasnier-Fauchet, F. and P. Nardon. 1986a. Comparison of sarcosine and methionine
sulfoxide levels in symbiotic and aposymbiotic larvae of two sibling species,
Sitophilus oryzae and Sitophilus zeamais (Coleptera: Curculionidae). Insect
Biochemistry 17(1): 17-20.
Gasnier-Fauchet, F. and P. Nardon. 1986b. Comparison of methionine metabolism in
symbiotic and aposymbiotic larvae of Sitophilus oryzae L. (Coloeptera:
Curculionidae)- n. Involvement of the symbiotic bacteria in the oxidation of
methionine. Comp. Biochem. Physiol. 58(1): 251-254.
Gauthier, V.L., R.N. Hoffinaster and M. Semel. 1981. History of Colorado potato beetle
control, pp. 13-34. IN J.H. Cashcomb and R. Casagrande (eds.), Advances in
Potato Pest Management. Hutchinson and Ross, Stroudsburg, PA. 672pp.
Geer, B.W. 1966. Utilization of D-amino acids for growth by Drosophila melanogaster
larvae. J. Nutr. 90: 31-39.
Giordana, B, M. Forcella, M.G. Leonardi, M. Casartelli, L. Fiandra, G.M. Hanozet and P.
Parenti. 2002. A novel regulatory mechanism for amino acid absorption in
lepidopteran larval midgut. J. Insect Physiol. 48: 585-592.
Giovanelli, J, S.H. Mudd and A.H. Datko. 1980. Sulfur amino acids in plants. IN: B.J.
Mifhn (ed.) The Biochemistry of Plants VoL 5, Academic Press, New York, pp.
453-505.
Giroux, S., R.M. Duchesne and D. Coderre. 1995. Predation of Leptinotarsa
decemlineata (Coleptera: Coccinellidae) by Coleomegilla maculata (Coleptera:
Coccinellidae): Comparative effectiveness of predator developmental stages and
effect of temperature. Environ. Entomol. 24: 748-754.
Glare, T.R, and M. O Callaghan, 1998. Environmental and Health Impacts of Bacillus
thuringiensis isrealensis. Report for the New Zealand Ministry of Health, 58pp.


o
Os
Figure 5-19. Relative Gain and Loss of Pay-offs to the Mexican Sugar Industry and Welfare Caused by Production Improvement and
HFCS Adoption


15
production (Sang and King 1961). Lack of methionine in the diet of the female may also
explain the transfer of methionine in the ejaculate of the male during fertilization
(Bownes and Partridge 1987). Methionine plays another role in insect biochemistry,
especially in juvenile hormone biosynthesis, inhibitory allatostatins, and storage proteins
known as hexamerins. Audsley et al. (1999) found that in vitro rates of juvenile hormone
synthesis in females of the tomato moth (Mamestra olercea (L.) (Lepidoptera:
Noctuidae)) were dependent on the concentration of methionine present in the incubation
medium. Tobe and Clarke (1985) found a direct relationship between methionine
concentration and juvenile hormone biosynthesis in the cockroach, Diploptera punctata
(Eschscholtz) (Blattodea: Blaberidae), further supporting the idea that methionine plays
an important role in insect biochemistry.
Storage proteins, or hexamerins, act as a storehouse for amino acids that can be
sequestered for later use in the developmental cycle (Pan and Telfer 1996). Many
Lepidoptera have been identified with hexamerins containing high concentrations of
methionine and are metabolized during the last larval stage, and presumably used for egg
production (Wheeler et al. 2000).
Methionine as a potential pesticide has not been overlooked entirely. Tzeng
(1988) tested a methionine and riboflavin mixture and found it successful in controlling
various pests, including the larvae of Culex spp. (Dptera: Culicidae). The mode of
action for this mixture was attributed to a photodynamic reaction and the production of
oxygen rich radicals (Tzeng et al. 1990). Their research led to the use of this methionine
compound as a control agent for sooty mold of strawberry (Tzeng and Devay 1989;
Tzeng et al. 1990) but not as an insecticide.


99
target ingests the methionine first as it feeds ensuring the overload CAATCH1 system
and eventually death.
As for the stored methionine, it is released from the storage proteins as needed to
synthesize juvenile hormone and allow for transformation in addition to other functions.
The remaining methionine is then used for protein synthesis in the tissues around the
ovaries to boost yolk production, as seen in the transfer of methionine from male to
female Drosophila species (Bownes and Partridge 1987). In the THW, the presence of
hexamerins with high methionine content may be an alternative to the male contribution
possibly found in its ejaculate. Methionine-rich hexamerins are common in Lepidoptera
and have been shown to provide the larvae a source of amino acids during the synthesis
of these proteins during the last stage of larval development (Wheeler et al. 2000). In
addition to the need for methionine for metabolism and reproduction, the release of
methionine may also in part account for the decrease in ion transport of the posterior
region of the midgut during larval molts and the wandering stage present before pupation.
Currently, little is known regarding the mechanisms involved with the decrease of ion
transport during these developmental stages (Lee et al. 1998). Clearly there appears to be
more to the role methionine plays in the development of some insects other than the
vague designation of essential amino acid.
Insects have evolved to deal with limiting resources, such as methionine, and have
successfully found effective strategies like hexamerin storage or alternate pathways to
deal with such problems. No attempt to link together all the aspects of the role of
methionine in a whole organism or system context. It appears that methionine actually
may play a role far more important than that of just an essential amino acid. From the


LIST OF TABLES
Table page
2-1 Cane Sugar Production in Selected Countries, 1997-2000 Average 25
2-2 World Sugar Consumption in 2000 27
2-3 Quota and Tariff Schedule Imposed on Mexican Sugar Exported to the U.S 27
4-1 Assumptions for Baseline (Status quo) Scenario 66
4-2 Assumptions for Mexican Sweetener Market Situations 67
4-3 Assumptions for U.S. Sugar Policies 69
4-4 Listing of Examined Scenarios 70
4-5 Strategies for the Sugar Trading Game 71
4-6 Data Sources for U.S. Demand 72
4-7 Data Sources for Mexican Demand 72
4-8 Data Sources for U.S. Supply 73
4-9 Data Sources for Mexico Supply 73
4-10 Data Sources for Miscellaneous 74
5-1 Summary of the U.S. Supply-Demand Analysis 91
5-2 Summary of the Mexican Supply-Demand Analysis 92
5-3 Impact of Changes in Mexican Sweetener Market on Pay-offs to the Industries and
Nations Welfare [Billion US$] 101
5-4 Impact of Changes in U.S. Price Stabilization Policy on Pay-offs to the Industries
and Nations Welfare [Billion US$] Flexible Quota Allocations 102
5-5 Impact of Changes in U.S. Quota Allocation Policy on Pay-offs to the Industries
and Nations Welfare [Billion US$] Minimum Quota Allocations 103
vii


70
surfactant that has wetting and spreading properties (Helena Chemicals 2002) and was
found to be compatible with solutions of L-methionine.
The objectives for this portion of the study were to examine the effects of a
methionine and Silwet L-77 mixture on a crop plant (eggplant) in terms of yield (both
fruit weight and total yield) and to evaluate this mixture as an insecticide under natural
conditions.
Materials and Methods
Preliminary Investigation of Silwet L-77 and L-methionine
Adult CPBs were obtained from the University of Florida Horticultural Unit,
Gainesville and held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a hardware
cloth (to facilitate cleaning) and held at 27C, 60% relative humidity and 16L/8D
photoperiod in FRIUs. Twenty-four adults were exposed used in each of the 5
treatments, with 4 replicates per treatment (nTOtai=120). Adults were used because of the
lack of sufficient numbers of larvae to test. Excised leaves were dipped in solutions of
deionized H2O containing different concentrations of methionine and Silwett L-77
(0.5% concentration), 0.1% L-methionine, 0.5% L-methionine, 1.0% L-methionine and
controls of deionized H2O and deionized H2O +Silwet L-77. The additional control
was to determine the possible insecticidal properties of Silwet L-77 alone and to make
sure the addition of this adjuvant did not affect mortality or deter feeding.
Plot Design
Eggplants {Solarium melongena L.,Classic variety) were grown and maintained
at the University of Florida Horticultural Unit, Gainesville, from 18 June to 04 November
2001. Eight, one hundred ft. rows of plants were used for this study, with two rows on
each side consisting of buffer rows and four rows in the middle used for the experiments.


TABLE OF CONTENTS
Elge
ACKNOWLEDGMENTS iii
LIST OF FIGURES viii
ABSTRACT xi
CHAPTERS
1 THE INTEGRATED PEST MANAGEMENT DILEMMA: ARE
CONVENTIONAL PESTICIDES THE ONLY ANSWER? 1
Introduction 1
Importance of IPM in Florida and Surrounding States 2
Problems Associated with Pesticide Misuse 4
Biorational Compounds- An Alternative to Chemical Pesticides 5
2 HISTORY OF THE USE OF AMINO ACIDS AS A MEANS TO CONTROL
INSECT PESTS 7
Non-Protein Amino Acids 7
Essential Amino Acids 10
The Cation-Anion Modulated Amino Acid Transporter with Channel
Properties (CAATCH1) System 9
Methionine 13
Research Objectives 16
3 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT
OF THE TOBACCO HORNWORM, Manduca sexta, UNDER
LABORATORY CONDITIONS 17
Introduction 17
Materials and Methods 18
Diets and Survivorship 18
Feeding and Development 20
Preference Tests 22
Data Analysis 24
Results 24
v


Ill
Rock, G.C., A. Khan, and E Hodgson. 1975. The nutritional value of seven D-amino
acids and a-keto acids for Argyrotaenia velutinana, Heliothis zea and Phormia
regina. J. Insect Physiol. 21:693-703.
Romoser, W.S. and J. G Stoffolano, Jr. 1998. The Science of Entomology, Sedition.
McGraw-Hill. Newyork, 605pp.
Rosen, D., F.D. Bennett, and J.L Capinera. 1996. Preface, pp. V-vi. IN D. Rosen, F.D.
Bennett and J.L. Capinaera (eds.), Pest Management in the Subtropics: Biological
Control- a Florida Perspective. Intercept Limited, Andover, UK. 737pp.
Rosenthal, G. A. 1977. The biological effects and mode of action of L-canavanine, a
structural analogue of L-arginine. Q. Rev. Biol. 52(2): 155-178.
Rosenthal, G.A. and D.L. Dahlman. 1975. Non-protein amino acid-insect interactions.
II. Effects of canaline-urea cycle amino acids growth and development of the
tobacco homworm, Manduca sexta (L.) (Sphingidae). Comp. Biochem. Physiol.
52:105-108.
Rosenthal, G.A. and D.L. Dahlman. 1988. Degradation of aberrant proteins by larval
tobacco homworm, Manduca sexta (L) (Sphingidae). Arch. Insect Biochem
Physiol. 8: 165-172.
Rosenthal, G. A. and D.L. Dahlman. 1991. Incorporation of L-canavanine into proteins
and the expression of its antimetabolic effects. J. Ag. and Food Chon. 39:987-
990.
Rosenthal, G.A., D.L. Dahlman, P.A. Crooks. S.N. Phuket, and L.S. Trifonov. 1995.
Insecticidal properties of some derivatives of L-canavanie. J. Agrie. Food Chem.
43:2728-2734.
Rosenthal, G.A., D.L. Dahlman and D.H. Janzen. 1976. A novel means for dealing with
L-canavanine, a toxic metabolite. Science 192: 256-258.
Rosenthal, G.A., D.L. Dahlman and D.H. Janzen. 1977. Degradation and detoxification
of canavanine by a specialized seed predator. Science 196:658-660.
Rosenthal, G.A., D.L. Dahlman and D.H. Janzen. 1978. L-canaline detoxification: A
seed predators biochemical defense. Science 202: 528-529.
Rosenthal, G.A., P. Nkomo and D.L. Dahlman. 1998. Effect of long-chained esters on
the insecticidal properties of L-canavanine. J. Agrie. Food Chem. 46(1): 296-299.
Royer, T.A., K.L. Giles, S.D. Kindier and N.C. Elliott. 2001. Developmental response
of three geographic isolates of Lysiphlebus testaceipes (Hymenoptera: Aphididae)
to temperature. Environ. Entomol. 30(4): 637-641.


Table 5-9. Indexed Pay-off Matrix for the Trade Policy Game Played by the Industry Coalition and the Government Coalition without


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81
Overall, it appears that L-methionine can be used in a natural setting to control
CPB larvae without affecting crop production. The adjuvant Silwett L-77 worked well
with L-methionine in controlling CPB larvae but not the adults. The lack of effectiveness
on the adults may be attributed to their ability to stop feeding and living off of reserves
acquired during the larval stage until suitable food sources can be found. It is unknown if
L-methionine, alone or in combination with Silwett L-77 adversely affects fecundity of
the adults.


I certify that I have read this study and that in my opinion it conforms to acceptable
standards of scholarly presentation and is fully adequate, in scope and quality, as a
dissertation for the degree of Doctor of Philosophy.
Thomas H. Spreen, Chair
Professor of Food and Resource Economics
I certify that I have read this study and that in my opinion it conforms to acceptable
standards of scholarly presentation and is fully adequate, in scope and quality, as a
dissertation for the degree of Doctor of Philosophy.
A.
Lisa A. House, Cochair
Associate Professor of Food and Resource
Economics
fc-S,
I certify that I have read this study and that in my opinion it conforms to acceptable
standards of scholarly presentation and is fully adequate, in scope and quality, as a
dissertation for the degree of Doctor of Philosophy.
(Lk'iZy O
Chris O. Andrew
Professor of Food and Resource Economics
I certify that I have read this study and that in my opinion it conforms to acceptable
standards of scholarly presentation and is fully adequate, in scope and quality, as a
dissertation for the degree of Doctor of Philosophy.
~~rriMLh
Terry L. McCoy
Professor of Political Science
I certify that I have read this study and that in my opinion it conforms to acceptable
standards of scholarly presentation and is fully adequate, in scope and quality, as a
dissertation for the degree of Doctor of Philosophy.
Kenneth L. Buhr
Assistant Professor of Agronomy


44
O 1
3 4 5
Days of Exposure
Figure 4-1. Mortality of Colorado potato beetle larvae exposed to excised eggplant
leaves treated with various concentrations of L-methionine (nTOtai=560).
Proline (1.0%) and Bit were included for comparison as positive and
negative controls. Data were adjusted using Abbotts formula for
control mortality.


120
the inelastic supply price elasticities in both countries imply that sugar production is not
responsive to price changes. Contrasted with the possible improvement in U.S. sugar
industry and welfare, Mexico does not benefit from changes in U.S. price stabilization
policy.
Alternative Sugar Policy by the United States
An alternative U.S. sugar policy to the current policy of price supports is sought
from the aggregate simulation results using game theory. Cooperative games among
industries and governments (the U.S. HFCS industry, the U.S. sugar industry, the
Mexican sugar industry, cost-adjusted U.S. welfare, and Mexicos welfare) are also
examined by assuming that both nations welfare is transferable and that total pay-offs
are redistributed among coalition members.
As expected, the Mexican government and sugar industry will always and
harmoniously choose a high production strategy and avoid any strategies that involve
HFCS adoption. By comparison, the U.S. sugar industry does not agree with the choice
made by the U.S. government: the U.S. sugar industry prefers price support to alternative
policies. When coalitions are formed, one case for country coalitions (United States vs.
Mexico) and the other for the govemment/industry coalitions (governments vs.
industries), the games reach different solutions: the former game results in U.S. sugar
production control strategy and the latter game results in U.S. government follows the
strategy of buying up excess sugar. This contrast indicates that the U.S. government
prefers to form a coalition with its own industry while the Mexican government prefers to
form a coalition with the U.S. government. For the Mexican government, Mexicos
welfare improves by having U.S. cooperation at the expense of its own sugar industry.
This implies that Mexico faces a choice either of improving the nations well-being or of


58
financial stress, vividly illustrated by the mill expropriation by the government in 2001,
and producing and exporting surplus sugar to the rest of the world at the same time.
Simulated Scenarios
Scenarios are formulated by considering alternative assumptions related to
Mexicos sweetener market situations affected by continued gains in the productivity of
the sugar industry, HFCS consumption, as well as a policy lever and U.S. sugar policy
levers. Each scenario carries a combination of Mexicos sweetener market situation and
U.S. sugar policy. To compare the impacts of scenarios, a baseline scenario is defined
where status quo is maintained (Table 4-1). In the baseline scenario, it is assumed that
shifts in sugar demand and supply in both countries continues at the average rates
observed in recent years and that the U.S. government maintains price support and
allocates quota among exporters in a flexible manner, abiding by the WTO minimum
import requirement.2
Assumptions related to Mexicos sweetener market situations are summarized in
Table 4-2. Four situations are proposed: higher sugar production, higher HFCS adoption,
a combination of both and introduction of tax on HFCS as a Mexicos alternative policy
lever. The rates of increase in production and HFCS adoption are defined relative to the
baseline. Impacts of Mexicos tax on HFCS is based on the forecast by Haley and Suarez
(2003) where Mexican HFCS consumption drops significantly in 2002 and 2003 due to a
tax imposed on beverages that contain HFCS. The impact of this tax is assumed
significant considering soft-drink manufacturers currently account for about one-third of
domestic sugar consumption in Mexico (Buzzanell, 2002). In all situations, U.S. demand
2 Current allocation system is based on historical trade shares (Skully, 1998).


4-3. Mean head capsule widths of Colorado potato beetle larvae exposed to excised
eggplant leaves treated with various concentrations of L-methionine
(niotai=320) 46
4-4. Total leaf area consumed by Colorado potato beetle larvae exposed to excised
eggplant leaves treated with various concentrations of L-methionine
(nrotar=320) 48
4-5. Mean leaf consumption by Colorado potato beetle in the preference tests 49
5-1. Bioassay setup for yellow fever mosquito larvae exposed to various concentrations
of amino acids and Bti 55
5-2. Mortality of yellow fever mosquito larvae exposed to various concentrations of
L-methionine (nTOtai=240) 57
5-3. Mortality of yellow fever mosquito larvae exposed to various concentrations of
D-methionine (nrotai=240) 58
5-4. Mortality of yellow fever mosquito larvae exposed to various concentrations of Tris-
buffered L-methionine (nrotai~240) 60
5-5. Mortality of YFM larvae exposed to various concentrations of Proline (nrotai=240) 61
5-6. Mortality of yellow fever mosquito larvae exposed to various concentrations of
L-leucine (nTotai=240) 62
5-7. Mortality of YFM larvae exposed to various concentrations of Beta-alanine
(nTotai=240) 63
5-8. Mean head capsule widths of yellow fever mosquito larvae exposed to various Tris
buffered (7.0 pH) concentrations of L-methionine (niotai=320) 64
5-9. Concentrations (%) resulting in 50% mortality (LC50) of yellow fever mosquito
larvae exposed to various amino acids after 10 days (nrotar=240 for each amino
acid) 65
6-1. Overview of the design layout used to study the effects of L-methionine and Silwett
L-77 solutions on yield of eggplant 72
6-2. Weed Systems, Inc. KQ 3L CO2 backpack back sprayer used for application of
L-methionine and Silwett L-77 solutions 73
6-3. Mortality of Colorado potato beetle adults exposed to excised eggplant leaves treated
with L-methionine and the adjuvant Silwett L-77 (nTotai=120) 75
6-4. Effects of L-methionine and Silwett L-77 on eggplant yield (A) and mean weight
in grams of fruit (B) from 09 June to 31 August 2001 76
IX


40
an excellent candidate for the evaluation of L-methionine as a possible means of
controlling this devastating pest.
Because little information is available on the insecticidal properties of
L-methionine, several baseline experiments were necessary to determine what
concentrations of this amino acid to test. Therefore, it was necessary to test L-methionine
and CPB interaction in a variety of ways including survivorship of both larvae and adults,
development of larvae when exposed to different concentrations of the amino acid, and
preference tests. The purpose of this portion of this study was to conduct bioassays to
determine if exposure to L-methionine was detrimental to the survival and development
of the CPB and to determine if L-methionine could be used to control this species.
Materials and Methods
Eggs of CPB were obtained under UDSA permit from the insectary of the New
Jersey Department of Agriculture and held in 26.4L x 19.2W x 9.5H (cm) clear plastic
boxes with a hardware cloth (to facilitate cleaning) and held at 27 C, 60% relative
humidity and 16L/8D photoperiod in FRIUs (Figure 3-1). Excised eggplant leafs were
placed in the chambers with the neonates and they were allowed to feed for 2 days after
eclosin before being transferred to experiments. A camel hair brush was used for
transferring the neonates to minimize the risk of damaging the larvae.
Survivorship
Larvae and adults of the CPB were tested in preliminary experiments with the
highest concentration (1.0% L-methionine (wt/wt)) observed in tests done on the THW in
the previous chapter. The diet for the larvae and adults consisted of excised eggplant
leaves (,Solarium melongena L.,Classic variety (Family: Solanaceae)) from plants
grown and maintained at the University of Florida, Department of Entomology and


Table 5-7. Indexed Pay-off Matrix for the Trade Policy Game without Coalitions [Baseline 100]
Mexico's strategies
Maintain the current policy
(status quo)
High production
High HFCS adoption
High production -
High HFCS adoption
U.S.
HFCS
U.S.
sugar
u.s.
adjusted
welfare
Mexican
sugar
Mexico
welfare
U.S.
HFCS
u.s.
sugar
U.S.
adjusted
welfare
Mexican
sugar
Mexico
welfare
U.S.
HFCS
U.S.
sugar
U.S.
adjusted
welfare
Mexican
sugar
Mexico
welfare
U.S.
HFCS
U.S.
sugar
U.S.
adjusted
welfare
Mexican
sugar
Mexico
welfare
u.s.
strategies
Maintain
price support
(status quo)
100
100
100
100
100
100
99.57
99.87
100.61
102.42
178.27
98.86
99.89
98.09
XXXXXXXXXXX"
XXXXXXXXXXXX
.XxxxxxXXXXX-
88.29
XxxxxxXNXXV
XXXXXXXXXXXX
XXXXXXXWXXX
178.27
99.03
99.92
98.05
97.11
Buying up
excess sugar
in the market
100
99.13
100.12
101.89
XXXXXXXXXXXXXX
CvXVWWWWXV
100
99.13
100.10
100.61
102.42
178.27
98.26
100.15
98.24
xx-xxxxxxxxx
wvvwww
VWVWWWW
kWWWWWV
vvvxvvvvx
11
JxxxxCxxxvxx'
178.27
98.58
100.14
98.06
97.11
Production
control with
Mexico
100
99.13
100.15
101.89
93.73
100
99.10
100.17
106.46
96.03
178.27
98.26
100.21
98.24
CxxxxxxXVXxXx
88.3
XxxxxxXXXXXX
AWXXVVW
XX.xxxXVXXXX
178.27
95.00
100.30
XXXXXXXXXXXXXXXXX
94.38
89.68


B-4 Exports of Products Made from Com in 2002
127
xi


67
Table 4-2. Assumptions for Mexican Sweetener Market Situations
Situations
(code)
Shifters
Assumptions
High
Mexican
Production
(P)
Production cost,
Downtime,
Sugar loss
Will improve an additional 1 percent to the
Baseline.
High HFCS
Adoption
(A)
HFCS
consumption
It is assumed that HFCS consumption will increase in
a linear fashion until it replaces 50 percentage of
indirect consumption of sugar in 2008. After 2008, its
share remains at 50 percent.
High
Mexican
Poduction -
High HFCS
adoption
(PA)
Production cost,
Downtime,
Sugar loss
The same as High Mexican Production
HFCS
consumption
The same as HFCS Adoption
Tax on
HFCS
(T)
HFCS
consumption
It is assumed that HFCS consumption for 2002
through 2004 will drop due to tax (Haley and Suarez,
2003) and remain at reduced consumption level (7
percent to the indirect consumption of sugar) for the
rest of the forecast horizon.3
Other shifters are the same as the Baseline.
3 A 20-percent tax on beverages that contain HFCS was introduced on January 1, 2002; suspended on
March 5 by the presidents decision; and then reimposed on July 16, 2002 with the decision by Mexicos
Supreme Court of Justice (USDA, 2002a).


9
proteins (produced by the assimilation of L-canavanine) into newly synthesized proteins;
the proteases involved do not efficiently degrade enough to prevent some damage from
occurring in the insect (Rosenthal and Dahlman 1986; 1988).
Surprisingly, L-canavanine also was shown to increase the effectiveness of
Bacillus thuringiensis in vivo by altering membrane properties, mainly gut permeability,
and active transport in the midgut of the THW (Felton and Dahlman 1984). However,
despite the possible synergistic relationship between the relatively safe Bt product and
this amino acid, no further research has been conducted to evaluate the combination for
future commercial use.
Other species of insects have also been tested for susceptibility to canavanine with
a variety of results. Larvae of Drosophilia melanogaster Meigen (Dptera:
Drosophilidae) showed no deleterious response to lower concentrations of canavanine,
but showed mortality increased at concentrations over 1,000 ppm (Harrison and Holiday
1967). Lower concentrations also were ineffective against adult Pseudosarcophaga
affinis (Fallen) (Dptera: Calliphoridae), with no effect on oocyte development (Hegdekar
1970). Dahlman et al. (1979) examined four species of muscoid flies and found greater
than 70% mortality at the higher concentration (800 ppm) and decreased pupal weights as
concentrations of canavanine increased.
Despite the toxicity of canavanine to some insects, others have evolved
detoxifying mechanisms to deal with high concentrations of this compound. Rosenthal et
al. (1978) attributed the detoxification of canavanine in the bruchid Caryedes brasiliensis
Thunberg (Coleptera: Bruchidae) to the beetles ability to convert canavanine to
canaline, another toxic amino acid. Canaline is metabolized through reductive
deamination to homoserine and ammonia, with the overall result being the detoxification


5-4 U.S. Sugar Import Forecast (Scenario 4 PA-S-F) 94
5-5 U.S. Sugar Import Forecast (Scenario 13 PA-S-M) 95
5-6 U.S. Sugar Import Forecast (Scenario 8 PA-B-F) 95
5-7 U.S. Sugar Import Forecast (Scenario 12 PA-C-F) 96
5-8 U.S. Sugar Import Forecast (Scenario 16 T-S-F) 96
5-9 Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 4
PA-S-F) 97
5-10 Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 8
PA-B-F) 97
5-11 Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 12
PA-C-F) 98
5-12 Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 16
T-S-F) 98
5-13 Forecasted Sugar Demand and Supply for both the United States and Mexico
(Scenario 1 Baseline) 99
5-14 Forecasted Sugar Demand and Supply for both the United States and Mexico
(Scenario 4 P-S-F) 99
5-15 Forecasted Equilibrium Sugar Prices in the United States and Mexico (Scenario 14
PA-B-M) 100
5-16 Forecasted Equilibrium Sugar Prices in the United States and Mexico (Scenario 15
PA-C-M) 100
5-17 Absolute Effects of Production Improvement and HFCS Adoption on Pay-off to the
Mexican Sugar Industry 104
5-18 Absolute Effects of Production Improvement and HFCS Adoption on Pay-off to
Mexicos Welfare 105
5-19 Relative Gain and Loss of Pay-offs to the Mexican Sugar Industry and Welfare
Caused by Production Improvement and HFCS Adoption 106
B-l Recent Com Production and Consumption for Selected Countries 126
B-2 Food and Industrial Com Use in the U.S., 1980-2002 126
B-3 Com Price (No.2 Yellow) in Chicago Market, 1981-1998 127
x


27
Table 2-2. World Sugar Consumption in 2000
Country/ region
National total
consumption2
Per capita
consumption2
Sweetener15
(1,000 MT)
Sugar
(1,000 MT)
Sweetener15
(kg)
Sugar
(kg)
India
26,234
18,101
25.59
17.66
United States
21,221
9,371
74.22
32.77
European Union (15 countries)
18,960
14,370
50.26
38.10
China
11,805
11,028
9.13
8.53
USSR, Former Area of
11,771
11,407
40.57
39.32
Brazil
10,007
9,620
57.99
55.75
Mexico
5,100
4,476
50.81
44.60
Japan
3,741
2,327
29.38
18.27
Philippines
2,135
1,975
27.68
25.61
Thailand
1,937
1,924
30.47
30.26
Canada
1,297
1,128
41.83
36.37
Australia
1,060
922
54.81
47.67
Cuba
711
710
63.24
63.18
World
168,632
133,401
27.49
21.75
a: Production + net import + change in stocks
b: Sugar (raw equivalent) plus other kinds of sweeteners
Source: FAO, 2003
Table 2-3. Quota and Tariff Schedule Imposed on Mexican Sugar Exported to the U.S.
Year
U.S. Import Quota (MT)
Over-Quota Tariff
(raw cane, cents/pound)
Mexico as a net surplus
producer
Mexico NOT as a net
surplus producer
1994
25,000
7,258
16.00 (Base)
1995
25,000
7,258
15.20
1996
25,000
7,258
14.80
1997
25,000
7,258
14.40
1998
25,000
7,258
14.00
1999
25,000
7,258
13.60
2000
250,000
7,258
12.09
2001
250,000
7,258
10.58
2002
250,000
7,258
9.97
2003
250,000
7,258
7.56
2004
250,000
7,258
6.04
2005
250,000
7,258
4.53
2006
250,000
7,258
3.02
2007
250,000
7,258
1.51
2008 and beyond
CO
0.00
Source: USD A, 1999.


129
EScost= (AQs/ACOST)*(COST/Qs). (C.5)
With supply price elasticity (E^), equation [C.4] is expressed as
e = (-if cost /E?P) *( pS/COST). (C.6)
Final forms of inverse linear function for sugar demand and supply for the U.S.
market derived from corresponding equations [4.1] and [4.5] are expressed as:
Pus, t = IUj + IU2*QDus, + IU3*GDP US,, + IU4*POP us,, (C.7a)
or Pus, t = IUj + IU2*Qus t + ShifterD us,, (C.7b)
Psus,, = IUU, + IUU2*QSus,, + IUU3*COSTus, + IUU4*RCVt. (C.8a)
or Psus,, = IUU, + IUU2*QSus,, + Shifted us,, (C.8b)
Note that the inverse linear demand function is re-specified on an annual base
rather than quarterly in order to balance the market with the supply equation and that the
inverse linear supply function is re-specified with the current price without lagged
quantity for simplicity. Coefficients for corresponding variables are presented in the
Table C-l.
Table C-l. Coefficients for
nverse Linear Functions -U.S.-
Inverse linear demand function
Inverse linear supply function
iu2
(l/EuP.usY{PUus/QUus.)
IUU2
(1/E?p.us)*(P*us/&us)
IU3
(-EUgdp, us/EUp, us)*
(PD us/GDP us)
IUU3
(-E?cost, us/E?p,us)*
(PS us/COSTus)
IU4
{-Eupop, us/Eup us)*
(P us/POP us)
IUU4
{-Efpcv/Efp, us)*(P\¡s/RCV)
In the case of Mexico, the equation for annual total sugar is expressed with
estimates of direct consumption of sugar as preliminary estimation showed statistically
insignificant estimates associated with price variables in total and indirect consumption
of sugar. By assuming that indirect consumption demand is totally inelastic (i.e., a
vertical demand curve in quantity-price space), total sugar demand is expressed with a


93
Figure 5-2. U.S. Sugar Import Forecast (Scenario 2 P-S-F)


40
compared to sugar from Mexico: the first sugar that enters into the United States over
quota must be from Mexico. The transfer cost of sugar from the rest of the world to the
United States includes the price of sugar, i.e. sugar from Mexico and the rest of the world
compete with each other to enter the U.S. market: the one with lower transfer cost enters
the market first. The last term in the equation [3.23] considers Mexicos sales of sugar to
the rest of the world. Equation [3.23] is rewritten in quadratic form as follows:
Max (1/2)* IU2*(Qus f + (fU¡ + Shifter0us)*Qus
+ (1/2)* IM2*(QTCmx f + (IM, + Shifter0mx)*Qtcmx
- (1/2)* IUU2*(QSus f + (IUUi + Shifter*US)*QSus
- (1/2)* IMM2*(Qsmx )2 + (IMMi + Shifter3mx)*Qsmx
(Tmx, us)*XmX, us
(Tus, MX )*X us, MX
(Tmx, us + OQTarMx, us ) *XXmX us
(Trow,us + Prow)*Xrow,us
+ Prow *Xmx, row (3.24)
Constraints are defined to balance the quantities shipped with the quantities
demanded as well as supplied; to impose a quota on imported sugar from Mexico; and to
impose a quota on imported sugar from the rest of the world, which is equivalent to the
U.S. minimum sugar import quota required under WTO agreements:
Qus X us, us XMX us XX mx, us X row, us ^ 0 (3.25)
QTCMX -Xmx.mx X us, mx ^ 0 (3.26)
- QSus + X us, us + X us, mx ^ 0 (3.27)
s
- Q MX + Xmx, MX + XMX, US + XXMX, US + XMX, ROW ^ 0
(3.28)


135
U.S. Department of Agriculture, Foreign Agricultural Service. New National
Sugar Policy. GAIN report #MX2031, Washington, DC, February 2002b.
U.S. Department of Agriculture, Economic Research Service. Sugar and Sweetener
Situation and Outlook Yearbook. Washington, DC, various issues, 2003a.
U.S. Department of Agriculture, Foreign Agricultural Service. Sugar: World
Market and Trade. Internet site: http://www.fas.usda.gov/htp/sugar/2003/may (Accessed
July 2003b).
U.S. Department of Agriculture, Economic Research Service. Feed Outlook
Rreport. Washington, DC, 2004.
U.S. Department of Commerce, U.S. Census Bureau. Internet site:
http://www.census.gov/ipc/www/idbsprd.html (Accessed February 2003).
U.S. Department of Labor, Bureau of Labor Statistics. Consumer Price Index.
Internet site: http://www.bls.gov/cpi/home.htm (Accessed November 2003).
Varan, Hal R. Microeconomic Analysis, 3rd edition. New York: W. W. Norton &
Company, 1992.


124
Mexico
United States
1995
Inflation rate hit 51% while the price
of sugar increased 25.7%.
1996
The government announced increases
in import duties on HFCS-42, HFCS-
55, and crystalline fructose to 12.5,
above the then-current rate of 10.5
percent. (December).
1996 farm bill
1997
SECOFI initiated anti-dumping
investigation (February) and imposed
temporary tariff on two grades of
HFCS (June).
SECIFI published the formula to
determine the wholesale price of
standard sugar (March).
1998

CRA (Com Refiners Association)
asked for the review of Mexicos anti
dumping actions under chapter of
NAFTA.
WTO established panel for Mexicos HFCS dumping case (January).
1999
2000
WTO panel ruled against Mexicos dumping case (January).
Vicente Fox was elected as president,
ending 71 years of authoritarian one-
party rule in Mexico (December).
2001
WTO Appellate Body turned down the
Mexicos appeal of HFCS dumping case.
Mexican government expropriated 27
sugar mills (September).
2002
A 20-percent tax on beverages that
contain HFCS was introduced on
January; suspended on March 5; and
reimposed on July 16.
National Sugar Policy for 2002 -
2006 (February)
Source: Created from Polopolus and Alvarez, 1991; Greene, 1998; Garcia Chaves et al.,
2002; Buzzanell, 2002; and various issues by USDA.


CHAPTER 2
SUGAR INDUSTRIES AND SWEETENER MARKETS IN THE UNITED STATES
AND MEXICO
Sugar, one of the basic commodities with a long history of utilization, is traded in
mature markets in many parts of the world with established business practices and
networks. The recent trend towards freer markets in the international trade area has not
left the industry unchanged. The sugar industries in the United States and Mexico are not
exceptions. They have experienced more changes in the face of this recent trend towards
rapid trade liberalization. In fact these two industries have become more economically
inseparable than ever before as the sweetener markets in the United States and Mexico
have been integrated under North American Free Trade Agreement (NAFTA). In this
chapter, sugar industries in both the United States and Mexico are examined in the
context of the sweetener market as well as the integrated market, paying close attention to
historical and political perspectives.
First, the Mexican sugar industry and sweetener market are introduced with
fundamental characteristics of the structure and government involvement. The status of
Mexico as a sugar exporter is also presented in conjunction with Mexicos relation to the
U.S. market under the provisions of NAFTA. Next, the development and adoption of
high fructose com syrup (HFCS) is presented. Lastly, the U.S. sugar industry and
sweetener market is introduced with emphasis on the current political environment
surrounding that market.
8


60
Days of Exposure
Figure 5-4. Mortality of yellow fever mosquito larvae exposed to various
concentrations of Tris-buffered L-methionine (nxotai=240). Data
were adjusted using Abbotts formula for control mortality.
Note the longer exposure because of the bioassay involving
neonates instead of 3rd instars. Note the overlap in some of the
trend lines on Day 1 with the 0.3% L-methionine and 0.5% L-
methionine treatments.


17
Development and Adoption of High Fructose Corn Syrup
Sweeteners are generally classified into two categories, caloric and non-caloric,
common sweeteners in the former group are sucrose, invert sugar, lactose, maltose, and
sorbitol; and aspartame and saccharine in the latter. Sucrose is found in various forms of
sugar such as raw sugar, granulated sugar and brown sugar derived form sugarcane or
sugar beets, or in honey and maple sugar. Invert sugar such as dextrose, glucose, fructose,
and HFCS are made form starch through chemical processes. HFCS is produced by
converting a portion of naturally occurring glucose in starch into fructose through a com
wet milling process (Congressional Research Service, 1999). Lactose, maltose and
sorbitol are found naturally in certain kinds of food and give food a sweet taste. Non
caloric sweeteners, sometimes called artificial sweeteners, such as aspartame, are often
used for special dietary purposes.
Commercially produced and rapidly adopted since the early 1970s in the United
States, HFCS became an important player in the sweetener market among sugar
substitutes (Figure 2-6). Production of HFCS has increased from 51,000 MT in 1970 to
nearly 8.7 million MT in 2001 (Figure 2-7). HFCS production expanded during the 1980s
as a substitute for sugar used in the soft-drinks. Today, about 75 percent of total HFCS
and 90 percent of HFCS-55 (55 percent fructose) supplied in the United States are
consumed in soft-drink market (Buzzanell, 2002; Congressional Research Service, 1999).
HFCS-42 (42 percent fructose), which is roughly 90 percent as sweet as sugar, is used
mainly in beverages (44 percent), processed food products (21 percent), and other
products including cereal and bakery products (Buzzanell, 2002; Congressional Research
Service, 1999). As a result, HFCS and two other corn-derived sweeteners, glucose syrup
and dextrose, accounted for 55 percent of total U.S. caloric sweetener use in recent years


23
Figure 3-3. Chambers used for tobacco homworm and Colorado potato beetle
preference tests. Two treatments (control and 1.0% L-
methionine) were used to determine if any larvae exhibited any
preference or avoidance to L-methionine. Treatments were
alternated in the chamber and neonates were released in the center
of the dish and allowed to search for food. The filter paper in the
bottom of the dish was moistened to prevent desiccation of the
leaf disks and the test specimens.


107
Griffin, M.L. and K.V. Yeargan. 2002. Oviposition site selection by the spotted lady
beetle Coleomegilla maclala (Coleptera: Coccinellidae): Choices among plant
species. Environ. Entomol. 31(1): 107-111.
Groden, E., F.A. Drummond, R.A. Casagrande and D.H. Haynes. 1990. Coleomegilla
maculata (Coleptera: Coccinellidae): Its predation upon the Colorado potato
beetle (Coleptera: Chrysomelidae) and its incidence in potatoes and surrounding
crops. J. Econ. Entomol. 83:1306-1315.
Gubler, DJ. 1998. Resurgent vector-borne diseases as a global health problem. Emerg.
Infect. Dis. 4(3): 442-450.
Gubler, D.J. and G.G. Clark. 1995. Dengue/Dengue Hemorrhagic Fever: The emergence
of a global health problem. Emerg. Infect. Dis. 1(2): 55-57.
Haag, K.H. and D.G. Boucias. 1991. Infectivity of insect pathogens against Neochetina
eichhorniae, a biological control agent of water hyacinth. Florida Ent. 74(1): 128-
133.
Haag, K.H. and D.H. Habeck. 1991. Enhanced biological control of water hyacinth
following limited herbicide use. Aquat. Plant Management 29: 24-28.
Harrison, J. and R. Holliday. 1967. Senescence and the fidelity of protein synthesis in
Drosophila. Nature 214: 990-993.
Hazzard, R.V., D.N. Ferro, R.G. van Driesche and A.F. Tuttle. 1991. Mortality of eggs of
Colorado potato beetle (Coleptera: Chrysomelidae) from predation of
Coleomegilla maculata (Coleptera: Coccinellidae). Environ. Entomol. 20: 841
848.
Hegdekar, B.M. 1970. Amino acid analogues as inhibitors of insect reproduction. J.
Econ. Entomol. 63: 1950-1956.
Heim, D.C., G.G. Kennedy and J.W. Van Duyn. 1990. Survey of insecticide resistance
among North Carolina Colorado potato beetle (Coleptera: Chrysomelidae)
populations. J. Econ. Entomol. 83(4): 1229-1235.
Helena Chemicals, Inc. 2002. Silwett L-77. Technical Data Sheet SL77080596.
Helena Chemicals, Inc. Internet URL: http://www.helenachemicalwest.
com/data/TDS/Silwett.pdf. Accessed April 2004
Hilbeck, A. and G.G. Kennedy. 1996. Predators feeding on Colorado potato beetle
insecticide-free plots and insecticide-treated commercial potato fields in eastern
North Carolina. Biol. Control. 6: 273-282.
Hoffmann, M.P. and A.C. Frodsham. 1993. Natural Enemies of Vegetable Insect Pests.
Cooperative Extension, Cornell University, Ithaca, NY. 63pp.


115
Insects. Lewis, along with fellow graduate student Jim Dunford, were awarded the
Outstanding Teacher Award by the Entomology and Nematology Student Organization of
the University of Florida for outstanding teaching accomplishments in the department.
While at the University of Florida, Lewis joined the U.S. Army Reserve as a
medical entomologist. He was assigned to the local Medical Detachments, and served
there from 2000 to 2004. Originally he had planned on graduating in 2003, but was
called to active duty with the 1469th Medical Detachment as a part of Operation Enduring
Freedom (OEF). Lewis was the OEF Theater entomologist, and served as the Executive
Officer (responsible for the deployment of personnel and equipment to South West Asia).
He was stationed at Kandahar Airfield, where he performed his duty and was awarded an
Army Commendation Medal for his work in protecting soldiers from health hazards and
diseases associated with the area. Lewis returned and continued his work toward
graduation.
Lewis was married in August 1992 to Karen Abbott, and is the father of Emilia
Irene (1994) and Bryan Scott (1997). Lewis plans on having a career in the military as a
medical entomologist, and all look forward to seeing the world and the rest of their
future.


36
Discussion
The initial studies involving the high concentrations of L-methionine (i.e., 3.0-10.0%,
which are outside the range normally encountered in nature) showed that a concentration
of 1.0% L-methionine was sufficient enough to provide good control of THW larvae
reared on both artificial and natural diets. The 0.1%L-methionine concentration
remained similar to that of the control for developmental and feeding trials (Figure 3-9),
indicating a level of methionine that can be tolerated to some extent, as seen in the low
mortality of this treatment. This is in stark contrast to the mortality seen in the excised
leaf trials in which the same concentration had over 60% mortality (Figure 3-6). One
possible explanation could be the amount of L-methionine present on the leaf disk being
low enough and ingested at a slower rate than that of the whole leaf, which was left in the
chamber with the larvae until the leaf was either completely consumed or too wilted for
the larvae to ingest.
The preference tests did show some preference towards control leaf disks over the
1.0%L-methionine treated disks as seen in the correlation analysis of the diet consumed
and the mean head capsule width of the larvae. Despite the lack of a statistical difference
between the amount consumed, the larvae could have fed on the treated disks and then
switched to the control disks based on a physiological cue. It is unclear if THW larvae
possess specialized sensory structures to detect amino acids like those found in other
Lepidoptera (Beck and Henee 1958; Dethier and Kuch 1971; Schoonhoven 1972), but the
possible switch from the methionine rich treatment to the control leaf disks does indicate
some sort of mechanism for detection. Del Campo and Renwick (2000) found THW
larvae were induced to feeding on plants outside of their normal diet when the plants


CHAPTER 4
EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE
COLORADO POTATO BEETLE, Leptinotarsa decemlineata, UNDER LABORATORY
CONDITIONS
Introduction
Leptinotarsa decemlineta (Say) (Coleptera: Chrysomelidae), the Colorado
potato beetle (CPB), is considered an economic pest throughout North America. The
larvae and adults of the CPB feed on a wide variety of solanaceous crop plants and are
responsible for $150 million in losses and control related costs (Durham 2000). To
further complicate matters, the CPB is resistant to numerous pesticides, including various
pyrethroids and carbamates (Bills et al. 2004). Historically, CPB management relied
heavily on chemical control methods that led to the development of resistance to different
pesticides in several areas of the eastern United States (Forgash 1985; Gauthier et al.
1981). Control of CPB without the use of chemicals is further complicated given the
species ability to develop resistance and the limitations on the use of resistant varieties of
potato (Ragsdale and Radcliffe 1999). The use of plant varieties that are resistant to CPB
and other pests also run the risk of developing tolerance to chemical pesticides in other
pest species (Sorenson et al. 1989). Despite the success of Bacillus thuringiensis-
tenebrionis (Btt) and the biocontrol agents Podisus maculiventris Say (Hemiptera:
Pentatomidae) and Edovum puttleri Grissel (Hymenoptera: Eulophidae), more biorational
alternatives are necessary for controlling CPB to prevent yet another devastating threat to
the potato industry because of this insects ability develop resistance and overcome
control methods (Boucher 1999; Ferro 1985; Tipping et al. 1999). This makes the CPB
39


I certify that I have read this study and that in my opinion it conforms to
acceptable standards of scholarly presentation and is fully adequate, in scope and quality,
as a dissertation for the degree of Doctor of Philosophy.
Associate Professor of Entomology and
Nematology
This dissertation was submitted to the Graduate Faculty of the College of
Agricultural and Life Sciences and to the Graduate School and was accepted as partial
fulfillment of the requirements for the degree of Doctor of Philosophy.
May 2004
Dean, College of Agricultural
Sciences
Dean, Graduate School


28
Figure 3-6. Mortality of tobacco homworm larvae exposed to various concentrations of
L-methionine (nTotai= 1,540) on excised eggplant leaves. Data were
adjusted using Abbotts formula for control mortality. Note the overlap in
trend lines for the 3.0% L-methionine-10.0% L-methionine concentrations
after Day 1.


99
Figure 5-13. Forecasted Sugar Demand and Supply for both the United States and
Mexico (Scenario 1 Baseline)
Figure 5-14. Forecasted Sugar Demand and Supply for both the United States and
Mexico (Scenario 4 P-S-F)


104
Leptinotarsa decemlineata (Coleptera: Chrysomelidae). Annals Ent. Soc Am.
60(3): 626-631.
Cook, R.J., W.L. Bruckart, J.R. Coulson, M.S. Goettel, R.A. Humber, R.D. Lumsden,
J.V. Maddox, M.L. McManus, L. Moore, S.F. Meyer, P.C. Quimby, Jr., J.P.
Stack, and J.L. Vaughn. 1996. Safety of microorganisms intended for pest and
plant disease control: A framework for scientific evaluation. Biological Control
7: 333-351.
Dadd, R.H. 1975. Alkalinity within die midgut of mosquito larvae with alkaline-active
digestive enzymes. J. Insect Physiol. 21:1847-1853.
Dadd, R.H. and D.L. Krieger. 1968. Dietary amino acid requirements of the aphid,
Myzus persicae. J. Insect Physiol. 14: 741-774.
Dahlman, D.L. 1980. Field tests ofL-canavanine for control of tobacco homworm. J.
Econ. Entomol, 73:279-281.
Dahlman, D.L., F. Herald and F.W. Knapp. 1979. L-canavanine effects on growth and
development of four species of Muscidae. J. Econ. Entomol. 72: 678-679.
Dahlman, D.L. and G.A. Rosenthal. 1975. Non-protein amino acid-insect alterations (1)
Growth effects and symptomology of L-canavanine consumption by tobacco
homworm, Manduca sexta (L). Comp. Biochem Physiol. 51: 33-36.
Dahlman, D.L. and G A. Rosenthal. 1976. Further studies on the effect of L-canavanine
on the tobacco homworm, Manduca sexta. Insect Physiol. 22:265-271.
Dahlman, D.L. and G.A. Rosenthal. 1982. Potentiation of L-canavanine-induced
developmental anomalies in the tobacco homworm, Manduca sexta, by some
amino acids. J. Insect Physiol. 28(10): 829-833.
Deedat,Y.D. 1994. Problems associated with the use of pesticides: An overview. Insect
Sci. Applic. 15(3): 247-251.
Del Campo, M., and J.A.A. Renwick. 2000. Induction of host specificity in larvae of
Manduca sexta: Chemical dependence controlling host recognition and
developmental rate. Chemecol. 10: 115-121.
Dethier, V.G. and J.H. Kuch. 1971. Electrophysiological studies of gustation in
lepidopterous larvae 1. Comparative sensitivity to sugars, amino acids and
glycosides. Z. Vergl. Phys. 72: 343-363.
Dietary Supplement Information Bureau. 2000. Methionine Dietary Supplement
Education Alliance. Internet URL: http://www.supplementinfo.org/index.htm
Accessed April 2004.


53
requirements of methionine in the amounts of 0.0007mg/ml for the YFM. This amino
acid also is considered essential for other species of mosquito in untraceable (in those
studies) amounts (Chen, 1958; Singh and Brown, 1957). Given the high alkalinity found
in the midgut of the YFM as well as other mosquito species, this physiological condition
indicates the possibility of the presence of the CAATCH1 system in larval mosquitoes
(Dadd, 1975).
The purpose of this portion of the study was to examine the survival and
development of YFM larvae exposed to water treated with excess L-methionine (adults
were not tested given the feeding nature). In addition to L-methionine, other amino acids
were tested in an effort to see if their response (i.e., survivorship) was similar CAATCH1
responses to methionine found by Feldman et al. (2000).
Materials and Methods
Bioassav
The bioassay experiments consisted of six treatments (control, 0.1%, 0.3%, 0.5%,
0.7% and 1.0%) each with four replicates. Both L-methionine and D-methionine were
tested along with proline, Beta-alanine and L-leucine to examine the other amino acids
that were found to be reactive to the CAATCH-1 system (Feldman et al., 2000).
Bt-isrealiensis (Aquabac @ a rate of 2.3 mL/m2; Biocontrol Network, Brentwood, TN)
and proline also were included in some trials of L-methionine to allow for comparison of
both positive and negative effects. Amino acids were weighed using a Denver
Instruments Co. XD2-2KD digital scale and added to glass quart jars containing 500ml of
deionized FLO. Concentrations were based on the proportion of lg/100ml for a 1%
solution and for corresponding concentrations. Solutions were allowed to sit at room


5
consumption was derived from domestically-produced sugar before 1994. Reflecting this
threatening trend of replacing domestic sugar consumption with HFCS, in 1996, the
Mexican government imposed tariffs on HFCS claiming that U.S. companies were
dumping HFCS at an unfair price and affecting the export volume and value of Mexican
sugar. This action evolved into a trade dispute between the United States and Mexico and
ended when the WTO panel ruled against Mexicos claim (Garcia Chaves et al., 2002 and
2004). Overall, NAFTA has not brought about significant changes in the U.S. sugar
market because the Mexican exporters have been unable to significantly expand
shipments to the United States. Rather, attention was poured into issue of HFCS and its
immediate impact on the Mexicos sweetener market.
In this study, the direction of U.S.-Mexico sugar trade is examined using
quantitative methods, with close attention to issues related to NAFTA and HFCS
adoption in Mexico. Demand and supply analyses in both countries and a bilateral trade
model using mathematical programming provide insights for the market balance in the
future including political implications. Aggregated results from various simulations on
the trade model are examined using a game theory analysis to investigate possible policy
recommendations through assessing gainers and losers in sugar trade.
Problem Statement
The future outlook for the U.S.-Mexico sweetener market needs to be
quantitatively analyzed in a manner that includes influential factors such as trade
agreements under NAFTA; trends in HFCS consumption in Mexico; and other related
economic and political issues in the sweetener markets.
Researchable Questions
The study attempts to answer the following set of questions.


100
Year
-U.S. price
- Mexico price
Figure 5-15. Forecasted Equilibrium Sugar Prices in the United States and Mexico
(Scenario 14 PA-B-M)
Year
U.S. price
-A Mexico price
Figure 5-16. Forecasted Equilibrium Sugar Prices in the United States and Mexico
(Scenario 15 PA-C-M)


134
Secretara de Agricultura, Ganadera, Desarrollo Rural, Pesca y Alimentacin
(SAGARPA), Sistema Integral de Informacin Agroalimentaria y Pesquera (SIAP).
Avance Comparativo Siembras y Cosechas: Perennes: Situacin al 31 de Mayo 2002 y
2003. Internet site: http://www.siap.sagarpa.gob.mx/ar_comdeagr.html (Accessed July
2003).
Skully, David W. Auctioning Tariff Quotas for U.S. sugar Imports. Special
article presented in Sugar and Sweetener SSS-223, Economic Research Service, USDA,
Washington, DC, May 1998.
Spreen, Thomas H., Mechel Paggi, Anouk Flambert, and Waldir Fernandes, Jr..
An Analysis of the EU Banana Trade Regime. Selected poster presented at the
American Agriculture Economics Association meeting, Tampa, FL, August 2000.
Electronic abstract at http://agecon.lib.umn.edU/aasa.html/#aaeaOO.
Takayama, T and G. G. Judge. Equilibrium Among Spatially Separated Markets:
A Reformulation. Econometrica 32: 510-24, 1964.
U.S. Department of Agriculture, Economic Research Service. Sugar Statistical
Compendium Stock #91006. Washington, DC, October 1991.
U.S. Department of Agriculture, Economic Research Service. Com Sweetener
Statistics Stock #94002. Washington, DC, September 1993.
U.S. Department of Agriculture, Economic Research Service. Agricultural
Outlook. Washington, DC, March 1997.
U.S. Department of Agriculture, Economic Research Service. Agricultural
Outlook. Washington, DC, September 1999.
U.S. Department of Agriculture, Economic Research Service. Sugar and Sweetener
Situation and Outlook Yearbook. Washington, DC, various issues, 2001a.
U.S. Department of Agriculture, Foreign Agricultural Service. Mexico
Expropriated 27 Sugar Mills. GAIN report #MX1161, Washington, DC, September
2001b.
U.S. Department of Agriculture, Foreign Agricultural Service. The North
American Trade Agreement. Internet site:
http://www.fas.usda.gov/info/factsheets/nafta.html (Accessed July 2001c).
U.S. Department of Agriculture, Economic Research Service. Sugar and Sweetener
Situation and Outlook Yearbook. Washington, DC, various issues, 2002a.


66
hours, and to 0.11% for 48-168 hours and remained constant since the trial lasted longer
because of the use of neonates instead of 3rd instars. The D-methionine treatments were
similar with 0.44% for 24 and 48 hours, 0.33% for 72 hours and 0.32% after 168 hours.
While not as striking as the others, Beta-alanine had a LC50 concentration of 1.1% after
24 hours, dropped to 0.5% after 48 hours and leveled off around at 0.35% after 72 and
168 hours. Probit analysis of the Proline and L-leucine treatments was not performed, as
the mortality associated with those treatments was too low (Figures 5-5 and 5-6).
Discussion
Although not commonly encountered, the D- form of methionine had virtually the
same effect as the L- form on larval mosquito mortality. The D-and L-methionine trials
showed that the D- form had lower mortality associated with it than the more reactive
L-counterpart. Insects do not commonly use the D- form of amino acids, although
D-methionine is metabolized by some orders to a limited extent (Ito and Inokuchi, 1981).
The YFM could be an example of this phenomenon.
Because of the nature of the CAATCH1 system in the alkaline midgut, buffering
may have acted to increase the effectiveness of the system. Buffering the solutions did
result in an increase in mortality, with even the lowest concentration of 0.1%
L-methionine exhibiting a two-fold increase with the buffered form (Figure 5-4).
Complete mortality was reached sooner with the buffered forms even for concentrations
that did not reach 100% in the unbuffered form. In a field setting, the addition of
L-methionine would be buffered naturally by the chemical properties of the bodies of
water to which it was applied and similar results would be expected.


CHAPTER 6
CONCLUSIONS AND IMPLICATIONS FOR POLICY
Conclusions and Implications for Policy
Sugar is a basic commodity with a long history of utilization and trade between
regions where excess demand and excess supply exists. This trade in the sugar market has
provided a classic example for study in the field of agricultural economics. Although its
economic value has diminished, as seen in the trend of the declining world prices and a
smaller percentage of income spent on sugar in developed countries, its bargaining power
continues as a powerful force in the political arena today. Emergence of a new substitute
commodity, high fructose com syrup (HFCS) produced predominantly by the United
States, has brought about changes in the climate of the sweetener markets, particularly
those tied to U.S. markets. These changes are expected to continue. Recent trends in the
international trade environment to move towards freer and borderless trade have also
accelerated changes in the market climate for sugar trade.
When examining sugar trade issues between the United States and Mexico,
Mexicos adoption of HFCS and the provisions of North American Free Trade
Agreement (NAFTA) related to sugar and HFCS play critical roles for shaping the
sweetener market balance. HFCS, already with a large share in the U.S. sweetener
market, has gained sizeable market power, as well as power in the political arena. The
HFCS industry supports the U.S. sugar program under the umbrella of the American
Sugar Alliance (ASA). Despite the efforts of critics, the U.S. sugar program remains
intact due largely to successful lobbying efforts by ASA. On the other hand, the growth
115


87
U.S. sugar industry will lobby against a production control strategy for fear that it may
lose up to five percent (high production-high adoption market situation). On the contrary,
the Mexican governments choice of strategy will be accepted by the industry in most
cases, except for the case of where the U.S. government buys sugar in the baseline
scenario. Since HFCS adoption harms both welfare and the industry, the Mexican
government will continue to struggle to suppress HFCS adoption in its market.
Results from the game played by two coalitions of countries (the United States and
Mexico) are shown in Table 5-8. In this game, the U.S. HFCS industry is excluded from
the game. As expected, the high production strategy becomes Mexicos pure strategy and
thus the game is solved when the U.S. government chooses a strategy: production control
policy (cells shaded with vertical stripes). This combination of strategies is coincidentally
the best choice for the U.S. coalition, but not for the Mexican coalition. The Mexican
government would prefer the U.S. government to choose either the price support or
buying up excess sugar strategies.
Results from the game played by the sugar industry coalition and the government
coalition are shown in Table 5-9. The high production strategy becomes a pure strategy
for Mexico when a decision is made by the government coalition. The solution of the
game is thus determined when the U.S. government chooses the strategy of buying up
excess sugar. This result differs from the game played by the country coalitions
mentioned above. By a government cooperating with the other government rather than
with the industry in its own country, the Mexican government improves its pay-off by US
$4.75 billion; the U.S. government loses by US $0.24 billion; and the Mexican sugar
industry loses US $1.27 billion (Table 5-4). Since the loss bom by the U.S. government is


89
government production control strategy for country coalitions (Table 5-11), Mexican
government high production strategy and the U.S. government buying up excess sugar
strategy for govemment/industry coalitions, and Mexican govemmet high production
strategy and the U.S. government buying up excess sugar strategy for grand coalition.
Yet, pooled gains become larger due to inclusion of gains from the U.S. HFCS industry,
the pay-off matrices offer additional factors that need to be taken into the policy
assessment process.
Results from the game played by country coalitions are shown in Table 5-11.
Although the solution is the same as the one without the U.S. HFCS industry included,
pay-offs expected from Mexican governments other strategies become more attractive to
the U.S. coalition. It is obvious that the U.S. coalition would receive a better pay-off if
Mexico chooses the HFCS adoption strategy. This fact gives the U.S. coalition a strong
incentive to influence Mexican governments choice of strategy.
Intensified conflict of interests between the industry coalition and the government
coalition is illustrated in Table 5-12. Without the HFCS industry in a coalition, the
industry coalition prefers the U.S. production control strategy to the buying up excess
sugar strategy for the sake of slight gains. When gains from the HFCS industry are
pooled, the industry coalition will lobby for strategies that involve Mexicos HFCS
adoption, no matter which strategy the U.S. government plays. This is possible only if the
industry coalition promises to compensate the Mexican sugar industry for the loss. If
redistribution of gains among industries is feasible, the Mexican sugar industry may
prefer being compensated to expecting protection from the Mexican governments
strategy. Ultimately, if the Mexican government chooses to overlook the impact on


44
The complementary slackness conditions indicate that if Mexico exports sugar to
*
the U.S. under-quota (XMx, us > 0), then the demand price in the United States should not
exceed the value of Mexican exporting sugar, which is equivalent to the sum of the
supply price, transportation cost and the marginal value of exporting sugar under-quota
(equation [3.45]). By the same token, if Mexico exports sugar to the United States over
quota (XXmx, us> 0), then the demand price in the United States should not exceed the
sum of the Mexican supply price, transportation cost, tariff imposed on over-quota sugar
and the marginal value of exporting sugar under-quota (equation [3.47]). The inequality
of the prices expressed in equation [3.49] accords with reality. If both over-quota export
from Mexico (XXMx, us) and import from the rest of the world (XMx, us ) are greater than
zero, the following must also hold from equations [3.47] and [3.48].
P5mx-Prow = row, us ¡mx, us -OQTarMx.us (3.50)
This equation implies that when the price difference between Mexico and the rest
of the world is equal to the difference in transfer cost (transportation cost and tariff), both
over-quota export from Mexico and import from the rest of the world occur at the same
time. In other words, Mexico would export over-quota only if the transportation cost
from the rest of the world is high enough to justify Mexico to do so.


Figure 3-1. Conceptual Framework for the Analysis in this Study


8
L-canavanine exhibits a range of insecticidal effects in artificial diets when
exposed to the THW. Dahlman (1977) demonstrated a reduction in consumption of
artificial diet containing less than 1% canavanine (w/v) which resulted in a lower body
mass and increased developmental time to the adult stage. Fecundity and fertility also
was affected by L-canavanine. Rosenthal and Dahlman (1975) showed that
concentrations as low as 0.5 mM L-canavanine in the diets of the THW resulted in the
reduction of ovarial mass of adults, while Palumbo and Dahlman (1978) showed that
concentrations of L-canavanine in agar-based diets resulted in the reduction of
chorionated oocyte production in concentrations between 1.0 and 2.0 mM.
Under natural conditions, L-canavanine was found to retard development, and
increased the susceptibility of exposed larvae to biotic and abiotic mortality factors
(Dahlman 1980). However, field applications of L-canavanine were shown to be
impractical because of the expense involved in synthesizing L-canavanine from its
source, the jack bean (Canavalia ensiformis (L.) DC. (Family: Fabaceae)).
Other sources of L-canavanine (i.e., analogues and homologues) were sought in
an attempt to find a more practical source of the amino acid. Structural homologues of
canavanine were examined and found to contribute to pupal deformities (and to a lesser
degree, to mortality) (Rosenthal et al. 1998). Long-chain esters of L-canavanine were
found to be more toxic than the parent compound when injected or added to an artificial
diet exposed to last instar of THW specimens (Rosenthal et al. 1998). Adding amino
acids other than arginine (the parent compound to L-canavanine) to diets containing L-
canavanine increased deformities and mortality of THW larvae and was attributed to the
structure and position of the functional groups on the added compounds (Dahlman and
Rosenthal 1982). Although the THW has an effective means of degrading aberrant


127
(NroTt-i^'or-ooC'O r^m^tw^vor^oo
OOOOOOOOOOOOOOOOOOONC'C'C'C''CSC'CVC'
C'C'C'O'O'CT'C'C'C'ONONO'ONONC'O'C'O'
Year
Figure B-3. Com Price (No.2 Yellow) in Chicago Market, 1981-1998
Source: USDA, 2004
Corn gluten feed
Com gluten meal
Corn oil, crude
Corn oil, fully refined
Modified starches derived from corn starch
Glucose syrup not containing fructose or containing in the dry
state less than 20% fructose
Corn meal
Corn starch
Glucose (dextrose)
Fructose syrup with 50%+ fructose
Chemically pure fructose
Fructose solids containing more than 50% fructose
Value [million US$]
Wrd00v7i.exe figure B-4. Exports of Products Made from Com in 2002
Source: Com Refiners Association, 2004


BIOGRAPHICAL SKETCH
Lewis Scotty Long was bom in Calhoun, Georgia on August 20,1971. He
graduated from Madisonville High School (Madisonville, Tennessee) in May 1989. On a
biology scholarship, Lewis attended Middle Tennessee State University (MTSU), where
he earned his BS in May 1994. On graduation, he took a job as an aquatic biologist for
Aquatic Resources Center (Franklin, Tennessee). Lewis worked there specializing in
taxonomy of mayflies, stoneflies, caddisflies, and freshwater molluscs (snails and
mussels). While still employed at Aquatic Resources Center, he started his graduate
studies in 1996 at MTSU and continued the work he had started during his undergraduate
years. In May of 1999, Lewis graduated with his MS. After receiving his MS, Lewis
moved to Florida and entered the PhD program at the University of Florida, Department
of Entomology and Nematology. He worked with Dr. Bill Peters (Florida A&M
University) on the worldwide taxonomic revision of an understudied group of mayflies.
However, Dr. Peters unexpectedly passed away in 2000, and Lewis took this unfortunate
event as a chance to broaden his expertise in entomology. In 2000, he took a part-time
job with Drs. James Cuda and Bruce Stevens on research that was in the patent process.
This was the research that Lewis undertook for his dissertation. Lewis also served as a
teaching assistant for the department for classes such as Bugs and People, Life Sciences
for Education Majors, Principles of Entomology, and Medical and Veterinary
Entomology. He served as primary instructor for Insect Classification and Immature
114


21
Figure 3-2. Setup for whole plant studies involving tobacco homworm. Top and
portions of the sides were replaced with fine mesh to allow for
airflow and to reduce condensation.


46
(a)
Qr
Mexican Market Mexico-U.S. Market U.S. Market
(b)
Figure 3-2. Two-country Trade Model (a) with Quota System and (b) without Quota
System.


48
real retail price of refined sugar (deflated by Consumer Price Index (CPI)) [cents /
pound], GDP is real per capita GDP (deflated by CPI [US$]), POP is population, QTRs
are dummy variables representing quarters of the year, DHFCS is dummy variable for
availability of high fructose com syrup (HFCS) in the sweetener market {DHFCS= 1 after
t=1975), and e, is an error term. In theory, the price of HFCS would be preferred as prices
of substitutes are expected to influence quantity demanded, however, limited availability
of the data made this impossible, therefore, the dummy was used. It is expected that the
elasticities associated with price (Ui), the first quarter (U5) (compared to the omitted
fourth quarter), and HFCS availability (Us) to be negative and the elasticities of the
remaining variables to be positive. Price elasticity is expected to be inelastic based on
previous research. Previously reported own-price elasticities for sugar are -0.141 by
Lopez (Lopez, 1990) and -0.73 by Petrolia and Kennedy (Petrolia and Kennedy, 2002).
In the latter estimation, the U.S. wholesale refined beet sugar price reported at the
Midwest Markets was used. In the process of regressions, serial correlation is anticipated
and corrected by the Yule-Walker Method with appropriate lags assigned.
Mexican Sweetener Demand Model
Estimation of Mexican sweetener demand was conducted in a similar manner to
that of the U.S. demand. The main difference was that the demand for two kinds of sugar
is estimated separately in the Mexican model. Traditionally the demand for sugar in
Mexico has been estimated by regressing the total consumption on sugar price and per-
capita income. Borrell (1991) estimated the price elasticity be -0.004 and the income
elasticity at 0.5. This model was appropriate in early 1990s because there was no HFCS
consumption in Mexico. In order to account for the entry of HFCS into the Mexican
sweetener market, the demand for sweetener is estimated by disaggregating sugar


4
and old technology also contribute to low productivity. Although the price of sugar at the
wholesale level has been privatized, the sugar price paid to growers is still controlled by a
government agency, and hence farmers have little incentive to grow sugar cane other than
to receive social benefits from the Mexican sugar program. At the national level, the
Mexican government faces a dilemma between gaining competitiveness in the
international market and maintaining social stability through offering employment and
financial supports to the livelihood of a large number of growers and related workers.
Overall, there has been little benefit to the Mexican sugar industry resulting from
NAFTA.
The U.S. sugar market, where a large quantity of sugar is traded by a large number
of sellers, has maintained commodity balance by assigning tariffs and import quotas to
foreign sellers and maintaining domestic price support through the U.S. sugar program.
As a result of GATT, the United States committed to accept a minimum import quota of
1.256 million MT of sugar in 1990; however, the U.S. sugar market has been maintained
unchanged until today through successful lobbying efforts by the American Sugar
Alliance (ASA), the sugar producers primary alliance.
In the meantime, HFCS had been gaining its share in the U.S. sweetener market
since the early 1970s when commercial production of HFCS became possible by the
advancement of wet-milling technology. Today, more than 50 percent of caloric
sweetener consumption in the United States is derived from com syrup including HFCS
(Congressional Research Service, Library of Congress, 1999). A similar phenomenon
appears to be beginning in Mexico. The implementation of NAFTA resulted in opening
the door for HFCS consumption in Mexico where nearly all caloric sweetener


LIST OF FIGURES
Figure page
2-1 Distribution of Individual Farm Size of Mexican Sugarcane Growers 24
2-2 Map of Mexicos Sugar Producing and Processing States 24
2-3 Annual Rainfall and Irrigation Rate in Sugarcane Fields 25
2-4 Mexican Sugar Production, 1988-2002 26
2-5 Main Use of Sugarcane Derivatives in Mexico 26
2-6 U.S. Refined Sugar and HFCS Use per Capita 28
2-7 U.S. HFCS Production 28
2-8 U.S. HFCS Supply 29
2-9 U.S. HFCS Export 29
2-10 Transition of U.S. HFCS Export 30
2-11 Consumption of Sugar and HFCS per Capita in Mexico 30
2-12 Transition of Prices of Sugar and HFCS 31
3-1 Conceptual Framework for the Analysis in this Study 45
3-2 Two-country Trade Model (a) with Quota System and (b) without Quota System. 46
4-1 Image of Model Calibration 65
4-2 Forecasted Indirect Sugar and HFCS Consumption in Mexico under Alternative
Scenarios 68
5-1 U.S. Sugar Import Forecast (Scenario 1 Baseline) 93
5-2 U.S. Sugar Import Forecast (Scenario 2 P-S-F) 93
5-3 U.S. Sugar Import Forecast (Scenario 3 A-S-F) 94
IX


54
goods. In addition, details of trade agreements are difficult to incorporate in these models.
The fact that there exist numerous bilateral and regional trade agreements, a spatial
equilibrium model has the advantage of enabling the various trade agreements to be
tailored into the model. Lastly but most importantly, it is critical to consider HFCS when
the sweetener markets in both the United States and Mexico are under scrutiny. Since this
study is focused on the bilateral relation between the United States and Mexico, a spatial
equilibrium is chosen to conduct a more microscopic analysis, incorporating detailed
trade agreements and the impacts from HFCS consumption.
Simplifying assumptions
Several simplifying assumptions are made to run the model. First, the quantities in
the models are raw sugar equivalent. In doing so, the price difference along the vertical
market channel is ignored and derived demand and supply curves are assumed to posses
the same slopes as demand and supply of refined sugar.
Second, changes in sugar stocks in both counties are also ignored and hence the
excess sugar supply from Mexico and the excess sugar demand from the United States
are captured as the difference between domestic sugar demanded and supplied in each
county, illustrated in the following. Formally, the U.S. sweetener market balance is
expressed with sugar production, consumption, import, export, and change in stock as
well as with those of HFCS:
Qsugar, t + SE,+ ST,+i+ HFCSd, =Qssugar, ,+ SI,+ STt.¡+ HFCS5, (4.9)
where Qsugar is the quantity of sugar demanded, Q5sugar is the quantity of sugar
supplied from domestic sugar production, SE is sugar export, SI is sugar import, ST,+i is
sugar stock carried over to the next year, STis sugar stock carried over from the
previous year, and HFCS and HFCSs are HFCS demand and supply related transactions,


72
Table 4-6. Data Sources for U.S. Demand
Data
Unit
Source
Consumption of sugar
(dependent variable)
1000
short
tons
Sugar Statistical Compendium by (Stock
#91006, 1970-1990) and Sugar and
Sweetener Situation and Outlook
Yearbook (SSS-2002, 1992-2002) by
Economic Research Service, U.S.
Department of Agriculture
Retail price of refined sugar
cents/
pound
Gross Domestic Product
(GDP)
US$
Statistic database by Organization for
Economic Co-operation and Development
(OECD)
Population
persons
International Database by U.S. Bureau of
the Census, U.S. Department of
Commerce
Data length: 1970-2002, quarter
y
Table 4-7.
Data Sources for Mexican Demand
Data
Unit
Source
Direct consumption of sugar
(dependent variable)
metric
tons
[MT]
Azcar S.A. de C.V. Estadistica
Azucareras (1970-1989), database by
Financiera Nacional Azucarera, S.N.C de
C.V. and Comit de la Agroindustria
Azucarera (COAAZUCAR) (1990-1999)
Indirect consumption of sugar
(dependent variable)
Total consumption of sugar
(dependent variable)
Retail price of standard sugar
pesos/
kg
Gross Domestic Product
(GDP)
pesos
Statistic database by Organization for
Economic Co-operation and Development
(OECD)
Population
persons
International Database by U.S. Bureau of
the Census, U.S. Department of
Commerce
Data length: 1970-1999


CHAPTER 1
THE INTEGRATED PEST MANAGEMENT DILEMMA: ARE CONVENTIONAL
PESTICIDES THE ONLY ANSWER?
Introduction
Integrated Pest Management (IPM), the sustainable approach to the management
of pest species using a combination of biological, chemical and cultural methods to
reduce economic, environmental, and public health risk, was a result of economic losses
associated with years of overuse of chemical control leading to resistance problems. The
use of IPM strategies have certainly decreased pesticide usage and encouraged the use of
methods that ensure a safer environment but many feel that it is not enough. After three
decades of research efforts in the United States, IPM as it was envisioned in the 1970s
was practiced on less than 8% of U.S. crop acreage based on Consumers Union
estimateswell short of the national commitment to implement IPM on 75% of the total
U.S. acreage by the end of the 1990s (Ehler and Bottrell 2000). This means that farm
practices have changed little since the national IPM initiative was established in 1994 to
implement biologically based alternatives to pesticides for controlling arthropod pests. It
should be noted that the low percentage of IPM practices on commercial U.S. farmland
may possibly be related to the lack of sufficient reporting means and actually may be
higher than believed when the local growers and homeowners are included. However,
the United States is considered the worlds largest user of chemical pesticides, accounting
for nearly 50% of total worldwide production and shows no sign of slowing (Deedat
1994). Pesticides remain the primary tool of pest consultants and farmers, because of the
lack of economic incentives to adopt alternative strategies that require more effort to
1


38
2004; Nester et al. 2002). Resistance in insects involves a variety of mechanisms and
many are the result of a combination of different pesticide classes. The CAATCH1
system is one that could be used in cases where the only alternative is by adding more
pesticides or at higher rates to break resistance. Further research is needed to determine
compatibility of the different Bt insecticides and L-methionine with each other for cases
in which Bt resistance is observed in natural populations. Given the safety of
L-methionine and the shorter time required for 100% mortality (when compared to Btk
results of this study), this compound could represent a viable alternative for pesticides
currently used in the management of the THW.


97
Year
Mexico price
U.S. price
Figure 5-9. Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets
(Scenario 4 PA-S-F)
Figure 5-10. Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets
(Scenario 8 PA-B-F)


To my parents and sister


61
States. Both policies abide by the minimum import requirement under WTO; however,
political feasibility is assumed to be quite different.
Scenarios to simulate are formulated by combining specific Mexicos sweetener
market situation and the U.S. sugar policy. A list of 16 scenarios is shown in Table 4-4.
Finally, special simulations are prepared in order to further examine the effects of
Mexicos production improvement and HFCS adoption on the Mexican sugar industry
and welfare. In addition to the assumption regarding Mexicos production improvement
(an additional 1 percent to the baseline) and HFCS adoption (share of indirect sweetener
consumption by HFCS at 50 percent) summarized in Table 4-2, simulations are
conducted by changing production improvement rate at additional 0.5 and 1.5 percent as
well as HFCS adoption to achieve a market share of 30, 40, and 45 percent.
Game Theory Analysis
In order to assess policy recommendations using aggregated results from the
various simulations, an analysis based upon game theory is introduced. The basis of the
game used in the study is a non-zero-sum game with mixed strategies. Non-zero-sum
means the sum of the pay-offs in each pair of strategies is not zero; in other words, one
players winning does not necessarily cause the other to lose. Mixed strategies means a
player chooses a strategy to play with probability (Morris, 1994; Mas-Colell et al., 1995).
Since the game is non-zero-sum, both cooperative and non-cooperative games are
considered. While a cooperative game allows players to make binding agreements about
how they will play or about sharing pay-offs, a non-cooperative game does not. In the
latter case, the game is played by two parties: the U.S. and the Mexican governments who
hold the strategies and make decisions on behalf of the economy as a whole. In the
former case, the game goes through the process of considering the possible pay-offs to


16
Discovery of novel means for controlling various insect pests is one tenant of
IPM. The amino acid methionine, an environmentally safe organic compound, appears to
be a candidate for further study. Before it can be considered for use in controlling insects
pests, several issues must be addressed, including the determination of concentrations
needed to provide effective control, compatibility with current application systems, safety
to nontarget organisms (i.e., beneficial or biological-control agents), and to phytotoxicity.
Research Objectives
Our overall goal was to evaluate the effects of L-methionine, and its amino acid
analogues, on the CAATCH1 system putatively in the midgut/hindgut as a means to
control different insect pests. The working hypothesis is that the L-methionine only
affects the CAATCH1 system and no other system, especially those involving Na+
channels or pumps (i.e., nervous tissue). The L-isomer of methionine was chosen
because of the inability of most insect species to utilize the D-isomer. Ideal targets for
this research are those pests that cause severe damage to agricultural systems and to
human health. Specific objectives were to
Examine the effects of L-methionine as an insecticide on the larvae of M. sexta
(Tobacco homworm), L. decemlineata (Colorado potato beetle) and A. aegypti
(Yellow-fever mosquito) under various conditions
Determine any adverse effects of L-methionine on plant health to ensure its safe
use in a cropping system
Examine the effects of L-methionine on various nontarget insect species to ensure
the environmental safety of L-methionine and thus its compatibility with natural
enemies in the context of IPM.


85
on Mexicos welfare differ from those on the industry (Figure 5-18). Production
improvement increases welfare, but not HFCS adoption, assuming that sugar is a primary
and preferred source of sweetener. While the gains to the Mexican sugar industry from
production improvement at 1.5 percent to the baseline (US$ 0.65 billion) can make up the
loss to the industry itself from HFCS adoption, even at a 50 percent share (US$ 0.42
billion), gains to Mexicos welfare cannot. The net loss to the nation from HFCS
adoption at 30 percent adoption share is forecasted US$ 5.12 billion (loss of US$ 5.41
billion from welfare and a gain of US$ 0.29 billion from the industry).1 This loss is far
greater than the sum of expected gains from production improvement at additional 1.5
percent, which is US$ 3.42 billion (gains of US$ 2.77 billion from welfare and US$ 0.65
billion from the industry). The Mexican government faces difficulties allowing faster
HFCS adoption to happen in the domestic market (Figure 5-19).
Game Theory Analysis
Results from the game theory analysis are summarized in Tables 5-6 through 5-13.
The analysis is based on the game setting played by the United States and Mexico with
multiple strategies that correspond to Mexicos market situation (the Mexican
governments strategy set) and the U.S. policy levers (the U.S. governments strategy set)
in order to assess gainers and losers from trade. Calculated actual pay-offs to five payees
(the U.S. HFCS industry, the U.S. sugar industry, Mexican sugar industry, U.S. cost-
adjusted welfare, and Mexicos welfare) for each combination of U.S. and Mexicos
strategies of the game are shown in Table 5-6. Values in the other tables are indexed
relative to the baseline scenario. Three different forms of coalitions are considered:
1 Assuming that industrys revenue and nations welfare can be added together.


85
Neochetina eichhorniae
Adults of the MWHW were used in this study since the larvae and pupae are
buried deep in plant tissue and therefore not likely to come into contact with methionine
that could be present in a body of water. Specimens were supplied by Hydromentia, Inc.
(Ocala, FL), from areas around South Florida. Weevils were maintained following the
procedures outlined by Haag and Boucias (1991), with small petri dishes fitted with
moistened filter paper and freshly cut water hyacinth leaves. Water hyacinth plants were
collected from Lake Alice on the campus of the University of Florida and maintained in
the University of Florida, Department of Entomology and Nematology greenhouse.
Treatments consisted of cut leaves dipped in deionized H2O (control) or solutions
containing 0.1% L-methionine, 0.5% L-methionine, 1.0% L-methionine or 1.0% proline.
Prior to weevil exposures, each leaf was inspected for feeding scars or damage
and noted to ensure the counts were based on current feeding. Each treatment consisted
of 4 replicates with n=5 per replicate (n=20 per treatment and total n=100). Weevils and
hyacinth leaves were held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a
hardware cloth (to facilitate cleaning) and maintained at 27 C, 60% relative humidity
and 16L/8D photoperiod in FRJUs. Fresh leaves were provided every 4 days; exposed
leaves were preserved in sealed plastic bags and placed in a refrigerator until scars could
be counted. Feeding damage was determined (with the use of an Olympus Tokyo Model
213598 stereo microscope) by the total number of scars present with each counted scar
marked with a fine tipped permanent marker (Figure 7-3).
Statistical analyses of the weevil data were performed using Minitab Version 12
(Minitab, Inc.; State College, PA). Feeding scars on control and treatment leafs were


Mean Amount of Leaf Material Consumed (ci)i
49
(Error Bars @ 95%; F(0.05)i,i8=5.98, F=1.64; P =0.217)
Control 1.0% L-methionine
Figure 4-5. Mean leaf consumption by Colorado potato beetle in the preference
tests. Error bars denote 95% SE, and treatments were found not to
be statistically different. No correlation between either Control or
Treatment Diet consumed and mean head capsule width was found
(Pearson Correlation Coefficient 0.466, P=0.175 and 0.665,
P=0.036, respectively).


26
Figure 3-5. Mortality of tobacco homworm larvae exposed to various concentrations of
L-methionine (nxotai-480) in artificial diet. Data were adjusted using
Abbotts formula to account for control mortality. Note the overlap in trend
lines for the 3.0% L-methionine-10.0% L-methionine concentrations after
Day 1 and the 0.3% L-methionine and 0.5% L-methionine treatments from
Day 1 to Day 10.


13
when sugar mills were privatized. Growers are paid by a fixed portion of the reference
sugar price calculated from aggregated sugarcane harvested and processed in the specific
mill. Details for the cane price setting formula are shown in equations [2.1] and [2.2]
(Garcia Chaves et al, 2004):
Cane price / ton = (KARBE/ ton of cane)*(Price of KARBE)*(0.57) (2.1)
KARBE/ ton of cane = (Pol) *(FF)*(FP)*(EBF)*(TF) (2.2)
where KARBE is kilogram of recoverable standard sugar basis (Pol 99.4 percent) for net
ton of cane; Pol is polarization of cane (apparent percentage of sucrose in cane); FF is the
fiber factor; FP is the purity factor; EBF is mill efficiency; and TF is the transformation
factor.
As seen in equation [2.1], currently growers are paid 57 percent of the wholesale
price per kilogram of standard sugar. Although in 1991 growers began to be paid
according to the quality of cane produced as opposed to solely on weight as decreed in
the amendments to Decreto Caero, the Sugarcane Growers Law, growers have little
incentive to produce higher quality cane. The cane price is capped at 57 percent of
average quality cane for the specific mill, not a price reflecting each growers sugarcane.
Thus, a main incentive for Mexican sugarcane growers is to receive social benefits and
medical services from the government as opposed to producing quality cane.
The wholesale price of sugar produced at mills has been liberalized since 1997;
however, it still quotes a reference price calculated based on the formula published by the
secretariat of Commerce (SECOFI). The price is determined by considering both the
recent domestic price and the expected export price, which is the composite of the U.S.
and world price (Garcia Chaves et al, 2004), as shown in equation [2.3],


77
Control
0.10%
0.50%
1.00%
20
Days after treatment
Figure 6-5. Mortality of Colorado potato beetle larvae on eggplants treated with
L-methionine and Silwett L-77. Mortality of larvae corrected using
Abbotts formula (Abbott, 1925). Analysis performed on arcsin
transformed data. Error bars denote 2 SE. Data points having by the
same letter are not statistically different (Tukeys MST, P=0.05)


Table 5-8. Indexed Pay-off Matrix for the Trade Policy Game Played by Two Coalitions of Countries without the U.S. HFCS industry


no
Nester, E.W., L.S. Thomashow, M. Metz and M. Gordon. 2002. One Hundred Years of
Bacillus thruingiensis: A Critica Scientific Assessment Report from the
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Onifade, A.A., O.O. Oduguwa, A.O. Fanimo, A.O. Abu, T.O. Olutunde, A. Arije and G.
M. Babatunde. 2001. Effects of supplemental methionine and lysine on the
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Technol. 78:191-194.
Palumbo, R.E. and D.L. Dahlman. 1978. Reduction of Manduca sexta fecundity and
fertility by L-canavanine. J. Econ. Entomol, 71:674-676.
Pan, M.L. and W.H. Telfer. 1996. Methionine-rich hexamerin and arylphorin as
precursor reservoirs for reproduction and metamorphosis in female Luna moths.
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Patterson, K.D. 1992. Yellow fever epidemics and mortality in the United States, 1693-
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Perfect, T.J. 1992. IPM in 2000, pp.47-53. IN A.A.S.A. Kadir and H.S. Barlow (eds.),
Pest Management and the Environment in 2000. CAB International, Oxford, UK.
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Quick, M. and B.R. Stevens. 2001. Amino acid transporter CAATCH1 is also an amino
acid-gated cation channel. J. Bio. Chan. 276(36): 33143-33418.
Racioppi, J.V. and D.L. Dahlman. 1980. Effects of L-canavanine on Manduca sexta
(Sphingidae: Lepidoptera) larval hemolymph solutes. Comp. Biochem. Physiol.
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Ragsdale, D. and E.B. Radcliffe. 1999. Colorado potato beetle management. University
of Minnesota Cooperative Extension Service. Internet URL: http://ipmworld.umn.
edu/aphidalert/CPB~DWR.html. Accessed April 2004.
Robertson, J.L. and H.K. Priesler. 1991. Pesticide bioassays with arthropods. CRC
Press, Inc. Boca Raton, 127pp.
Rock, G.C. 1971. Utilization of D-isomers of the dietary, indispensable amino acids by
Argyrotaenia velutinana larvae. J. Insect Physiol. 17:2157-2168.
Rock, G.C. and E. Hodgson. 1971. Dietary amino requirements for Heliothis zea
determined by dietary deletion and radiometric techniques. J. Insect. Physiol. 17:
1087-1097.
Rock, G.C., BG. Lign, and E. Hogson. 1973. Utilization of methionine analoges by
Argyrotaenia velutinana larvae. Arm. Entol. Soc. Amer. 66(1): 177-179.


pests. The tobacco horn worm (THW), Colorado potato beetle (CPB) and the yellow
fever mosquito (YFM) were tested and found to be susceptible to concentrations greater
than 0.1%. Diets, both natural and artificial, containing this compound resulted in the
complete mortality of THW and also in the natural diet for CPB. Development and
feeding rates were also affected by the addition of L-methionine to diets for THW and
CPB. Survivorship and developmental rates of YFM were also affected by the addition
of this amino acid to the larval habitat.
In Chapter 6 it was found that the field application of L-methionine under natural
conditions was able to control CPB. It was also determined that L-methionine was
compatible with Silwett L-77, a commonly used adjuvant, and showed no detrimental
effects on crop yield of eggplant.
Finally, the application of a compound such as L-methionine has to be able to
control the pests that it is used against and not have an effect on beneficial organisms that
may come into contact with this compound. Chapter 7 detailed the results of tests that
involved various beneficial insects from different feeding guilds (herbivore, predator and
parasitoid) showed that L-methionine does not appear to pose a threat to nontarget
organisms.
One aspect of the use of a compound like L-methionine that is very important is
the relative safety. The health hazards related to the contamination of the environment
with pesticides are well documented and in the recent years have resulted of the review
and removal of several insecticides from commercial and private use. The use of
L-methionine as an insecticide would alleviate the dangers associated with other
pesticides. The approved use as a nutritional supplement for livestock feed is a testament


TABLE OF CONTENTS
page
ACKNOWLEDGMENTS iv
LIST OF TABLES vii
LIST OF FIGURES ix
ABSTRACT xii
CHAPTER
1 INTRODUCTION 1
Background 1
Problem Statement 5
Researchable Questions 5
Objectives 6
Organization of the Study 6
2 SUGAR INDUSTRIES AND SWEETENER MARKETS IN THE UNITED
STATES AND MEXICO 8
The Mexican Sugar Industry and Sweetener Market 9
Mexicos Sugarcane Production 9
Mexicos Sugar Production 10
The Mexican Sugar Industry and Government Involvement 12
Mexicos Sugar Consumption 14
Mexico as a Sugar Exporter 15
Development and Adoption of High Fructose Corn Syrup 17
The U.S. Sugar Industry and Sweetener Market 19
3 CONCEPTUAL AND THEORETICAL FRAMEWORK 32
Conceptual Framework 32
Theoretical Framework 35
Sweetener Market Analysis 35
Sweetener demand 35
Sweetener supply 36
U.S.-Mexico Bilateral Sugar Trade System 37
v


51
The higher concentrations of L-methionine that produced mortality similar to the
Btt is encouraging considering the occurrence of resistance to this compound seen in
many pest insect species because of reduced receptor activity and binding (Bills et al.
2004; Nester et al. 2002). Resistance in insects involves a variety of mechanisms and
many are the result of exposure to a combination of different pesticide classes. The
Methionine-CAATCHl system could be exploited in cases where the only alternative is
applying different pesticides or using higher rates to break resistance. Further research is
needed to determine compatibility with Bt and L-methionine for cases in which resistance
is observed in natural populations. Given the safety of L-methionine and the shorter time
required for 100% mortality (when compared to Btt), this compound could represent a
new biorational tool for the management of the CPB.


92
Table 5-2. Summary of the Mexican Supply-Demand Analysis
Country
Mexico
Dependent
variables
Independent^^
variables
Demand
Supply
Direct
consumption
of sugar
Indirect
consumption
of sugar
Total
consumption
of sweeteners
Production
of sugar
Number of
observations
30
30
30
13
Degree of freedom
26
25
25
6
Real retail price of
standard sugar 1}
-0.0734
(-1.93) *
-0.003535
(-0.07)
-0.0215
(-0.98)
Real per capita GDP
i)
-0.2595
(-1.37)
0.9213
(2.96) **
0.4363
(3.33) **
Population
1.1194
(8.51) **
1.4175
(5.39) **
1.2486
(10.92) **
Dummy variable for
availability of HFCS
-0.1025
(-1.58)
-0.0250
(-0.88)
Real wholesale price
of standard sugar in
the previous year 1)
0.2152
(2.52) *
Real production cost
per ton of sugar 1 ]
-0.3228
(-7.89) **
Downtime
-0.4275
(-6.77) **
Loss of sugar during
the process
-0.2112
(-1.64)
Duration of the
harvest
1.0010
(9.13)**
Trend
0.5435
(12.75) **
Constant
-3.4499
(-2.72) **
-20.5135
(-6.90) **
-11.9624
(-8.91) **
11.8453
(20.89) **
Total R2
0.9344
0.9729
0.9905
0.7380
Durbin-Watson 2)
1.9217(1)
1.9961 (2)
1.5933 (1)
1.5357 (2)
1.1648 (3)
1.8715(1)
1.5834 (2)
2.9128
* and **: Significant at 90% and 95% confident level, respectively.
1) Deflated by CPI.
2) Values are after corrected by Yule-Walker method. ( ) corresponds to the order of
lag assigned.
3) Indicates neither positive nor negative correlation.


55
respectively. Qsugar, is expressed in annual base converted from quarterly base. Since
neither supply nor demand of HFCS is estimated, HFCS and HFCSs are treated constant.
This is justified by the fact that HFCS use in the United States has remained stable in
recent years, accounting for roughly half of total U.S. caloric sweetener use. In addition,
changes in stock have also remained stable given the nature of commodity demand and
production practices, ST,.¡ and ST,+¡ are set zero. SE is also set zero as sugar export from
the United States is negligible given the production capacity. Equation [4.9] is then
simplified and the quantity of forecasted sugar import in the United States (SI,) is
expressed with forecasted quantities of sugar demanded and supplied (Q sugar, i
and <25sugar, t) as:
SI, = Q sugar, t QSSUGAR, t (4.10)
Mexicos sweetener market balance is expressed in a similar fashion. The
differences are sugar demand in Mexico is estimated by total, direct and indirect sugar
consumption and Mexico imports HFCS from the United States to meet its domestic
demand for sweeteners. By ignoring stock changes and sugar import, the sweetener
market balance is expressed as:
QDCSUGAR, t + QICSUGAR, + HFCSd ,+ SE, = Qssugar, + HFCSs, (4.11)
where Q sugar is the quantity of demanded by households (direct consumption),
Q,Csugar is the quantity of sugar demanded by bulk users (indirect consumption), HFCS
is the quantity of HFCS demanded by bulk users, SE is sugar export, Qssugar is the
quantity of sugar supplied from domestic sugar production, and HFCSs is the quantity of
HFCS supplied from domestic production and import from the United States. The study
is interested in forecasting sugar surplus available for exporting to the United States


91
Figure 7-4. Mortality of Neochetina eichhorniae on treated water hyacinth
leaves. Data corrected for control mortality using Abbotts
formula.


ACKNOWLEDGMENTS
I thank Jim Cuda and Bruce Stevens for giving me the financial and intellectual
freedom that made this work possible. I want to thank Jim for housing me in his lab and
providing the facilities to perform this work, and Bruce for allowing me to take his initial
work and elaborate on it as well as including me as a co-inventor of the research
presented. Most of all, I would like to express my sincere appreciation to Judy Gillmore.
Without her support and help this research would not have been completed. Judy was
integral in every aspect of this endeavor and put up with more than her fair share of my
research. I extend heartfelt thanks to George Gerencser, James Maruniak, Simon Yu,
and Susan Webb for serving as members of my supervisory committee. I would like to
also thank Jim Lloyd, Jerry Butler, and Carl Barfield for all the experiences and
knowledge shared. Finally, I want to express my deepest, eternal gratitude to my fellow
graduate students Jim Dunford and Heather Smith, for providing support and guidance
that only colleagues, intellectual equals, and close friends can give. I can only hope to
repay them for their help by providing the same amount of support for their endeavors as
they did mine.
m


2
special safeguards to protect import-sensitive crops, including sugar, which are defined
under side agreements between Mexico and the United States (USDA, 2001c).
When trading sugar with the United States, the Mexican sugar industry faces two
counteracting conditions under the NAFTA regime: increased access to the U.S. market
which would facilitate sugar exports at favorable prices; and the pressure of increased
imports of HFCS from the United States which have been gaining an increasing share of
Mexicos sweetener market since 1994. Under the provisions of NAFTA, both an over
quota tariff for Mexican sugar which enters into the United States and a tariff on exported
HFCS which enters the Mexican market are regulated in such a way that both tariffs will
be reduced to zero by 2008 and 2004, respectively. In addition to the rules of the tariffs,
Mexican sugar is subject to U.S. import quota allocations. Mexico is allowed to access
two kinds of quotas, depending on Mexicos domestic balance in the sweetener market: if
Mexicos sugar production exceeds its sweetener consumption (the sum of sugar and
HFCS consumption in two consecutive years -net surplus sweetener producer status),
Mexico receives 25,000 MT of sugar import quota; and if not, Mexico receives 7,258 MT
of quota. Additionally beginning in 2000, the sugar import quota expands from 25,000
MT to 250,000 Mt as long as Mexico satisfies the conditions of a net surplus sweetener
producer. Mexico can export over the 7,258 MT quota without attaining net surplus
sweetener, but any sugar exported in this scenario would be subject to taxation in the
form of tariffs as mentioned above. In 2008 when all the restrictions, i.e. both tariffs and
quotas, are lifted, Mexico will have free and unlimited access to the U.S. sugar market.
In Mexico, the sugar industry has played an important role in the economy and the
politics of the country. In spite of experiencing drastic economic and political changes,


55
Figure 5-1. Bioassay setup for yellow fever mosquito larvae exposed to various
concentrations of amino acids and Bti. Jars contained 500mL of
solution and were covered with screen to prevent the escape of
emerging adults.


CHAPTER 2
HISTORY OF THE USE OF AMINO ACIDS AS A MEANS TO CONTROL INSECT
PESTS
Non-Protein Amino Acids
One avenue of pest management explored in the field of biorational pesticides is
the use nonprotein amino acids. Secondary plant materials such as these serve many
functions in insect-plant relationships from attractants and repellents to crude insecticides
(Dahlman 1980). Only a few nonprotein amino acids have been examined as a potential
means to control insect pests. L-canavanine and its by-product of detoxification, L-
canaline, have been studied extensively, with a variety of effects ranging from
developmental deformities to aberrant adult behavior (Dahlman and Rosenthal 1975;
1976; Rosenthal et al. 1995). L-canavanine is found mainly in leguminous plants,
including several economic species (Bell 1978; Felton and Dahlman 1984). It is believed
that plants produce this allelochemical for protection against feeding by phytophagous
insects and herbivores (Rosenthal 1977). The mode of action for canavanine can be
traced to several metabolic processes, including disruption of DNA/RNA and protein
synthesis, arginine metabolism, uptake, anomalous canavanyl protein formation, and the
reduction of active transport of K+ in the midgut epithelium (Kammer et al. 1978;
Racioppi and Dahlman 1980; Rosenthal 1977; Rosenthal et al. 1977; Rosenthal and
Dahlman 1991). In contrast, canaline possesses neurotoxic characteristics with an
unknown mode of action (Kammer et al. 1978). The species of choice for studies
involving nonprotein amino acids has been the tobacco homworm (THW), Manduca
sexta (L.) (Lepidoptera: Sphingidae).
7


4EMPIRICAL MODELS AND DATA SOURCE
.47
Empirical Models 47
U.S. Sweetener Demand Model 47
Mexican Sweetener Demand Model 48
U.S. Sweetener Supply Model 50
Mexican Sweetener Supply Model 52
U.S.-Mexico Bilateral Sugar Trade Model 53
Simplifying assumptions 54
Model calibration 56
Simulated Scenarios 58
Game Theory Analysis 61
Sources of Data 63
5 EMPIRICAL RESULTS AND INTERPRETATION 75
Demand and Supply Analyses 75
Bilateral Sugar Trade Analysis 77
Game Theory Analysis 85
6 CONCLUSIONS AND IMPLICATIONS FOR POLICY 115
Conclusions and Implications for Policy 115
Impact of Changes in Trade Regime 116
Mexicos Export Potential 117
The Impact of Changes in Mexican Market Situation 117
The Impact of Changes in U.S. Sugar Policy 118
Alternative Sugar Policy by the United States 120
Limitation of the Study and Suggestions for Future Research 122
APPENDIX
A MAJOR EVENTS IN THE SUGAR INDUSTRY HISTORY IN MEXICO AND
THE U.S. 123
B CORN STATISTICS 125
C DERIVATION OF INVERSE LINEAR EQUATIONS 128
BIOGRAPHICAL SKETCH 136
vi


12
sugarcane fields and poor road conditions also contribute to longer transportation time,
and hence decreased sugarcane quality.
Mexicos sugar production was approximately 4.8 million MT, raw equivalent, in
2002, ranking it seventh among all cane sugar-producing nations; Brazil and India are by
far the largest cane sugar-producing nations, followed by China, the United States,
Thailand and Australia (Table 2-1). Production in Mexico has been increasing for the past
few decades (Figure 2-4).
The Mexican Sugar Industry and Government Involvement
Mexicos sugar production accounts for 0.5 to 0.7 percent of its gross domestic
product (Garcia Chaves et al., 2002; Farm Foundation, 2003). Since privatizing in the late
1980s, mills have neither successfully accumulated capital nor renewed their equipment
leaving the industry financially vulnerable. In 2001, the Mexican government
expropriated 27 mills, which represented approximately 50 percent of sugar production in
Mexico. In February 2002, the Government of Mexico announced a National Sugar
Policy for 2002 2006 which included a series of short- and long- term measures to help
Mexicos ailing sugar industry with the main objective of regulating the sugar market and
making the sugar sector profitable (USDA, 2002b). Today the sugar industry remains
important in Mexico because it is considered crucial for maintaining social stability due
to the large number of growers and related workers.
Among the public organizations that deal with the Mexican sugar industry,
Commite de la Agroindustria Azucarera (COAAZUCAR) plays an important role by
monitoring and compiling sugarcane and sugar production data at each mill. Although the
industry has been privatized, COAAZUCAR is in charge of determining the cane price. It
took over the task from the former body, Azcar, S.A. which was dismantled in 1991


82
forecasted larger exports in 2008 and 2009, which consequently brings a lower U.S. sugar
price accompanied by a higher cost of the price support. Overall, U.S. welfare becomes
worse off with changes in Mexican sweetener market for two reasons: (1) reduced
producer surplus caused by a lower producer price as a result of increased Mexican
export and (2) increased net costs even though increased tariff revenue is expected as
mentioned above. The Mexican sugar industry gains from expanding sugar production,
but the gains are dissipated when HFCS is adopted at higher rates, given the assumption
that HFCS price held constant. In Mexicos HFCS tax scenario, the Mexican sugar
industry gains not from exporting to the U.S. market but from domestic sales at higher
prices. All the entities except for U.S. HFCS industry benefit from this policy; however,
the policy lever may not be acceptable in the international trade environment. In fact, the
Mexican government swung its decisions in the past: a 20-percent tax on beverages that
contain HFCS was introduced on January 1, 2002; suspended on March 5 by the
presidents decision; and then reimposed on July 16, 2002 with the decision by Mexicos
Supreme Court of Justice (USDA, 2002a).
The impact of changes in the U.S. price stabilization policy is illustrated in Table 5-
4. The compared three scenarios (Scenario 4, 8, and 12) are based on the assumption that
Mexico increases sugar production as well as HFCS adoption and that the U.S.
government allocates quotas between Mexico and the rest of the world in a flexible
manner. Alternative sugar policies to the price support only bring about improvement in
U.S. welfare; sugar industries in both countries and Mexicos welfare become worse off,
posing a larger negative impact on the Mexican sugar industry and welfare than the sugar
industry in its own country. When the U.S. government switches sugar policy from


34
(Error Bars @ 95%; F(o.os)7,i52=2 .37, F=\8.2; P<0.001)
Figure 3-11. Total leaf area consumed by tobacco homworm larvae exposed to
excised eggplant leaves treated with various concentrations of L-
methionine (nrotai=320). Proline (1.0%) and Btk were included for
comparison as positive and negative controls. Error bars denote 2 SE.
Bars within treatments having the same letter are not statistically
different (Tukeys MSTP, P<0.001).


19
sweetener market over the past several years. The Mexican sugar industry has struggled
to supply sugar to the domestic market at a price competitive with HFCS. Prior to 1994,
nearly all of the caloric sweetener consumption was derived from domestically produced
sugar; however, implementation of NAFTA resulted in opening the door for HFCS
consumption in Mexico. As a result, Mexican per capita sugar consumption has
decreased slightly since 1991, while per capita sweetener consumption has been
increasing (Figure 2-11). Soft-drink manufacturers are believed to account for about one-
third of the total sugar domestically demanded (Buzzanell, 2002). Currently, Mexicos
HFCS consumption accounts for approximately 12 percent of total consumption of
sweetener (approximately 25 percent of indirect sugar consumption) in 2001.
In response to this threatening trend of replacing domestic sugar consumption with
U.S. produced HFCS, the Mexican government imposed tariffs in 1996 on HFCS, based
on a claim that U.S. companies were dumping HFCS at an unfair price and affecting the
export volume and value of Mexican sugar. This action evolved into a trade dispute
between the United States and Mexico and ended when the WTO panel ruled against
Mexicos claim (Garcia et al., 2002 and 2004). Combined with a slump in production that
occurred in 1999 and 2000, the Mexican sugar industry underwent an economic crisis. In
September 2001, the Mexican government expropriated 27 of 60 Mexicos functioning
sugar mills in order to maintain the industry (USDA, 2001b). Major events in sugar
industry history in both Mexico and in the United States are summarized in Appendix A.
The U.S. Sugar Industry and Sweetener Market
Sugar production in the United States comes from two sources: sugarcane and
sugar beets. The main sugarcane production regions are Florida, Hawaii, Louisiana, and
Texas where the climate is tropical or semi-tropical. Louisiana and Florida produce


130
curve that has the same slope as direct consumption of sugar with the intercept shifted by
adding the vertical indirect consumption demand curve. In this way, the demand equation
is expressed in terms of total consumption of sugar (Qrc,) with significant estimates {M2
in equation [4.2]), incorporating the effect of change in indirect consumption of sugar as
a consequence of the change in HFCS consumption.
Pmx, = IM,+ IM2*QTCMx.t + IM3*GDPMx + IM4*POPmx, IM2*QICmx, <
(C.9a)
or P MX, 1 IM, + IM2 *QtcMx + Shifter0mx, t (C.9b)
PSMX, < = IMM, + 1MM2*QSmx, + IMM3*COSTmx, + IMM4*DT,
+ IMM5*SUGLOSS, + IMM6*DURTN, (C. 10a)
or PSmx, = IMM, + IMM2 *Qs mx, 1 + Shifter^mx, t (C.lOb)
and the coefficients are presented in Table C-2.
Table C-2. Coefficients for Inverse Linear Functions -Mexico-
Inverse linear demand function
Inverse linear supply function
IM2
(1/ EIX p mx)*(P MX/QIL mx)
imm2
(1/ ES P Mx) *(PS MX /QS mx)
IM3
(-EGDP, MX /EP MX )*
(Pmx/GDPmx)
IMM3
(~ESCOST, MX/ESPMX)*
(P5MX/COSTMX)
im4
(-Epop, mx /E p} mx)*
(PMX/POPMX)
imm4
{-ESdt/ Esp mx) *(PSmx /DT)
imm5
("E?SUGLOSS / ESp mx) *
(PS mx /S U GLOSS)
imm6
(.-E?durtn/E?p mx)*
(P^mx/DURTN)


78
When Mexico expands sugar production (Scenario 2), net surplus sweetener
producer status will still not be attained (Figure 5-2). Yet, Mexico will generate enough
surplus sugar to export over-quota (before 2008) and quota-free (after 2008), resulting in
significant impacts on the U.S. market. In total, Mexican sugar will take up about one-
third (Scenario 1) or more than half (Scenario 2) of the U.S. minimum import
requirement at peak in 2008. The amount of export will decline in later years due to
expanding domestic sugar consumption in Mexico.
In Scenario 3, when Mexico adopts HFCS at a higher rate, Mexicos sugar export
swells as a result of substitution between sugar and HFCS in the domestic market. This
result includes direct impacts on the Mexican sweetener market as well as extended
impacts on the U.S. sweetener market (Figure 5-3). Although Mexico will not attain net
surplus sweetener producer status, over-quota export will reach over 1.2 million MT by
2007 and will remain over one million MT until 2014. This export quantity will take up
almost the entire U.S. minimum import requirement and as a result, sugar export from the
rest of the world will be marginalized. Similar results are drawn when increases in
Mexicos production and HFCS adoption are combined (Scenario 4). A slightly larger
scale of Mexicos exports than Scenario 3 is shown in Figure 5-4.
The policy followed by the U.S. government in its allocation of its import quota
has a large impact on sugar exports from both Mexico and the rest of the world. The
aforementioned large-scale export of Mexican sugar is possible only if the U.S. imports
the minimum amount of sugar and allocates sugar quotas in a flexible manner among
exporters. This allocation method may cause friction with the other countries that export
sugar to the United States since Mexican sugar has potential to take up a large portion of


95
body of the host, and not through direct contact with the foliage where the compound was
applied. There is a possibility for the parasitoid having higher methionine requirements;
based on filarial worm infected Aedes aegypti (L.) (Dptera; Culicidae) females and the
associated drop in methionine levels in the haemolymph (Jaffe and Chrin 1979). This
makes alternatives such as L-methionine safe for use around beneficial insects like the
greenbug parasitoid.
Overall, the results indicate that the PSLB (C. maclala), the MWHW (JV.
eichhorniae) and the GBP, (L. tes tace ipes) were not adversely affected by exposure to
L-methionine in excess concentrations in a variety of artificial and natural diets.
Survivorship and feeding rates were not statistically different between control and
treatment groups for each species. From these data, it can be concluded that
L-methionine is safe for use with beneficial insects and could be considered biorational
in that it showed no adverse effects on non-target species. It also should be stressed that
additional testing on other beneficial insects would be, on a case by case basis, necessary
to examine the safety and biorational qualities of L-methionine.


ANALYSIS OF U.S.-MEXICO SUGAR TRADE: IMPACTS OF THE NORTH
AMERICAN FREE TRADE AGREEMENT (NAFTA) AND PROJECTIONS FOR THE
FUTURE
By
DAISUKE SANO
A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT
OF THE REQUIREMENTS FOR THE DEGREE OF
DOCTOR OF PHILOSOPHY
UNIVERSITY OF FLORIDA
2004


11
The Cation-Anion Modulated Amino Acid Transporter With Channel Properties
(CAATCHl) System
Cation-Anion modulated Aminoacid Transporter with Channel properties
(CAATCHl) is a recently cloned insect-membrane protein isolated from larval
midgut/hindgut nutritive absorptive epithelium. This membrane protein exhibits a unique
polypeptide and nucleotide sequence related to, but different from, mammalian Na+-, Cf-
coupled neurotransmitter transporters (Feldman et al. 2000). Using a unique PCR-based
strategy, the gene encoding CAATCHl was cloned from the digestive midgut of THW
larvae. The unique biochemical, physiological, and molecular properties of CAATCHl
indicate that it is a multifunction protein that mediates thermodynamically uncoupled
amino acid uptake, functions as an amino acid-modulated gated alkali cation channel, and
is likely a key protein in electrolyte and organic-solute homeostasis of pest insects (Quick
and Stevens 2001). In the presence of no amino acids, the cations K+ and Na+ are
transported through the membrane via the channel (Figure 2A). When exposed to
proline, the amino acid is transported through the membrane with an increase in cation
flow, especially Na+ (Figure 2B). However, when exposed to methionine, the amino acid
transport is stopped and cation flow is altered, mainly the increased flow of K+ and the
decreased flow of Na+ (Figure 2C). The CAATCHl system works in alkaline conditions,
at a pH optimum ~ 9.5. This alkaline condition is found in the midgut of several species
(Nation 2001) and has been attributed to a variety of causes, from the detoxification of
plant allelochemicals to amino acid uptake (Giordana et al., 2002; Leonardi et al. 2001).


34
The U.S.-Mexico bilateral trade system involves sugar exported from Mexico to the
United States and HFCS exported from the United States to Mexico. Mexican sugar is
exported to a variety of markets, but the primary destination is the U.S. sweetener market.
Once Mexico exports the amount of sugar to the United States allowed under the
U.S.import system, excess sugar is exported to the world sugar market. In total, more
than 30 countries exported sugar to the United States under the allocated tariff-rate quota
in 2002 (USDA, 2002a). Under the conditions of NAFTA, Mexico could be allocated
250,000 MT of the U.S. import quota given certain conditions (successful attainment of
net surplus sweetener producer status), which would increase Mexicos share of the total
U.S. import sugar quota allocation to 20 percent. A two-country trade model with quota
imposed by a large country importer is illustrated in Figure 3-2 (a). By imposing a quota
on Mexican excess supply of sugar (ESmx), the quantity exported to the United States is
limited to Qq instead of Qf. Consequently, the price for imported sugar in the United
Staes increases to Pq> us and the price for exported sugar from Mexico decreases to Pq, mx-
This causes welfare loss in Mexico (area abed) due to lower sugar export price and
generates quota revenue in the United States (area efgh) collected by the U.S. quota
holders or the government. Beginning in 2008 Mexico will have free access to the U.S.
market. The same trade model without the quota system is illustrated in Figure 3-2 (b).
Trade without distortion brings an increase in welfare in both countries. For simplicity,
the producer price support policy is excluded from both Figures (a) and (b).


16
Another advantage Mexico attained is the preferred trade conditions under the
North American Free Trade Agreement (NAFTA) implemented in 1994. NAFTA created
a freer trade environment among Mexico, the United States and Canada by eliminating
tariffs. In terms of agricultural trade between Mexico and the United States, many tariffs
were eliminated immediately, while others were scheduled to be phased out over periods
of 5 to 15 years (USDA, 2001a). Mexico benefited from exporting its surplus sugar to the
U.S. market at a higher price and at a lower tariff rate which will be reduced each year
and eventually gives Mexico free and unlimited access to the U.S. market beginning in
2008 (Table 2-3).
The NAFTA agreement is a double-edged sword to the Mexican sugar industry,
however, creating counteracting conditions in the Mexican sweetener market: one is the
increased access to the U.S. market and the other increased access of high fructose com
syrup (HFCS) from the United States. The quantity of duty-free sugar exported from
Mexico is limited by quotas which vary depending on Mexicos balance in the domestic
sweetener market not the sugar market; Mexico receives a larger quota if its domestic
sugar production exceeds domestic consumption of sweetener, including HFCS (called
net surplus sweetener producer status) in two consecutive years. At the same time, the
agreement gives U.S. HFCS producers free access to the Mexican sweetener market
beginning in 2004 as a tariff imposed on HFCS imported from the United States is also
being phased out. If Mexico fails to attain net surplus sweetener producer status, it
receives a sugar import quota of only 7,258 MT rather than 250,000 MT and thus most of
Mexican sugar exported to the U.S. market is subject to over-quota tariffs until 2008
when all the restrictions on Mexican sugar are lifted.


109
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Number M2108. JT Baker Inc. Internet URL:httpV/www jtbakcr.com
/msds/englishhtml/M2108.htm. Accessed April 2004.
Marrone, P.G. and S.C. Macintosh. 1993. Resistance to Bacillus Ihuringiensis and
resistance management, pp. 221-236. IN, P.F. Entwistle, J.S. Cory, M.J. Bailey
and S. Higgs (eds.), An Environmental Biopesticide: Theory and Practice. John
Wiley and Sons New York. 330pp.
McPherson, R.M. and D.C. Jones. 2002. Tobacco Insects: Summary of losses from
insect damage and costs of control in Georgia-2001. University of Georgia
Integrated Pest Management. Internet URL: http://entomology.ent.uga.edu/IPM/
si 01/tobacco.htm. Accessed April 2004.
Melangeli, C., G.A. Rosenthal and D.L. Dahlman. 1997. The biochemical basis for L-
canavanine tolerance by the tobacco budworm Heliothis virescens (Noctuidae).
Proc. Natl. Acad. Sci. USA. 94: 2255-2260.
Mitchell, B.K. 1974. Behavioral and electrophysiological investigations on the
responses of larvae of the Colorado potato beetle (Leptinotarsa decent!ineata) to
amino acids. Ent. Exp. & Appl. 17:255-264.
Mitchell, B.K. and L.M. Schoonhoven. 1974. Taste receptors in Colorado potato beetle
larvae. J. Insect Physiol. 20:1787-1793.
Mittler, T.E. 1967a. Effect of amino acid and sugar concentrations on the food uptake of
the aphid Myzus persicae. Ent Exp. & Appl. 10: 39-51.
Mittler, T.E. 1967b. Gustation of dietary amino acids by the aphid Myzus persicae. Ent.
Exp. & Appl. 10: 87-96.
Munyaneza, J. and J.J. Obrycki. 1998. Development of three populations of
Coleomegilla maculata (Coleptera: Coccinellidae) feeding on eggs of Colorado
potato beetle (Coleptera: Chrysomelidae). Environ. Entomol. 27: 117-122.
Nakajima,N. S. Hiradate and Y. Fujii. 2001. Plant growth inhibitory activity of L-
canavanine and its mode of action. J. Chem. Ecol. 27(1): 19-31.
Nation, J. 2001. Insect Physiology and Biochemistry. CRC Press, Boca Raton. 496pp.
Neishman, O.N. and K. Vulinec. 2001. Florida Crop/Pest Management Profiles:
Eggplant CIR 1264. Pesticide Information Office, Food Science and Human
Nutrition Department, Florida Cooperative Extension Service, Institute of Food
and Agricultural Science, University of Florida. Internet URL:
http://www.edis.ifas.ufl.edu/ BODY_PI045Jitm. Accessed April 2004.


42
+ WmX (QSMX X MX, MX X MX, US XX MX, US ~ X MX, ROW )
+ A (Quota X mx, us )
+ o (X row, us + XX mx, us USMin)
(3.31)
where V, W, A, and erare Kuhn-Tucker multiplier associated with each constraint
representing the imputed marginal value of price of sugar demanded, supplied, that of
Mexican sugar exported under-quota and that of over-quota, respectively. Note that A is
positive in sign and cris negative, reflecting the way their associated constraints are
defined. Kuhn-Tucker conditions are expressed as follows:
=IU2*QDus +IUI+ShifterDus Vus < 0, *QDUS = 0, Qus > 0
(3.32)
= -QU2 *QSus +IUU,+ Shifter*us )+Wus< 0, *QSUs = 0, QSUs > 0
(3.33)
=IM2*QICmx +/Mi+ShifterDmx -Vmx ^ 0, *QKmx 0, QTCmx ^ 0
(3.34)
= -(IMM2 *QSmx +IMM, -t-Shifter*mx )+WMX < 0, *QSmx = 0, QSmx > 0
(3.35)
(3.36)
(3.37)


37
were treated with an extract from potato foliage suggesting induced host preference,
attraction, and dependence on this compound in the extent of sustained feeding and
development. A combination of sensory structures may be involved for the detection of
specific amino acids and host plant compounds, which may explain the selection of
methionine depleted host plants to avoid problems with the CAATCH1 system present in
the midgut of the THW.
The difference in the LC50 for the artificial and natural diets was striking
considering the concentrations were the same. One possible explanation is the
L-methionine on the natural diet was more readily available than that found in the
artificial diet. With the artificial diet, the L-methionine is presumably spread throughout
the diet and would therefore take longer for the THW to ingest enough to adversely affect
the CAATCH1 system. In contrast, the L-methionine was found on the surface of the
leaf in higher concentrations than that of the artificial diet and was also freely available
once ingested. Thus, larvae were exposed to a higher concentration of L-methionine with
less work to digest, resulting in lower survivorship in the same period of time.
The 1.0%L-methionine concentration had the same mortality, feeding and
developmental rates for THW, as did the Btk treatments (Figure 3-9). The 0.3%
L-methionine, 0.5% L-methionine and 0.7% L-methionine treatments were virtually the
same for mortality (Figure 3-9), developmental rate (Figure 3-10) and total leaf material
consumed (Figure 3-11) and statistically the same as the 1.0% L-methionine
concentration and the Btk treatment. The similar mortality rate observed for the higher
concentrations of L-methionine and Btk is encouraging considering the resistance to Bt
seen in many insect species because of reduced receptor activity and binding (Bills et al.


103
Bills, P.S., D. Mota-Sanchez and M. Whalon. 2004. The Database of Arthropods
Resistant to Pesticides. Michigan State University Center for Integrated Plant
Systems. Internet URL: http://www.cips.msu.edu/resistance/nndb/. Accessed
April 2004.
Boucher, T. J. 1999. Using IPM on CPB saves money, insecticides. Yankee Grower
1(2): 7-9.
Bourgis, F., S. Roje, M.L. Nuccio, D.B. Fisher, M.C. Tarczynski, C. Li, C. Herschbach,
H. Rennenberg, M.J. Pimenta, T. Shen, D.A. Gage and A.D. Hanson. 2000. S-
methymethionine has a major role in pholem, sulfer transport and is synthesized
by a novel methyltransferase. Pp. 283-284. IN C. Brunold (ed.), Sulfiir Nutrition
and Sulfur Assimilation in Higher Plants. Paul Haupt, Bern, Switzerland. 427pp.
Bownes, M. and L. Partridge. 1987. Transfer of molecules from ejaculate to females in
Drosophila melanogaster and Drospohila pseudoobscura. J. Insect Physiol.
33(12): 941-947.
Brogdon, W.G and J.C. McAllister. 1998. Insecticide resistance and vector management.
Emerging Infect. Diseases 4(4): 605-613.
Capinera, J.L, F.D. Bennett and D. Rosen. 1994. Introduction: Why biological control
and IPM are important to Florida, pp.3-8. In D. Rosen, F.D. Bennett and J.L.
Capinera (eds.), Pest Management in the Subtropics: Biological Control- a Florida
Perspective. Intercept Limited, Andover, UK. 737pp.
Center, T.D. 1994. Biological control of weeds, Chapter 23. pp.481-521. IN: D. Rosen,
F.D. Bennett, J.L. Capinera, (eds.), Pest Management in the Subtropics:
Biological Control-The Florida Experience. Intercept, Ltd., Andover, Hampshire,
UK. 737pp.
Center, T.D., F.A. Dray and V.V. Vandriver, Jr. 1998. Biocontrol with insects: The
water hyacinth weevils. Florida Cooperative Extension Service, Institute of Food
and Agricultural Sciences, University of Florida. Internet URL:
http://edis.ifas.ufl.edu/scripts/htmlgen.exe?body&DOCUMUMENT_AG01.
Accessed April 2004.
Centers for Disease Control (CDC). 2003. Malaria: General Information. Centers for
Disease Control. Internet URL: http://www.cdc.gov/travel/malinfo.htm. Accessed
April 2004.
Chen, P.S. 1958. Studies on the protein metabolism of Culexpipens L.-I. Metabolic
changes of free amino acids during larval and pupal development. J. Ins. Physiol.
2:38-51.
Cibula, A.B., R.H. Davidson, F.W. Fisk and J.B. LaPidus. 1967. Relationship of free
amino acids of some Solanaceous plants to growth and development of


41
Nematology green and shade houses. Excised leaves were dipped in solutions of
deionized H2O containing different concentrations of methionine and held in the clear
plastic boxes and held at the aforementioned environmental conditions (Figure 3-1).
Additional treatments of proline (1.0%) and Bt-tenebrionis (Novodor FC @12.4 mL/L;
Valent Biosciences, Libertyville, IL) were included as positive and negative controls,
respectively. Survivorship data were pooled from several different trials for data
analysis.
Feeding and Development
To test L-methionine on the developmental rates of CPB, larvae were exposed to
excised eggplant leaves dipped in different concentrations of L-methionine under the
same conditions as the survivorship trials. Additional treatments of proline (1.0%) and
Btt were included as positive and negative controls, respectively. Leaves were scanned
photometrically using the Cl 203 Area Meter with conveyor attachment (CID, Inc.,
Camas, WA) before exposure to the larvae and measuring after leaf consumption. The
difference in leaf areas resulting from the missing leaf tissue was assumed to be the
amount eaten by the developing larvae. Larval head capsule widths were measured at the
time of death or the end of the trial (using an Olympus Tokyo Model 213598
stereomicroscope with an ocular micrometer) as an evaluation of larval development.
Preference Tests
It was unknown if the additional methionine acted to attract or repel larvae.
Neonate larvae were used in the choice tests to determine if there was a preference
between the Control (deionized H20) and the treatments (1.0% L-methionine). Leaves
were obtained from potted plants maintained in the outdoor shade house. The tests


63
Days of Exposure
Figure 5-7. Mortality of yellow fever mosquito larvae exposed to various
concentrations of Beta-alanine (nrotai=240). Data were adjusted
using Abbotts formula for control mortality.


91
Table 5-1. Summary of the U.S. Supply-Demand Analysis
Country
United States
Dependent
variables
Demand
Supply
Independent
variables
Consumption
of sugar
Production of
sugar
Production of
beet sugar
Production
of cane
sugar
Number of
observations
132
43
43
43
Degree of freedom
124
36
36
36
Real retail price of
refined sugar u
-0.2323
(-4.23) **
Real per capita GDP
i)
0.1378
(0.29)
Population
-0.6470
(-0.71)
Dummy variable
(Quarter=l)
-0.0707
(-5.26) **
Dummy variable
(Quarter=2)
0.0136
(1.18)
Dummy variable
(Quarter=3)
0.0941
(10.14) **
Dummy variable for
availability of HFCS
-0.1853
(-3.89) **
Trend
0.0067
(2.94) **
0.0069
(3.20) **
0.0091
(3.08) **
Real retail price of
refined sugar in the
previous year !)
0.1471
(2.35) **
0.2195
(2.19) **
0.0711
(1.44)
Real total farm
production expenses
-0.2316
(-2.19) **
-0.3986
(-2.74) **
-0.0830
(-1.34)
Sugar recovery rate
-0.2351
(-0.57)
-0.5829
(-1.55)
0.1863
(0.58)
Production in the
previous year
0.6204
(5.26) **
0.6755
(5.37) **
0.3672
(3.24) **
Constant
19.8376
(1.52)
5.9033
(2.65) **
7.8421
(3.22) **
5.0365
(6.12)**
Total R~
0.8645
0.8950
0.7253
0.9489
Durbin-Watson 2)
2.0563 (1)
2.5487 (2)
2.0495 (1)
2.0325 (2)
2.0968 (1)
2.0525 (2)
2.0399 (1)
1.9725 (2)
* and **: Significant at 90% and 95% confident level, respectively.
1) Deflated by CPI.
2) Values are after corrected by Yule-Walker method. ( ) corresponds to the order of
lag assigned.


21
sugar mill, this company is able to produce bagged sugar from sugarcane in a single plant
location.
Sugar is one of several commodities protected in the Farm Bill, which includes
rice, cotton, dairy, tobacco, peanuts, grain such as wheat and com, and soybeans (Alvarez
and Polopolus, 2002b). The sugar program operates through a loan program and market
stabilization price (MSP) without production or acreage restraints, differentiating it from
other programs that include target prices or deficiency payments along with export
enhancement programs (Alvarez and Polopolus, 2002b). Loans are issued as non
recourse loans1 and are available to processors of domestically grown sugarcane at a rate
of 18 cents per pound and to processors of domestically grown sugar beets at 22.9 per
pound of sugar, respectively (Haley and Suarez, 2002). Since there are no production
restraints, import quotas and tariffs are the main policy instruments utilized to comply
with the provision that the program has to operate no cost to the government (Alvarez
and Polopolus, 2002b).2 In 1990, the United States committed to accept a minimum
import quota of 1.256 million MT of sugar as a result of GATT.
The U.S. sweetener market has maintained a stable commodity balance, unlike in
Mexico, even after NAFTA was implemented and during the trade dispute with Mexico
over HFCS dumping. One of the reasons is the successful effort by the American Sugar
Alliance (ASA), the sugar producers alliance in the United States, which lobbies for the
U.S. sugar program. The ASA is a strong coalition that includes sugarcane producers,
sugar beet producers, and sugar processors, as well as com producers and HFCS
1 As long as the raw sugar tariff-quota is set higher than 1.5 million short tons (Haley and Suarez, 2002).
2 The no cost provision was not included in Food and Agriculture Improvement and Reform Act (FAIR)
in 1996 (Haley and Suarez, 2002).


CHAPTER 5
EMPIRICAL RESULTS AND INTERPRETATION
This chapter presents and discusses the empirical results for each of three analyses:
(1) demand and supply analysis for both the U.S. and Mexican sugar markets, (2)
bilateral sugar trade analysis, and (3) game theory analysis.
Demand and Supply Analyses
Results for the U.S. demand and supply analysis are summarized in Table 5-1. In
the demand equation, signs of estimates associated with each significant variable were as
expected. Significant estimates at the 95 percent confidence level were associated with
price, the dummy variables for quarter 1 and 3, and the dummy variable for HFCS
availability. The estimated price elasticity of demand was inelastic. The significant
estimate associated with the HFCS dummy variable implies that HFCS replaces sugar as
a substitute in the market to some degree.
In the supply equation for the United States, estimates associated with trend and
production in the previous year (autoregressive term) were significant at the 95 percent
confidence level for all three models, i.e. total, beet and cane sugar supply regression
models. Estimates associated with sugar recovery rate were insignificant in all models.
Estimates associated with price and cost were significant at the 95 confidence level for
total and beet sugar supply regressions, but not for the cane sugar supply equation. Two
possible reasons why cane sugar production does not respond to the refined sugar price
but beet sugar production does are: (1) cane sugar has two steps in the refinery process
while beet sugar has one and (2) sugarcane is a perennial crop while sugar beets is an
75


Table 5-3. Impact of Changes in Mexican Sweetener Market on Pay-offs to the Industries and Nations Welfare [Billion US$]
Scenarios
United States
Mexico
HFCS
industry
revenue
Sugar
industry
revenue
Welfare
(A)
Adjusted
welfare
(A+B-C-D)
Tariff revenue
from Mexican
sugar
(B)
Cost of
price
support
(C)
Cost of
buying up
excess sugar
(D)
Net cost
(B-C-D)
Sugar
industry
revenue
Welfare
Baseline
(status quo)
5.684
(100)
35.713
(100)
354.486
(100)
354.370
(100)
0.093
(100)
0.208
(100)
0
( )
0.116
(100)
21.737
(100)
74.443
(100)
Scenario 2
("P S F")
35.560
(99.57)
353.941
(99.85)
353.902
(99.87)
0.159
(170.79)
0.198
(95.11)
0
(-)
0.040
(34.39)
21.869
(100.61)
76.242
(102.42)
Scenario 3
("A-S-F")
10.132
(178.27)
35.306
(98.86)
353.878
(99.83)
353.975
(99.89)
0.350
(377.20)
0.254
(121.75)
0
(-)
-0.096
(-83.19)
21.322
(98.09)
65.726
(88.29)
Scenario 4
("PA S F")
35.367
(99.03)
353.981
(99.86)
354.080
(99.92)
0.297
(320.15)
0.198
(95.11)
0
(-)
-0.099
(-85.43)
21.314
(98.05)
72.293
(97.11)
Scenario 16
("T S F")
1.591
(27.99)
36.502
(102.21)
354.477
(100.00)
354.477
(100.03)
0.000
(0.00)
0.000
(0.00)
0
(-)
0.000
(0.00)
25.853
(118.94)
79.104
(106.26)
P = Mexico high production; A= Mexico high HFCS adoption; PA= Mexico high production-high HFCS adoption; T= Mexico tax on
HFCS; S= U.S. price support; B= U.S. buying up excess sugar; C= U.S. production control; F= U.S. flexible quota allocation; and M=
U.S. minimum quota allocations to the rest of the world.


28
-6 Total HFCS use per capita Refined sugar use per capita
Figure 2-6. U.S. Refined Sugar and HFCS Use per Capita
Source: USD A, 1993
Figure 2-7. U.S. HFCS Production
Source: USDA, 1993 and 2001a.


10
of the two antimetabolites. This process actually increases the nitrogen intake from the
foodstuff (from the increase of ammonia) (Rosenthal et al. 1976; Rosenthal et al. 1977).
Another insect, the tobacco budworm (Heliothis virescens (Fab.) (Lepidoptera:
Noctuidae)) was able to metabolize far more canavanine then the bruchid beetle larva
ever takes in during its development, suggesting that generalists may have more than a
single detoxification mechanism for compounds they may encounter (Berge et al. 1986).
Metabolism of L-canavanine by the tobacco budworm was attributed to the gut enzyme
canavanine hydrolase, and may have been the result of feeding on canavanine-containing
plants of the Fabaceae (Melangeli et al. 1997).
Essential Amino Acids
In despite of the extensive toxicological research conducted on nonprotein amino
acids, another group of amino acids, the essential ones, has been overlooked. One reason
this avenue for research has not been pursued is that we do not want to give pests
convenient access to an integral part of their diet. The fear of creating a super insect
(that has been provided with compounds that actually aid in its development) is a rational
one. Mittler (1967a; 1967b) found an increase in gustation in Myzus persicae (Sulzer)
(Hemiptera: Aphididae), with amino acid levels as low as 0.2% concentration in a
sucrose solution. Likewise, Sugarman and Jakinovich (1986) found increased gustatory
response to both D-and L-methionine by Periplaneta americana (L) (Blattodea:
Blattidae) adults. Another reason that essential amino acids have not been examined for
use as a pesticide is the knowledge regarding the limited mode of action these compounds
could be involved with (i.e., an active site or systemic response). Recent studies on the
membrane proteins of insects show the possibility of a biophysiological system that can
be exploited for insect control with certain essential amino acids.


126
300,000
250,000
H
s 200,000

o
d 150,000
QJ
E
J2 100,000
o
>
50,000
0
A China production
U.S. production
* European Union production
0 U.S. consumption
A Chiifti consumption
1 Brazil consumption
1999/2000 2000/2001 2001/2002 2002/2003
Year
Figure B-l. Recent Com Production and Consumption for Selected Countries
Source: Com Refiners Association, 2004
Figure B-2. Food and Industrial Com Use in the U.S., 1980-2002
Source: Com Refiners Association, 2004


122
Limitation of the Study and Suggestions for Future Research
HFCS production and consumption are not quantitatively estimated and integrated
into the trade model. This limits the effects on the price and quantity of both sugar and
HFCS from substitution between the two commodities. In the trade model, the rest of the
world is treated as a single homogenous region; however, a country or region such as
Brazil or the European Union that embraces significant sugar demand and supply could
have an individual impact on the U.S. market. Including changes in sugar stocks in each
region as well as flows of sugar-containing goods across borders into the bilateral model
is expected to enhance the quality and depth of the results drawn from the model.


CHAPTER 6
FIELD EVALUATION OF L-METHIONINE AS AN INSECTICIDE
Introduction
The role of methionine in animal systems is well known and only recently
understood in plants. Methionine is required for protein synthesis; it is a precursor to
several important biochemical compounds including ethylene and polyamines, sulfate
uptake and assimilation, and also acts as an activator of threonine-synthase (Giovanelli et
al. 1980; Droux et at. 2000; Bourgis et at. 2000; Zeh et at. 2001). Recently, research has
focused on the transgenic modification of crop plants to overproduce methionine in order
to increase their nutritional quality without affecting other biochemical processes (Zeh et
al 2001). However, little work has been conducted on the effects of exogenous
methionine and it became important to understand the role of externally applied
methionine on plant health.
Furthermore, the application of L-methionine to plants exposed to natural
conditions presents additional problems in terms of how long the residue remains on the
plant. Observations of other experiments using L-methionine revealed the tendency of
this compound to crystallize after the aqueous portion evaporated forming a brittle, crusty
coating that is easily removed. This coating does not appear to interfere with respiration
and transpiration at the concentrations studied (1% and lower). To prevent the loss of
L-methionine from the plants in a natural setting, the adjuvant Silwett L-77 (Helena
Chemical; Collierville, TN) was included in this portion of the study in an effort to
increase residual activity on the plant. Silwet L-77 is a nonionic organosilicate
69


86
coalition of countries, coalition of the sugar industries, and grand coalition that includes
all five entities. Since the inclusion of the U.S. HFCS industry into a coalition is a
determinant factor, both coalitions with and without the U.S. HFCS industry are also
considered.
Pay-offs to the U.S. HFCS industry and the government of Mexicos welfare
fluctuate more than the other three payees (Table 5-6). Among the five strategies facing
Mexico, introduction of tax on HFCS returns the best pay-offs to the Mexican sugar
industry and welfare and thus the Mexican government would always choose this
strategy. Yet, this policy lever may not be acceptable in the international trade
environment. In the following games, Mexicos tax strategy is excluded.
Indexed pay-offs are shown in Table 5-7. Values in corresponding cells are
categorized in four ways: no change (100), gain (over 100, cells shaded in gray), slight
loss (over 95 and under 100), and loss (less than 95, cells shaded with stripes). In this
game, there is neither dominant strategy (a strategy that is chosen over other strategies)
nor pure strategy (a single strategy chosen by the government that brings about
improvement in the industry or industries as well as welfare in its own country: this
strategy is played with probability of 1 for either U.S. or Mexican government). By
inspecting the tendency of pay-offs, some interesting contrasts among payees rise to the
surface without solving mathematically. In the United States, the government always
prefers a production control strategy to the other two but the sugar industry is always
better off with the price support strategy, no matter which strategy Mexico chooses. As
Mexico increases sugar production or HFCS adoption, the U.S. sugar industry likely
loses while the U.S. government and the HFCS industry never become worse off. The


CHAPTER 1
INTRODUCTION
This chapter introduces the research problem for U.S.-Mexico sugar trade.
Background, the researchable questions, and objectives are provided, followed by the
organization of the dissertation.
Background
Trade issues surrounding the world sugar market are often seen as classic examples
in agricultural economics, yet the market still provides us with important questions today.
In the case of the U.S.-Mexico sugar trade, the main issues boil down to two aspects: the
provisions of North American Free Trade Agreement (NAFTA) and the role of high
fructose com syrup (HFCS), a substitute for sugar, in sweetener markets. In the following
section, a summary of this area, focusing on these two aspects, is provided.
NAFTA was implemented in 1994, creating a freer trade environment among
Mexico, the United States, and Canada by eliminating tariffs. Among other regional trade
agreements involving North America, Latin and Caribbean countries, it is the least
ambitious on paper of the major trade agreements, but it has been the most successful
adhering to the negotiated schedule in lowering tariffs (McCoy, 2002). In terms of
agricultural trade between Mexico and the United States, many tariffs were eliminated
immediately while others being phased out over periods of 5 to 15 years (USDA, 2001c).
In addition to a transition period of up to 15 years for certain products, NAFTA has
1


5
such as Dengue fever, yellow fever, and West Nile virus to name just a few, put countless
millions more at risk. It would be dangerous to think that these diseases only occur in
underdeveloped countries and not the United States. Integrated Pest Management
practices also should be adopted for controlling the medical and veterinarian important
insect vectors of these and other diseases.
Biorational Compounds: An Alternative to Traditional Chemical Insecticides
One way to reduce this reliance on traditional chemical pesticides and delay
resistance is by increasing the variety and use biorational compounds. Biorational
compounds are effective against selected pest species but are innocuous to nontarget or
beneficial organisms; and have limited affect (if any) on biological control agents
(Stansly et al. 1996). Biorational compounds include detergents, oils, pheromones,
botanical products, microbes, and systemic and insect growth regulators (Perfect 1992;
Wienzierl et al. 1998). Their safety lies in the low toxicity of the compound to nontarget
organisms and the compounds short residual activity in the field. For example, Bacillus
thuringiensis isrealensis (Bti) currently is one of the most widely used microbial
pesticides for controlling aquatic dipteran pests (i.e., mosquitoes and black flies) because
of its selectivity to this group and minimal nontarget effects (Glare and OCallaghan
1998). However, resistance to Bt products has occurred in many species of lepidoptera
from overuse of Bacillus thuringiensis kurstaki, and in some mosquito species to Bti, thus
showing the need for alternatives to these compounds that are still effective (Brogdon and
McAllister 1998; Marrone and Macintosh 1993). In addition to resistance, other
problems are associated with the use of microbial control agents. Cook et al. (1996)
discussed potential hazards, not properly identified in the planning stages, of
displacement of native microorganisms, allergic responses in susceptible humans and


Bioassays involving the tobacco homworm, Colorado potato beetle (CPB),
Leptinotarsa decemlineata, and the yellow fever mosquito (YFM), Aedes aegypti, were
conducted to determine the insecticidal properties of this compound. L-methionine in
artificial and natural diets resulted in the mortality of 50 to 100% in concentrations of
0.3% and higher for THW and CPB. Feeding rates and larval development also were
affected with reduced levels (>0.1%) of L-methionine. Bioassay trials involving YFM
larvae were similar, concentrations greater than 0.1% L-methionine produced mortality
rates of 70 to 100%. All three species responded better to higher concentrations of L-
methionine than to Bacillus thuringiensis, the most commonly used and commercially
available biorational pesticide.
Field trials and non-target tests also were performed to determine L-methionine
effectiveness under natural settings and safety to other organisms. Eggplant yield was
not reduced by the application of L-methionine, which effectively controlled CPB larvae
on the plants. Furthermore, several beneficial insects that were tested (a predator, a
herbivore, and a parasitoid) were not affected by the addition of L-methionine to their
diets.
Based on these results, L-methionine was found to be effective in controlling
selective agriculturally and medically important insect pest species, yet posed little threat
to the crop plants applied to or to non-target organisms. The use of L-methionine as a
pesticide, its relationship with insects and its possible use in delaying insecticide
resistance were also examined.
xu


100
synthesis of homocysteine to produce methionine to the presence of methionine rich
hexamerins and allophorins and protein synthesis, the role of methionine in plant-insect
interactions may be larger than originally theorized.
The production of methionine overproducing plants could also be used in future
IPM strategies. Preliminary results indicate that genetically modified plants do produce
enough methionine to affect the survivorship of caterpillars feeding on the plant
(unpublished data). This could be used in crops in which improved nutritional quality is
important as well as the insecticidal properties of the additional methionine. However,
there appears to be a sublethal level (0.1%) of L-methionine in which THW and CPB can
tolerate and survive with little mortality (Figures 3-9 and 4-1). Any system that makes
use of a crop that can overproduce compounds like L-methionine would have to be able
to express levels greater than this level to avoid any resistance/tolerance.
This research has also provided more possibilities for the use of compounds such
as L-methionine in the YFM portion of this study. The amino acid Beta-alanine provided
similar levels of control, as did the methionine trials (Figure 5-7). Although unexpected
(as discussed in Chapter 5), it shows that there are several other systems that can possibly
be exploited in controlling some insects.
Further research is necessary to determine if the combination of a compound like
methionine and a pesticide already in use would result in the increase in toxicity or the
decrease in the concentration of pesticide used. If compatibility between methionine and
Bacillus thuringiensis does exists, then it is possible that resistance could be broken in a
given population. For example, if a population of THW started to show resistance to
Bacillus thuringiensis kurstaki then methionine could be used to remove both susceptible


43
dX
^ = -Tmx, us + Vus -WMX <0, *Xmx, us -O, X Mx, us ^ 0
MX,US
dX
MX,US
(3.38)
dX
- -Tus, MX + Vus WMX ^ 0, *X us, MX -0, X US,MX 0
US, MX
dx
US, MX
(3.39)
dxx
= -Tmx, us OQTqtmx, us +Vus -Wmx + <7< 0, -777 *Xmx, us -0,
MX,US
dX
MX,US
X MX, US ^ 0
(3.40)
dX
= -Trow, us -Prow + Vus +cr< 0, 777 *Xrow, us -0, Xrow, us ^ 0
ROW,US
dx
ROW,US
(3.41)
dX
= Prow Wmx ^ 0, *Xmx, row -0, Xmx, row ^ 0
MX,ROW
dX
MX,ROW
(3.42)
By solving equations above, the following conditions must hold at equilibrium:1
Vus = P us Wus PS us
(3.43)
Vmx = P mx Wmx = PS mx
(3.44)
P us B5 mx + Tmx, us+A
(3.45)
P MX P$ US "b Tus, MX
(3.46)
P us P* mx "b Tmx, us + OQTavMx, us -&
(3.47)
P us

(3.48)
PROW ^ P MX
(3.49)
1 Assuming non-zero production and consumption in both the U.S. and Mexico.


9
The Mexican Sugar Industry and Sweetener Market
The Mexican sugar industry has a long history of playing an important role in the
nations economy and policy. A large number of small-scale sugarcane growers and
antiquated sugar milling facilities still remain as the driving force of an industry under the
protection of the government.
Mexicos Sugarcane Production
Most regions of Mexico have a suitable climate for sugarcane production, except
for the northern region of the country where the climate tends to be cooler and drier.
Sugarcane production is widely spread across the southern and coastal regions of the
country under different environmental conditions. Production occurs at altitudes that vary
from sea level to over 1,300 meters (4,333 feet) above sea level; annual average
temperature from 17 to 35 C (from 63 to 95 F); and annual rainfall from 500 mm to
over 3,000 mm (from 20 to over 118 inches) rainfall.
In the 2001/02 crop season, total net sugarcane production in Mexico was 41.5
million MT, with a yield of 4.9 million MT of raw sugar (COAAZUCAR, 2003a).
Among producing regions, the state of Veracruz has the largest production accounting for
38.5 percent of national production (COAAZUCAR, 2003a). The total area harvested in
Mexico was 610,121 ha in 2002 (COAAZUCAR, 2003a), making sugarcane the second
largest agricultural crop by area, following only coffee (maize, wheat, alfalfa, beans, and
oranges follow sugarcane) in 2002 (SAGARPA, 2003). Although sugarcane is a
relatively low maintenance crop, sometimes referred to as the lazy mans crop,
varieties have been developed with higher disease resistance, higher sucrose content, and
lower fiber content, yielding better sugar production. In order to avoid poor yields,
sugarcane fields are usually replanted every six or seven years (Greene, 1998).


87
Results
Coleomesilla maculata
There was virtually no difference between the control and treatment groups for
either the artificial or natural diet tests after correction for control mortality. Mortality
was slightly higher for the control groups than the 1.0% L-methionine treatment (Figures
7-1 and 7-2). Further analysis was not necessary because of the identical numbers.
Neochetina eichhorniae
Total mortality for the treatments was less than 20% for all treatments, with the
individual treatments having similar results (Figure 7-4). Feeding damage ranged
between 2,000 and 4,000 scars per treatment and an average of 10.7 to 16.9 scars per
survivor during the course of the experiment (Figure 7-5). No statistical differences were
observed between the treatment and control groups
Lvsiphlebus testaceipes
In total, 188 and 232 aphid mummies with exit holes were found on treatment and
control plants, respectively. Means for each treatment were not statistically different for
each collection period or overall based on One-way ANOVA (Figure 7-6) with the only
exception being the second and last collection period.
Discussion
In general, L-methionine did not have the same toxic effect on the non-target
organisms tested when compared to the pest species exposed to the compound in
previous chapters. The pink spotted ladybird beetle adults actually showed the least
amount of susceptibility to L-methionine. Survival of the adult beetles was higher in the
1.0% L-methionine treatments than the control for both the artificial and natural diet


116
of HFCS was not welcomed in Mexico. Mexico, a traditional net sugar exporter is now
facing pressures from importing a sugar substitute. Although NAFTA promised Mexico
favored opportunities to export sugar to the U.S. market, implementation of NAFTA also
resulted in opening the door for HFCS consumption in Mexico. The consequences of this
threatening trend of increased HFCS consumption was inscribed on the sweetener history
when Mexico lost the HFCS dumping case with the United States in 2001.
This study attempted to answer five specific questions: (1) What was the impact of
changes in trade regime on the U.S. and Mexican sweetener market since NAFTA was
implemented in 1994?; (2) How much sugar surplus can Mexico generate, how much
sugar will cross the border both under- and over-quota, and what will happen after 2008
when all the restrictions are eliminated on Mexican sugar?; (3) What will be the impact
of changes in the Mexican market situation and how much influence will HFCS adoption
cause in both the U.S. and Mexican sweetener markets?; (4) What will be the impact of
changes in U.S. sugar policy on both the United States and Mexico?; and (5) Is there
alternative sugar policy for the United States to current price support?. In the following,
answers for each question are summarized.
Impact of Changes in Trade Regime
NAFTA brought about mixed impacts on the United States and Mexico and some
are different from what was expected. The Mexican sugar industry has benefited little in
the past ten years under the NAFTA regime. By comparison, NAFTA did not bring about
drastic change to the U.S. sugar market: expanded exports from Mexico have failed to
materialize. Rather, attention was poured into issues of HFCS and its immediate impact
on Mexicos sweetener market. As seen in the trade dispute over HFCS, the Mexican
government struggled to suppress HFCS adoption in its market.


13
Several amino acids were found to initiate the blocking action of ion flow through the
CAATCH1 protein, including threonine, leucine, and methionine, with the latter
producing the greater response, based on CAATCH1 research (Feldman et al. 2000;
Stevens et al. 2002; Quick and Stevens 2001).
Methionine
The amino acid methionine is considered essential in the diets of many organisms.
Methionine is considered an indispensable amino acid in humans. Because the body does
not synthesize it, uptake of methionine must occur in the diet. The recommended daily
allowance of methionine for a healthy lifestyle ranges from 13 to 27 mg/kg/day for
infants to full-grown adults (Young and El-Khoury 1996). This amino acid is linked to a
decrease in histamine levels, increased brain function, and is found in a variety of
sources; with the highest concentration in various seeds, greens, beef, eggs, chicken, and
fish (Dietary Supplement Information Bureau 2000). Recently, research has centered on
the genetic modification of crop plants to overproduce methionine to increase its
nutritional quality (Zeh et al. 2001). Wadsworth (1995) discussed using methionine as a
feed supplement, as an aid in the therapy of ketosis in livestock, and as a treatment for
urinary infections in domestic pets. Onifade et al. (2001) examined the use of housefly
larvae as protein foodstuffs, and found an increase in body weight gain and erythrocyte
counts in rats whose diets were supplemented with fly larvae and methionine. Likewise,
Koo et al. (1980) suggested dry face fly pupae could be used as a dietary supplement and
foodstuff extender for poultry because of the high concentration of methionine. The
environmental safety of methionine is well known, as it poses no risk to vertebrates due
to a rather high oral LD50 of 36g/kg~' observed in rats (Mallinckrodt Baker 2001) and also


101
and resistant alike because of the difference in mode of action. Once the population was
reduced, and the corresponding resistant genotype, Btk could be used once more at a
lower concentration, closer to that of the susceptible population. This system could also
be used for the reduction of Bt toxin resistance in the CPB and YFM if the compounds
are compatible.
In conclusion, it appears that L-methionine can be used as an insecticide to
control insect pests of economic and medical importance. The target site (CAATCH1) is
known and found in the midgut/hindgut (presumed) in at least three pest species (tobacco
homworm, Colorado potato beetle and the yellow fever mosquito) and possibly more.
The compound (L-methionine) is a safe compound that is already used for livestock feed
supplements, has very low mammalian toxicity, and is compatible with insecticide
application systems. Non-target organisms were not affected with the application of L-
methionine, further supporting its use as a biorational insecticide. With increasing
resistance to current insecticides in the study organisms, alternatives such as L-
methionine are needed now more than ever to further support of Integrated Pest
Management strategies.


75
Figure 6-3. Mortality of Colorado potato beetle adults exposed to excised eggplant
leaves treated with L-methionine and the adjuvant Silwett L-77
(nTotai=120). Data corrected for control mortality using Abbotts formula.
Note the overlap in trend lines for the Control treatments and 0.1%L-methk>nine
treatment.



PAGE 1

ANALYSIS OF U.S.-MEXICO SUGAR TRADE: IMPACTS OF THE NORTH AMERICAN FREE TRADE AGREEMENT (NAFTA) AND PROJECTIONS FOR THE FUTURE By DAISUKE SANO A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2004

PAGE 2

Copyright 2004 by Daisuke Sano

PAGE 3

To my parents and sister

PAGE 4

ACKNOWLEDGMENTS I would like to express my gratitude to Dr. Thomas H. Spreen, chair of my supervisory committee, for providing me with resources throughout my program of studies. My dissertation would not have been possible without his guidance, unflagging enthusiasm and helpful ideas. I also would like to express my gratitude to Dr. Lisa A. House, cochair of my supervisory committee, for her generous and patient support during my course of studies. Under her supervision, my research has been completed in an efficient and successful manner. Special appreciation is extended to Dr. Chris O. Andrew for his encouragement throughout the program and for broadening my horizons. His inspiration has been a compass after the direction of my career had been shifted. I am also deeply grateful to Dr. Luis R. Garcia for providing me with insights and detailed data which were indispensable for this study; to Dr. Terry L. McCoy for his contributing to the depth of my study; and to Dr. Kenneth L. Buhr for his sharing suggestions for the breadth of my study. Kind support from Dr. Jeffrey R. Burkhardt and staff in the department for their various services are also appreciated. I cannot fail to thank John S. Lander, who opened my eyes to studying overseas, for his unconditional support and wisdom. Friendship from my colleagues in the program is a cherished treasure. Last but not least, the financial support received from the Food and Resource Economics Department throughout my program has been greatly appreciated. iv

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TABLE OF CONTENTS page ACKNOWLEDGMENTS iv LIST OF TABLES vii LIST OF FIGURES ix ABSTRACT xii CHAPTER 1 INTRODUCTION 1 Background 1 Problem Statement 5 Researchable Questions 5 Objectives 6 Organization of the Study 6 2 SUGAR INDUSTRIES AND SWEETENER MARKETS IN THE UNITED STATES AND MEXICO 8 The Mexican Sugar Industry and Sweetener Market 9 Mexico's Sugarcane Production 9 Mexico's Sugar Production 10 The Mexican Sugar Industry and Government Involvement 12 Mexico's Sugar Consumption 14 Mexico as a Sugar Exporter 15 Development and Adoption of High Fructose Com Syrup 17 The U.S. Sugar Industry and Sweetener Market 19 3 CONCEPTUAL AND THEORETICAL FRAMEWORK 32 Conceptual Framework 32 Theoretical Framework 35 Sweetener Market Analysis 35 Sweetener demand 35 Sweetener supply 36 U.S.-Mexico Bilateral Sugar Trade System 37 V

PAGE 6

4 EMPIRICAL MODELS AND DATA SOURCE 47 Empirical Models 47 U.S. Sweetener Demand Model 47 Mexican Sweetener Demand Model 48 U.S. Sweetener Supply Model 50 Mexican Sweetener Supply Model 52 U.S.-Mexico Bilateral Sugar Trade Model 53 Simplifying assumptions 54 Model calibration 56 Simulated Scenarios 58 Game Theory Analysis ^1 Sources of Data ^-^ 5 EMPIRICAL RESULTS AND INTERPRETATION 75 Demand and Supply Analyses 75 Bilateral Sugar Trade Analysis 77 Game Theory Analysis ^5 6 CONCLUSIONS AND IMPLICATIONS FOR POLICY 115 Conclusions and Implications for Policy 115 Impact of Changes in Trade Regime 116 Mexico's Export Potential 1 17 The Impact of Changes in Mexican Market Situation 1 17 The Impact of Changes in U.S. Sugar Policy 118 Alternative Sugar Policy by the United States 120 Limitation of the Study and Suggestions for Future Research 122 APPENDIX A MAJOR EVENTS IN THE SUGAR INDUSTRY HISTORY IN MEXICO AND THE U.S 123 B CORN STATISTICS 125 C DERIVATION OF INVERSE LINEAR EQUATIONS 128 BIOGRAPHICAL SKETCH 136 vi

PAGE 7

LIST OF TABLES Table ms& 2-1 Cane Sugar Production in Selected Countries, 1997-2000 Average 25 2-2 World Sugar Consumption in 2000 27 2-3 Quota and Tariff Schedule Imposed on Mexican Sugar Exported to the U.S 27 4-1 Assumptions for Baseline (Status quo) Scenario 66 4-2 Assumptions for Mexican Sweetener Market Situations 67 4-3 Assumptions for U.S. Sugar Policies 69 4-4 Listing of Examined Scenarios 70 4-5 Strategies for the Sugar Trading Game 71 4-6 Data Sources for U.S. Demand 72 4-7 Data Sources for Mexican Demand 72 4-8 Data Sources for U.S. Supply 73 4-9 Data Sources for Mexico Supply 73 410 Data Sources for Miscellaneous 74 51 Summary of the U.S. Supply-Demand Analysis 91 5-2 Summary of the Mexican Supply-Demand Analysis 92 5-3 Impact of Changes in Mexican Sweetener Market on Pay-offs to the Industries and Nation's Welfare [Billion US$] 101 5-4 Impact of Changes in U.S. Price Stabilization Policy on Pay-offs to the Industries and Nation's Welfare [BilHon US$] Flexible Quota Allocations 102 5-5 Impact of Changes in U.S. Quota Allocation Policy on Pay-offs to the Industries and Nation's Welfare [Billion US$] Minimum Quota Allocations 103 vii

PAGE 8

5-6 Pay-off Matrix for the Trade Policy Game [Billion US$] 107 5-7 Indexed Pay-off Matrix for the Trade Policy Game without Coalitions [Baseline = 100] 108 5-8 Indexed Pay-off Matrix for the Trade Policy Game Played by Two Coalitions of Countries without the U.S. MFCS industry [Baseline=100] 109 5-9 Indexed Pay-off Matrix for the Trade Policy Game Played by the Industry Coalition and the Government Coalition without the U.S. HFCS industry [Baseline=100] HO 5-10 Indexed Pay-off Matrix for the Trade Policy Game Played by the Grand Coalition without the U.S. HFCS industry [Baseline=100] 11 1 5-11 Indexed Pay-off Matrix for the Trade Policy Game Played by Two Coalitions of Countries with the U.S. HFCS industry [Baseline=100] 112 5-12 Indexed Pay-off Matrix for the Trade Policy Game Played by the Industry Coalition and the Government Coalition with the U.S. HFCS industry [Baseline=100] 113 5-13 Indexed Pay-off Matrix for the Trade Policy Game Played by the Grand Coalition with the U.S. HFCS industry [Baseline=100] 1 14 C-1 Coefficients for Inverse Linear Functions -U.S.129 C-2 Coefficients for Inverse Linear Functions -Mexico130 viii

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LIST OF FIGURES Figure page 2-1 Distribution of Individual Farm Size of Mexican Sugarcane Growers 24 2-2 Map of Mexico's Sugar Producing and Processing States 24 2-3 Annual Rainfall and Irrigation Rate in Sugarcane Fields 25 2-4 Mexican Sugar Production, 1988-2002 26 2-5 Main Use of Sugarcane Derivatives in Mexico 26 2-6 U.S. Refined Sugar and HFCS Use per Capita 28 2-7 U.S. HFCS Production 28 2-8 U.S. HFCS Supply 29 2-9 U.S. HFCS Export 29 2-10 Transition of U.S. HFCS Export 30 2-11 Consumption of Sugar and HFCS per Capita in Mexico 30 212 Transition of Prices of Sugar and HFCS 31 31 Conceptual Framework for the Analysis in this Study 45 32 Two-country Trade Model (a) with Quota System and (b) without Quota System. 46 41 Image of Model Calibration 65 42 Forecasted Indirect Sugar and HFCS Consumption in Mexico under Alternative Scenarios 68 51 U.S. Sugar Import Forecast (Scenario 1 "Baseline") 93 5-2 U.S. Sugar Import Forecast (Scenario 2 "P-S-F") 93 5-3 U.S. Sugar Import Forecast (Scenario 3 "A-S-F') 94 ix

PAGE 10

5-4 U.S. Sugar Import Forecast (Scenario 4 "PA-S-F") 94 5-5 U.S. Sugar Import Forecast (Scenario 13 "PA-S-M") 95 5-6 U.S. Sugar Import Forecast (Scenario 8 "PA-B-F") 95 5-7 U.S. Sugar Import Forecast (Scenano 12 "PA-C-F") 96 5-8 U.S. Sugar Import Forecast (Scenario 16 "T-S-F") 96 5-9 Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 4 "PA-S-F') 97 5-10 Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 8 "PA-B-F") 97 5-11 Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 12 "PA-C-F") 98 5-12 Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 16 'T-S-F") 98 5-13 Forecasted Sugar Demand and Supply for both the United States and Mexico (Scenario 1 "Baseline") 99 5-14 Forecasted Sugar Demand and Supply for both the United States and Mexico (Scenario 4 "P-S-F") 99 5-15 Forecasted Equilibrium Sugar Prices in the United States and Mexico (Scenario 14 "PA-B-M") 100 5-16 Forecasted EquiHbrium Sugar Prices in the United States and Mexico (Scenario 15 "PA-C-M") 100 5-17 Absolute Effects of Production Improvement and HFCS Adoption on Pay-off to the Mexican Sugar Industry 104 5-18 Absolute Effects of Production Improvement and HFCS Adoption on Pay-off to Mexico's Welfare 105 5-19 Relative Gain and Loss of Pay-offs to the Mexican Sugar Industry and Welfare Caused by Production Improvement and HFCS Adoption 106 B-I Recent Com Production and Consumption for Selected Countries 126 B-2 Food and Industrial Com Use in the U.S., 1980-2002 126 B-3 Com Price (No.2 Yellow) in Chicago Market, 1981-1998 127

PAGE 11

B-4 Exports of Products Made from Com in 2002 127 xi

PAGE 12

CHAPTER 1 INTRODUCTION This chapter introduces the research problem for U.S. -Mexico sugar trade. Background, the researchable questions, and objectives are provided, followed by the organization of the dissertation. Background Trade issues surrounding the world sugar market are often seen as classic examples in agricultural economics, yet the market still provides us with important questions today. In the case of the U.S.-Mexico sugar trade, the main issues boil down to two aspects: the provisions of North American Free Trade Agreement (NAFTA) and the role of high fructose com syrup (HFCS), a substitute for sugar, in sweetener markets. In the following section, a summary of this area, focusing on these two aspects, is provided. NAFTA was implemented in 1994, creating a freer trade environment among Mexico, the United States, and Canada by eliminating tariffs. Among other regional trade agreements involving North America, Latin and Caribbean countries, it is the least ambitious on paper of the major trade agreements, but it has been the most successful adhering to the negotiated schedule in lowering tariffs (McCoy, 2002). In terms of agricultural trade between Mexico and the United States, many tariffs were eliminated immediately while others being phased out over periods of 5 to 15 years (USDA, 2001c). In addition to a transition period of up to 15 years for certain products, NAFTA has 1

PAGE 13

special safeguards to protect import-sensitive crops, including sugar, which are defined under side agreements between Mexico and the United States (USDA, 2001c). When trading sugar with the United States, the Mexican sugar industry faces two counteracting conditions under the NAFTA regime: increased access to the U.S. market ^ which would facilitate sugar exports at favorable prices; and the pressure of increased imports of MFCS from the United States which have been gaining an increasing share of Mexico's sweetener market since 1994. Under the provisions of NAFTA, both an overquota tariff for Mexican sugar which enters into the United States and a tariff on exported HFCS which enters the Mexican market are regulated in such a way that both tariffs will be reduced to zero by 2008 and 2004, respectively. In addition to the rules of the tariffs, Mexican sugar is subject to U.S. import quota allocations. Mexico is allowed to access two kinds of quotas, depending on Mexico's domestic balance in the sweetener market: if Mexico's sugar production exceeds its sweetener consumption (the sum of sugar and HFCS consumption in two consecutive years -"net surplus sweetener producer status"), Mexico receives 25,000 MT of sugar import quota; and if not, Mexico receives 7,258 MT of quota. Additionally beginning in 2000, the sugar import quota expands from 25,000 MT to 250,000 Mt as long as Mexico satisfies the conditions of a net surplus sweetener producer. Mexico can export over the 7,258 MT quota without attaining net surplus sweetener, but any sugar exported in this scenario would be subject to taxation in the form of tariffs as mentioned above. In 2008 when all the restrictions, i.e. both tariffs and quotas, are lifted, Mexico will have free and unlimited access to the U.S. sugar market. In Mexico, the sugar industry has played an important role in the economy and the politics of the country. In spite of experiencing drastic economic and political changes.

PAGE 14

3 including NAFTA, devaluation, privatization of the sugar cane processing industry in the 1990s, and several changes in the policy regime, sugar production has shown steady expansion over the past 10 years: Mexican sugar production expanded from 3.2 million MT in 1990 to 4.7 million MT in 2000 (COAAZUCAR, 2003a). A significant amount of surplus sugar destined to export has been generated since 1995, ranging from 200,000 MT in 1995 to over 1.1 million MT in 1998 (COAAZUCAR, 2003a). These records may appear favorable; however, Mexico stood to benefit little from NAFTA. From 1996 through 1999 Mexico successfully received a 25,000 MT import quota as a result of attaining net surplus producer status, yet it did not enjoy the expanded quota (250,000 MT) from 2000 through 2002 (USDA, 2003a), the amount equivalent to 20 percent of the U.S. minimum sugar import requirement under GATT, because Mexico's production fell short relative to its sweetener consumption. This indicates that Mexico missed the opportunity to export sugar under-quota even though it generated a significant surplus. Combined with a slump in production that occurred in 1999 and 2000, the Mexican sugar industry underwent an economic crisis. In September 2001, the Mexican government expropriated 27 of 60 of Mexico's functioning sugar mills in order to maintain the industry (USDA, 2002b). Today, the circumstances surrounding the sugar industry remain unfavorable. At the industry level, many mills are financially vulnerable and suffer from low efficiency of production due to old technology or poor infrastructure. Foreign investment has not been successfully encouraged to provide capital for needed investments in new capital equipment. At the farm level, production efficiency is low due to fragmented farmland, which is a result of the Ejido system (Mexico's agrarian law) and social security program specifically tailored to sugar cane growers. Lack of credit

PAGE 15

4 and old technology also contribute to low productivity. Although the price of sugar at the wholesale level has been privatized, the sugar price paid to growers is still controlled by a government agency, and hence farmers have little incentive to grow sugar cane other than to receive social benefits from the Mexican sugar program. At the national level, the Mexican government faces a dilemma between gaining competitiveness in the international market and maintaining social stability through offering employment and financial supports to the livelihood of a large number of growers and related workers. Overall, there has been little benefit to the Mexican sugar industry resulting from NAFTA. The U.S. sugar market, where a large quantity of sugar is traded by a large number of sellers, has maintained commodity balance by assigning tariffs and import quotas to foreign sellers and maintaining domestic price support through the U.S. sugar program. As a result of GATT, the United States committed to accept a minimum import quota of 1.256 million MT of sugar in 1990; however, the U.S. sugar market has been maintained unchanged until today through successful lobbying efforts by the American Sugar Alliance (ASA), the sugar producers' primary alliance. In the meantime, HFCS had been gaining its share in the U.S. sweetener market since the early 1970s when commercial production of HFCS became possible by the advancement of wet-milling technology. Today, more than 50 percent of caloric sweetener consumption in the United States is derived from com syrup including HFCS (Congressional Research Service, Library of Congress, 1999). A similar phenomenon appears to be beginning in Mexico. The implementation of NAFTA resulted in opening the door for HFCS consumption in Mexico where nearly all caloric sweetener

PAGE 16

5 consumption was derived from domestically-produced sugar before 1994. Reflecting this threatening trend of replacing domestic sugar consumption with HFCS, in 1996, the Mexican government imposed tariffs on HFCS claiming that U.S. companies were dumping HFCS at an unfair price and affecting the export volume and value of Mexican sugar. This action evolved into a trade dispute between the United States and Mexico and ended when the WTO panel ruled against Mexico's claim (Garcia Chaves et al., 2002 and 2004). Overall, NAFTA has not brought about significant changes in the U.S. sugar market because the Mexican exporters have been unable to significantly expand shipments to the United States. Rather, attention was poured into issue of HFCS and its immediate impact on the Mexico's sweetener market. In this study, the direction of U.S. -Mexico sugar trade is examined using quantitative methods, with close attention to issues related to NAFTA and HFCS adoption in Mexico. Demand and supply analyses in both countries and a bilateral trade model using mathematical programming provide insights for the market balance in the future including political implications. Aggregated results from various simulations on the trade model are examined using a game theory analysis to investigate possible policy recommendations through assessing gainers and losers in sugar trade. Problem Statement The future outlook for the U.S.-Mexico sweetener market needs to be quantitatively analyzed in a manner that includes influential factors such as trade agreements under NAFTA; trends in HFCS consumption in Mexico; and other related economic and political issues in the sweetener markets. Researchable Questions The study attempts to answer the following set of questions.

PAGE 17

6 1. What was the impact of changes in the trade regime in the U.S. and Mexican sweetener market since NAFTA was implemented in 1994? 2. How much surplus sugar can Mexico generate and how much sugar will cross the border both underand over-quota? What will happen after 2008 when all the restrictions are eliminated on Mexican sugar? 3. What will be the impact of changes in Mexico's market on both the United States and Mexico? How much influence will HFCS adoption cause in both the U.S. and Mexican sweetener markets? 4. What will be the impact of changes in U.S. sugar policy on both the United States and Mexico? 5. Is there alternative sugar policy for the United States to current price support? Objectives The primary objective of the study is to develop a bilateral trade model of the U.S.Mexico sugar industry that reflects provisions of NAFTA, as well as related market conditions in order to forecast the outlook of the sweetener market through various simulations, encompassing hypothetical changes in Mexican sweetener situations and U.S. sugar policy. Secondly, the study aims to provide policy recommendations by examining aggregated results from these simulations, paying attention to identify gainers and losers under different scenarios. By doing so, the study hopes to illustrate conflicts of interest among the various players in the U.S.-Mexico sweetener market. Organization of the Study The remainder of the dissertation is organized as follows. In chapter 2, the sugar industries in both the United States and Mexico are introduced in the context of the sweetener market in each country as well as the integrated market, paying close attention to historical and political perspectives. In chapter 3, the conceptual and theoretical

PAGE 18

7 framework employed to analyze U.S. -Mexico sugar trade is presented. In chapter 4 and 5, empirical procedures as well as data set used in the study and the results form the empirical study are presented. Lastly, conclusions and implications for policy are discussed in chapter 6.

PAGE 19

CHAPTER 2 SUGAR INDUSTRIES AND SWEETENER MARKETS IN THE UNITED STATES AND MEXICO Sugar, one of the basic commodities with a long history of utilization, is traded in mature markets in many parts of the world with established business practices and networks. The recent trend towards freer markets in the international trade area has not left the industry unchanged. The sugar industries in the United States and Mexico are not exceptions. They have experienced more changes in the face of this recent trend towards rapid trade liberalization. In fact these two industries have become more economically inseparable than ever before as the sweetener markets in the United States and Mexico have been integrated under North American Free Trade Agreement (NAFTA). In this chapter, sugar industries in both the United States and Mexico are examined in the context of the sweetener market as well as the integrated market, paying close attention to historical and political perspectives. First, the Mexican sugar industry and sweetener market are introduced with fundamental characteristics of the structure and government involvement. The status of Mexico as a sugar exporter is also presented in conjunction with Mexico's relation to the U.S. market under the provisions of NAFTA. Next, the development and adoption of high fructose com syrup (HFCS) is presented. Lastly, the U.S. sugar industry and sweetener market is introduced with emphasis on the current political environment surrounding that market. 8

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9 The Mexican Sugar Industry and Sweetener Market The Mexican sugar industry has a long history of playing an important role in the nation's economy and policy. A large number of small-scale sugarcane growers and antiquated sugar milling facilities still remain as the driving force of an industry under the protection of the government. Mexico's Sugarcane Production Most regions of Mexico have a suitable climate for sugarcane production, except for the northern region of the country where the climate tends to be cooler and drier. Sugarcane production is widely spread across the southern and coastal regions of the country under different environmental conditions. Production occurs at altitudes that vary from sea level to over 1,300 meters (4,333 feet) above sea level; annual average temperature from 17 to 35 C (from 63 to 95 F); and annual rainfall from 500 mm to over 3,000 mm (from 20 to over 118 inches) rainfall. In the 2001/02 crop season, total net sugarcane production in Mexico was 41.5 million MT, with a yield of 4.9 million MT of raw sugar (COAAZUCAR, 2003a). Among producing regions, the state of Veracruz has the largest production accounting for 38.5 percent of national production (COAAZUCAR, 2003a). The total area harvested in Mexico was 610,121 ha in 2002 (COAAZUCAR, 2003a), making sugarcane the second largest agricultural crop by area, following only coffee (maize, wheat, alfalfa, beans, and oranges follow sugarcane) in 2002 (SAGARPA, 2003). Although sugarcane is a relatively low maintenance crop, sometimes referred to as the "lazy man's crop," varieties have been developed with higher disease resistance, higher sucrose content, and lower fiber content, yielding better sugar production. In order to avoid poor yields, sugarcane fields are usually replanted every six or seven years (Greene, 1998).

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10 The total number of sugarcane growers is reported as approximately 158,000 (COAAZICAR, 2003b), which is equivalent to roughly 2 percent of the total labor force in the agricultural sector (INEGI, 2003). If related workers such as sugarcane cutters, cane-transport employees, factory workers and administrative, and technical and management personnel are included, total employment in the sugar sector exceeds 1,000,000 (Garcia Chaves et al., 2002) and accounts for more than 14 percent of agricultural labor. Land area per grower ranges from less than 1 ha, which accounts for 3.6 percent of the total sugarcane area, to over 15 ha, which accounts for 17.5 percent of the land, averaging 3.9 ha per grower (COAAZUCAR, 2003b). When the number of growers is allotted to each land size category, a skewed distribution is revealed along the land scale spectrum with many small-land holders and a few large-land holders (Figure 2-1). A large number of small-scale sugarcane growers were created as a result of the Mexican revolution and the sugar program that evolved after the revolution: the communal land (ejido) has been divided and distributed among farmers since the revolution and the Mexican sugar program offers social security and medical services to each grower proving to be a large incentive for farmers to grow sugarcane. Mexico's Sugar Production There are 60 operating sugar mills located across 15 states in the nation (Figure 22). Sugar mills are responsible not only for milling sugarcane, but also supervising sugarcane cultivation and organizing the harvest. This includes inspecting and advising on cultivation, scheduling harvest dates, pooling and arranging laborers, and providing trucks and drivers for the harvest.

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11 In the 2001/02 crop season, average sugarcane yield was 70.32 MT per hectare in Mexico, yielding 4,872,388 MT of sugar (COAAZUCAR, 2003a). Imgation is one of the factors which influences cane yield; however, irrigation systems are found only in the area where less rainfall is expected (Figure 2-3): 30 percent of total sugarcane area has no irrigation system and 25 percent has full irrigation system (COAAZUCAR, 2003c). Harvest is the most labor-intensive part of sugarcane production; the harvest season lasts for about six months starting between November and January and ending in June in most regions, depending on weather and size of enterprise. Harvest competes for grower's labor with other winter crops since many growers are also engaged in production of crops such as maize, vegetables and citrus. Most of the harvest is carried out manually; only 9 percent of total sugarcane processed at mills is harvested by machine; 27 out of 60 mills do not employ a machine harvester at all; however, a cane loader is used in most cases (COAAZUCAR, 2003d). Since the mills own machine harvesters, cane loaders, and trucks, growers do not need to own them; however, it means growers have no means to harvest and sell their sugarcane without the mill's assistance and coordination. Similar situations regarding grower's capacity in harvest are found in other crops such as citrus. Upon harvest, sugarcane is bought by mills from growers and processed into sugar. Sugarcane quality is vital to the sugar production process; high sucrose content cane leads to high sugar production. Yet, it is often the case in Mexico that trucks endure long waiting times to unload cane due to limited milling capacities. The average wait time observed in 2001 was almost 30 hours across mills (COAAZUCAR, 2003e). The longer trucks wait, the lower the quality of sugarcane becomes. Scattered and fragmented

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sugarcane fields and poor road conditions also contribute to longer transportation time, and hence decreased sugarcane quality. Mexico's sugar production was approximately 4.8 million MT, raw equivalent, in 2002, ranking it seventh among all cane sugar-producing nations; Brazil and India are by far the largest cane sugar-producing nations, followed by China, the United States, Thailand and Australia (Table 2-1). Production in Mexico has been increasing for the past few decades (Figure 2-4). The Mexican Sugar Industry and Government Involvement Mexico's sugar production accounts for 0.5 to 0.7 percent of its gross domestic product (Garcia Chaves et al., 2002; Farm Foundation, 2003). Since privatizing in the late 1980s, mills have neither successfully accumulated capital nor renewed their equipment leaving the industry financially vulnerable. In 2001, the Mexican government expropriated 27 mills, which represented approximately 50 percent of sugar production in Mexico. In February 2002, the Government of Mexico announced a National Sugar Policy for 2002 2006 which included a series of shortand longterm measures to help Mexico's ailing sugar industry with the main objective of regulating the sugar market and making the sugar sector profitable (USDA, 2002b). Today the sugar industry remains important in Mexico because it is considered crucial for maintaining social stability due to the large number of growers and related workers. Among the public organizations that deal with the Mexican sugar industry, Commite de la Agroindustria Azucarera (COAAZUCAR) plays an important role by monitoring and compiling sugarcane and sugar production data at each mill. Although the industry has been privatized, COAAZUCAR is in charge of determining the cane price. It took over the task from the former body, Azucar, S.A. which was dismantled in 1991

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13 when sugar mills were privatized. Growers are paid by a fixed portion of the reference sugar price calculated from aggregated sugarcane harvested and processed in the specific mill. Details for the cane price setting formula are shown in equations [2.1] and [2.2] (Garcia Chaves et al, 2004): Cane price / ton = (KARBE/ ton of cane)*(Price of KARBE)*(0.57) (2.1) KARBE/ ton of cane = (Pol) *(FF)*(FP)*(EBF)*(TF) (2.2) where KARBE is kilogram of recoverable standard sugar basis (Pol 99.4 percent) for net ton of cane; Pol is polarization of cane (apparent percentage of sucrose in cane); FF is the fiber factor; FP is the purity factor; EBF is mill efficiency; and TF is the transformation factor. As seen in equation [2.1], currently growers are paid 57 percent of the wholesale price per kilogram of standard sugar. Although in 1991 growers began to be paid according to the quality of cane produced as opposed to solely on weight as decreed in the amendments to Decreto Canero, the Sugarcane Growers Law, growers have little incentive to produce higher quality cane. The cane price is capped at 57 percent of average quality cane for the specific mill, not a price reflecting each grower's sugarcane. Thus, a main incentive for Mexican sugarcane growers is to receive social benefits and medical services from the government as opposed to producing quality cane. The wholesale price of sugar produced at mills has been liberalized since 1997; however, it still quotes a reference price calculated based on the formula published by the secretariat of Commerce (SECOFI). The price is determined by considering both the recent domestic price and the expected export price, which is the composite of the U.S. and world price (Garcia Chaves et al, 2004), as shown in equation [2.3].

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Pr = a.Pn + (l-a)*P'x (2-3) where Pr is wholesale price per kilogram of standard sugar to be used as the reference for cane payment during the harvest; a is expected portion of harvest to be consumed nationally (a equals one if expected consumption is greater than expected production); Pn is reference price for standard domestic sugar calculated by comparing the OctoberSeptember average price of the previous year with the current year; (1-a ) is expected surplus as a portion of production; and P% is expected export price of sugar, which is calculated by a weighed average of the U.S. price (Contract No. 14) and the world price (Contract No. 11) with corresponding export quantities. The Instituto Medical y Seguro Social (IMSS), the Mexican Social Security Institute, provides both pension and medical services to all the employees in Mexico as well as to small farmers who grow sugarcane (Greene, 1998). It is often the case that IMSS clinics are located to the next to the mills (Greene, 1998). Borrel (1991) as well as Buzzanell and Lord (1995) have pointed out this special relationship as a source of inefficiency in the Mexican sugar industry. Mexico's Sugar Consumption Since Mexico is neither an importer of sugar nor producer of sugar beets, sugar consumed in the country is derived solely from sugarcane grown domestically. The main use of sugarcane derivatives are shown in Figure 2.5. Sugarcane requires two steps in the refining process to obtain the refined sugar used by households and industries. Raw sugar, which is the product of the extraction process, is either stored or exported to other countries where refinery facilities are available. Mexico consumes two kinds of refined sugar, called standard sugar and white sugar. Standard sugar has a slight impurity.

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15 whereas white sugar is as purely refined as ordinary refined sugar found elsewhere. Both kinds of refined sugar are indistinctly consumed by households as well as domestic bulk sugar users such as soft-drink manufacturers and confectionaries. This unique aspect is different from the U.S., where a purer form of sugar, equivalent to white sugar in Mexico, is what most households purchase. Molasses, the heavy dark viscous liquid residue discharged by the centrifugal from which no more sugar can be obtained by simple means (Polopolus and Alvarez, 1991) is utilized in rum making in Mexico. Mexico's national total sugar consumption was approximately 4,500,000 MT, raw equivalent, in 2001 (FAO, 2003), making it the seventh largest sugar consumption country/ region in the world (Table 2-2). Per capita sugar consumption was 44.6 kg (98.5 lbs.), raw equivalent, in 2000; relatively high among other major sugar-producing countries (Table 2-2). When other kinds of sweeteners such as HFCS are included, the largest per capita consumer is the United States, followed by Cuba, Brazil, Australia, and Mexico (Table 2-2). Mexico as a Sugar Exporter Surplus raw sugar is either exported to the world market, primarily to the U.S. market due to higher price, or stored as stock. The magnitude of sugar exports from Mexico depends on the size of surplus determined by domestic production-consumption balance and the quota limitation imposed on all sugar imports entering the U.S. market. Geographically Mexico holds a sugar export advantage to the United States. One of the main shipping ports in Mexico is Veracruz, located facing the Gulf of Mexico, only 830 miles from New Orleans and 1 130 miles from South Florida where sugar refinery facilities are located. Furthermore, the state of Veracruz produces approximately 40 percent of Mexico's domestic production of sugar.

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16 Another advantage Mexico attained is the preferred trade conditions under the North American Free Trade Agreement (NAFTA) implemented in 1994. NAFTA created a freer trade environment among Mexico, the United States and Canada by eliminating tariffs. In terms of agricultural trade between Mexico and the United States, many tariffs were eliminated immediately, while others were scheduled to be phased out over periods of 5 to 15 years (USDA, 2001a). Mexico benefited from exporting its surplus sugar to the U.S. market at a higher price and at a lower tariff rate which will be reduced each year and eventually gives Mexico free and unlimited access to the U.S. market beginning in 2008 (Table 2-3). The NAFTA agreement is a double-edged sword to the Mexican sugar industry, however, creating counteracting conditions in the Mexican sweetener market: one is the increased access to the U.S. market and the other increased access of high fructose com syrup (HFCS) from the United States. The quantity of duty-free sugar exported from Mexico is limited by quotas which vary depending on Mexico's balance in the domestic sweetener market not the sugar market; Mexico receives a larger quota if its domestic sugar production exceeds domestic consumption of sweetener, including HFCS (called "net surplus sweetener producer status") in two consecutive years. At the same time, the agreement gives U.S. HFCS producers free access to the Mexican sweetener market beginning in 2004 as a tariff imposed on HFCS imported from the United States is also being phased out. If Mexico fails to attain net surplus sweetener producer status, it receives a sugar import quota of only 7,258 MT rather than 250,000 MT and thus most of Mexican sugar exported to the U.S. market is subject to over-quota tariffs until 2008 when all the restrictions on Mexican sugar are lifted.

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: 17 Development and Adoption of High Fructose Corn Syrup Sweeteners are generally classified into two categories, caloric and non-caloric; common sweeteners in the former group are sucrose, invert sugar, lactose, maltose, and sorbitol; and aspartame and saccharine in the latter. Sucrose is found in various forms of sugar such as raw sugar, granulated sugar and brown sugar derived form sugarcane or sugar beets, or in honey and maple sugar. Invert sugar such as dextrose, glucose, fructose, and HFCS are made form starch through chemical processes. HFCS is produced by converting a portion of naturally occurring glucose in starch into fructose through a com wet milling process (Congressional Research Service, 1999). Lactose, maltose and sorbitol are found naturally in certain kinds of food and give food a sweet taste. Noncaloric sweeteners, sometimes called artificial sweeteners, such as aspartame, are often used for special dietary purposes. Commercially produced and rapidly adopted since the early 1970s in the United States, HFCS became an important player in the sweetener market among sugar substitutes (Figure 2-6). Production of HFCS has increased from 51,000 MT in 1970 to nearly 8.7 million MT in 2001 (Figure 2-7). HFCS production expanded during the 1980s as a substitute for sugar used in the soft-drinks. Today, about 75 percent of total HFCS and 90 percent of HFCS-55 (55 percent fructose) supplied in the United States are consumed in soft-drink market (Buzzanell, 2002; Congressional Research Service, 1999). HFCS-42 (42 percent fructose), which is roughly 90 percent as sweet as sugar, is used mainly in beverages (44 percent), processed food products (21 percent), and other products including cereal and bakery products (Buzzanell, 2002; Congressional Research Service, 1999). As a result, HFCS and two other corn-derived sweeteners, glucose syrup and dextrose, accounted for 55 percent of total U.S. caloric sweetener use in recent years

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18 (Congressional Research Service, 1999). Due to its liquid form, HFCS is considered as a close substitute of sugar, though not a perfect one. Crystalline fructose, fine white crystals of pure fructose is slightly sweeter than sugar and is another potential substitute for sugar; however, crystalline fructose is more expensive than sugar and behaves differently from sugar in most baking and other manufactured food uses thus limiting its use as a sugar substitute (USDA, 1997). HFCS is predominantly produced in the United States, which accounted for 74 percent of the world HFCS production in 2001(Buzzanel, 2002). HFCS is produced in wet-milling faciUties located in com growing regions in the U.S. HFCS producers outside the United States and their recent production levels are: Japan (766,000 MT), Canada (400,000 MT), Argentina (312,000 MT, estimated), Mexico (291,000 MT, estimated), and European Union (293,000 MT) (Buzzanel, 2002). In the United States, most HFCS is supplied and consumed domestically and only a small fraction is exported to Mexico and Canada, the NAFTA member economies (Figure 2-8 and 2-9). The difference in trade between these two countries is that Canada and the United States exchange a similar amount of HFCS across the border while Mexico is a net buyer. Although the quantity exported to Mexico, 122,800 MT in 2001, accounts for only 1.5 percent of total HFCS demanded in the United States, the quantity accounts for 60 percent of total HFCS export from the United States (Figure 2-10). This amount is equivalent to about 40 percent of Mexico's domestic production ability (291,000 MT, Buzzanell, 2002). The introduction of HFCS into the Mexican sugar market brought about significant changes in the sugar consumption pattern in Mexico. Although HFCS is traded at a higher price than sugar in Mexico, it has been gaining an increasing share of the Mexican

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19 sweetener market over the past several years. The Mexican sugar industry has struggled to supply sugar to the domestic market at a price competitive with HFCS. Prior to 1994, nearly all of the caloric sweetener consumption was derived from domestically produced sugar; however, implementation of NAFTA resulted in opening the door for HFCS consumption in Mexico. As a result, Mexican per capita sugar consumption has decreased slightly since 1991, while per capita sweetener consumption has been increasing (Figure 2-1 1). Soft-drink manufacturers are believed to account for about onethird of the total sugar domestically demanded (Buzzanell, 2002). Currently, Mexico's HFCS consumption accounts for approximately 12 percent of total consumption of sweetener (approximately 25 percent of indirect sugar consumption) in 2001. In response to this threatening trend of replacing domestic sugar consumption with U.S. produced HFCS, the Mexican government imposed tariffs in 1996 on HFCS, based on a claim that U.S. companies were dumping HFCS at an unfair price and affecting the export volume and value of Mexican sugar. This action evolved into a trade dispute between the United States and Mexico and ended when the WTO panel ruled against Mexico's claim (Garcia et al., 2002 and 2004). Combined with a slump in production that occurred in 1999 and 2000, the Mexican sugar industry underwent an economic crisis. In September 2001, the Mexican government expropriated 27 of 60 Mexico's functioning sugar mills in order to maintain the industry (USDA, 2001b). Major events in sugar industry history in both Mexico and in the United States are summarized in Appendix A. The U.S. Sugar Industry and Sweetener Market Sugar production in the United States comes from two sources: sugarcane and sugar beets. The main sugarcane production regions are Florida, Hawaii, Louisiana, and Texas where the climate is tropical or semi-tropical. Louisiana and Florida produce

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20 approximately 48 and 45 percent of the total sugarcane production in 2001, respectively (USDA, 2001a). Florida's sugar production from sugarcane made up 50 percent of total cane sugar production in the United States in 2001 (USDA, 2002a), exceeding that of Louisiana (40 percent) due to a higher sugar recovery rate from cane. Sugar beet production regions are classified into four regions: Great Lakes, Upper Midwest, Great Plains, and Far West. The Upper Midwest, which includes Minnesota and North Dakota, produces approximately 47 percent of total sugar beet production; the Far West, which includes California, Idaho, Oregon and, Washington, produces approximately 26 percent of total sugar beet production in 2001 (USDA 2002a). Total sugar production from sugarcane and sugar beets was 4,017,000 short tons (3,615,000 MT) and 4,000,000 short tons (3,600,000 MT), respectively (USDA, 2002a). The proportion of sugar produced in the United States from sugarcane and sugar beets is about equal. Compared to Mexico, sugarcane production in the United States is regionally concentrated and highly mechanized as well as vertically integrated. In the case of the Florida's sugar industry, sugarcane is grown areas concentrated in flat land in south Florida. Sugarcane growing activities such as planting, harvesting and transporting harvested crop are fully mechanized, unlike Mexico. Six raw sugar mills, which are located near the sugarcane fields, possess an average daily processing capacity of 20,750 tons of sugarcane (Alvarez and Polopolus, 2002a). Two sugar refineries are located adjacent to two sugar mills. U.S. Sugar, the country's largest sugar producer operating in Florida, owns a fully integrated cane sugar refinery facility that manages not only sugar refining but also packaging and warehousing. With the facility built next to the existing

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21 sugar mill, this company is able to produce bagged sugar from sugarcane in a single plant location. Sugar is one of several commodities protected in the Farm Bill, which includes rice, cotton, dairy, tobacco, peanuts, grain such as wheat and com, and soybeans (Alvarez and Polopolus, 2002b). The sugar program operates through a loan program and market stabilization price (MSP) without production or acreage restraints, differentiating it from other programs that include target prices or deficiency payments along with export enhancement programs (Alvarez and Polopolus, 2002b). Loans are issued as nonrecourse loans' and are available to processors of domestically grown sugarcane at a rate of 18 cents per pound and to processors of domestically grown sugar beets at 22.9 per pound of sugar, respectively (Haley and Suarez, 2002). Since there are no production restraints, import quotas and tariffs are the main policy instruments utilized to comply with the provision that the program has to operate "no cost" to the government (Alvarez and Polopolus, 2002b).^ In 1990, the United States committed to accept a minimum import quota of 1.256 million MT of sugar as a result of GATT. The U.S. sweetener market has maintained a stable commodity balance, unlike in Mexico, even after NAFTA was implemented and during the trade dispute with Mexico over HFCS dumping. One of the reasons is the successful effort by the American Sugar Alliance (ASA), the sugar producer's alliance in the United States, which lobbies for the U.S. sugar program. The ASA is a strong coalition that includes sugarcane producers, sugar beet producers, and sugar processors, as well as com producers and HFCS As long as the raw sugar tariff-quota is set higher than 1.5 million short tons (Haley and Suarez, 2002). ^ The "no cost" provision was not included in Food and Agriculture Improvement and Reform Act (FAIR) in 1996 (Haley and Suarez, 2002).

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, 22 manufacturers (Moss and Schmitz, 2002). The other entity in the U.S. sweetener market is Coahtion for Sugar Reform (CSR), the industrial sugar user's coahtion. CSR opposes U.S. sugar poHcy, but has been unsuccessful at bringing a lower sugar price to the market from which industrial sugar users as well as consumers would benefit. Many studies have shown that producers clearly gain while consumers and industrial users lose in the U.S. sugar program: an analysis conducted by the General Accounting Office (GAO, 2000) indicates that food manufacturers could be substantial gainers from elimination of the sugar program (Moss and Schmitz, 2002). A history of sugar and HFCS prices are shown in Figure 2-12. Both sugar and HFCS prices exhibit a continuing declining trend. The decline in both U.S. domestic and export prices of the HFCS price is due to sophistication of wet-milling technology in combination with decreases in the tariff schedule under NAFTA for the latter. HFCS has been marketed at a lower price than raw sugar in the U.S. market. Although the U.S. HFCS industry sells at a lower price than sugar, it still benefits from the sugar program because the price of sugar is maintained higher than it would be without the program. This structure confirms why the HFCS industry has been supporting the sugar program as a member of ASA; however, an opportunity for the HFCS industry to increase marketing overseas such as in Mexico may weaken the incentive to support the sugar program: if the sugar price drops as a result of Mexican sugar flowing into the U.S. market through the large import quota promised under NAFTA, the industry has to weigh both costs and benefits to the industry (Moss and Schmitz, 2002). Although the economic implications of the price differential between sugar and HFCS are identified, estimation of a quantitative relationship for HFCS supply is not

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23 straightforward. According to the analysis by Moss and Schmitz (2002), the relationship between sugar and HFCS prices has changed over time: HFCS price responded to wholesale sugar price from 1983 to 1996, but not from 1997 to 2001. Another analysis by Evans and Davis (2002) indicates that the estimated cross price elasticity of HFCS with respect to sugar demand was found to be insignificant. This implies that the HFCS price is set below the sugar price in order to attract bulk sweetener users but its behavior remains ambiguous and arbitrary. Furthermore, other research reported that HFCS price is not correlated with the price of com (Offenbach, 1995), but others found that the HFCS price responded to the price of com from 1983 to 1996 but not from 1997 to 2001 (Moss and Schmitz, 2002). Given the insignificant relationship between com price and HFCS supply mentioned above, the indirect impact of changes in com production on HFCS price would likely be small; however, any drastic changes in com program or large changes in com export may eventually affect HFCS prices. Com statistics such as recent com production for selected countries, industrial use of com in the U.S., and the com price in the U.S. are illustrated in Appendix B.

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24 G O u
PAGE 36

25 100 T 90 80 — 70 60 4 06 G 50 o 'S 40 30 i-i 20 10 0 1-T m-' m — mimm — — ^ — 500 1,000 1.500 2,000 2,500 Annual Rainfall [mm] 3,000 3.500 4,000 Figure 2-3. Annual Rainfall and Irrigation Rate in Sugarcane Fields Source: COAAZUCAR, 2003c Table 2-1. Cane Sugar Production in Selected Countries, 1997-2000 Average. Country Area Sugar Cane Sugar Sugarcane Harvested Production Yield Yield Recovery (1,000 ha) (1,000 MT) (MT/ha) (MT/ha) Rate (%) Brazil 4,914 18,339 68.28 3.73 5.47 India 4,092 17,233 69.41 4.21 6.07 Cuba 1,086 3,814 32.69 3.51 10.77 China 1,064 6,532 75.08 6.14 8.18 Pakistan 1,029 3,064 46.57 2.98 6.38 Thailand 923 5,468 56.16 5.92 10.55 Mexico 627 4,807 76.46 7.66 10.02 Australia 409 5,281 90.94 12.91 14.22 United States 360 3,811 88.00 10.59 12.03 Colombia 391 2,227 86.25 5.70 6.61 Philippines 324 1,796 81.06 5.55 6.87 South Africa 315 2,684 72.57 8.54 11.76 Wodd 19,307 90,340 65.12 4.68 7.19 Source: FAO 2003, USDA 2002a, USDA 2003b

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26 6.000.000 H g 5.000,000 O 4,000.000 u 3 3.000.000 o £ 2,000,000 u W) 1,000,000 O — o o o o o o ^ — — — rj (S Year Figure 2-4. Mexican Sugar Production, 1988-2002 Source: COAAZUCAR 2003a Sugarcane Cane juice IRa^v sugar Iiitemational market or stored l*Refined sugar Standai'd sugar 1^ White sugar (purer) Other grades Molasses Alcohol (rum) Other by-products Domestic market t Figure 2-5. Main Use of Sugarcane Derivatives in Mexico Source: Adapted from Polopolus and Alvarez, 1991

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27 Table 2-2. World Sugar Consumption in 2000 National total Per capita consumption" consumption" Coiintrv/ rppion \^WUllll jl l^glWll ITSweetener Sugar b Sweetener Sugar (1,000 MT) (1,000 MT) (kg) (kg) India 26,234 18,101 25.59 17.66 United States 21,221 9,371 74.22 32.77 European Union (15 countries) 18,960 14,370 50.26 38.10 China 11,805 11,028 9.13 8.53 USSR, Former Area of 11,771 11,407 40.57 39.32 Brazil 10,007 9,620 57.99 55.75 Mexico 5,100 4,476 50.81 44.60 Japan 3,741 2,327 29.38 18.27 Philippines 2,135 1,975 27.68 25.61 Thailand 1,937 1,924 30.47 30.26 Canada 1,297 1,128 41.83 36.37 Australia 1,060 922 54.81 47.67 Cuba 711 710 63.24 63.18 World 168,632 133,401 27.49 21.75 a: Production + net import + change in stocks b: Sugar (raw equivalent) plus other kinds of sweeteners Source: FAO, 2003 Table 2-3. Quota and Tariff Schedule Imposed on Mexican Sugar Exported to the U.S. Year U.S. Import Quota (MT) Over-Quota Tariff (raw cane, cents/pound) Mexico as a net surplus producer Mexico NOT as a net surplus producer 1994 25,000 7,258 16.00 (Base) 1995 25,000 7,258 15.20 1996 25,000 7,258 14.80 1997 25,000 7,258 14.40 1998 25,000 7,258 14.00 1999 25,000 7,258 13.60 2000 250,000 7,258 12.09 2001 250,000 7,258 10.58 2002 250,000 7,258 9.97 2003 250,000 7,258 7.56 2004 250,000 7,258 6.04 2005 250,000 7,258 4.53 2006 250,000 7,258 3.02 2007 250,000 7,258 1.51 2008 and beyond oo 0.00 Source: USD A, 1999.

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28 Year •Total HFCS use per capita Refined sugar use per capita Figure 2-6. U.S. Refined Sugar and HFCS Use per Capita Source: USD A, 1993 Figure 2-7. U.S. HFCS Production Source: USD A, 1993 and 2001a.

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29 Import (mostly from Canada) 1.6% \ MiiSjimf^ U.S. production 98.4% Figure 2-8. U.S. HFCS Supply Source: USDA, 2002a Export to ^Export to B Domestic consumption 97.5% Figure 2-9. U.S. HFCS Export Source: USDA, 2002a

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30 400.0 Year •U.S. total export U.S. export to Mexico Figure 2-10. U.S. HFCS Export, 1992-2001 Source: USDA, 2002a -Consumption of sugar per capita — — Consumption of sweetener per capita Figure 2-11. Consumption of Sugar and HFCS per Capita in Mexico Source: Garcia Cliaves et al., 2004

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31 70 T 1995 19% 1997 1998 1999 2(XX) Year -B—Worldrawa^ price -•— US rawsu^ price, city fee paid -A— US spot price for HPCS42, Mdv\est markrts -A— Uiit valiE of HKSecpoted to Nfexioo Figure 2-12. Prices of Sugar and HFCS, 1994-2000 Source: USD A, 2001a

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CHAPTER 3 CONCEPTUAL AND THEORETICAL FRAMEWORK In this chapter the conceptual and theoretical framework employed to analyze trade between the United States and Mexico in the sugar market is discussed. In the conceptual framework, factors that influence the market and the trade system are specified and linked. Based on the conceptual framework, theoretical foundations are established in two parts: an analysis of the sugar market for each country and an analysis of the market balance in the U.S.-Mexico bilateral sugar trade system. Conceptual Framework The sugar market, one of the oldest and most common agricultural commodity markets, is built upon sugarcane and sugar beet production and the resulting production of processed sugar from these raw materials. Being an essential commodity for a daily diet, sugar has been traded across borders for a long period of time. More recently, there have been major changes to the trading pattern due to the emergence of an alternative in the market. High fructose com syrup (HFCS), now the most widely adopted sugar substitute, expanded the sugar market into a sweetener market. This is particularly true in the case of the United States, where HFCS now occupies nearly half of the sweetener market, and is becoming the case in Mexico as a result of the two markets becoming more closely linked by the North American Trade Agreement (NAFTA). Understanding this linkage between the United States and Mexico holds important clues to analyze economic and political impacts in the sweetener markets. To illustrate this linkage, the 32

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33 flow of the goods in each market and factors that influence markets are illustrated in Figure 3-1. In Mexico, sugar distributed in the market is supplied entirely by domestic sugar production, which is also provided entirely by domestic sugar cane growers. Sugarcane production depends on inputs such as labor from Mexican farmers, land, and agricultural chemicals, technology to grow and harvest sugarcane, and other factors such as weather and government support. Grower's behavior is also influenced by their relationship with mills. Sugar processing depends on inputs such as labor from mill workers, harvested sugarcane, and energy such as petroleum to run the facilities, technology to produce sugar from sugarcane, infrastructure, and government support. Note that government plays an important role to support both grower's and mill's activities. HFCS is primarily supplied by domestic production and the remained is imported from the United States. Sugar is consumed by households and bulk users (such as soft-drink manufacturers), while HFCS is consumed only by bulk users. Determinants of sweetener demand are income, tastes and preferences, price and other factors such as population growth. In the United States, domestic sugar supply is derived from domestic production and supplemented by import from various origins. Unlike Mexico, domestic sugar is derived from both sugarcane and sugar beet production. Sugar mills produce sugar not only from domestically produced sugarcane and sugar beets, but also from imported raw sugar. HFCS is supplied only from domestic com sweetener manufacturers and supplies roughly half of caloric sweetener consumption in the United States. HFCS is also exported, and Mexico is one of the main destinations.

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34 The U.S. -Mexico bilateral trade system involves sugar exported from Mexico to the United States and HFCS exported from the United States to Mexico. Mexican sugar is exported to a variety of markets, but the primary destination is the U.S. sweetener market. Once Mexico exports the amount of sugar to the United States allowed under the U.S. import system, excess sugar is exported to the world sugar market. In total, more than 30 countries exported sugar to the United States under the allocated tariff-rate quota in 2002 (USDA, 2002a). Under the conditions of NAFTA, Mexico could be allocated 250,000 MT of the U.S. import quota given certain conditions (successful attainment of net surplus sweetener producer status), which would increase Mexico's share of the total U.S. import sugar quota allocation to 20 percent. A two-country trade model with quota imposed by a large country importer is illustrated in Figure 3-2 (a). By imposing a quota on Mexican excess supply of sugar (ESmx), the quantity exported to the United States is limited to Qq instead of Qf. Consequently, the price for imported sugar in the United Staes increases to Pq, us and the price for exported sugar from Mexico decreases to Pq, mx. This causes welfare loss in Mexico (area abed) due to lower sugar export price and generates quota revenue in the United States (area efgh) collected by the U.S. quota holders or the government. Beginning in 2008 Mexico will have free access to the U.S. market. The same trade model without the quota system is illustrated in Figure 3-2 (b). Trade without distortion brings an increase in welfare in both countries. For simplicity, the producer price support policy is excluded from both Figures (a) and (b).

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35 Theoretical Framework Sweetener Market Analysis The sweetener market in a country that consumes both sugar and HFCS can be expressed as follows: SUGAR =fl (PsUGAR„Zl) (3-1) HFCS =f2 (Phfcs„Z2) (3-2) &SUGAR = h,(PsUGAR, Wj) (3.3) (/hfcs = h2 (Phfcs,. W2) (3.4) SUGAR = SUGAR (3-5) (2^ HFCS = (^HFCS (3-6) where is aggregate quantity demanded, Q' is aggregate quantity supplied, F is a price, Z and W are vectors of other factors that influence aggregate demand and supply of sugar or HFCS, respectively. Equations [3.5] and [3.6] depict market clearing conditions for each commodity. Sweetener demand Sweetener demand is defined based on consumer demand theory derived from utility maximizing behavior. Following Varian's demonstration (1992), aggregate sugar demand is derived from maximizing utility of aggregated consumers, including industry sugar users, by purchasing sugar: max u ( QsuGAR Qx) s.t. QsuGAR *PsuGAR + Qx*Px = m (3.7) where Q is the quantity consumed, X represents all other goods consumed, P is a price, and m is national income. By solving maximization problem, the aggregate demand function is expressed as:

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36 QSUGAR (PsUGAR Px, m). (3.8) Since the demand function is homogeneous of degree zero, it can be normahzed by either price. Normalize by Px and thus the aggregate demand for sugar is expressed as a function of real price of sugar and real income. By treating real income as one of demand shifters, aggregate demand is expressed as shown in equation [3.1]. Similarly the aggregate demand for HFCS can be expressed as a function of HFCS price and a vector that affects aggregate HFCS demand. Note that in reality, HFCS is consumed directly by industry users and its price is observed by them; households as final consumers consume HFCS indirectly through HFCS-contained goods. Sweetener supply Sweetener supply is defined based on firm supply theory derived from profit maximizing behavior. Although in reality sugar supply consists of two steps of production in reality, sugarcane and sugar production, a simple aggregate sugar supply equation at industry level is derived rather than two equations. This is suitable for two reasons: a single industry supply equation makes simulations in the bilateral trade model in the following procedure simple, and it is the industry supply price that the government is interested in supporting. Suppose the industry faces a cost function given by: where Visa set of other factors including input prices. The industry maximizes profits, assuming the market is competitive (the industry as price taker): QsuGAR (PsuGAR / Px, / Px), (3.9) C=C{QsUGAR,V) (3.10)

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37 max n = Psugar Qsugar C (Qsugar., V ). (3.11) The first-order condition gives the supply relationship for the industry by equating output price to marginal cost. Expressed in a general form with a vector of other factors that influence aggregate supply: ffsuGAR (Psugar. Wi). (3.12) Similarly HFCS supply from industry {(^hfcs ) is theoretically expressed as a function of the HFCS price and the supply shifters; however, the quantitative relationship is expected to be insignificant: HFCS pricing is ambiguous and arbitrary as discussed in the previous chapter and thus the estimated relation does not likely represent the associated marginal cost curve. U.S.-Mexico Bilateral Sugar Trade System A spatial equilibrium model is used to portray the U.S.-Mexico bilateral sugar trade system, following the Takayama and Judge formulation (1964). The model provides the optimal equilibrium price as well as quantities demanded and supplied at the equilibrium through maximization of welfare in each region, i.e. the sum of the consumer and producer surplus, given the demand and supply equations and the transportation cost among regions. Let 5, (K,) represent the inverse supply function (price-dependent form) for sugar in region i; Y\ represents the quantity produced in region i; Dj {Qj) represents the inverse demand function (price-dependent form) for sugar in region j; and Qj represents the quantity consumed in region j. MaxXj/),(e,)^e, -Y^js,(Y,)dZ, -YX^R.X,^ -ZE7T/?,XX, (3.13) i=\ 1=1 /=i i=i 1=1 >=i s.t. X(^, +^,)^i',>Vj = l,..,/ (3.14)

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38 2;(X,, + XX,,)>e,,V; = l,...,7 (3.15) 1=1 X„ 0 where Xy is the quantity of sugar shipped from supply region i to demand region j under quota Hmit, TRij is per unit transfer costs associated with X,;,, XXy is the quantity of sugar shipped region i to demand region j over quota limit, TTRij is per unit transfer costs associated with Xy. TRij is a compound transfer cost that includes per unit transportation cost (Tij) and per-unit tariffs imposed on exported sugar under quota limit [Tanj ). Similarly, TTRij is a compound transfer cost that includes per unit transportation cost (Ty) and per-unit tariffs imposed on exported sugar over the quota limit (over-quota tariff, OQTar.j ). TR,j = T.j + Tarij (3.17) TTR,j = T,j + OQTanj (3.18) Let specific inverse linear demand and supply functions for United States and Mexico be defined as follows: P^us = lU, + lU2*(fvs + Shifter'' us (3.19) I^us = IUU, + IUU2*Q^us+ Shifter^ us (3.20) P'^MX = Ml + /M2*(2%x + Shifter'' MX (3.21) P^Mx = IMMi + IMM2 *Q^Mx + Shifter^ MX (3 .22) where P,and Q'' represent the price and the quantity demanded in each country, respectively; F^ and Q'^ represent the price and the quantity supplied in each country, respectively; Shifter'' and Shifter^ represent the demand and supply shifters in each

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39 country, respectively; Wi, lUUi, IMi, and IMMi represent the intercepts in each inverse Hnear equation; and IU2, IUU2, IM2, and IMM2 represent coefficients associated with each quantity variable. The objective function of the U.S.-Mexico bilateral sugar trade system is expressed with equations [3.19], [3.20], [3.21], and [3.22] with constraints such as necessary conditions, i.e. demand-incoming shipment and supply-outgoing shipment balance as well as those specific to the bilateral trade model such as Mexico's quota allocation under NAFTA and U.S. price support: Max j {lUi + IU2*Q^ us + Shifter^ us) us + j iIM, + IM2*Q^^ MX + Shifter^ Mx)dQ^^ MX j (lUUi + IUU2*Q^us + Shifter^ us) d^us J (IMM, + IMM2*Q^MX + Shifter^ Mx) d^MX (Tmx. us )*X MX. us (Tus. Mx)*X us, MX (Tmx us + OQTarMx us)*XXmx, us (Trow, us Prow )*Xrow, us + Prow *Xmx row (3.23) where US, MX, ROW represent Mexico, the United States, and the rest of the world, respectively; Ty is per unit transportation cost from i to j; OQTarMx. us is per unit per unit over-quota tariff imposed on Mexican sugar shipped to the United States; and Prow is world price of sugar. Note that the transportation cost within the country is assumed to be zero. The over-quota import from the rest of the world to the United States {XXrow. us) is ignored because of the high tariff rate imposed on the sugar from the rest of the world.

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40 compared to sugar from Mexico: the first sugar that enters into the United States overquota must be from Mexico. The transfer cost of sugar from the rest of the world to the United States includes the price of sugar, i.e. sugar from Mexico and the rest of the world compete with each other to enter the U.S. market: the one with lower transfer cost enters the market first. The last term in the equation [3.23] considers Mexico's sales of sugar to the rest of the world. Equation [3.23] is rewritten in quadratic form as follows: Max iV2)*IU2*(Q^usf +iIUi + Shifter'' us )*Q'' us + (l/2)*/M2*fQ%x)' + (/M/ + Shifter'' mx)*Q''' MX (1/2)* IUU2*( us f + UUU,+ Shifter^ us)*Q^ us (1/2)* IMM2*(Q'mx )' + (IMM, + Shifter^ mx)*Q' MX (Tmx, us )*X MX, us (TuS. MX )*X us, MX (Tmx, us + OQTarMx, us)*XXmx, us (Trow, us + Prow )*Xrow, us + Prow *Xmx, ROW (3-24) Constraints are defined to balance the quantities shipped with the quantities demanded as well as supplied; to impose a quota on imported sugar from Mexico; and to impose a quota on imported sugar from the rest of the world, which is equivalent to the U.S. minimum sugar import quota required under WTO agreements: Q us -Xus, us Xmx. us XX MX, us Xrow, t/s ^ 0 (3.25) Q^^mx -Xmx.mx X us, MX ^0 (3.26) Q^us + Xus,us+ X us, MX ^ 0 (3.27) Q^MX + Xmx, MX + Xmx, us + XX MX, us + Xmx. row ^ 0 (3.28)

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41 X MX. usQuota < 0 (3.29) Xrow. us + XX MX. us USMin > 0 (3.30) where Quotais the quota Mexico is assigned according to the NAFTA provisions and USMin is the quota allocated to the rest of the world (the U.S. minimum sugar import quota less the quota assigned to Mexico). Equation [3.25] and [3.26] satisfy domestic demand; equation [3.27] and [3.28] regulate the quantities shipped from supply regions; and equations [3.29] and [3.30] define quotas imposed on Mexican sugar and from the rest of the world. The implications of the model can be examined by writing the first-order conditions obtained through using Kuhn-Tucker theorem. From equations from [3.24] to [3.30] the Lagrangian form (L) is given as: L= i\/2)*IU2*(Q''usf +(IUi + Shifter^ us. )*Q'' us + (l/2)*/M2*(!2''W + (/M/ + Shifter'' MX )*Q''' MX (1/2)* IUU2*(Q^usf + iIUU,+ Shifter^ vs)*^ us (1/2)* IMM2*(Q'Mxf + (IMMj + Shifter' MX )*Q' MX (Tmx, us )*X MX. us (Tus.Mx)*X US.MX {Tmx, us + OQTarMx, us)*XXmx. us (Trow, us + P row )*Xrow, us + Prow *Xmx, row + Vus (X us, us + Xmx, us + XX MX, us + Xrow, us us ) + Vmx(XmX,MX + X USMX Q^^Mx) + Wus (Q^us Xus.us -X us, Mx)

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42 + Wa/x (Q^mx Xmx. MX Xmx. us XX MX, us Xmx. row ) + A (Quota -X MX, us) + a(XRow, us + XX MX us USMin) (3.31) where V, W, A, and crare Kuhn-Tucker multiplier associated with each constraint representing the imputed marginal value of price of sugar demanded, supplied, that of Mexican sugar exported under-quota and that of over-quota, respectively. Note that A is positive in sign and cris negative, reflecting the way their associated constraints are defined. Kuhn-Tucker conditions are expressed as follows: % =1^2*0^ us +IU,+Shifier''us Vus < 0. ^ *Q''us = 0, Q^'vs > 0 (3.32) -(IUU2*^us +IUU,+Shifter^us )+Wus < 0, *Q^us = 0, Q'us > 0 ^Qus ^Qus (3.33) ^ =IM2*Q'''mx +IMi+Shijief^Mx -Vmx < 0. *Q'''mx = 0, G%x > 0 (3.34) -{IMM2*Q'mx +IMMi+Shifter^Mx)+WMx < 0 *Q'mx = 0, Q'mx > 0 oQmx ^Qmx (3.35) = Vus Wus < 0, *X us, us =0, X us, us>0 (3.36) ^X[js,us ^Xus,us =Vmx -Wmx < 0, *Xmx MX =0, Xmx. mx>0 (3.37) ^

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43 = -Tmx, us + Vus -Wmx /I ^ 0, *Xmx, us =0, Xmx, as ^ 0 = -Tus. MX + ^us -Wmx ^ 0, — — *X us, mx -0, X vsmx ^ 0 (3.38) (3.39) = -Tmx. us OQTarMx, us +Vus -Wmx + cr< 0, —~ — *Xmx, us =0, ^XX dX Xmx. us>0 (3.40) = -Trow, us -Prow + Vus +<7< 0, — — *Xrow. us -0, Xrow. us^O ROW, us ROW, us = Prow Wmx ^ 0, *Xmx, row =0 Xmx, row ^ 0 (3.41) MX ,ROW MX MOW (3.42) By solving equations above, the following conditions must hold at equilibrium:' Vus = P^ us = Wus = P^ us (3.43) Vmx = P^MX = Wmx = P" mx (3.44) P^us
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44 The complementary slackness conditions indicate that if Mexico exports sugar to the U.S. under-quota {Xm, us > 0), then the demand price in the United States should not exceed the value of Mexican exporting sugar, which is equivalent to the sum of the supply price, transportation cost and the marginal value of exporting sugar under-quota (equation [3.45]). By the same token, if Mexico exports sugar to the United States overquota (XX MX, us>0), then the demand price in the United States should not exceed the sum of the Mexican supply price, transportation cost, tariff imposed on over-quota sugar and the marginal value of exporting sugar under-quota (equation [3.47]). The inequality of the prices expressed in equation [3.49] accords with reality. If both over-quota export from Mexico {XX mx. us ) and import from the rest of the world (Xmx. us ) are greater than zero, the following must also hold from equations [3.47] and [3.48]. P^m Prow = tRow. us Imx. us OQTarMx. us (3-50) This equation implies that when the price difference between Mexico and the rest of the world is equal to the difference in transfer cost (transportation cost and tariff), both over-quota export from Mexico and import from the rest of the world occur at the same time. In other words, Mexico would export over-quota only if the transportation cost from the rest of the world is high enough to justify Mexico to do so.

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46 Figure 3-2. Two-country Trade Model (a) with Quota System and (b) without Quota System.

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CHAPTER 4 EMPIRICAL MODELS AND DATA SOURCE Based upon theoretical framework developed in chapter 3, this chapter focuses on the empirical procedures and data set used in the analysis of the U.S.-Mexican sweetener market. The empirical model consists of three components: (1) demand and supply analysis models for both the U.S. and Mexican sugar markets; (2) a bilateral sugar trade analysis model; and (3) game theory analysis. These models are ordered sequentially so as to utilize the results from the former. After the trade model is introduced, assumptions associated with the model, the methods of model calibration, and simulated scenarios are presented. Results from simulations on the trade model are aggregated to assess policy recommendations. Lastly, an overview of the data set used in the empirical models is provided. Empirical Models U.S. Sweetener Demand Model Regression analysis is used to estimate sweetener demand utilizing quarterly timeseries data. The model is specified as a double-log (natural log) form that allows interpretation of estimated coefficients as elasticities associated with each variable. Based on the basic form of demand equation shown in equation [3.1], the U.S. sugar demand equation is specified as: SUGAR, t) = U]+ U2*In(I^ SUGAR. ,) + V3*ln(GDP,} + V4*In(P0P,} + U5*QTR1+ U6*QTR2+ U7*QTR3+ U8*DHFCS + e, (4.1) where (2^ is quantity of sugar demanded in each quarter in year t [1000 short tons], is 47

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48 real retail price of refined sugar (deflated by Consumer Price Index (CPI)) [cents / pound], GDP is real per capita GDP (deflated by CPI [US$]), POP is population, QTRs are dummy variables representing quarters of the year, DHFCS is dummy variable for availability of high fructose com syrup (HFCS) in the sweetener market {DHFCS=\ after t=1975), and e,is an error term. In theory, the price of HFCS would be preferred as prices of substitutes are expected to influence quantity demanded, however, limited availability of the data made this impossible, therefore, the dummy was used. It is expected that the elasticities associated with price {Ui), the first quarter {U5) (compared to the omitted fourth quarter), and HFCS availability {Us) to be negative and the elasticities of the remaining variables to be positive. Price elasticity is expected to be inelastic based on previous research. Previously reported own-price elasticities for sugar are -0.141 by Lopez (Lopez, 1990) and -0.73 by Petrolia and Kennedy (Petrolia and Kennedy, 2002). In the latter estimation, the U.S. wholesale refined beet sugar price reported at the Midwest Markets was used. In the process of regressions, serial correlation is anficipated and corrected by the Yule-Walker Method with appropriate lags assigned. Mexican Sweetener Demand Model Estimation of Mexican sweetener demand was conducted in a similar manner to that of the U.S. demand. The main difference was that the demand for two kinds of sugar is estimated separately in the Mexican model. Traditionally the demand for sugar in Mexico has been estimated by regressing the total consumption on sugar price and percapita income. Borrell (1991) estimated the price elasticity be -0.004 and the income elasticity at 0.5. This model was appropriate in early 1990s because there was no HFCS consumption in Mexico. In order to account for the entry of HFCS into the Mexican sweetener market, the demand for sweetener is estimated by disaggregating sugar

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49 consumption into: sugar consumed by households, referred to as "direct consumption"; and sugar consumed by bulk sugar users, referred to as "indirect consumption". Only indirect consumption of sugar is expected to be influenced by HFCS consumption as they are the only consumers of HFCS. In addition to estimating sugar demand, it would have been preferable to directly estimate HFCS consumption, but this was precluded because of the relatively short time series available. Instead, total consumption of sweeteners (the sum of sugar and HFCS consumption) is analyzed by regressing on sugar price and other variables. A dummy variable that represents the availability of HFCS in the market is also added to the regression forms of indirect consumption of sugar and total consumption of sweeteners where substitution between sugar and HFCS occurs in the market. In(Q^suGAR. ,) = M, + M2*ln(P^ SUGAR. ,} + M,*In(GDP,) + M4*In(P0P,)+ u, (4.2) SUGAR, t) = M5 + M6*In(P" SUGAR, ,) + M7*ln(GDP,) + M8*In(P0P,) + U9*DHFCS + v, (4.3) In(Q^^ SUGAR, t) = Mio + Mu*ln(P^ SUGAR, t) + Mj2*In(GDP,) + M]3*In(P0P,) + Ui4*DHFCS + w, (4.4) where (2 is direct consumption of sugar [MT], (2 is indirect consumption of sugar [MT], is total consumption of sweeteners [MT], is real retail price of standard sugar (deflated by CPI [pesos / Kg]), GDP is real per capita GDP (deflated by CPI [pesos]), POP is population, DHFCS is dummy variable for availability of high fructose com syrup (HFCS) in the sweetener market {DHFCS=l after t=1992), and i<,,v,,w,are error terms. It is expected that the elasticities associated with price (M2, Me, and M77 ) and

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50 HFCS availability (M9 and Mu) to be negative and the others to be positive. Price elasticities were expected to be inelastic. Serial correlation is anticipated and corrected by the Yule-Walker Method with appropriate lags assigned. U.S. Sweetener Supply Model Regression analysis is used to estimate the U.S. sweetener supply utilizing timeseries data. Aggregate sugar supply is analyzed in three parts, i.e. total sugar supply, sugar supply from sugarcane and sugar supply from sugar beets. Sugarcane and sugar beet production are estimated separately as they have different production regions and a different refining process. Sugar production is used as the dependent variable. It is assumed that domestic sugar production is the primary source of sugar supply and carried-over stock from the previous periods is considered constant over the estimated time span. The model is specified as a double-log (natural) form and estimated coefficients can be read as elasticities associated with each variable. lni(^ TOTAL SUGAR. ,) = UU l + UU2nn(P\oTAL SUGAR, + UU3*In(C0ST,) + UU4*In(RCVT0TAL SUGAR, t) + UU 5*In(Q^ TOTAL SUGAR, t-l) + UU6*TREND + eet (4.5) In(Q^ CANE SUGAR.,) = UU7+ UU8*ln(P ''cANE SUGAR, + UU9*ln(C0ST,) + UUio*In(RCVcANE SUGAR.,) + UUn*In(Q^ CANE SUGAR., -1) + UU,2*TREND + UU, (4.6) In(Q BEET SUGAR, t) = UUu + UUj4*In(P 'beet sugar, ,-i) + UUj5*In(COST0 + UUi6*In(RCV beet SUGAR, ,) + UUi7*In(Q \eetsugar. ,-i) + UUi8*TREND + vv, (4.7) where Q represents the total sugar quantity produced, sugar quantity produced from sugarcane, and sugar quantity produced from sugar beets [1000 short tons] in the three

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51 equations, respectively; P represents the real retail refined sugar price in the previous year (deflated by CPI), real raw sugar price at NY sugar exchange in the previous year (deflated by CPI), and real wholesale refined beet sugar price in the previous year (deflated by CPI [cents / pound]), respectively; COSTh real total farm production expenses deflated by CPI (used as a proxy of sugar production cost [US$]); RCVs are sugar recovery rates during sugar refining process [%]; TREND is a trend variable that represents technology advancement; and ee, uu,, vv, are error terms. Prices in the previous year are used assuming that decision-making on sugar production relies on sugar crop production, recognizing growers decide their production plan with the price realized in the previous year. Retail refined sugar price is used for total sugar production estimation due to lack of wholesale price data. Total sugar recovery rate is expressed as average of recovery rate of cane sugar and beet sugar computed by weighing each production onto each recovery rate. Autoregressive term (production in the previous year) is added in the regression to calculate long-run elasticities. Elasticities associated with price production cost {UU3, UUg, and UU15) are expected to be negative and the others to be positive. Price elasticities are expected to be inelastic. Previously reported own-price elasticities for the U.S. sugar supply at industry level are 0.14 for cane sugar and 0.34 for beet sugar by Petrolia and Kennedy (2002), using the U.S. wholesale refined beet sugar price reported at midwest markets. Price elasticities for land allowed to sugar production estimated by Lopez (1990) were 0.103 for cane and 0.246 for beets. Both authors reported higher elactisities for beet sugar.

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^ i wf f • : • ,?rit— • • '-^^ "^JPFT^T*." 52 Mexican Sweetener Supply Model Analysis of the aggregate supply of sugar in Mexico is conducted in a similar manner as the U.S. supply analysis except that Mexico produces sugar only from sugarcane. In((f,) = MMi + MM2*In(P ^,.1) + MM3*In(C0ST,) + MM4*In(DT,) + MM5*In(SUGL0SS,) + MM6*In(DURTN,) +MM7*TREND, +ww, (4.8) where Q are total sugar quantity produced [MT], F is real wholesale standard sugar price in the previous year (deflated by CPI [pesos / Kg]), COST is real average production cost per ton of sugar realized at sugar mill deflated by CPI [pesos], DT is average mill operation downtime ratio observed at sugar mill [%], SUGLOSS is average sugar loss ratio observed during sugar production process at mill [%], DURTN is average duration of the harvest in each season [days], TREND is a trend variable that represents technology advancement; and ww, is error term. Price in the previous year is used assuming that decision-making on sugar production relies on sugar crop production, recognizing growers decide their production plan with the price realized in the previous year. An autoregressive term (production in the previous year) is not added because of limited length of data available for regression. It is expected that the elasticities associated with price production cost (MM.?), downtime (MM4), and sugar loss (MM5) to be negative and the others to be positive. Price elasticities are expected to be inelastic. Previously reported ownprice elasticities for Mexican sugar supply at industry level is 0.67 by Petrolia and Kennedy (2002). Borrel (1991) estimated two sugarcane price elasticities by regressing sugarcane yield on sugar cane price in an Almon polynomial distributed lag model and by regressing sugarcane acreage on sugarcane price. From the

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53 former regression, statistically significant estimates were 0.0286 (lag=3) and 0.0294 (lag=4) and from the latter, the estimate was not statistically significant. U.S.-Mexico Bilateral Sugar Trade Model Based on the theoretical framework presented in chapter 3, a bilateral trade model of U.S.-Mexico sugar trade is developed and prepared for the empirical study. First, linear inverse demand and supply equations (price endogenous) are formulated using estimated elasticities from the previous analysis following the procedures by Spreen et al. (2000) in order to formulate the objective function in the trade model. Detailed derivation is noted in Appendix C. Although the double-log form is used in the demand and supply analysis because of its statistical advantages in estimation, the linear form of demand and supply equations are used in the trade analysis model for the following reasons: with the linear form, welfare can be measured, whereas welfare cannot be measured using log form equations due to the nature of the mathematical attributes; using the linear form makes it possible for the demand or supply curves to shift with their slopes held constant; and interpretation of Kuhn-Tucker conditions presented in chapter 3 is facilitated. Sugar trade models have been developed by Koo and Taylor (2000) and Petrolia and Kennedy (2000). The former model incorporates sugar production, consumption and stock changes in seventeen sugar producing and consuming countries and the latter encompasses the United States, Mexico, and Cuba. In these models, market equilibrium is solved based on the market clearing condition, i.e. the sum of all countries' excess demand for sugar becomes zero as sugar price adjusts. These models are relatively simple and may be suitable for macroscopic analysis since they are able to include many economies without making the models large and complicated; however, the solutions derived from these models pay little attention to country's welfare or transfer costs of

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54 goods. In addition, details of trade agreements are difficult to incorporate in these models. The fact that there exist numerous bilateral and regional trade agreements, a spatial equilibrium model has the advantage of enabling the various trade agreements to be tailored into the model. Lastly but most importantly, it is critical to consider HFCS when the sweetener markets in both the United States and Mexico are under scrutiny. Since this study is focused on the bilateral relation between the United States and Mexico, a spatial equilibrium is chosen to conduct a more microscopic analysis, incorporating detailed trade agreements and the impacts from HFCS consumption. Simplifying assumptions Several simplifying assumptions are made to run the model. First, the quantities in the models are raw sugar equivalent. In doing so, the price difference along the vertical market channel is ignored and derived demand and supply curves are assumed to posses the same slopes as demand and supply of refined sugar. Second, changes in sugar stocks in both counties are also ignored and hence the excess sugar supply from Mexico and the excess sugar demand from the United States are captured as the difference between domestic sugar demanded and supplied in each county, illustrated in the following. Formally, the U.S. sweetener market balance is expressed with sugar production, consumption, import, export, and change in stock as well as with those of HFCS: Q"" SUGAR. ,+ SE,+ ST.^i + HFCS'' =Q' sugar. t+ SI ,+ ST,.i + HFCS' (4.9) where Q'^ sugar is the quantity of sugar demanded, sugar is the quantity of sugar supplied from domestic sugar production, SE is sugar export, SI is sugar import, ST,+i is sugar stock carried over to the next year, ST,.i is sugar stock carried over from the previous year, and HFCS'^ and HFCS^ are HFCS demand and supply related transactions,

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55 respectively. Q'^ sugar. I '^s expressed in annual base converted from quarterly base. Since neither supply nor demand of HFCS is estimated, HFCS^ and HFCS^ are treated constant. This is justified by the fact that HFCS use in the United States has remained stable in recent years, accounting for roughly half of total U.S. caloric sweetener use. In addition, changes in stock have also remained stable given the nature of commodity demand and production practices, ST,.] and 57,+/ are set zero. SE is also set zero as sugar export from the United States is negligible given the production capacity. Equation [4.9] is then simplified and the quantity of forecasted sugar import in the United States ( 57, ) is D expressed with forecasted quantities of sugar demanded and supplied ( Q sugar, t andQ^ SUGAR, t) as: si, = Q SUGAR. Q 'sugar, (4.10) Mexico's sweetener market balance is expressed in a similar fashion. The differences are sugar demand in Mexico is estimated by total, direct and indirect sugar consumption and Mexico imports HFCS from the United States to meet its domestic demand for sweeteners. By ignoring stock changes and sugar import, the sweetener market balance is expressed as: Q'^'^sugar. + Q"^sugar. t + MFCS'", + SE, = Q^sugar, + HFCS^ (4.11) where Q sugar is the quantity of demanded by households (direct consumption), (^^ sugar is the quantity of sugar demanded by bulk users (indirect consumption), HFCS^ is the quantity of HFCS demanded by bulk users, SE is sugar export, sugar is the quantity of sugar supplied from domestic sugar production, and HFCS^ is the quantity of HFCS supplied from domestic production and import from the United States. The study is interested in forecasting sugar surplus available for exporting to the United States

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56 D S considering Mexico's increasing demand for HFCS. Since MFCS'' and MFCS' are not estimated directly, HFCS demand {MFCS^ ) is forecasted based on the associated scenarios with details provided in the following section.' Once HFCS demand is forecasted, the quantity of excess sugar destined to export is calculated by considering the substitution between HFCS and indirect sugar consumption, which occurs at industry (bulk users) level. Let Q'^ sugar, r be HFCS forecast-adjusted indirect sugar consumption, then the sugar export forecast ( SE, ) is expressed with forecasted quantities of sugar demanded and supplied {Q^ sugar, t, Q^^ sugar, t ,and Q'^ sugar, t) as: SE, = Q ^ sugar, I (Q ^'^ sugar, t + Q sugar, t ) (4. 12) In equation [4.34], it is also assumed that HFCS is consumed only to supplement sugar consumption and its demand is met by readily available HFCS from domestic production and import from the United States. Accordingly, NAFTA provisions are defined, depending on the Mexico's domestic sweetener balance; Mexico receives a larger quota for the following year if is attained net surplus producer status for two successive years: Q '''' SUGAR, t + Q sugar, + MFCS'" < Q ^ SUGAR, t (4.13) otherwise, Mexico receives a smaller quota until 2008 when access to the U.S. market is unlimited. Model calibration The model is calibrated by positioning the intercepts of inverse linear demand and supply equations and by adjusting average transportation cost from the rest of the worid Alternatively HFCS demand can be forecasted balancing the total consumption of sweetener equation with direct and indirect consumption sugar equations, yet it resulted in a poor forecast.

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57 to the United States in order to iterate for solution from 2002 through 2015 using the mathematical programming software package GAMS. Intercepts of inverse linear demand and supply equations in quantity-price space {lUi, lUUi, IMi, and /MM; in equations [4.14] through [4.17], respectively) are calibrated with the actual values realized in 2001 (base year) from equations [C.7b], [C8b], [C9b] and [C. 10b] presented in Appendix C: lUi = us. 2001 (IU2*Q^us. 2001 + Shifter^ us. 2001 ) (4. 14) lUU] = P'us. 2001 (IUU2*Q^us.200i+ Shifter^ us. 2001 ) (4.15) IM J =P" MX. 2001 -(IM2*Q^^ MX. 2001 + Shifter'' MX. 200j) (4.16) IMM, = P^MX. 2001 (IMM2*Q^MX. 2001 + Shifter^ MX. 2001)(4. 17) By calibrating intercepts, both excess supply in Mexico (quantity presented as "A" on ESmx curve in Figure 4-1) and excess demand in the United States (quantity presented as "B" on EDus curve in Figure 4-1) are set to correspond to the actual volume realized in 2001. Slopes for both inverse demand and inverse supply curves in both countries are held constant, yet these curves shift over the forecast horizon according to the scenarios proposed in the following section. • • In order to represent Mexican sugar supply and export capacity adequately, the model is further calibrated by adjusting the average transportation cost from the rest of the world to the United States, based on the relationship expressed in equation [3.50] in chapter 3. To do so, the transportation cost is calibrated so that Mexico exports sugar over-quota at the minimum amount. This calibration procedure resulted in a rather high transportation cost from the rest of the world to the United States; however, it insinuates the irrational behavior of the Mexican sugar industry which has been suffering from

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58 financial stress, vividly illustrated by the mill expropriation by the government in 2001, and producing and exporting surplus sugar to the rest of the world at the same time. Simulated Scenarios Scenarios are formulated by considering alternative assumptions related to Mexico's sweetener market situations affected by continued gains in the productivity of the sugar industry, HFCS consumption, as well as a policy lever and U.S. sugar policy levers. Each scenario carries a combination of Mexico's sweetener market situation and U.S. sugar policy. To compare the impacts of scenarios, a baseline scenario is defined where status quo is maintained (Table 4-1). In the baseline scenario, it is assumed that shifts in sugar demand and supply in both countries continues at the average rates observed in recent years and that the U.S. government maintains price support and allocates quota among exporters in a flexible manner, abiding by the WTO minimum import requirement.^ Assumptions related to Mexico's sweetener market situations are summarized in Table 4-2. Four situations are proposed: higher sugar production, higher HFCS adoption, a combination of both and introduction of tax on HFCS as a Mexico's alternative policy lever. The rates of increase in production and HFCS adoption are defined relative to the baseline. Impacts of Mexico's tax on HFCS is based on the forecast by Haley and Suarez (2003) where Mexican HFCS consumption drops significantly in 2002 and 2003 due to a tax imposed on beverages that contain HFCS. The impact of this tax is assumed significant considering soft-drink manufacturers currently account for about one-third of domestic sugar consumption in Mexico (Buzzanell, 2002). In all situations, U.S. demand ^ Current allocation system is based on historical trade shares (Skully, 1998).

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59 and supply are held at "Baseline" level. The duration of harvest is held constant because it is partly affected by the weather. Also, the assumption where Mexico's HFCS adoption and tax on HFCS occur at the same time is not considered since the scenario becomes incompatible to simulate. The scenarios associated with "HFCS adoption" are consistent with an increasing trend since 1994 when NAFTA went into effect and speculation of expansion of HFCS production in Mexico. If Mexico continues to increase HFCS consumption, it may follow a similar path as was seen in the United States in early 1980s when soft-drink manufacturers decided to switch to HFCS from sugar. HFCS manufacturers, who are mostly the U.S. corporations, see this phenomenon as a business opportunity in Mexico. With HFCS capital-intensive facilities, existing HFCS plants in Mexico are operated by the firms based in the United States. The "HFCS Adoption" case assumes that HFCS will be adopted in a linear fashion until HFCS consumption is 50 percent of total indirect sweetener consumption in 2008 and its share remains constant for the rest of the forecast horizon. The 50 percent share of indirect consumption of sweetener is equivalent to about 27 percent of total consumption of sweeteners. In 2001 (base year), share of HFCS in indirect sweetener consumption and total sweetener consumption were 25.3 percent and 1 1.6 percent, respectively. Comparisons of HFCS and indirect sugar consumption forecast for "Baseline" scenario, "HFCS adoption", and "Tax on HFCS" situations are illustrated in Figure 4.2. Assumptions related to U.S. sugar policy levers are summarized in Table 4-3. Two kinds of policy levers are considered: one is to stabilize the demand price and the other is to allocate quotas to exporters. For the former, two possible sugar policies are used as

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60 alternatives to the current price support. In one policy, instead of directly supporting sugar price, the government indirectly supports a sugar price through buying up excess sugar in the market to assure that sugar price will not fall due to excess supply from overseas. In this assumption, the government buys up as much sugar as the net sugar import, i.e. total sugar import less minimum import requirement. In the bilateral trade model, the cost of buying up excess sugar is not included in the objective function, assuming that the government does not spend ex ante cost. Also for simplicity, sugar storage costs incurred by the government are ignored. The second policy option is to introduce sugar production controls in both the United States and Mexico. This option requires cooperation from the Mexican sugar industry: defection by either party will result in an unsuccessful outcome. In this way, both the United States and Mexico are assumed to control their sugar production according to forecasted demand, avoiding excess supply of sugar. In other words, the idea implies that the United States is willing to import sugar from Mexico as long as Mexico cooperates to reduce its sugar production to meet the sugar demand in both countries and also that Mexico can avoid excess surplus sugar which cannot be sold anywhere except in the world market. This sugar policy does not involve financial support from the government: the cost of the program is zero. U.S. policy levers related to quota allocations to exporters are treated with two different approaches: the U.S. government allocates import quotas in a flexible manner between Mexico and the rest of the world (status quo); and minimum quotas are maintained (the remainder of the minimum import requirement less that allocated to Mexico) to the rest of the world no matter how much Mexico exports to the United

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61 States. Both policies abide by the minimum import requirement under WTO; however, political feasibility is assumed to be quite different. Scenarios to simulate are formulated by combining specific Mexico's sweetener market situation and the U.S. sugar policy. A list of 16 scenarios is shown in Table 4-4. Finally, special simulations are prepared in order to further examine the effects of Mexico's production improvement and HFCS adoption on the Mexican sugar industry and welfare. In addition to the assumption regarding Mexico's production improvement (an additional 1 percent to the baseline) and HFCS adoption (share of indirect sweetener consumption by HFCS at 50 percent) summarized in Table 4-2, simulations are conducted by changing production improvement rate at additional 0.5 and 1.5 percent as well as HFCS adoption to achieve a market share of 30, 40, and 45 percent. Game Theory Analysis In order to assess policy recommendations using aggregated results from the various simulations, an analysis based upon game theory is introduced. The basis of the game used in the study is a non-zero-sum game with mixed strategies. Non-zero-sum means the sum of the pay-offs in each pair of strategies is not zero; in other words, one player's winning does not necessarily cause the other to lose. Mixed strategies means a player chooses a strategy to play with probability (Morris, 1994; Mas-Colell et al., 1995). Since the game is non-zero-sum, both cooperative and non-cooperative games are considered. While a cooperative game allows players to make binding agreements about how they will play or about sharing pay-offs, a non-cooperative game does not. In the latter case, the game is played by two parties: the U.S. and the Mexican governments who hold the strategies and make decisions on behalf of the economy as a whole. In the former case, the game goes through the process of considering the possible pay-offs to

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62 two coalitions formed among five parties (payees), i.e. three industries (the U.S. HFCS industry, the U.S. sugar industry and the Mexican sugar industry) and two countries; however, the final decisions are made by the governments who hold the strategies. Each country plays with multiple strategies that correspond to Mexico's market situations and the U.S. policy levers presented in the previous section (Table 4-5). In this game setting, a combination of strategies formulates a scenario. In the game, it is assumed that if the U.S. government introduces production control, Mexico always cooperates: a defection by either party is not considered. Also, the U.S. government chooses only flexible quota allocations to the rest of the world for simplicity. In the case of Mexico's strategies, HFCS adoption is treated as a strategy although it is neither a positive strategy nor controllable by the Mexican government. Pay-offs from each scenario are calculated for each payee. Pay-offs to the industries are expressed as present values of accumulated revenue between 2002 and 2015, assuming a three percent discount rate each year. The HFCS price is held constant at the average U.S. export price to Mexico realized between 1992 and 2001. Pay-offs to each country are expressed as present values of accumulated welfare, i.e. the sum of consumer and producer surplus. In doing so, changes in welfare are calculated only from the sugar market, assuming that sugar is a primal source of sweetener. Sugar cannot be substituted with HFCS for certain products due to the liquid form of HFCS. Also, sugar is preferred for certain products to HFCS due to flavor given to the final products. U.S. welfare is adjusted with tariff revenue from Mexico, the cost of the sugar program and the cost of buying up excess sugar. The cost of the sugar program is calculated by multiplying the price difference between the support price (loan rate for raw sugar, 18

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63 cents per pound) and the U.S. equilibrium price with the production quantity guaranteed at the loan rate. This cost is captured by comparing between scenarios with and without the price support, ceteris paribus. The cost of buying up excess sugar is calculated by multiplying the U.S. net sugar import, i.e. total sugar import less 1,256,000 MT of minimum import requirement with the U.S. equilibrium price. In the case of the policy to buy excess sugar policy, sugar storage costs incurred by the government are ignored. Note that since the values were converted in terms of U.S. dollars prior to simulations, the exchange rate realized in the base year (2001) was implicitly used for calculating payoffs. All the games proposed are solved through a two-players pay-off matrix (twodimensions) with the two governments are the decision-makers. When coalitions are formed, their individual pay-offs are pooled, assuming that the total pay-off is redistributed among them (Morris, 1994). In doing so, it is also assumed that industry revenue and each nation's welfare can be added together. After pay-offs are calculated for each party or coalition, these values are indexed as a relative gain or loss to the payoffs in baseline scenario to facilitate the decision process. Mixed-strategy games are first simplified by eliminating dominated strategies (strategies that played with zero probability) and then solved through maximizing the expected pay-off to each player from the game (Morris, 1994; Varian, 1992). Sources of Data Data for the Mexican sugar industry was obtained from the website of Comite de la Agroindustria Azucarera (COAAZUCAR, Sugar Agro-Industry Committee). The committee is in charge of monitoring sugar cane and sugar production at each mill as well as determining the cane price paid to farmers in the country. The latter task was

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64 taken over from the former government body after the privatization of the miUing sector. The committee carries an extensive data set regarding not only physical production and price but also detailed productivity and efficiency indicators such as sugar and fiber content in cane, mill downtime and sugar production loss during the process across 60 operating mills. Data for the U.S. sugar industry was obtained from the Sugar and Sweetener Situation and Outlook Yearbook and other publications by the Economic Research Service, USDA. Historic data for population were obtained form the website of the U.S. Bureau of the Census; and those for GDP, exchange rates and consumer price index were taken from OECD documents. Data sources used in the study are summarized in Tables 4-6 through 4-10. For demand analysis, aggregate time-series data at the national level are used in the regressions for both the United States and Mexico. Missing data were found in U.S. sugar consumption and related prices in 1991 and filled with average values of two adjacent years in order to maintain the data continuity. For the supply analysis, aggregate time-series data at industry level are used in the regression for both the United States and Mexico. No missing data were found in the data set. Nominal values of old Mexican pesos before devaluation found in the data are converted into current pesos. Local units are preserved during the estimation of elasticities in demand and supply analyses. In bilateral trade model, different local units are converted into common units such as metric tons and US dollars so that the model can achieve the equilibrium point in the system.

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65

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66 Table 4-1. Assumptions for Baseline (Status quo) Scenario Category Shifters Country Assumptions GDP United States and Will increase at the average real GDP annual growth rate realized between 1997 and 2001 (2.23 percent for the United States and 2.91 percent for Mexico). Market situations Population Mexico Will increase similarly as the forecast by the U.S. Bureau of the Census. (Annual growth rate: 0.88 percent for the United states and 1.15 percent for Mexico). Production cost Will decrease at the average annual reduction rate realized between 1997 and 2001 (1.10 percent for the United States and 1.80 percent for Mexico). Recovery rate United States Will increase at the average annual improvement rate realized between 1997 and 2001 (0.48 percent). Downtime Mexico Will decrease at the average annual reduction rate realized betweenl997 and 2001 (1.00 percent). Sugar loss Will decrease at the average annual reduction rate realized between nl997 and 2001 (2.00 percent). Duration of harvest Held constant HFCS consumption Will be consumed maintaining the same share in indirect consumption of sweetener realized in 2001 (25.3 percent) U.S. sugar Price support npi_ T T O A • i. The U.S. government maintains the pnce support (current policy) at 18 cents per pound (loan rate). policy Flexible allocation The U.S. government allocates quotas in a flexible manner between Mexico and the rest of the world.

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67 Table 4-2. Assumptions for Mexican Sweetener Market Situations Situations (code) Shifters Assumptions High Mexican Production (P) Production cost, Downtime, Sugar loss Will improve an additional 1 percent to the "Baseline". High MFCS Adoption (A) HFCS consumption It is assumed that HFCS consumption will increase in a linear fashion until it replaces 50 percentage or indirect consumption of sugar in 2008. After 2008, its share remains at 50 percent. High Mexican Poduction High HFCS adoption (PA) Production cost. Downtime, Sugar loss The same as "High Mexican Production" HFCS consumption The same as "HFCS Adoption" Tax on HFCS (T) HFCS consumption It is assumed that HFCS consumption for 2002 through 2004 will drop due to tax (Haley and Suarez, 2003) and remain at reduced consumption level (7 percent to the indirect consumption of sugar) for the rest of the forecast horizon.^ Other shifters are the same as the Baseline. A 20-percent tax on beverages that contain HFCS was introduced on January 1, 2002; suspended on March 5 by the president's decision; and then reimposed on July 16, 2002 with the decision by Mexico's Supreme Court of Justice (USDA, 2002a).

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68 6,000,000 5,000,000 H 4,000,000 g 3,000,000 e ^ 2,000,000 1,000,000 Year Indirect consumption of sugar (Baseline) — Indirect consumption of sugar (Tax on HFCS) -A— Indirect consumption of sugar (High HFCS adoption) 0 HFCS consumption (Baseline) -B— HFCS consumption (Tax on HFCS) -A — HFCS consumption (High HFCS adoption) Figure 4-2. Forecasted Indirect Sugar and HFCS Consumption in Mexico under Alternative Scenarios

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69 Table 4-3. Assumptions for U.S. Sugar Policies Categories Policies (code) Assumptions Stabilization of the demand price Price support (status quo) (S) rill T T A A A i A The U.S. government mamtams the pnce support at 18 cents per pound (loan rate, 396.48 US$ per MT). Buying up excess sugar in the market (B) The U.S. government abandons the price support and buys up excess sugar in the market instead. Production control (C) The U.S. government abandons the price. Instead, the U.S. and Mexican governments collectively control the sugar production in such a way that the sum of quantities demanded in two countries is primarily met by the sum of the quantities supplied from two countries. Quota allocations Flexible allocation (status quo) (F) The U.S. government allocates quotas in a flexible manner between Mexico and the rest of the world. Minimum quota allocations for the rest of the world (M) The U.S. government reserves the minimum quotas (the remaining minimum import requirement less allocated to Mexico) for the rest of the world.

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70 Table 4-4. Listing of Examined Scenarios Scenario Mexican market U.S. sugar policies number (code) situations Stabihzation of the demand price Quota allocations 1 (baseline) Basehne (B) Price support Flexible 2 (P-S-F) High production (P) (S) allocations (F) 3 (A-S-F) HFCS adoption (A) 4 (HA-S-F) TT* 1 J Til 0 1 High prod.-HFCS adop. (PA) 5 (B-B-F) Basehne (B) Buying up excess 6 (P-B-F) T T' 1 J i High production (P) sugar in the market (B) 7 (A-B-F) HFCS adoption (A) 8 (PA-B-F) High prod.-HFCS adop. (PA) 9 (B-C-F) Baseline (B) Production control 10 (P-C-F) High production (P) (C) 11 (A-C-F) HFCS adoption (A) 12 (PA-C-F) High prod.-HFCS adop. (PA) 13 (PA-S-M) TT" 1 1 Til '^"1 1 High prod.-HFCS adop. Price support (S) Minimum quotas allocation to the 14 (^r A-D-iVij (PA) Buying up excess sugar in the market (B) rest of the world (M) 15 (PA-C-M) Production control (C) 16 (T-S-F) Tax on HFCS (T) Price support (S) Flexible allocations (F)

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71 Table 4-5. Strategies for the Sugar Trading Game Country Strategies United States Strategy 1 Maintains price support (status quo) Flexible quota allocations among Strategy 2 Abandons price support and buys up excess sugar in the market Mexico and the rest of the world Strategy 3 Introduces production control with Mexico Mexico Strategy 1 Maintains the current policy (status quo) Strategy 2 Higher sugar production Strategy 3 Higher HFCS adoption Strategy 4 Higher sugar production and higher HFCS adoption Strategy 5 Introduces tax on HFCS

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72 Table 4-6. Data Sources for U.S. Demand Data Unit Source Consumption of sugar (dependent variable) 1000 short tons Sugar Statistical Compendium by (Stock #91006, 1970-1990) and Sugar and Sweetener Situation and Outlook Yearbook (SSS-2002, 1992-2002) by Economic Research Service, U.S. Department of Agriculture Retail price of refined sugar cents / pound Gross Domestic Product (GDP) US$ Statistic database by Organization tor Economic Co-operation and Development (OECD) Population persons International Database by U.S. Bureau of the Census, U.S. Department of Commerce Data length: 1970-2002, quarter y Table 4-7. 1 Data Sources for Mexican Demand Data Unit Source Direct consumption of sugar (dependent variable) metric tons [MT] Azucar S.A. de C.V. Estadistica Azucareras (1970-1989), database by Financiera Nacional Azucarera, S.N.C de C.V. and Comite de la Agroindustria Azucarera (CO A AZUCAR) (1990-1999) Indirect consumption of sugar (dependent variable) Total consumption of sugar (dependent vanable) Retail price of standard sugar pesos / kg Gross Domestic Product (GDP) pesos Statistic database by Organization for Economic Co-operation and Development (OECD) Population persons International Database by U.S. Bureau of the Census, U.S. Department of Commerce Data length: 1970-1999

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73 Table 4-8. Data Sources for U.S. Supply Data Unit Source Production of sugar, total (dependent variable) 1000 short tons Sugar Statistical Compendium by (Stock #91006, 1970-1990) and Sugar and Sweetener Situation and Outlook Yearbook (SSS-2002, 1980-2002) by Economic Research Service, U.S. Department of Agriculture Production of cane sugar (dependent variable) Production of beet sugar (dependent variable) Retail price of refined sugar Wholesale price of refined cents / pound Sugar recovery rate, total percent Beet sugar recovery rate Cane sugar recovery rate Total farm production expenses million US$ Database by Economic Research Service, U.S. Department of Agriculture Data length: 1960-2002 Table 4-9. Data Sources for Mexico Supply Data Unit Source Production of sugar (dependent variable) MT Database by Comite de la Agroindustria Azucarera (COAAZUCAR) Wholesale price of standard sugar pesos / kg Cost of sugar production per ton of sugar produced pesos / ton of sugar Downtime observed at mills percent Sugar loss during the process percent Duration of the harvest days Data length: 1988-2000

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74 Table 4-10. Data Sources for Miscellaneous Data Source U.S. Consumer Price Index (CPI) (19821984=100) Database by the Bureau of Labor Statistics, U.S. Department of Labor Mexico Consumer Price Index (CPI) (1994=100) Database by the Banco de Mexico Exchange rate (US$Mexican pesos) Quota-tariff rates Agricultural Outlook (1999) by Economic Research Service, U.S. Department of Agriculture Sugar per unit transportation cost (bagged, seaborne freight rate) Personal communication with an exporter of sugar Loan rate for raw sugar in the U.S. Haley and Suarez (2002) Guarantee price for raw sugar in Mexico Database by Comite de la Agroindustria Azucarera (COAAZUCAR)

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CHAPTER 5 EMPIRICAL RESULTS AND INTERPRETATION This chapter presents and discusses the empirical results for each of three analyses: (1) demand and supply analysis for both the U.S. and Mexican sugar markets, (2) bilateral sugar trade analysis, and (3) game theory analysis. Demand and Supply Analyses Results for the U.S. demand and supply analysis are summarized in Table 5-1. In the demand equation, signs of estimates associated with each significant variable were as expected. Significant estimates at the 95 percent confidence level were associated with price, the dummy variables for quarter 1 and 3, and the dummy variable for HFCS availability. The estimated price elasticity of demand was inelastic. The significant estimate associated with the HFCS dummy variable implies that HFCS replaces sugar as a substitute in the market to some degree. In the supply equation for the United States, estimates associated with trend and production in the previous year (autoregressive term) were significant at the 95 percent confidence level for all three models, i.e. total, beet and cane sugar supply regression models. Estimates associated with sugar recovery rate were insignificant in all models. Estimates associated with price and cost were significant at the 95 confidence level for total and beet sugar supply regressions, but not for the cane sugar supply equation. Two possible reasons why cane sugar production does not respond to the refined sugar price but beet sugar production does are: (1) cane sugar has two steps in the refinery process while beet sugar has one and (2) sugarcane is a perennial crop while sugar beets is an 75

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76 annual crop. These results reflect these differences and imply that sugar beet production is more sensitive to price changes. Although cane sugar production is assumed to respond to raw sugar price, the coefficient of the price variable is not significant. Estimated price elasticities were both inelastic for total sugar supply and beet sugar supply as anticipated. The long-run supply price elasticities were calculated dividing the estimated price elasticities (short-run) by (1X) where X represents the estimates for production in the previous year (autoregressive term). Computation yielded all inelastic long-run price elasticities: 0.3875 and 0.6764 for total sugar and beet sugar, respectively. Results for the demand and supply analysis for Mexico are summarized in Table 52. In the demand equation, signs of significant estimates associated with each variable were consistent with a priori expectations. The only statistically significant estimate among the three price elasticities was direct consumption, and it was inelastic. The population variable accounted for most of the explanatory power of consumption in all models. Significant estimates associated with GDP in indirect sugar and total sweetener consumption indicate that consumers tend to consume more sugar through sugarcontaining products as their income increases. In the supply equation for Mexico, the signs of estimates associated with each variable corresponded with a priori expectations. The estimate associated with price was inelastic. While reduction in production cost and factory downtime indicated an increase in production, the length of sugarcane harvest duration was almost perfectly correlated to sugar output from the mills. The coefficient for the variable representing sugar loss during the process was not significantly related to sugar output implying that the degree of sugar loss was not as critical as other factors such as production cost and factory

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77 downtime for mills trying to improve their production efficiency. A positive and significant estimate associated with the trend variable indicates technology related to sugar production at sugar mills had been advancing during the time period covered in this study (1988-2000). V Bilateral Sugar Trade Analysis The results from the bilateral sugar trade analysis are divided into two parts: U.S. sugar import forecasts and the comparison of pay-offs among industries and countries. Results and discussion are guided by comparing the impacts from changes in Mexican sweetener market situations and from changes in U.S. sugar policy. U.S. sugar import forecasts for the selected eight scenarios (scenario 1, 2, 3, 4, 8, 12, 13, and 16 in Table 4-4) are shown in Figures 5-1 through 5-8. Comparisons among scenarios 1, 2, 3, 4, and 16 illustrate the impacts from changes in the Mexican sweetener market situations; comparisons among scenarios 4, 8, and 12 illustrate the impacts from changes in U.S. sugar price stabilization policy; and comparison between scenarios 4 and 13 illustrates the impacts from changes in U.S. quota allocation policy. The results from the baseline scenario (Scenario 1) shows that if status quo surrounding the sugar industries in both countries is maintained over the forecast horizon, Mexico will not likely attain a net surplus sweetener producer status and hence will miss the opportunity to benefit from exporting sugar under a larger quota allocation (250,000 MT) under NAFTA (Figure 5-1). This is due to growing domestic sugar demand relative to domestic sugar production. For comparison, forecasts for Mexican sugar consumption and production in 2008 by this study are 5.4 million and 5.8 million MT, respectively and those by Koo and Taylor (2000) are 5.3 million and six million MT, respectively.

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78 When Mexico expands sugar production (Scenario 2), net surplus sweetener producer status will still not be attained (Figure 5-2). Yet, Mexico will generate enough surplus sugar to export over-quota (before 2008) and quota-free (after 2008), resulting in significant impacts on the U.S. market. In total, Mexican sugar will take up about onethird (Scenario 1) or more than half (Scenario 2) of the U.S. minimum import requirement at peak in 2008. The amount of export will decline in later years due to expanding domestic sugar consumption in Mexico. In Scenario 3, when Mexico adopts HFCS at a higher rate, Mexico's sugar export swells as a result of substitution between sugar and HFCS in the domestic market. This result includes direct impacts on the Mexican sweetener market as well as extended impacts on the U.S. sweetener market (Figure 5-3). Although Mexico will not attain net surplus sweetener producer status, over-quota export will reach over 1.2 million MT by 2007 and will remain over one million MT until 2014. This export quantity will take up almost the entire U.S. minimum import requirement and as a result, sugar export from the rest of the world will be marginalized. Similar results are drawn when increases in Mexico's production and HFCS adoption are combined (Scenario 4). A slightly larger scale of Mexico's exports than Scenario 3 is shown in Figure 5-4. The policy followed by the U.S. government in its allocation of its import quota has a large impact on sugar exports from both Mexico and the rest of the world. The aforementioned large-scale export of Mexican sugar is possible only if the U.S. imports the minimum amount of sugar and allocates sugar quotas in a flexible manner among exporters. This allocation method may cause friction with the other countries that export sugar to the United States since Mexican sugar has potential to take up a large portion of

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79 the U.S. quota. Thus, allocating such a large portion of the quota to one country may not be a feasible policy option in the United States. In Scenario 13, it is conjectured that the U.S. government maintains minimum quotas (the remainder of the minimum import requirement less allocated to Mexico) for the rest of the world no matter how much Mexico exports. As shown in Figure 5-5, Mexico's over-quota and quota-free export will be dampened because of Mexico's comparative disadvantage to the rest of the world, while the export from the rest of the world remains over 1.2 million MT over the entire forecast horizon. By contrast, a change in the U.S. government's policy on stabilization of the domestic price does not pose much impact on sugar exports to the U.S. market. With the same Mexican sweetener market situations (high production-high HFCS adoption), the U.S. policy options of price support, buying up excess sugar in the market, and production controls are compared (Figures 5-4, 5-6 and 5-7). The results indicate that all three scenarios bring about the similar trends in Mexican sugar export. Yet, U.S. production control with Mexico causes an overall increase in quota-free export after 2008. This is because the Mexican sugar price is maintained lower relative to the price support or buying up excess sugar scenarios (Figures 5-9, 5-10, and 5-11). The prices of sugar in both countries will converge in the integrated market without a U.S. price support (Figure 5-10) while the difference in prices are kept after 2008 with production control (Figure 5-11): the price difference corresponds to the transportation cost from Mexico to the United States. Mexico's tax on HFCS brings about very different results. In this scenario, Mexico is unable not only to export either under-quota, over-quota or quota-free but also

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80 to generate a sugar surplus after 2006 due to expanding domestic sugar consumption accelerated by higher sugar consumption by bulk users who chose sugar over HFCS. Mexico cannot produce enough sugar to meet domestic demand after 2008 and will import sugar from the United States, resulting in a higher domestic sugar price than that of the U.S. (Figure 5-12). This extreme case would occur only if the impact of tax on HFCS lingers over the forecast horizon as assumed in this study; however, the chance for Mexico to enjoy exporting sugar would be slim. In spite of fluctuating imports from Mexico and the rest of the world, U.S. domestic sugar consumption and production will remain relatively unchanged. Sugar demand and supply forecasts for both the United States and Mexico for baseline and high productionhigh HFCS adoption scenarios are shown in Figures 5-13 and 5-14, respectively. In either scenario, U.S. demand and supply are forecasted to remain approximately at nine million and 8.8 million MT over the forecast horizon, respectively. The results from the various simulations showed that the U.S. sugar price will gradually decline but will not dip below the support price level (396.48 US$ per MT) before 2008 if the United States accepts most of the imported sugar from Mexico rather than from the rest of the world by allocating quotas in a flexible manner to the exporters, no matter which U.S. demand price stabilization policy is in place (Figures 5-9, 5-10, and 5-11). This implies that the Mexican sugar price contributes to maintain a high sugar price in the integrated U.S.-Mexico sugar market; in other words, accommodating Mexican sugar can act as an alternative form of price support in the United States. On the other hand if the U.S. government maintains minimum quotas for the rest of the world no matter how much Mexico exports and abandons the price support, U.S. equilibrium price

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81 will fall below the support price (Figures 5-15 and 5-16). This is a result of cheaper sugar from the rest of the world flowing into the U.S. market. In reality, the U.S. sugar price will face downward pressure from importing world sugar as well as political pressure from the rest of the world, considering the likelihood that many sugar exporting countries will not easily give up their existing shares of the U.S. import quotas. Pay-offs to the industries and countries also portray interesting contrasts among scenarios. The results from three sets of selected scenarios are summarized in Tables 5-3, 5-4, and 5-5. In these tables, the present values of accumulated pay-offs are expressed in billion of dollars and those values are indexed relative to the baseline scenario inside the brackets. The impact of changes in Mexican sweetener market on pay-offs to the industries and the two nations' welfare is illustrated in Table 5-3. The listed five scenarios are based on the assumptions that the U.S. government maintains price support and allocates quotas between Mexico and the rest of the world in a flexible manner. It is clear that the U.S. HFCS industry will become better off if Mexico adopts HFCS: revenue for the U.S. HFCS industry increases by 78 percent; and that the industry will become worse off if Mexico introduces taxes on HFCS: revenue for the industry decreases by 72 percent. The U.S. sugar industry does not gain from either Mexico's increase in sugar production or HFCS adoption since either change in Mexican sweetener market generates a larger Mexican sugar surplus that is destined for the U.S. market. Tariff revenue to the U.S. government from Mexican sugar increases as Mexico increases sugar production or HFCS adoption. Large tariff revenue from high HFCS adoption scenario, which is unexpectedly larger than the high production-high HFCS adoption scenario, is due to

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82 forecasted larger exports in 2008 and 2009, which consequently brings a lower U.S. sugar price accompanied by a higher cost of the price support. Overall, U.S. welfare becomes worse off with changes in Mexican sweetener market for two reasons: (1) reduced producer surplus caused by a lower producer price as a result of increased Mexican export and (2) increased net costs even though increased tariff revenue is expected as mentioned above. The Mexican sugar industry gains from expanding sugar production, but the gains are dissipated when HFCS is adopted at higher rates, given the assumption that HFCS price held constant. In Mexico's HFCS tax scenario, the Mexican sugar industry gains not from exporting to the U.S. market but from domestic sales at higher prices. All the entities except for U.S. HFCS industry benefit from this policy; however, the policy lever may not be acceptable in the international trade environment. In fact, the Mexican government swung its decisions in the past: a 20-percent tax on beverages that contain HFCS was introduced on January 1, 2002; suspended on March 5 by the president's decision; and then reimposed on July 16, 2002 with the decision by Mexico's Supreme Court of Justice (USDA, 2002a). The impact of changes in the U.S. price stabilization poHcy is illustrated in Table 54. The compared three scenarios (Scenario 4, 8, and 12) are based on the assumption that Mexico increases sugar production as well as HFCS adoption and that the U.S. government allocates quotas between Mexico and the rest of the world in a flexible manner. Alternative sugar policies to the price support only bring about improvement in U.S. welfare; sugar industries in both countries and Mexico's welfare become worse off, posing a larger negative impact on the Mexican sugar industry and welfare than the sugar industry in its own country. When the U.S. government switches sugar policy from

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83 supporting price directly to buying up excess sugar in the marlcet, the cost of the program is expected to diminish, resulting in welfare improvement. Yet, if the U.S. government were to switch policies from the price support to buying up excess sugar, the timing to do so will be important so as to minimize the cost incurred by the government; the cost of buying up excess sugar will rise immediately after policies are switched while the cost of the price support will not because the U.S. sugar price will be maintained relatively high in the early stage of the forecast horizon. In practice, storage costs need to be considered. When the United States controls sugar production with Mexico, welfare improves by increased consumer surplus and tariff revenue combined with zero program cost incurred by the government. This scenario also demonstrates that U.S. welfare becomes better off at the expense of the sugar industries in both countries and Mexico's welfare. When the policy of buying up excess sugar and the production control policy are compared, the U.S. government could pursue the latter in light of the nation's welfare; however, the gain is very small compared to the loss bom by industries and Mexico. Both alternative policies do not satisfy pareto optimality and the overall loss outweighs the gain by the United States as a nation. The impact of changes in U.S. price stabilization policy is illustrated in Table 5-5 with the assumption that the U.S. government maintains the minimum quotas for the rest of the world no matter how much Mexico exports. When the U.S. government introduces alternative policies to price support, U.S. welfare gains but both the U.S. and Mexican sugar industries as well as Mexico's welfare lose to a larger degree. An extreme result comes from Scenario 13. In this scenario, the U.S. sugar price support becomes extremely costly if the U.S. government reserves the minimum quota for the rest of the

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84 world. As a result of cheap sugar from the rest of the world flowing into the U.S. market, the U.S. price will fall far below the support price. The loss bom by the Mexican sugar industry particularly stands out: nearly a half of its expected revenue disappears due to restricted access to the U.S. market. When fostering the sugar industry in its own country and Mexico as a neighboring trade partner, the U.S. government needs to accept sugar from Mexico more than from the rest of the world. This requires sensitive negotiations among exporters. Overall, the Mexican sugar industry and welfare are more prone to changes in the Mexican sweetener market and U.S. sugar policy than the U.S. sugar industry and U.S. welfare. Among tested changes, Mexico's HFCS adoption and U.S.'s quota allocation policy hold the greatest effects. Effects of Mexico's production improvement and HFCS adoption on the Mexican sugar industry and welfare are illustrated in Figures 5-17, 5-18, and 5-19. The rate of production improvement ranges from the average rate realized between 1997 and 2001 (baseline) to additional 0.5, I.O, and 1.5 percent to the average rate; the HFCS adoption ranges from 25 percent share of indirect sweetener consumption (baseline) to 30, 40, 45, and 50 percent (HFCS adoption situation). The effects on the Mexican sugar industry are shown in Figure 5-17. Expected accumulated revenue increases as sugar production improves an additional 1 and 1.5 percent above the average rate. An increase in HFCS adoption also brings about an increase in revenue until the share attained by HFCS climbs up to 40 percent. When HFCS share reaches 50 percent, revenue shrinks compared to the baseline. Increased revenue caused by HFCS adoption is due to higher domestic sugar price and decreased revenue is due to decreased quantity of sugar demanded. The effects

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85 on Mexico's welfare differ from those on the industry (Figure 5-18). Production improvement increases welfare, but not HFCS adoption, assuming that sugar is a primary and preferred source of sweetener. While the gains to the Mexican sugar industry from production improvement at 1.5 percent to the baseline (US$ 0.65 billion) can make up the loss to the industry itself from HFCS adoption, even at a 50 percent share (US$ 0.42 billion), gains to Mexico's welfare cannot. The net loss to the nation from HFCS adoption at 30 percent adoption share is forecasted US$ 5.12 billion (loss of US$ 5.41 billion from welfare and a gain of US$ 0.29 billion from the industry).' This loss is far greater than the sum of expected gains from production improvement at additional 1.5 percent, which is US$ 3.42 billion (gains of US$ 2.77 billion from welfare and US$ 0.65 billion from the industry). The Mexican government faces difficulties allowing faster HFCS adoption to happen in the domestic market (Figure 5-19). Game Theory Analysis Results from the game theory analysis are summarized in Tables 5-6 through 5-13. The analysis is based on the game setting played by the United States and Mexico with multiple strategies that correspond to Mexico's market situation (the Mexican government's strategy set) and the U.S. policy levers (the U.S. government's strategy set) in order to assess gainers and losers from trade. Calculated actual pay-offs to five payees (the U.S. HFCS industry, the U.S. sugar industry, Mexican sugar industry, U.S. costadjusted welfare, and Mexico's welfare) for each combination of U.S. and Mexico's strategies of the game are shown in Table 5-6. Values in the other tables are indexed relative to the baseline scenario. Three different forms of coalitions are considered: Assuming that industry's revenue and nation's welfare can be added together.

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86 coalition of countries, coalition of the sugar industries, and grand coalition that includes all five entities. Since the inclusion of the U.S. HFCS industry into a coalition is a determinant factor, both coalitions with and without the U.S. HFCS industry are also considered. Pay-offs to the U.S. HFCS industry and the government of Mexico's welfare fluctuate more than the other three payees (Table 5-6). Among the five strategies facing Mexico, introduction of tax on HFCS returns the best pay-offs to the Mexican sugar industry and welfare and thus the Mexican government would always choose this strategy. Yet, this policy lever may not be acceptable in the international trade environment. In the following games, Mexico's tax strategy is excluded. Indexed pay-offs are shown in Table 5-7. Values in corresponding cells are categorized in four ways: no change (100), gain (over 100, cells shaded in gray), slight loss (over 95 and under 100), and loss (less than 95, cells shaded with stripes). In this game, there is neither dominant strategy (a strategy that is chosen over other strategies) nor pure strategy (a single strategy chosen by the government that brings about improvement in the industry or industries as well as welfare in its own country: this strategy is played with probability of I for either U.S. or Mexican government). By inspecting the tendency of pay-offs, some interesting contrasts among payees rise to the surface without solving mathematically. In the United States, the government always prefers a production control strategy to the other two but the sugar industry is always better off with the price support strategy, no matter which strategy Mexico chooses. As Mexico increases sugar production or HFCS adoption, the U.S. sugar industry likely loses while the U.S. government and the HFCS industry never become worse off. The

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87 U.S. sugar industry will lobby against a production control strategy for fear that it may lose up to five percent (high production-high adoption market situation). On the contrary, the Mexican government's choice of strategy will be accepted by the industry in most cases, except for the case of where the U.S. government buys sugar in the baseline scenario. Since HFCS adoption harms both welfare and the industry, the Mexican government will continue to struggle to suppress HFCS adoption in its market. Results from the game played by two coalitions of countries (the United States and Mexico) are shown in Table 5-8. In this game, the U.S. HFCS industry is excluded from the game. As expected, the high production strategy becomes Mexico's pure strategy and thus the game is solved when the U.S. government chooses a strategy: production control policy (cells shaded with vertical stripes). This combination of strategies is coincidentally the best choice for the U.S. Coalition, but not for the Mexican coalition. The Mexican government would prefer the U.S. government to choose either the price support or buying up excess sugar strategies. Results from the game played by the sugar industry coalition and the government coalition are shown in Table 5-9. The high production strategy becomes a pure strategy for Mexico when a decision is made by the government coalition. The solution of the game is thus determined when the U.S. government chooses the strategy of buying up excess sugar. This result differs from the game played by the country coalitions mentioned above. By a government cooperating with the other government rather than with the industry in its own country, the Mexican government improves its pay-off by US $4.75 bilHon; the U.S. government loses by US $0.24 billion; and the Mexican sugar industry loses US $1.27 billion (Table 5-4). Since the loss bom by the U.S. government is

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88 relatively small, the choice by the Mexican government is made at the expense of its own sugar industry. On the contrary, the industry coalition would lobby for the U.S. production control strategy, assuming that the Mexican sugar industry promises to compensate the loss of US $0.01 billion (US $10 million) bom by the U.S. sugar industry with expected gain of US $1.27 billion (Table 5-6). With this contrast of the results between a country coalition (Table 5-8) and a government/industry coalition (Table 5-9), the U.S. government would likely form a coalition with its own industry while the Mexican government would try to form a coalition with the U.S. government. Results from the game played by the grand coalition that includes all players except the U.S. HFCS industry are shown in Table 5-10. The solution is the Mexican government high production strategy and the U.S. government buying excess sugar strategy. Although this solution does not satisfy pareto optimality for all four payees, the overall loss is minimal: the loss incurred by the U.S. sugar industry is US$ 0.31 billion (Table 5-6) and is theoretically compensated by the total gain of US$ 2.29 billion (US$ 0.36 billion from the U.S. government, US$ 0.13 billion from the Mexican sugar industry, and US$ 1.80 billion from Mexico's welfare. Table 5-6) This solution corresponds to the government/industry coalition seen in Table 5-9. In other words, if the U.S. government sees the benefit from pooling gains with Mexico rather than with its industry and if redistribution of gains is possible among governments and industries, no one loses from the game. Such an arrangement and agreement in practice would be expected to be difficult to reach and implement. When the U.S. HFCS industry is included in coalitions, the same solutions are reached: the combination of Mexican government high production strategy and U.S.

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89 government production control strategy for country coalitions (Table 5-11), Mexican government high production strategy and the U.S. government buying up excess sugar strategy for government/industry coalitions, and Mexican govemmet high production strategy and the U.S. government buying up excess sugar strategy for grand coalition. Yet, pooled gains become larger due to inclusion of gains from the U.S. HFCS industry, the pay-off matrices offer additional factors that need to be taken into the policy assessment process. Results from the game played by country coalitions are shown in Table 5-11. Although the solution is the same as the one without the U.S. HFCS industry included, pay-offs expected from Mexican government's other strategies become more attractive to the U.S. coalition. It is obvious that the U.S. coalition would receive a better pay-off if Mexico chooses the HFCS adoption strategy. This fact gives the U.S. coalition a strong incentive to influence Mexican government's choice of strategy. Intensified conflict of interests between the industry coalition and the government coalition is illustrated in Table 5-12. Without the HFCS industry in a coalition, the industry coalition prefers the U.S. production control strategy to the buying up excess sugar strategy for the sake of slight gains. When gains from the HFCS industry are pooled, the industry coalition will lobby for strategies that involve Mexico's HFCS adoption, no matter which strategy the U.S. government plays. This is possible only if the industry coalition promises to compensate the Mexican sugar industry for the loss. If redistribution of gains among industries is feasible, the Mexican sugar industry may prefer being compensated to expecting protection from the Mexican government's strategy. Ultimately, if the Mexican government chooses to overlook the impact on

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90 welfare and let its domestic sugar industry be taken care of by the U.S. sugar and HFCS industry, there will be little reason for the Mexican government to cooperate with the United States; however, Mexico's abandoning support to the sugar and its related industries may trigger political and social instability. Lastly, the results from the game played by the grand coalition are shown in Table 5-13. Although the solution of the game is the same as one without the U.S. HFCS industry, the pay-offs from Mexican government high production-high HFCS adoption strategy are almost as good. This result implies that adopting HFCS in the Mexican market is not necessarily unbeneficial as long as Mexico increases sugar production, under the assumption that gains to the grand coalition are redistributed among countries and industries. The challenge is how to put this into practice.

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91 Table 5-1. Summary of the U.S. Supply-Demand Analysis Country United States \. Dependent ^\ variables Independent variables ^\ Demand Supply Consumption of sugar Production of sugar Production of beet sugar Production of cane sugar Number of observations 132 43 43 43 Degree of freedom 124 36 36 36 Real retail price of refined sugar -0.2323 (-4.23) ** Real per capita GDP 1) 0.1378 (0.29) Population -0.6470 (-0.71) Dummy variable (Quarter=l) -0.0707 (-5.26) Dummy variable (Quarter=2) 0.0136 (1.18) Dummy variable (Quarterns) 0.0941 (10.14)** Dummy variable for availability of HFCS -0.1853 (-3.89) ** Trend 0.0067 (2.94) ** 0.0069 (3.20) ** 0.0091 (3.08) ** Real retail price of refined sugar in the 1 ) previous year 0.1471 (2.35) ** 0.2195 (2.19)** 0.0711 (1.44) Real total farm |)roduction expenses -0.2316 (-2.19)** -0.3986 (-2.74) ** -0.0830 (-1.34) Sugar recovery rate -0.2351 (-0.57) -0.5829 (-1.55) 0.1863 (0.58) Production in the previous year 0.6204 (5.26) ** 0.6755 (5.37) ** 0.3672 (3.24) ** Constant 19.8376 (1.52) 5.9033 (2.65) ** 7.8421 (3.22) ** 5.0365 (6.12) ** Total R' 0.8645 0.8950 0.7253 0.9489 Durbin-Watson '^^ 2.0563 (1) 2.5487 (2) 2.0495 (1) 2.0325 (2) 2.0968 (1) 2.0525 (2) 2.0399 (1) 1.9725 (2) and **: Significant at 90% and 95% confident level, respectively. 1) Deflated by CPl. 2) Values are after corrected by Yule-Walker method. ( ) corresponds to the order of lag assigned.

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92 Table 5-2. Summary of the Mexican Supply-Demand Analysis Country Mexico Dependent variables Independem\^ variables Demand Supply Direct consumption of sugar Indirect consumption of sugar Total consumption of sweeteners Production of sugar Number of observations 30 30 30 13 Degree of freedom 26 25 25 6 Real retail pnce or standard sugar '* -0.0734 (-1.93)* -0.003535 (-0.07) -0.0215 (-0.98) Real per capita GDP 1) -0.2595 (-1.37) 0.9213 (2.96) ** 0.4363 (3.33) ** Population 1.1194 (8.51) ** 1.4175 (5.39) ** 1.2486 (10.92) ** Dummy variable for availability oi RbCS -0.1025 (-1.58) -0.0250 (-0.88) Real wholesale price of standard sugar in the previous year 0.2152 (2.52) Real production cost per ton of sugar -0.3228 (-7.89) ** Downtime -0.4275 (-6.77) ** Loss or sugar dunng the process -0.2112 (-1.64) Duration of the harvest 1.0010 (9.13)** Trend 0.5435 (12.75) ** -3.4499 (-2.72) ** -20.5135 (-6.90) ** -11.9624 (-8.91)** 11.8453 (20.89) ** Total R^ 0.9344 0.9729 0.9905 0.7380 Durbin-Watson 1.9217(1) 1.9961 (2) 1.5933 (1) 1.5357 (2) 1.1648 (3) 1.8715(1) 1.5834 (2) 2.9128 and **: Significant at 90% and 95% confident level, respectively. 1) Deflated by CPI. 2) Values are after corrected by Yule-Walker method. ( ) corresponds to the order of lag assigned. 3) Indicates neither positive nor negative correlation.

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93 o o o s > 1,600 1.400 1,200 1,000 800 600 H 400 200 0 J] Jl Year S Mexico under-quota export I Mexico over-quota export Mexico quota-free export 0The rest of the world export to the U.S. Figure 5-1. U.S. Sugar Import Forecast (Scenario 1 "Baseline") 1,600 1 1,400 oooooooo — — — — — — oooooooooooooo Year @ Mexico under-quota export Mexico quota-free export Mexico over-quota export The rest of the world export to the U.S. Figure 5-2. U.S. Sugar Import Forecast (Scenario 2 "P-S-F')

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94 1,600 T „ 1,400 H ^ 1,200 § 1,000 o CNC<-/TtU-)\Ot-~00CNO — (NC^. TflO OOOQOOOQ — — — — — — OOOOOOOOOOOOOO CSCNCN(N(N(Nr-4CN{NtN(NCNr-)(N Year Q Mexico under-quota export Mexico quota-free export Mexico over-quota export H The rest of the world export to the U.S. Figure 5-3. U.S. Sugar Import Forecast (Scenario 3 "A-S-F") o o o S "o 1,600 1,400 1,200 1,000 800 600 400 200 0 CN 't o o o o o o
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95 ] Mexico under-quota export I Mexico over-quota export Year Mexico quota-free export O The rest of the world export to the U.S. Figure 5-5. U.S. Sugar Import Forecast (Scenario 13 "PA-S-M") 1,600 1,400 1,200 o 1,000 o ^ 800 a > 600 400 200 a. 00 Gv O O O — o o o tS (N (N o O o O tN Year ^ Mexico under-quota export Mexico over-quota export o ts Mexico quota-free export The rest of the world export to the U.S. Figure 5-6. U.S. Sugar Import Forecast (Scenario 8 "PA-B-F")

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96 o o o 1,800 1,600 1,400 1,200 1,000 800 600 400 200 0 — j 8 o o o o >n vo t~oo ov o o o o o o o o o o n CS IN fS CN o O O O o o Year I Mexico under-quota export I Mexico over-quota export Mexico quota-free export 0The rest of the world export to the U.S. Figure 5-7. U.S. Sugar Import Forecast (Scenario 12 "PA-C-F") H o o o > 2,500 2,000 1,500 1,000 500 ro r00 OV O CN >o O O o O o o o O O O o O o o o O o O O O O O CM CN (N (N Year M Mexico under-quota export Mexico quota-free export Mexico over-quota export ^ The rest of the world export to the U.S. Figure 5-8. U.S. Sugar Import Forecast (Scenario 16 "T-S-F")

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97 450 Figure 5-9. Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 4 "PA-S-F") 450 Figure 5-10. Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 8 "PA-B-F")

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98 U.S. price — A — Mexico price Figure 5-11. Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 12 "PA-C-F") U.S. price — 4t — Mexico price Figure 5-12. Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets (Scenario 16 "T-S-F")

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99 10, 9 o o o o > 000 000 ^ 8,000 000 000 000 000 S 3,000 .000 ,000 0 -Bs o Year U.& demand Mexico demand — H — U.S. Supply Mexico Supply Figure 5-13. Forecasted Sugar Demand and Supply for both the United States and Mexico (Scenario 1 "Baseline") Year U.S. demand • Mexico demand — B — U.S. Supply Mexico Supply Figure 5-14. Forecasted Sugar Demand and Supply for both the United States and Mexico (Scenario 4 "P-S-F")

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100 cs o o o o o o cs o o vo o o r4 o o 1 1 00 Ov O o o — o o o (N (S fS o o o o o Year U.S. price • Mexico price Figure 5-15. Forecasted Equilibrium Sugar Prices in the United States and Mexico (Scenario 14 "PA-B-M") 450 ^ 100 50 0 -1 1 1 1 1 1 1 1 1 1 1 1 1 1 — §0000000 — — — — — 0000000000000 (NCSO
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101 09 o o 00 X) B 'S 3 U O > (A C U U ^ ?^ O 1) o u ^ 2 3 ^ > + is hi 3 Si c r5 T3 > 00 i3 U c Ecu O 'u C u o C/2 ^ o o 2 O — o O 1 00 o 2 o 2 d ^ o in w 00 CO O O O O CO ^ o 9 d -H O (N — O On Tt r<-i O m o I 00 — O C\ —1 o d !:: O 0\ 00 CO 0\ ON 00 ro On U-) o\ ro 0\ On 00 O .5 CT lU 3 OQ o 'S c _; 00 (-1 Sso On ON — O 00 O d ^ ro d ro r~On On 00 rn On IT) On 00 ^ ro 00 oo r<-) ON in ON ro "-^ ro On oi ON o CN On cn ON TlO 00 00 — ON — "1 iri O On ON (N O d ro O ^ 00 CN O ON ON IT) On ro ^ 00 NO On 00 rn ON V-) On NO iri O On ON ~; 00 o t-.2 ^ c3 CO c u < GO r o C u o PL, I I < NO O (N ~: NO ON O 1^ ro in ON < o6 in —1 < -a c C II H -3 o 00 U S O o n 3 a, tr (U cs 00 -N 2 II o — 2 a o. c .2 J= 3 o o x: II < 'c _o O U a, 00 D II U i-T ?3 3 1/3 [/3 (/3
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102 o i3 b 3 1^ o o O o pie 2; ^ CO q ON oo On So d St •a u 'S s 9 :2 o -H O ON m ON ^ O 00 CO C w Q XI X O I O I o u 3 u ^ 5 00 ^ 00 —I ON — ; O On 3 C O > ^ 1 <^ 2 CO ST' O 2 d w 0\ CM O O o ^ 2 u ,03 ^ NO 00 o o 03 3 OO h 3 > (L) o o -a NO O ON ON [1^ 3 O 3h c u S o NO O NO CN O NO O ON ON d On (N O d o O 00 o ON oo ON ^ IT) On 00 NO On 00 On ON o d o "1 00 o r~ 00 O o5 00 ^:^ ^ 8 O ^ ^ 8 c u 00 o 3 CO 3 PL, 00 o 1 2 1 'i 00 *c ca c 1 c < (U CJ 00 00 O •c u 00 r

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103 ^ico Welfare 74.443 (100) 69.095 (92.82) 69.882 (93.87) 64.393 (86.50) Me; Sugar industry revenue 21.737 (100) 12.159 (55.94) 14.532 (66.86) 13.983 (64.33) Net cost (B-C-D) 0.116 (100) 5.979 (5,168.80) 1.820 (1,573.52) -0.200 (-172.54) Cost of buying up excess sugar (D) o ^ 8 O 1 Cost of price support (C) 0.208 (100) 6.112 (2,931.51) O 1 O 1 ed States Tariff revenue from Mexican sugar (B) 0.093 (100) 0.133 (142.87) 0.180 (194.04) 0.200 (215.06) Unit 1 Adjusted welfare (A+B-C-D) 354.370 (100) 338.136 (95.42) 354.819 (100.13) 357.277 (100.82) Welfare (A) 354.486 (100) 344.115 (97.07) 356.639 (100.61) 357.077 (100.73) Sugar industry revenue 35.713 (100) 33.480 (93.75) 27.886 (78.08) 26.516 (74.25) HFCS industry revenue 5.684 (100) 10.132 (178.27) Scenarios Baseline (status quo) Scenario 13 ("PA S M") Scenario 14 ("PA B M") Scenario 15 ("PA C M")

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104

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105

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106

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107 Mexico welfare 72.29 72.29 66.76 on option Mexican sugar 21.31 21.31 20.52 h product HFCS ad U.S. adjusted welfare 354.08 354.85 355.44 Hig High U.S. 35.37 35.21 33.93 U.S. HFCS 10.13 10.13 10.13 Mexico welfare 65.726 65.739 65.739 option Mexican sugar 21.322 21.355 21.355 HFCS ad( U.S. adjusted welfare 353.97 354.89 355.12 strategies High U.S. sugar 35.31 35.09 35.09 U.S. HFCS 10.13 10.13 10.13 Mexico's Mexico welfare 76.24 76.24 71.49 § Mexican sugar 21.87 21.87 23.14 gh produci U.S. adjusted welfare 353.90 354.73 354.97 X U.S. 35.56 35.40 35.39 U.S. HFCS 5.68 5.68 5.68 Mexico welfare 74.44 69.77 69.77 n the current policy [status quo) Mexican sugar 21.74 22.15 22.15 U.S. adjusted wciiore 354.37 354.79 354.92 Maintaii U.S. sugar 35.71 35.40 35.40 U.S. HrL.o 5.68 5.68 5.68 Maintain price support (status quo) Buying up excess sugar in the market Production control with Mexico U.S. strategies V2 X lo ts i2 on 5 u = 5-1 CO CO U 0\ IT) 00 00 CO o in NO ON >n t^ t-"n m in 00 NO o o CX 3 Q. O" CO aj — H

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109

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110 'ob o o .id o 2 ^ c o Q. O T3 ea on U CO o 3 o 00 s u B c ^ c '3 o c C3 00 1/5 3 U c/l — X c a 1) on T5 — o i! 00 on U T3 C C3 ^ 2 00 3 U pa u .s ^ 00 C o o i3 o X

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111 c o c o o a V3 00 jz S .2? o c fc— C3 O tl ^ t/) -a -s 1 -S a ^ ^ S ^ on on U On as ON q 00 On 60 o o c g D. O T3 a U •T3 C 60 3 3 c -S 1 -S 2i ^ 3 53 X =: as as 00 ON OO ON C/3 00 U o "o a. 1) o 3 u a .E ^ 2 '3 2 T3 C 60 cn 3 3 C o o 1) 1) ON as as as 00 •S .2, ,=5 00 00 U o o '3 ^ 1> o 3 ^-^ ^ Q. 3 1/5 cr => v= .E ^ S3 '5 S5 o ^ c 3 Da U 3 (U -s o w o cj h 2 o X

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CHAPTER 6 CONCLUSIONS AND IMPLICATIONS FOR POLICY Conclusions and Implications for Policy Sugar is a basic commodity with a long history of utihzation and trade between regions where excess demand and excess supply exists. This trade in the sugar market has provided a classic example for study in the field of agricultural economics. Although its economic value has diminished, as seen in the trend of the declining world prices and a smaller percentage of income spent on sugar in developed countries, its bargaining power continues as a powerful force in the political arena today. Emergence of a new substitute commodity, high fructose com syrup (HFCS) produced predominantly by the United States, has brought about changes in the climate of the sweetener markets, particularly those tied to U.S. markets. These changes are expected to continue. Recent trends in the international trade environment to move towards freer and borderless trade have also accelerated changes in the market climate for sugar trade. When examining sugar trade issues between the United States and Mexico, Mexico's adoption of HFCS and the provisions of North American Free Trade Agreement (NAFTA) related to sugar and HFCS play critical roles for shaping the sweetener market balance. HFCS, already with a large share in the U.S. sweetener market, has gained sizeable market power, as well as power in the political arena. The HFCS industry supports the U.S. sugar program under the umbrella of the American Sugar Alliance (ASA). Despite the efforts of critics, the U.S. sugar program remains intact due largely to successful lobbying efforts by ASA. On the other hand, the growth 115

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116 of HFCS was not welcomed in Mexico. Mexico, a traditional net sugar exporter is now facing pressures from importing a sugar substitute. Although NAFTA promised Mexico favored opportunities to export sugar to the U.S. market, implementation of NAFTA also resulted in opening the door for HFCS consumption in Mexico. The consequences of this threatening trend of increased HFCS consumption was inscribed on the sweetener history when Mexico lost the HFCS dumping case with the United States in 2001. This study attempted to answer five specific questions: (1) What was the impact of changes in trade regime on the U.S. and Mexican sweetener market since NAFTA was implemented in 1994?; (2) How much sugar surplus can Mexico generate, how much sugar will cross the border both underand over-quota, and what will happen after 2008 when all the restrictions are eliminated on Mexican sugar?; (3) What will be the impact of changes in the Mexican market situation and how much influence will HFCS adoption cause in both the U.S. and Mexican sweetener markets?; (4) What will be the impact of changes in U.S. sugar policy on both the United States and Mexico?; and (5) Is there alternative sugar policy for the United States to current price support?. In the following, answers for each question are summarized. Impact of Changes in Trade Regime NAFTA brought about mixed impacts on the United States and Mexico and some are different from what was expected. The Mexican sugar industry has benefited little in the past ten years under the NAFTA regime. By comparison, NAFTA did not bring about drastic change to the U.S. sugar market: expanded exports from Mexico have failed to materialize. Rather, attention was poured into issues of HFCS and its inmiediate impact on Mexico's sweetener market. As seen in the trade dispute over HFCS, the Mexican government struggled to suppress HFCS adoption in its market.

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117 Mexico's Export Potential The opportunity for Mexico to export sugar to reach the 250,000 MT expanded quota (which would be equivalent to approximately 20 percent of the U.S. minimum import requirement) seems unlikely to be enjoyed due to Mexico's restricted production compared to its sweetener consumption. Yet, Mexico possesses a large potential to export over-quota as well as quota-free when all restrictions are lifted. The magnitude of overquota and quota-free export depends on the expansion rate of Mexico's sugar production and HFCS adoption. Specifically, if Mexico adopts HFCS at an increasing rate, a considerable amount of surplus sugar destined to export will be generated, which consequently poses a direct impact on the Mexican sugar industry and an extended impact on the U.S. market. The Impact of Changes in Mexican Market Situation Mexico's sugar production, HFCS adoption, and a tax on HFCS are examined as factors that change the Mexican sweetener market situation. The impact from these three factors on Mexico is eminent. The Mexican sugar industry and welfare become better off from higher sugar production which can be achieved through reduction of production costs and mill downtime; however, this positive impact dissipates once HFCS is adopted at higher rates. The negative impact from adopting HFCS on Mexico originates with the imbalance between domestic sugar production and sweetener consumption, thus making Mexico unable to attain a net sweetener surplus producer. Mexico then faces a large sugar surplus that cannot be sold anywhere but the U.S. market with tariff (over-quota) or the world market. Simulations showed that the sugar industry can tolerate a higher rate of HFCS adoption if the gains from high production exceed the losses from adopting HFCS, whereas the nation's welfare cannot. This implies that the Mexican government faces

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118 difficulty in allowing faster HFCS adoption to happen and thus will continue to struggle to suppress HFCS adoption in its market. The impact from these three factors on the United States is also significant; however, U.S. domestic sugar production and consumption will remain relatively unchanged, although various simulations showed fluctuating U.S. import from Mexico and the rest of the world caused by changes in the Mexican sweetener situation. One notable result is that when Mexico's high production and high HFCS adoption is combined, Mexico's sugar export takes up the entire U.S. minimum import requirement, leaving sugar export from the rest of the world marginalized. This scenario results in not only reduced revenue to the U.S. sugar industry as well as reduced U.S. welfare, but also triggers political pressures from the other sugar exporters whose shares in the U.S. market are at stake. Mexico's HFCS adoption turns out to be beneficial only to the U.S. HFCS industry, but Mexico's tax on HFCS benefits all except for the U.S. HFCS industry. In the latter case, Mexico will not be able to generate sugar surplus and thus export either underquota, over-quota or quota-free; however, gains in producer surplus outweigh the losses in consumer surplus in both countries due to high prices of domestic sugar. This policy lever may not be acceptable in today's trade environment and in fact the decision by the Mexican government has changed back and forth in the past. The Impact of Changes in U.S. Sugar PoHcy Quota allocation and price stabilization policies are examined as possible changes in the U.S. sugar policy. The impact of a quota allocation policy poses a large impact on both Mexico and the U.S. markets. For Mexico, the aforementioned large scale of sugar export is possible only if the U.S. government allocates quotas among exporters in a

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119 flexible manner; if not Mexico's export will be dampened because of its comparative disadvantage to the rest of the world. For the United States, accommodating Mexican sugar can serve as an alternative form of price support because of the higher price of Mexican sugar compared to that from the rest of the world; otherwise the U.S. price will likely dip below the U.S. support price level caused by a large quantity of sugar from the rest of the world flowing into the U.S. market at a lower price. In this sense, sugar trade agreements were mutually beneficial for both the United States and Mexico. Two alternative price stabilization policies are examined in the study: buying up excess sugar in the market and production controls. Simulation results show some improvement in both U.S. sugar industry revenues and cost-adjusted U.S. welfare as a result of reduction in sugar program costs as well as an increase in tariff revenue from Mexican sugar, yet overall positive effects are minimal. It is noteworthy, however, that maintaining the price support may become extremely costly when combined with the situation in where the U.S. government promises to accept a large amount of sugar from the rest of the world. If the U.S. government switched policies from the price support to buying up excess sugar, the timing to do so would be important so as to minimize the cost incurred by the government: the cost of buying up excess sugar will rise immediately after policies are switched whereas the cost of the price support will not because the U.S. sugar price will be maintained relatively high in the early stage of the forecast horizon. When 'buying up excess sugar policy' and 'production control policy' are compared, losses in sugar industry revenues in both countries and in Mexico's welfare are larger for the latter policy, given the similar magnitude of gains in U.S. welfare. In general, these alternative policies still would be more effective than any price-led sugar policy because

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120 the inelastic supply price elasticities in both countries imply that sugar production is not responsive to price changes. Contrasted with the possible improvement in U.S. sugar industry and welfare, Mexico does not benefit from changes in U.S. price stabilization policy. Alternative Sugar Policy by the United States An alternative U.S. sugar policy to the current policy of price supports is sought from the aggregate simulation results using game theory. Cooperative games among industries and governments (the U.S. HFCS industry, the U.S. sugar industry, the Mexican sugar industry, cost-adjusted U.S. welfare, and Mexico's welfare) are also examined by assuming that both nation's welfare is transferable and that total pay-offs are redistributed among coalition members. As expected, the Mexican government and sugar industry will always and harmoniously choose a high production strategy and avoid any strategies that involve HFCS adoption. By comparison, the U.S. sugar industry does not agree with the choice made by the U.S. government: the U.S. sugar industry prefers price support to alternative policies. When coalitions are formed, one case for country coalitions (United States vs. Mexico) and the other for the government/industry coalitions (governments vs. industries), the games reach different solutions: the former game results in U.S. sugar production control strategy and the latter game results in U.S. government follows the strategy of buying up excess sugar. This contrast indicates that the U.S. government prefers to form a coalition with its own industry while the Mexican government prefers to form a coalition with the U.S. government. For the Mexican government, Mexico's welfare improves by having U.S. cooperation at the expense of its own sugar industry. This implies that Mexico faces a choice either of improving the nation's well-being or of

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121 protecting the sugar industry consisting of a large number of growers and related workers. When the U.S. HFCS industry is included in a coalition, a conflict of interest between the governments and industries becomes intensified. When pay-off to the U.S. HFCS industry is pooled into a coalition, the industry coalition (the U.S. HFCS industry, the U.S. sugar and Mexican sugar industries) will lobby for the strategy that involves Mexico's HFCS adoption. Although the game has reached a different solution through government's decision making, this is true because the industries recognize a better payoff to the coalition. This contrast implies that the U.S. HFCS and the U.S. sugar industries have a strong incentive to influence Mexico's choice of strategy. Furthermore, since the Mexican sugar industry theoretically becomes better off from being compensated by the industry coalition, it may be tempted to allow HFCS adoption rather than expecting protection from its own government. The results from games with a grand coalition indicate that there is no solution that satisfies pareto optimality. If redistribution of pooled revenues from industries and welfare is feasible, there are solutions that minimize the total losses for all entities. In summary, in no case did the game reach a solution that included the current U.S. policy of price support as the best policy; alternative policies examined in the study are better sugar policies to pursue. Also, the stance of the U.S. HFCS industry on sugar policy can be influential. If the U.S. HFCS industry is included in the decision-making process, a sugar policy that differs from that chosen by the governments can be lobbied for with enticing side agreements within the coalition.

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122 Limitation of the Study and Suggestions for Future Research HFCS production and consumption are not quantitatively estimated and integrated into the trade model. This limits the effects on the price and quantity of both sugar and HFCS from substitution between the two commodities. In the trade model, the rest of the world is treated as a single homogenous region; however, a country or region such as Brazil or the European Union that embraces significant sugar demand and supply could have an individual impact on the U.S. market. Including changes in sugar stocks in each region as well as flows of sugar-containing goods across borders into the bilateral model is expected to enhance the quality and depth of the results drawn from the model.

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APPENDIX A MAJOR EVENTS IN THE SUGAR INDUSTRY HISTORY IN MEXICO AND THE U.S. Mexico United States 1910 The Mexican Revolution 1917 Farmers ownership of Ejido land started (Article 27, Mexican Constitution) 1972 HFCS-42 production started. 1977 HFCS-55 production started. 1980 Coca Cola and PepsiCo replaced 50% of sugar use with HFCS-55. 1988 Government started privatizing sugar mills and dismantling AZUCAR S.A. 1989 Crystalline fructose production started. 1990 A minimum import quota of 1.256 million MT (raw value) of sugar was established in agreement with the implementation of GATT (October). 1990 Farm bill All major brands of soft drinks utilized 100% HFCS as the nutritive sweetener ingredient. 1991 COAAZUCAR was formed (amendments in Decreto Cafiero). Sugarcane growers started to be paid by sucrose content instead of by the weight of sugar cane (54 % of the wholesale price of standard sugar based on KARBE system). 1992 Agrarian reform (Article 27) allowed to sell/rent Ejido land. 1994 NAFTA agreement (January) Devaluation (December) Sugarcane growers started to be paid 57% of the wholesale price of standard sugar based on KARBE system. 123

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124 Mexico United States 1995 Inflation rate hit 57% while the price of sugar increased 25.7%. 1996 The government announced increases in import duties on HFCS-42, HFCS55, and crystalline fructose to 12.5, above the then-current rate of 10.5 percent. (December). 1996 farm bill 1997 SECOFI initiated anti-dumping investigation (February) and imposed temporary tanff on two grades of HFCS (June). SECIFI published the formula to determine the wholesale price of standard sugar (March). 1995 • CRA (Com Refiners' Association) asked for the review of Mexico's antidumping actions under chapter of NAFTA. WTO established panel for Mexico's HFCS dumping case (January). 1 QOO 2000 WTO panel ruled against Mexico's dumping case (January). Vicente Fox was elected as president, ending 71 years of authoritarian oneparty rule in Mexico (December). 2001 WTO Appellate Body turned down the Mexico's appeal of HFCS dumping case. Mexican government expropriated 27 sugar mills (September). 2002 A 20-percent tax on beverages that contain HFCS was introduced on January; suspended on March 5; and reimposed on July 16. National Sugar Policy for 2002 2006 (February) Source: Created from Polopolus and Alvarez, 1991; Greene, 1998; Garcia Chaves et al., 2002; Buzzanell, 2002; and various issues by USDA.

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APPENDIX B CORN STATISTICS Figure B-I shows the recent com production and consumption for selected countries. Currently, the United States is the largest producer as well as consumer of com, followed by China. However, in the 2002/2003 season, China became a net producer. Trends show that production in China has been increasing for the last three seasons while that of U.S. has been decreasing. Figure B-2 shows the transition of food and industrial com use in the United States during the past two decades. The largest industrial use for com is for fuel alcohol, representing approximately 41 percent of total use in 2002. HFCS is the second largest use, representing approximately 24 percent of total use. HFCS use has been level in the last five years, while fuel alcohol use exhibits rapid growth. Figure B-3 shows transition of the U.S. com price (No.2 Yellow) in Chicago market during the past two decades. The price shows some fluctuation around 2.5 US dollars per bushel. Figure B-4 shows U.S. exports of products made from com in 2002 expressed in million US dollars. Com gluten feed, com gluten meal, and com oil are the dominant products made from com. When three kinds of fructose (fructose solids containing more than 50% fructose. Chemically pure fructose, and fructose syrup with 50%+ fructose) are aggregated, the total value becomes fourth largest, following com oil. If HFCS is already saturated in the United States, it will have better marketing opportunities overseas; however, the business size still will be small relative to that for the leading products. 125

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126 300,000 250,000 H S 200,000 o o o ^ 150,000 B 3 100,000 > 50,000 0 1999/2000 2000/2001 2001/2002 Year 2002/2003 China production -U.S. consumption U.S. production -Chiifc consumption European Union production Brazil consumption Figure B-1. Recent Com Production and Consumption for Selected Countries Source: Com Refiners Association, 2004 2,500 Figure B-2. Food and Industrial Com Use in the U.S., 1980-2002 Source: Com Refiners Association, 2004

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127 Ji 4.5 Year Figure B-3. Com Price (No.2 Yellow) in Chicago Market, 1981-1998 Source: USDA, 2004 Corn gluten feed Corn gluten meal Corn oil, crude Corn oil, fully refined Modified starches derived from corn starch Glucose syrup not containing fructose or containing in the dry state less than 20% fructose Corn meal Corn starch Glucose (dextrose) Fructose syrup with 50%+ fructose Chemically pure fructose Fructose solids containing more than 50%* fructose 0 50 100 150 200 250 300 350 Value [million US$] Wrd00v71.exe Figure B-4. Exports of Products Made from Com in 2002 Source: Com Refiners Association, 2004

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APPENDIX C DERIVATION OF INVERSE LINEAR EQUATIONS Assuming that markets in both countries are competitive and that the demand and supply equations linear in quantity, the demand and supply functions are expressed in the form ofP^ = a + b*Q^ (b<0) and = c + d*Q^ (d>0), respectively. By definition, the supply price flexibility {Tfp)\s given as: rfp = d*(Q'/P'). (C.I) Since the supply price flexibility is the reciprocal of price elasticity (Efp), the slope of the inverse supply equation is derived from equation [C.I]: d= rfp*(P^/^)=(l/Efp)*(P^/Q^). (C.2) Similarly, the slope of the inverse demand function of the form = a + b*^ is given: b=Tfp *(I^/Q^) =(l/E^p)*{P^/Q^) (C.3) where rf pis the price flexibility for the inverse linear demand function and e'^p is the price elasticity for the linear demand function. In the case of other variables in supply and demand functions, the slopes in the inverse linear functions are also expressed with estimated elasticities. Suppose that the supply function is expressed with cost variables as = c + d*Q^+ e*COST. The slope associated with cost is expressed by definition: e = AP^/ACOST (C.4) Estimated elasticity associated with cost is: 128

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129 E^cosT^ (AQ^/ACOST)*(COST/Q^). (C.5) With supply price elasticity (Efp), equation [C.4] is expressed as e = i-EfcosT /E^p) *(P^/COST). (C.6) Final forms of inverse linear function for sugar demand and supply for the U.S. market derived from corresponding equations [4.1] and [4.5] are expressed as: = IU, + IU2 *Q''us. t + Wi *GDP us, + IU4 *POP us, t (C.7a) or P^us, = IUi + IU2*Q^us. + Shifter'' us. t (C.7b) P^us, t = lUUi + IUU2 *^us, + IUU3 *COST us, t + IUU4 *RCV,, (C.8a) or P^us. = lUUi + IUU2*Q\s. + Shifter' us. t (C.8b) Note that the inverse linear demand function is re-specified on an annual base rather than quarterly in order to balance the market with the supply equation and that the inverse linear supply function is re-specified with the current price without lagged quantity for simplicity. Coefficients for corresponding variables are presented in the Table C-1. Table C-1. Coefficients for Inverse Linear Functions -U.S.Inverse linear demand function Inverse linear supply function W2 (l/E''p,usnP^us/Q''us) IUU2 (l/E'p.us)*iP'us/Q'us) IU3 {-E GDP, us /E p,us)* (P^us/GDPus) IUU3 {-E^cosT. us / E^p.us)* (P'us/COSTus) IU4 (-E POP. us/ E p^ us)* (P^ us/POP us) IUU4 {-E^RCv/E^p. us)*(P'us/RCV) In the case of Mexico, the equation for annual total sugar is expressed with estimates of direct consumption of sugar as preliminary estimation showed statistically insignificant estimates associated with price variables in total and indirect consumption of sugar. By assuming that indirect consumption demand is totally inelastic (i.e., a vertical demand curve in quantity-price space), total sugar demand is expressed with a

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curve that has the same slope as direct consumption of sugar with the intercept shifted by adding the vertical indirect consumption demand curve. In this way, the demand equation is expressed in terms of total consumption of sugar (Q^*^, ) with significant estimates (M2 in equation [4.2]), incorporating the effect of change in indirect consumption of sugar as a consequence of the change in HFCS consumption. P^Mx. = IM, + IM2*Q'^MX.t + IM3*GDPmx + IM4*P0Pmx. t /Ms*^'^ (C.9a) or P^MXt = IMi + IM2 *Q'^mx t + Shifter^ MX, (C.9b) P^Mx,t = IMMi + IMM2*Q^Mx I + IMMs*COSTmx t + IMM4*DT, + IMM5*SUGL0SS, + IMM6*DURTN, (C. 10a) or P'mx, = MM, + IMM2 *Q'mx t + Shifter' mx t (C.lOb) and the coefficients are presented in Table C-2. Table C-2. Coefficients for Inverse Linear Functions -MexicoInverse linear demand function Inverse linear supply function IM2 (l/E''^P,MXnP^Mx/Q"Mx) IMM2 (l/EfpMxYiP'Mx/^Mx) IM3 {-e gdp, mx /e p mx )* (P'^mx/GDPmx) IMM3 i-E^cosT, mx/ E^p,mx)* (P'mx/COSTmx) IM4 {-E POP, MX /E p,mx)* (P^mx/POPmx) IMM4 {-E'dt/E'p_mx)*(P'mx/DT) IMM5 (-E'suGLOss / E^p. mx) (P^Mx/SUGLOSS) IMMs {-E'durtn/ E^p, mx)* (P'mx/DURTN)

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LIST OF REFERENCES Alvarez, Jose and Leo Polopolus. "The Florida Sugar Industry." EDIS document SC042, Food and Resource Economics Department, University of Florida, Gainesville, FL, June 2002a. Alvarez, Jose and Leo Polopolus. "The Sugar Program: Description and Debate." EDIS document SC020, Food and Resource Economics Department, University of Florida, Gainesville, FL, June 2002b. Azucar S.A. de C.V. Estadii'tica Azucareras. "Ventas de azucar el pais por clase, destino y tipo de operacion 1970-1990." Mexico, DF, 1990. Borrel, Brent. "The Mexican Sugar Industry." International Economics Department, Policy, Research and External Affairs Working Paper Series No.596. Washington, DC: Worid Bank, February 1991. Buzzanell, Peter. "The U.S.-Mexico High Fructose Com Syrup (MFCS) Trade Dispute." In Schmitz, Andrew, Thomas H. Spreen, William A. Messina, Jr., and Charles B. Moss, Sugar and Related Sweetener Markets: International Perspectives (53-64). New York: GABI Publishing, 2002. Buzzanell, Peter, and Ron Lord. "Mexico: Sugar and Com Sweetener, an Update." Sugar and Sweetener S&O V 20(2), June 1995. Comite de la Agroindustria Azucarera (COAAZUCAR). "Desarrollo Operative Campo-Fabrica 1996/2002." Internet site: http://www.sagarpa.gob.mx/Coaazucar/menu2/nacional.htm (Accessed May 2003a). Comite de la Agroindustria Azucarera (COAAZUCAR). "Rangos de Superficie y Numero de Caneros a Nivel Nacional Zafra 2000/2001." Intemet site: http://www.sagarpa.gob.mx/Coaazucar/menu6/ann09.htm (Accessed May 2003b). Comite de la Agroindustria Azucarera (COAAZUCAR). "Superficie de Riego y Temporal y su Produccion de Cafia Zafra 2000/2001." Intemet site: http://www.sagarpa.gob.mx/Coaazucar/menu6/sprt03.htm (Accessed May 2003c). Comite de la Agroindustria Azucarera (COAAZUCAR). "Avance del Tipo de Cosecha, Numero de Cortadores, Cosechadoras Integrales y Alzadoras al 17 de Febrero del 2001 Zafra 2000/2001." Intemet site: http://www.sagarpa.gob.mx/Coaazucar/menu4/ult_estim_prod.htm (Accessed May 2003d). 131

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132 Comite de la Agroindustria Azucarera (COAAZUCAR). "Acciones de Cosecha de Cafia que Reflejan Calidad Zafra 2000/2001." Internet site: http://www.sagarpa.gob.nix/Coaazucar/menu4/ult_estim_prod.htm (Accessed May 2003e). Comite de la Agroindustria Azucarera (COAAZUCAR). "Resultados Economicos del Campo Cafieros del Campo de las Zafras 1987/2002." Internet site: http://www.sagarpa.gob.mx/Coaazucar/menu3/indexl.htm (Accessed May 2003f). Comite de la Agroindustria Azucarera (COAAZUCAR). "Relacion Historica de los Precios de la Caiia de Azucar como Promedio Nacional." Internet site: http://www.sagarpa.gob.mx/Coaazucar/menu7/com_precio.htm (Accessed May 2003g). Congressional Research Service, Library of Congress, Agriculture: A Glossary of Terms, Programs, and Laws, 2nd Edition, Washington, 1999. Com Refiners Association. "Food and Industrial Com Use 1980 to Present." Intemet site: http://www.com.org/web/foodseed.htm (accessed May 27, 2004). Diario Oficial de la Federacion. Mexico, DP. Secretaria de Medio Ambiente y Recursos Naturales. March 26, 1997. Farm Foundation. 'Trade Dispute in an Unsettled Industry: Mexican Sugar." Intemet site: http://farmfoundation.org/flags/schwedel.pdf (Accessed August 2003). Food and Agriculture Organization of the United Nations Statistics (FAO). Intemet site: http://apps.fao.org/page/collections?subset=agriculture (Accessed Febmary 2003). Garcia Chaves, Luis R., Thomas H. Spreen and Gretchen Greene. "Structural Reform and Implications for Mexico's Sweetener Market." In Schmitz, Andrew, Thomas H. Spreen, William A. Messina, Jr., and Charles B. Moss, Sugar and Related Sweetener Markets: International Perspectives (81-100). New York: GABI Publishing, 2002. Garcia Chaves, Luis R., Gretchen Greene, Thomas H. Spreen, Daisuke Sano and Chris O. Andrew. Transitions in the Mexican Sugar Industry: An Analysis of the Production and Marketing System. Lake Alfred: Florida Science Source, 2004. General Accounting Office (GAO). Sugar Program: Supporting Prices Has Increased Users' Costs While Benefiting Producers. RCED-00-126, Washington DC, 2000. Greene, Gretchen. "Transitions in the Mexican Sugar Industry." Ph.D. dissertation. University of Florida, 1998. Haley, S. and N.R. Suarez. Sugar and Sweetener Outlook. Washington, DC: USDA, January 2003.

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133 Haley, S. and N.R. Suarez. "U.S. Sugar Policy and Prospects for the U.S. Sugar Industry." In Schmitz, Andrew, Thomas H. Spreen, William A. Messina, Jr., and Charles B. Moss, Sugar and Related Sweetener Markets: International Perspectives (53-64). New York: GABI Publishing, 2002. Institute Nacional de Estadistica, Geografia e Informatica (INEGI). "Social and demographic statistics." Internet site: http://www.inegi.gob.mx/difusion/ingles/fiesoc.html (Accessed July 2003). Koo, Won W. and Richard D. Taylor. "2000 Outlook of the U.S. and World Sugar Markets." Agricultural Economics Report No.444, Northern Plains Trade Research Center, North Dakota State University, Fargo, ND, July 2000. Lopez, Rigoberto A. "Economic Surplus in the U.S. Sugar Market." Northeastern Journal of Agricultural Economics. 19(1): 28-36, April 1990. Mas-Colell, Andreu, Michael D. Whinston and Jerry R. Green. Microeconomic Theory, New York: Oxford University Press, 1995. McCoy, Terry L. "Latin American Sweetener Markets: Economic Reform and Regional Integration." In Schmitz, Andrew, Thomas H. Spreen, William A. Messina, Jr., and Charles B. Moss, Sugar and Related Sweetener Markets: International Perspectives (81-100). New York: GABI Publishing, 2002. Morris, Peter. Introduction to Game Theory, New York: SpringerVerlag, 1994. Moss, Charles B. and Andrew Schmitz. "Coalition Structures and U.S. Sugar Policy." In Schmitz, Andrew, Thomas H. Spreen, William A. Messina, Jr., and Charles B. Moss, Sugar and Related Sweetener Markets: International Perspectives (53-64). New York: GABI Publishing, 2002. Offenbach, Lisa A. Effects of Sugar and Ethanol Related Policies on the Market for High Fructose Com Syrup. Ph.D. Thesis. Department of Agricultural Economics. Kansas State University, Manhattan, KS, 1995. Organisation for Economic Co-operation and Development (OECD). "Statistics Portal." Internet site: http://www.oecd.org/statsportal/ (Accessed November 2003). Petrolia, Daniel R. and P. Lynn Kennedy. "A Partial-Equilibrium Simulation of Increasing the U.S. Tariff-Rate Sugar Quota for Cuba and Mexico." Selected paper presented at the American Agriculture Economics Association meeting. Long Beach, CA, July 2002. Polopolus, L and J. Alvarez. Marketing Sugar and Other Sweeteners. New York: Elsevier Science Publishing Company Inc., 1991.

PAGE 145

134 Secretana de Agricultura, Ganaderia, Desarrollo Rural, Pesca y Alimentacion (SAGARPA), Sistema Integral de Informacion Agroalimentaria y Pesquera (SIAP). "Avance Comparativo Siembras y Cosechas: Perennes: Situacion al 31 de Mayo 2002 y 2003." Internet site: http://www.siap.sagarpa.gob.mx/ar_comdeagr.html (Accessed July 2003). Skully, David W. "Auctioning Tariff Quotas for U.S. sugar Imports." Special article presented in Sugar and Sweetener SSS-223, Economic Research Service, USDA, Washington, DC, May 1998. Spreen, Thomas H., Mechel Paggi, Anouk Flambert, and Waldir Femandes, Jr.. "An Analysis of the EU Banana Trade Regime." Selected poster presented at the American Agriculture Economics Association meeting, Tampa, FL, August 2000. Electronic abstract at http://agecon.lib.umn.edU/aasa.html/#aaeaOO. Takayama, T and G. G. Judge. "Equilibrium Among Spatially Separated Markets: A Reformulation." Econometrica 32: 510-24, 1964. U.S. Department of Agriculture, Economic Research Service. "Sugar Statistical Compendium Stock #91006." Washington, DC, October 1991. U.S. Department of Agriculture, Economic Research Service. "Com Sweetener Statistics Stock #94002." Washington, DC, September 1993. U.S. Department of Agriculture, Economic Research Service. "Agricultural Outlook." Washington, DC, March 1997. U.S. Department of Agriculture, Economic Research Service. "Agricultural Outlook." Washington, DC, September 1999. U.S. Department of Agriculture, Economic Research Service. Sugar and Sweetener Situation and Outlook Yearbook. Washington, DC, various issues, 2001a. U.S. Department of Agriculture, Foreign Agricultural Service. "Mexico Expropriated 27 Sugar Mills." GAIN report #MX1 161, Washington, DC, September 2001b. U.S. Department of Agriculture, Foreign Agricultural Service. "The North American Trade Agreement." Internet site: http://www.fas.usda.gov/info/factsheets/nafta.html (Accessed July 2001c). U.S. Department of Agriculture, Economic Research Service. Sugar and Sweetener Situation and Outlook Yearbook. Washington, DC, various issues, 2002a.

PAGE 146

135 U.S. Department of Agriculture, Foreign Agricultural Service. "New National Sugar Policy." GAIN report #MX2031, Washington, DC, February 2002b. U.S. Department of Agriculture, Economic Research Service. Sugar and Sweetener Situation and Outlook Yearbook. Washington, DC, various issues, 2003a. U.S. Department of Agriculture, Foreign Agricultural Service. "Sugar: World Market and Trade." Internet site: http://www.fas.usda.gov/htp/sugar/2003/may (Accessed July 2003b). U.S. Department of Agriculture, Economic Research Service. Feed Outlook Rreport. Washington, DC, 2004. U.S. Department of Commerce, U.S. Census Bureau. Internet site: http://www.census.gov/ipc/www/idbsprd.html (Accessed February 2003). U.S. Department of Labor, Bureau of Labor Statistics. "Consumer Price Index." Internet site: http://www.bls.gov/cpi/home.htm (Accessed November 2003). Varian, Hal R. Microeconomic Analysis, ^'^ edition. New York: W. W. Norton & Company, 1992.

PAGE 147

BIOGRAPHICAL SKETCH Daisuke Sano was bom on January 27, 1967, in Japan. He received the Bachelor of Science in 1989 and the Master of Science in 1991 in applied biochemistry from the University of Tsukuba in Japan. He served the Ministry of Agriculture, Forestry and Fisheries of Japan as Technical Officer from 1992 to 2000; meanwhile he received the Master of Science in food and resource economics from the University of Florida in 1999 funded by a scholarship from Japan International Cooperation Agency (JICA). He received the Doctor of Philosophy in food and resource economics from the University of Florida in 2004. 136

PAGE 148

I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. Thomas H. Spreen, Chair Professor of Food and Resource Economics I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. Lisa A. House, Cochair Associate Professor of Food and Resource Economics I certify that 1 have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quahty, as a dissertation for the degree of Doctor of Philosophy. Chris O. Andrew Professor of Food and Resource Economics I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. „ Terty L. M^Coy Professor of Political Science I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. Kenneth L. Buhr Assistant Professor of Agronomy

PAGE 149

This dissertation was submitted to the Graduate Faculty of the College of Agricultural and Life Sciences and to the Graduate School and was accepted as partial fulfillment of the requirements for the degree of Doctor of Philosophy. August 2004 Dean, College of Agncu Sciences Dean, Graduate School


Mean Number of Parasitized
Mummies/Plant
93
(Error bars @ 95%; 1=4.41: F=3.25; P =0.005)
16
10/22/02 10/29/02 11/5/02 11/12/02 11/19/02 11/26/02
Figure 7-6. Lysephlebius testiceipes parasitized aphids on cotton plants treated with
L-methionine. Ten plants were used for each treatment and held in the
shade house at the University of Florida, Department of Entomology and
Nematology from 22 October to 25 November 2002. No statistical
differences were observed except for the second and final collection date.


68
Year
Indirect consumption of sugar (Baseline)
Indirect consumption of sugar (Tax on HFCS)
Indirect consumption of sugar (High HFCS adoption)
HFCS consumption (Baseline)
HFCS consumption (Tax on HFCS)
HFCS consumption (High HFCS adoption)
Figure 4-2. Forecasted Indirect Sugar and HFCS Consumption in Mexico under
Alternative Scenarios


APPENDIX C
DERIVATION OF INVERSE LINEAR EQUATIONS
Assuming that markets in both countries are competitive and that the demand and
supply equations linear in quantity, the demand and supply functions are expressed in the
form of P = a + b*(2(b<0) and P5 = c + d*Qs (d>0), respectively. By definition, the
supply price flexibility (rfP) is given as:
rfP = d^tf/P3). (C.l)
Since the supply price flexibility is the reciprocal of price elasticity (E3p), the slope of the
inverse supply equation is derived from equation [C.l]:
d = if ?*(!*/&) =(1/ESP)*(PS/QS). (C.2)
Similarly, the slope of the inverse demand function of the form PD a + b*Q is given:
b= tfp *(PD/QD) =(1/Edp)*(Pd/Qd) (C.3)
where rfp is the price flexibility for the inverse linear demand function and Ep is the
price elasticity for the linear demand function.
In the case of other variables in supply and demand functions, the slopes in the
inverse linear functions are also expressed with estimated elasticities. Suppose that the
supply function is expressed with cost variables as P5 = c + d*Qs+ e*COST. The slope
associated with cost is expressed by definition:
e = APs/ACOST. (C.4)
Estimated elasticity associated with cost is:
128


20
Department of Entomology and Nematology green and shade houses. Excised leaves
were dipped in solutions of deionized H2O containing different concentrations of
methionine; depending on the experiment and exposed to larvae in the same rearing
chambers as the artificial diet trials under the same conditions. Survivorship data were
pooled from several different trials for data analysis.
In total, 64 potted eggplants were used for the whole-plant portion of the study.
Plants were held in FRIUs under the same conditions as the artificial and excised leaf
trials, in 38H x 15D (cm) plexigls cylinders (Figure 3-2). Four THW neonates were
placed on each plant for a total of 64 larvae (16 replicates) per treatment (nx0tai=256
larvae). The treatment of L-methionine was applied to the test plants (using a hand-held
sprayer calibrated to deliver approximately 10 mL of solution to each plant) before the
addition of larvae.
Feeding and Development
To test L-methionine on the developmental rates of THW, larvae were exposed to
excised eggplant leaves dipped in solutions containing the same concentrations of L-
methionine used in the artificial diet trials. Additional treatments of proline (1.0%) and
Bt-kurstaki (Dipel 86% WP at 3.5 grams/liter; Bonide, Oriskany, NY) were included as
positive and negative controls, respectively. Leaves were scanned photometrically using
the Cl 203 Area Meter with conveyor attachment (CID, Inc.; Camas, WA) to measure
leaf consumption before and after exposure to larvae. The difference in leaf areas
resulting from the missing leaf tissue was assumed to be the amount eaten by the
developing larvae. Larval head capsule widths were measured at the time of death or the


22.6
22.2
$ 22.0
I -
So 214
0> =
£ £, 21.4
1 21.2
I 21.0
^ 20.8
11 l 1 1 1 1 1
P
im
(
roduction Production Propduction Production HFCS adoption HFCS adoption HFCS adoption HFCS adoption HFCS adoption
provement, improvement imptovement improvement at 25% at 30% at 40% at 45% at 50%
average additional 0.5% additional 1% additional 1.5% (Baseline)
baseline)
Figure 5-17. Absolute Effects of Production Improvement and HFCS Adoption on Pay-off to the Mexican Sugar Industry


Copyright 2004
by
Lewis Scotty Long


25
Figure 2-3. Annual Rainfall and Irrigation Rate in Sugarcane Fields
Source: COAAZUCAR, 2003c
Table 2-1. Cane Sugar Production in Selected Countries, 1997-2000 Average.
Country
Area
Harvested
(1,000 ha)
Sugar
Production
(1,000 MT)
Cane
Yield
(MT/ha)
Sugar
Yield
(MT/ha)
Sugarcane
Recovery
Rate (%)
Brazil
4,914
18,339
68.28
3.73
5.47
India
4,092
17,233
69.41
4.21
6.07
Cuba
1,086
3,814
32.69
3.51
10.77
China
1,064
6,532
75.08
6.14
8.18
Pakistan
1,029
3,064
46.57
2.98
6.38
Thailand
923
5,468
56.16
5.92
10.55
Mexico
627
4,807
76.46
7.66
10.02
Australia
409
5,281
90.94
12.91
14.22
United States
360
3,811
88.00
10.59
12.03
Colombia
391
2,227
86.25
5.70
6.61
Philippines
324
1,796
81.06
5.55
6.87
South Africa
315
2,684
72.57
8.54
11.76
World
19,307
90,340
65.12
4.68
7.19
Source: FAO 2003, USDA 2002a, USDA 2003b


66
Table 4-1. Assumptions for Baseline (Status quo) Scenario
Category
Shifters
Country
Assumptions
GDP
United
States
and
Will increase at the average real GDP annual
growth rate realized between 1997 and 2001
(2.23 percent for the United States and 2.91
percent for Mexico).
Market
situations
Population
Mexico
Will increase similarly as the forecast by the
U.S. Bureau of the Census. (Annual growth
rate: 0.88 percent for the United states and 1.15
percent for Mexico).
Production
cost
Will decrease at the average annual reduction
rate realized between 1997 and 2001 (1.10
percent for the United States and 1.80 percent
for Mexico).
Recovery rate
United
States
Will increase at the average annual
improvement rate realized between 1997 and
2001 (0.48 percent).
Downtime
Mexico
Will decrease at the average annual reduction
rate realized between 1997 and 2001 (1.00
percent).
Sugar loss
Will decrease at the average annual reduction
rate realized between nl997 and 2001 (2.00
percent).
Duration of
harvest
Held constant
HFCS
consumption
Will be consumed maintaining the same share
in indirect consumption of sweetener realized in
2001 (25.3 percent)
U.S.
sugar
Price support
The U.S. government maintains the price
support (current policy) at 18 cents per pound
(loan rate).
policy
Flexible allocation
The U.S. government allocates quotas in a
flexible manner between Mexico and the rest of
the world.


77
downtime for mills trying to improve their production efficiency. A positive and
significant estimate associated with the trend variable indicates technology related to
sugar production at sugar mills had been advancing during the time period covered in this
study (1988-2000).
Bilateral Sugar Trade Analysis
The results from the bilateral sugar trade analysis are divided into two parts: U.S.
sugar import forecasts and the comparison of pay-offs among industries and countries.
Results and discussion are guided by comparing the impacts from changes in Mexican
sweetener market situations and from changes in U.S. sugar policy.
U.S. sugar import forecasts for the selected eight scenarios (scenario 1, 2, 3, 4, 8,
12, 13, and 16 in Table 4-4) are shown in Figures 5-1 through 5-8. Comparisons among
scenarios 1, 2, 3,4, and 16 illustrate the impacts from changes in the Mexican sweetener
market situations; comparisons among scenarios 4, 8, and 12 illustrate the impacts from
changes in U.S. sugar price stabilization policy; and comparison between scenarios 4 and
13 illustrates the impacts from changes in U.S. quota allocation policy.
The results from the baseline scenario (Scenario 1) shows that if status quo
surrounding the sugar industries in both countries is maintained over the forecast horizon,
Mexico will not likely attain a net surplus sweetener producer status and hence will miss
the opportunity to benefit from exporting sugar under a larger quota allocation (250,000
MT) under NAFTA (Figure 5-1). This is due to growing domestic sugar demand relative
to domestic sugar production. For comparison, forecasts for Mexican sugar consumption
and production in 2008 by this study are 5.4 million and 5.8 million MT, respectively and
those by Koo and Taylor (2000) are 5.3 million and six million MT, respectively.


This dissertation was submitted to the Graduate Faculty of the College of
Agricultural and Life Sciences and to the Graduate School and was accepted as partial
fulfillment of the requirements for the degree of Doctor of Philosophy.
August 2004
Dean, College of Agncu
Sciences
Dean, Graduate School


EVALUATION OF THE AMINO ACID METHIONINE FOR BIORATIONAL
CONTROL OF SELECTED INSECT PESTS OF ECONOMIC AND MEDICAL
IMPORTANCE
By
LEWIS SCOTTY LONG
A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT
OF THE REQUIREMENTS FOR THE DEGREE OF
DOCTOR OF PHILOSOPHY
UNIVERSITY OF FLORIDA
2004


108
Ito, T. and T. Inokuchi. 1981. Nutritive effects of D-amino acids on the silkworm,
Bombyx mor. J. Insect Physiol. 27(7): 447-453.
Jaffe, J.J. and L.R. Chrin. 1979. De novo synthesis of methionine in normal and Brugia-
infected Aedes aegypti. J. Parasitol. 65(4): 550-554.
Jones, D.C and R.M. MacPherson. 1997. Tobacco Insects: Summary of losses from
insect damage and costs of control in Georgia -1997. University of Georgia,
Waroell School of Forest Resources and College of Agricultural and
Environmental Sciences Internet URL: http://www.bugwood.org/tobbaco97.htm.
Accessed April 2004.
Kaldy, M.S. and A.M. Harper. 1979. Nutrient constituents of a grain aphid,
Metopolophium dirhodum (Homoptera: Aphididae), and its host, oats (Avena
sativa). Canadian Entomol. 111(7): 787-790.
Kammer, A.E., D. L. Dahlman and G.A. Rosenthal. 1978. Effects of the non-protein
aminoacids L-canavanine and L-canaline on the nervous system of the moth
Manduca sexta (L). J. Exp. Biol. 75: 123-132.
Kasting, R.. G.R.F. Davis amd A.J. McGinnis. 1962. Nutritionally essential and non-
essential amino acids for the prairie grain wireworm, Ctenicera desctructor
Brown, determined with Glucose-U-C. J. Insect Physiol. 8: 589-596.
Knutson, A., Boring IE, E.P., Michaels, Jr., G.J., and Gilstrap, F. 1993. Biological
Control of Insect Pests in Wheat Texas Agrie. Ext. Service Publ. B-5044 8pp.
Koo, S.I., T.A. Currin, M.G. Johnson, E.W. King and D.E. Turk. 1980. The nutritional
value and microbial content of dried face fly pupae (Musca autumnal is (DeGeer))
when fed to chicks. Poultry Sci. 59: 2514-2518.
Koyama, K. 1985. Nutritional physiology of the brown rice planthopper Nilaparvata
lugens StAl (Hemiptera: Delphacidae). II. Essential amino acids for nymphal
development Appl. Ent Zool. 20(4): 424-430.
Koyama, K. and J. Mitsuhashi. 1975. Essential amino acids for the growth of the smaller
brown planthopper, Laodelphax striatellus FallSn (Hemiptera: Delphacidae).
Appl. Ent Zool. 10(3): 208-215.
Lee, K., F.M. Horodyski, and M.E. Chamberlin. 1998. Inhibition of midgut ion transport
by allatotropin (Mas-AT) and Manduca FLRFamides in the tobacco homworm
Manduca sexta. J. Exp. Biol. 201:3067-3074.
Leonardi, M.G., M. Casartelli, L. Fiandra, P. Parenti and B. Giordana. 2001. Role of
specific activators of intestinal amino acid transport in Bombyx mor larval growth
and nutrition. Arch. Insect Biochem. Physiol. 48: 190-198.


59
exposure (Figures 5-4). The 1.0%L.methionme treatment caused 100% mortality after 2 days
while the Bti treatment took 3 days to reach the same level of control. The proline
treatment caused less than 10% mortality.
In contrast to methionine, survival of YFM larvae exposed to proline and
L-leucine was higher, with only approximately 20% mortality for the higher 0.7% proline
and 1.0% proline concentrations (Figure 5-5) and less than 3% mortality with the highest
L-leucine concentration (Figure 5-6). Beta-alanine mortality was similar to the
L-methionine treatments with between 75% and 83% mortality for the 0.5% Beta-alanine
thru 1.0% Beta-alanine concentrations, respectively, greater than 40% mortality with the
0.3% Beta-alanine, and less than 5% mortality for the 0.1% Beta-alanine concentrations
(Figure 5-7).
Growth and Development
Developmental rates of YFM larvae resulted in three distinct groups, with the
control and proline treatments, producing virtually identical results; both were
statistically different from the 0.1%L-methionine treatment and the remaining
L-methionine treatments (Figure 5-8). The Bti treatment was statistically the same as the
0.3% L-methionine to 1.0% L-methionine treatments, with very little growth taking
place.
Probit analysis for unbuffered L-methionine (nrotai=40 for 5 treatments; 0.1%,
0.3%, 0.5%, 0.7% and 1.0%) revealed an overall LC50 of 0.19% concentration for the
YFM after 7 days of exposure (Figure 5-9). The LC50 of 1.2% for 24 hours dropped to
0.41% after 48 hours and to 0.24% after 72 hours. When the L-methionine treatments
(same concentrations) were buffered to a pH Of 7.0, the values dropped to 0.64% for 24


g of L-methionine/g of Leaf Material
L-methionine Concentration (%)
Y = 8.65E-04 + 4.76E-03X
R-Sq = 98.6 %
95% Confidence Intenal
+/- 2SE (SE=.001997)
Figure 3-4. Amount of L-methionine present on leaf surface after treatment.
Excised leaves were weighed, dipped into various concentrations
of L-methionine, allowed to dry, and then re-weighed. Difference
assumed to be the amount of L-methionine remaining on leaf
surface (T=22.43, and P<0.001).


LIST OF FIGURES
Figure Pagi
2-1. The CAATCH1 system identified from the midgut of the tobacco homworm 15
3-1. Rearing chamber for tobacco homworm and Colorado potato beetle larvae used in
the artificial and excised leaf diet tests 19
3-2. Setup for whole plant studies involving tobacco homworm 21
3-3. Chambers used for tobacco homworm and Colorado potato beetle preference tests 23
3-4. Amount of L-methionine present on leaf surface after treatment 25
3-5. Mortality of tobacco homworm larvae exposed to various concentrations of
L-methionine (n-rotar^O) in artificial diet 26
3-6. Survivorship of THW larvae exposed to various concentrations of L-methionine
(nTota 1,540) on excised eggplant leaves 28
3-7. Mortality of tobacco homworm larvae exposed to various concentrations of
L-methionine (nrotai=256) on whole plants 29
3-8. Concentrations (%) of L-methionine required for the mortality of 50% of test
population of tobacco homworm after 9 days exposure (nT0tai=1,540; n=180
for 3.0% L-methionine 10.0% L-methionine, n=200 for remainder) 30
3-9. Mortality of tobacco homworm larvae exposed to various concentrations of L-
methionine (ntota 160) on excised eggplant leaves for feeding and
development trials 32
3-10. Mean head capsule widths of tobacco homworm larvae exposed to excised eggplant
leaves treated with various concentrations of L-methionine (nTOtai=320) 33
3-11. Total leaf area consumed by tobacco homworm larvae exposed to excised eggplant
leaves treated with various concentrations of L-methionine (nrotai=320) 34
3-12. Mean leaf consumption by tobacco homworm in the preference tests 35
4-1. Mortality of Colorado potato beetle larvae exposed to excised eggplant leaves treated
with various concentrations of L-methionine (nTOtai=560) 44
4-2. Concentrations (%) of L-methionine concentrations required for the mortality of
50% of the test population of Colorado potato beetle after 8 days exposure
(niotai=220) 45
vui


70
Table 4-4. Listing of
examined Scenarios
Scenario
Mexican market
U.S. sugar policies
number
(code)
situations
Stabilization of the
demand price
Quota allocations
1
(baseline)
Baseline
(B)
Price support
Flexible
2
(P-S-F)
High production
(P)
(S)
allocations
(F)
3
(A-S-F)
HFCS adoption
(A)
4
(HA-S-F)
High prod.-HFCS adop.
(PA)
5
(B-B-F)
Baseline
(B)
Buying up excess
6
(P-B-F)
High production
(P)
sugar in the market
(B)
7
(A-B-F)
HFCS adoption
(A)
8
(PA-B-F)
High prod.-HFCS adop.
(PA)
9
(B-C-F)
Baseline
(B)
Production control
10
(P-C-F)
High production
(P)
(C)
11
(A-C-F)
HFCS adoption
(A)
12
(PA-C-F)
High prod.-HFCS adop.
(PA)
13
(PA-S-M)
High prod.-HFCS adop.
Price support
(S)
Minimum quotas
allocation to the
14
(PA-B-M)
(PA)
Buying up excess
sugar in the market
(B)
rest of the world
(M)
15
(PA-C-M)
Production control
(C)
16
(T-S-F)
Tax on HFCS
(T)
Price support
(S)
Flexible
allocations (F)


74
Data Analysis
Data from the fruit and the CPB experiments were analyzed with ANOVA using
Minitab Version 12. Survivorship of CPB was corrected using Abbotts formula (Abbott
1925) to account for control mortality, mean separation was performed using Tukeys
multiple comparison procedure (Zar 1999). Data for both the eggplant weight mean per
treatment and also mean number of eggplants per treatment were analyzed using paired
t-test.
Results
Effects of L-methionine and Silwett L-77 on CPB Adults Under Laboratory Conditions
Little mortality was observed with the adult CPB at the 1.0% L-methionine
concentration (Figure 6-3). The 0.5% L-methionine concentration had the highest
mortality of all the treatments at approximately 20% with the other treatments showing
no adverse effects after correction for control mortality.
Effects of L-methionine and Silwett L-77 on yield
In total, 735 eggplants were collected during the course of this study from 09 June
to 31 August 2001. Mean weight and yield of eggplants between the treatments were not
statistically different from each other (Figures 6-4). Control plants produced 195 fruits
with a mean weight of 276.9 grams, followed by the 0.1% treatment with 191 fruits at
281.2 grams. The 0.5% and 1.0% treatments yielded 175 and 174 fruits with mean
weights of 295.7 grams and 283.6 grams, respectively.
Survival of CPB larvae
No statistical difference in survivorship of CPB larvae was observed between the
three treatments for the first day after exposure (Figure 6-5) but treatment differences


18
interaction in a variety of ways, including artificial diet, natural diet (excised leaves,
whole plant, and choice tests. The purpose of this portion of this study was to determine
whether L-methionine was detrimental to the survival and development of the THW and
to determine if L-methionine could be used to control this species.
Materials and Methods
Eggs of THW were obtained from the insectary of North Carolina State
University, and were held in 26.4L x 19.2W x 9.5H (cm) clear plastic rearing chambers
with a hardware cloth (to facilitate cleaning) (Figure 3-1). Florida Reach-In Units
(FRIUs) were used to control the environment for the rearing containers (Walker et al.
1993) Containers were held at 27 C, 60% relative humidity, and a 16L:8D photoperiod
in FRIUs with either artificial or natural diet (excised eggplant leaves or whole plants)
depending on the pending experiment. Neonates were allowed to feed for 2 days after
eclosin before being transferred to treatment groups. A camel hair brush was used for
transferring larvae, to minimize the risk of damage.
Diets and Survivorship
The artificial diet was prepared using the procedures outlined in Baumhover et al.
(1977) with the inclusion of L-methionine for the treatment concentrations of 0.1%,
0.3%, 0.5%, 1.0%, 3.0%, 5.0% and 10.0% (wt/wt). The artificial diet was changed on a
regular basis to prevent desiccation and fungal growth. Larvae were exposed to the
artificial diet in the clear plastic rearing chambers with a hardware cloth, and kept in the
FRIUs programmed with the aforementioned environmental constants.
Natural diets consisted of excised eggplant leaves (Solarium melongena
L.,Classic variety) of potted plants grown and maintained at the University of Florida,


95
Year
Mexico under-quota export
Mexico over-quota export
Mexico quota-free export
^ The rest of the world export to the U.S.
Figure 5-6. U.S. Sugar Import Forecast (Scenario 8 PA-B-F)


27
treatment after 10 days of exposure. The 0.1% L-methionine concentration had lowest
larval mortality with approximately 30% observed for the trial.
The excised leaf trials exhibited higher mortality rates associated with the
treatments than did the artificial diet trials. Again, complete mortality was observed with
the 3.0% L-methionine thru 10.0% L-methionine concentrations after 1 day of exposure
(Figure 3-6). Greater than 90% in the 0.5% L-methionine and 1.0% L-methionine
treatments, followed by 80% mortality in the 0.3% L-methionine treatment, and greater
than 60% mortality occurred in the 0.1% L-methionine treatment after 8 days.
Whole plant trials produced results similar to the excised leaf trials with greater
than 90% larval mortality observed with the 1.0% L-methionine treated plants after 14
days (Figure 3-7). Mortalities exceeding 20% and 60% were observed for the 0.1%
L-methionine and 0.5% L-methionine treatments, respectively, during the same time
interval.
PROBIT analysis of a sample size of n-rota 1,540 for 7 treatments (0.1%
L-methionine, 0.3% L-methionine, 0.5% L-methionine, 1.0% L-methionine, 3.0%
L-methionine, 5.0% L-methionine and 10.0% L-methionine) revealed an overall LC50 of
0.66% (32.3 mg/g leaf material) concentration for the artificial diet and 0.074% (4.39
mg/g leaf material) concentration for the natural diet after 9 days of exposure (Figure
3-8). The LC50 for the THW exposed to artificial diet was approximately half the value
of that for the natural diet for the 24 to 72 hour exposure period. The LC50 for the
artificial diet of 1.08% (52.3 mg/g leaf material) for 24 h dropped to 1.0% (48.5 mg/g leaf
material) after 48 h and to 0.57% (28.0 mg/g leaf material) after 72 h. As for the natural
diet, the LC50 of 0.53% (26.1 mg/g leaf material) was found to be lower than the artificial


ACKNOWLEDGMENTS
I would like to express my gratitude to Dr. Thomas H. Spreen, chair of my
supervisory committee, for providing me with resources throughout my program of
studies. My dissertation would not have been possible without his guidance, unflagging
enthusiasm and helpful ideas.
I also would like to express my gratitude to Dr. Lisa A. House, cochair of my
supervisory committee, for her generous and patient support during my course of studies.
Under her supervision, my research has been completed in an efficient and successful
manner.
Special appreciation is extended to Dr. Chris O. Andrew for his encouragement
throughout the program and for broadening my horizons. His inspiration has been a
compass after the direction of my career had been shifted. I am also deeply grateful to Dr.
Luis R. Garcia for providing me with insights and detailed data which were indispensable
for this study; to Dr. Terry L. McCoy for his contributing to the depth of my study; and to
Dr. Kenneth L. Buhr for his sharing suggestions for the breadth of my study. Kind
support from Dr. Jeffrey R. Burkhardt and staff in the department for their various
services are also appreciated. I cannot fail to thank John S. Lander, who opened my eyes
to studying overseas, for his unconditional support and wisdom. Friendship from my
colleagues in the program is a cherished treasure.
Last but not least, the financial support received from the Food and Resource
Economics Department throughout my program has been greatly appreciated.
IV


36
Qsugar (Psugar Px, m).
(3.8)
Since the demand function is homogeneous of degree zero, it can be normalized by either
price. Normalize by Px :
(3.9)
Qsugar (Psugar/Px, m/Px),
and thus the aggregate demand for sugar is expressed as a function of real price of sugar
and real income. By treating real income as one of demand shifters, aggregate demand is
expressed as shown in equation [3.1]. Similarly the aggregate demand for HFCS can be
expressed as a function of HFCS price and a vector that affects aggregate HFCS demand.
Note that in reality, HFCS is consumed directly by industry users and its price is
observed by them; households as final consumers consume HFCS indirectly through
HFCS-contained goods.
Sweetener supply
Sweetener supply is defined based on firm supply theory derived from profit
maximizing behavior. Although in reality sugar supply consists of two steps of
production in reality, sugarcane and sugar production, a simple aggregate sugar supply
equation at industry level is derived rather than two equations. This is suitable for two
reasons: a single industry supply equation makes simulations in the bilateral trade model
in the following procedure simple, and it is the industry supply price that the government
is interested in supporting.
Suppose the industry faces a cost function given by:
C C (Qsugar, V)
(3.10)
where Visa set of other factors including input prices. The industry maximizes profits,
assuming the market is competitive (the industry as price taker):


24
larval head capsule measurements made using the same procedures described in the
Feeding and Development section.
Data Analysis
Sample sizes of all experiments were chosen according to the guidelines
recommended by Robertson and Preisler (1991) for optimal sample size and data
analysis. Probit analysis and determination of mean Lethal Concentration (LC50) were
performed using PROBIT Version 1.5 (Ecological Monitoring Research Division,
USEPA) after Abbotts correction for control mortality (Abbott 1925). Survival data
were normalized to the previous value when control mortality was greater than the
treatment mortality, to produce a smoother trend line. Statistical analysis was performed
on the data using Minitab Version 14 (Minitab, Inc.; State College, PA). Analysis of the
data included One-way ANOVA and separation of significant means using Tukeys
Multiple Comparison and Pearson Correlation was performed on the choice trial data to
examine possible relationships between development and consumption of treated leaf
material (Zar 1999). Regression analysis using lest squares were performed on the leaf
weights before and after the L-methionine treatment for the equation used to convert %
concentration to mg/g plant material (Figure 3-4).
Results
Diets and Survivorship
The artificial diet resulted in 100% mortality of THW larvae for the 3.0%
L-methionine to 10.0% L-methionine treatment after only one day of exposure (Figure
3-5). Approximately 80% mortality was observed in the 1.0% L-methionine treatment
after 4 days, and 50% mortality for both the 0.3% L-methionine and 0.5% L-methionine


42
consisted of 4 leaf disks (30 mm diameter) dipped into the Control solution and placed
into the chamber alternately with four leaf disks (30 mm diameter) dipped into the
treatment solution and replicated with a total of 10 chambers. Each chamber consisted of
a large petri dish (25.0 cm diameter x 9.0 cm depth) lined with a Seitz filter disk. The
filter disk was moistened routinely with deionized H2O to prevent the leaf disks from
desiccation (Figure 3-3). Chambers were held in FRIUs at the same environmental
constants described previously. The leaf disks also were scanned photometrically and
larval head capsule measurements made using the same procedures described in the
Feeding and Development section.
Data Analysis
Sample sizes of all experiments were chosen according to the guidelines
recommended by Robertson and Preisler (1991) for optimal sample size and data
analysis. Probit analysis and determination of mean Lethal Concentration (LC50) were
performed using PROBIT Version 1.5 (Ecological Monitoring Research Division,
USEPA) after Abbotts correction for control mortality (Abbott 1925). Survival data
were normalized to the previous value when control mortality was greater than the
treatment mortality, to produce a smoother trend line. Statistical analysis was performed
on the data using Minitab Version 14 (Minitab, Inc.; State College, PA). Analysis of the
data included One-way ANOVA and separation of significant means using Tukeys
Multiple Comparison and Pearson Correlation was performed on the choice trial data to
examine possible relationships between development and consumption of treated leaf
material (Zar 1999).


81
will fall below the support price (Figures 5-15 and 5-16). This is a result of cheaper sugar
from the rest of the world flowing into the U.S. market. In reality, the U.S. sugar price
will face downward pressure from importing world sugar as well as political pressure
from the rest of the world, considering the likelihood that many sugar exporting countries
will not easily give up their existing shares of the U.S. import quotas.
Pay-offs to the industries and countries also portray interesting contrasts among
scenarios. The results from three sets of selected scenarios are summarized in Tables 5-3,
5-4, and 5-5. In these tables, the present values of accumulated pay-offs are expressed in
billion of dollars and those values are indexed relative to the baseline scenario inside the
brackets.
The impact of changes in Mexican sweetener market on pay-offs to the industries
and the two nations welfare is illustrated in Table 5-3. The listed five scenarios are based
on the assumptions that the U.S. government maintains price support and allocates quotas
between Mexico and the rest of the world in a flexible manner. It is clear that the U.S.
HFCS industry will become better off if Mexico adopts HFCS: revenue for the U.S.
HFCS industry increases by 78 percent; and that the industry will become worse off if
Mexico introduces taxes on HFCS: revenue for the industry decreases by 72 percent. The
U.S. sugar industry does not gain from either Mexicos increase in sugar production or
HFCS adoption since either change in Mexican sweetener market generates a larger
Mexican sugar surplus that is destined for the U.S. market. Tariff revenue to the U.S.
government from Mexican sugar increases as Mexico increases sugar production or
HFCS adoption. Large tariff revenue from high HFCS adoption scenario, which is
unexpectedly larger than the high production-high HFCS adoption scenario, is due to


62
two coalitions formed among five parties (payees), i.e. three industries (the U.S. HFCS
industry, the U.S. sugar industry and the Mexican sugar industry) and two countries;
however, the final decisions are made by the governments who hold the strategies.
Each country plays with multiple strategies that correspond to Mexicos market
situations and the U.S. policy levers presented in the previous section (Table 4-5). In this
game setting, a combination of strategies formulates a scenario. In the game, it is
assumed that if the U.S. government introduces production control, Mexico always
cooperates: a defection by either party is not considered. Also, the U.S. government
chooses only flexible quota allocations to the rest of the world for simplicity. In the case
of Mexicos strategies, HFCS adoption is treated as a strategy although it is neither a
positive strategy nor controllable by the Mexican government.
Pay-offs from each scenario are calculated for each payee. Pay-offs to the
industries are expressed as present values of accumulated revenue between 2002 and
2015, assuming a three percent discount rate each year. The HFCS price is held constant
at the average U.S. export price to Mexico realized between 1992 and 2001. Pay-offs to
each country are expressed as present values of accumulated welfare, i.e. the sum of
consumer and producer surplus. In doing so, changes in welfare are calculated only from
the sugar market, assuming that sugar is a primal source of sweetener. Sugar cannot be
substituted with HFCS for certain products due to the liquid form of HFCS. Also, sugar is
preferred for certain products to HFCS due to flavor given to the final products. U.S.
welfare is adjusted with tariff revenue from Mexico, the cost of the sugar program and
the cost of buying up excess sugar. The cost of the sugar program is calculated by
multiplying the price difference between the support price (loan rate for raw sugar, 18


78
were observed thereafter. By Day 4 the 1.0% and 0.5% treatment were the only
treatments that were statistically different from the control. There was substantial
unexplained attrition of CPB larvae in the field for all treatments, which leveled off by
Day 3. Data from day 5 was discounted because of the onset of a severe cold front that
made it difficult to separate the effects of the weather from the treatments affects.
Discussion
The results of the field studies show that, using conventional application
techniques, a mixture of methionine and Silwett L-77 did not appear to affect eggplant
yield. Furthermore, the same combination produced substantial control of CPB larvae
under natural field conditions after four days. Dahlman (1980) found that L-canavanine,
a non-protein amino acid, could be used in the same manner for control of THW on
tobacco, but the widespread use of this compound was limited by the cost ($107.85 for lg
L-canavanine versus $3.35 for lg of L-methionine (Fisher Scientific International 2004)),
adverse effect on plant development (Nakajima et al. 2001), and toxicity to vertebrates
(Rosenthal 1977). Although complete coverage of the plant was not feasible,
approximately 2.5 grams to 7.5 grams of L-methionine was applied to the plants in each
of the treatment plots. Each plant, based on the amount applied, received approximately
7.5xl06 pg for the 1.0% L-methionine treatment, 3.8xl05 pg for the 0.5% L-methionine
treatment and 2.5x104 pg for the 0.1% L-methionine treatment. This compares to only 4pg of
L-canavanine, which resulted in decreased size, fecundity, and mortality of THW under
field conditions (Dahlman 1980). It should be noted that the toxicity of L-canavanine is
well documented and has a different mode of action than L-methionine and cannot be


Table 5-12. Indexed Pay-off Matrix for the Trade Policy Game Played by the Industry Coalition and the Government Coalition with


38
Yj(Xu + XXij)>Qj,Vj = l...,J (3.15)
1=1
X¡j < Quotdy, Vi = 1,...,/, V/ = 1,...,7 (3.16)
Yi,QrX,JandXXl}>{)
where X¡j is the quantity of sugar shipped from supply region i to demand region j under
quota limit, TRy is per unit transfer costs associated with Xy, XX¡j is the quantity of sugar
shipped region i to demand region j over quota limit, TTRy is per unit transfer costs
associated with Xy. TRy is a compound transfer cost that includes per unit transportation
cost (T¡j) and per-unit tariffs imposed on exported sugar under quota limit (Tary ).
Similarly, TTRy is a compound transfer cost that includes per unit transportation cost ('Ty)
and per-unit tariffs imposed on exported sugar over the quota limit (over-quota tariff,
OQTary).
TRy = Ty + Tary (3.17)
TTRy = Ty +OQTary (3.18)
Let specific inverse linear demand and supply functions for United States and
Mexico be defined as follows:
Pus = IU, + IU2*QDus + shifter us
(3.19)
p\js = IUU, + IUU2*QSus + Shifters us
(3.20)
PMX = IM, + IM2 *QTCmx + Shifter0 MX
(3.21)
PSMX = IMM, + IMM2*Qsmx + Shifter5 MX
(3.22)
where P/;and QDrepresent the price and the quantity demanded in each country,
respectively; Ps and ^ represent the price and the quantity supplied in each country,
respectively; Shifter and Shifter^ represent the demand and supply shifters in each


14
in its use as a feed supplement for livestock under the trade name of Alimet (Novus,
Inc., St. Louis, MO).
In addition to vertebrates, methionine also is considered an essential amino acid
for insects (Nation 2001). Based on research on nutritional requirements for insects, the
amount of methionine needed in a diet for survival ranged from as little as 0.0007 mg/mL
(for Aedes aegypti (L.) (Diptera: Culicidae) to as high as 100 mg/mL (for Heliothis zea
(Broddie) (Lepidoptera: Noctuidae)) (Dadd and Krieger 1968; Eymann and Friend 1985;
Friend et al. 1957; Kaldy and Harper 1979; Kasting et al. 1962; Koyama 1985; Koyama
and Mitsuhashi 1975; Rock and Hodgson 1971; Singh and Brown 1957). Methionine
occurs naturally as the L-isomer while the D-isomer (an optical enantiomer) is toxic to
many insects and is not found in nature (Anand and Anand 1990). A few exceptions are
known, (mainly Diptera and Lepidoptera) that actually are capable of metabolizing the
normally unusable D-isomer (Dimond et al. 1958; Geer 1966; Rock 1971; Rock et al.
1973; Rock et al. 1975). The requirement for small amounts of this amino acid (as
compared to other amino acids) may be a result of the ability for some insects to
synthesize methionine from cysteine (a common sulfur containing amino acid) thus
reducing the need to take in exogenous sources of methionine. Jaffe and Chrin (1979)
found that A. aegypti adults were able to synthesize methionine from homocysteine with
the aid of a methionine synthetase. They found this enzyme similar to those common in
other metazoans, and found that the levels of methionine synthetase increased with the
presence of filarial parasites. They hypothesized that this increase in methionine
synthetase was a result of the parasite depleting the host of methionine.
Fertility and fecundity also have been associated with methionine in some insects
(mainly D. melanogaster,) with the possibility if it being a limiting factor during egg


CHAPTER 4
EMPIRICAL MODELS AND DATA SOURCE
Based upon theoretical framework developed in chapter 3, this chapter focuses on
the empirical procedures and data set used in the analysis of the U.S.-Mexican sweetener
market. The empirical model consists of three components: (1) demand and supply
analysis models for both the U.S. and Mexican sugar markets; (2) a bilateral sugar trade
analysis model; and (3) game theory analysis. These models are ordered sequentially so
as to utilize the results from the former. After the trade model is introduced, assumptions
associated with the model, the methods of model calibration, and simulated scenarios are
presented. Results from simulations on the trade model are aggregated to assess policy
recommendations. Lastly, an overview of the data set used in the empirical models is
provided.
Empirical Models
U.S. Sweetener Demand Model
Regression analysis is used to estimate sweetener demand utilizing quarterly time-
series data. The model is specified as a double-log (natural log) form that allows
interpretation of estimated coefficients as elasticities associated with each variable. Based
on the basic form of demand equation shown in equation [3.1], the U.S. sugar demand
equation is specified as:
In(QDSUGAR, i) = U,+ U2*In(PD sugar,,) + U3*In(GDPt) + U4*In(POPt)
+ U5*QTR1+ U6*QTR2+ U7*QTR3+ U8*DHFCS + e, (4.1)
where Q is quantity of sugar demanded in each quarter in year t [1000 short tons], P is
47


80
to generate a sugar surplus after 2006 due to expanding domestic sugar consumption
accelerated by higher sugar consumption by bulk users who chose sugar over HFCS.
Mexico cannot produce enough sugar to meet domestic demand after 2008 and will
import sugar from the United States, resulting in a higher domestic sugar price than that
of the U.S. (Figure 5-12). This extreme case would occur only if the impact of tax on
HFCS lingers over the forecast horizon as assumed in this study; however, the chance for
Mexico to enjoy exporting sugar would be slim.
In spite of fluctuating imports from Mexico and the rest of the world, U.S. domestic
sugar consumption and production will remain relatively unchanged. Sugar demand and
supply forecasts for both the United States and Mexico for baseline and high production-
high HFCS adoption scenarios are shown in Figures 5-13 and 5-14, respectively. In either
scenario, U.S. demand and supply are forecasted to remain approximately at nine million
and 8.8 million MT over the forecast horizon, respectively.
The results from the various simulations showed that the U.S. sugar price will
gradually decline but will not dip below the support price level (396.48 US$ per MT)
before 2008 if the United States accepts most of the imported sugar from Mexico rather
than from the rest of the world by allocating quotas in a flexible manner to the exporters,
no matter which U.S. demand price stabilization policy is in place (Figures 5-9, 5-10, and
5-11). This implies that the Mexican sugar price contributes to maintain a high sugar
price in the integrated U.S.-Mexico sugar market; in other words, accommodating
Mexican sugar can act as an alternative form of price support in the United States. On the
other hand if the U.S. government maintains minimum quotas for the rest of the world no
matter how much Mexico exports and abandons the price support, U.S. equilibrium price


89
100
80
g
13 60
S 40
20
0
Figure 7-2. Mortality of Coleomegilla maculata adults after exposure to L-
methionine treated cotton plant leaves infested with aphids. Data
corrected for control mortality using Abbotts formula.
Control- ND
1.0% L-methionine ND
Survivorship of 1.0%L-methionine Grp> Control Grp
4 5 6 7 8 9
Days After Exposure
10 11 12


56
Data Analysis
Sample sizes of all experiments were selected according to the guidelines of
Robertson and Preisler (1991) for optimal sample size and data analysis. Probit analysis
and determination of mean Lethal Concentration (LC50) were performed using PROBIT
Version 1.5 (Ecological Monitoring Research Division, USEPA) after Abbotts
correction for control mortality (Abbott 1925). Probit analysis was performed on
different concentrations (0.1%, 0.3%, 0.5%, 0.7% and 1.0%) of L-methionine, Tris-
buffered L-methionine, D-methionine, Beta-alanine, proline and L-leucine for 24,48, 72
and 168 hours (the end of the trials). Survival data were normalized to the previous value
when control mortality was greater than the treatment mortality, to produce a smoother
trend line. Statistical analyses were performed on the data using Minitab Version 12.
Analysis (Minitab, Inc; State College, PA) of the data included One-way ANOVA and
separation of means using Tukeys Multiple Comparison test (Zar 1999).
Results
Bioassav
Mortality of YFM larvae in both the unbuffered L-and D-methionine trials was
similar with low or no mortality, at the 0.1% concentrations (Figures 5-2 and 5-3). The
0.3% concentration had lower mortality with D-methionine (45%) than L-methionine
(75%) and greater than 80% mortality for the 0.5% concentration for both isomers.
Higher concentrations of both D-and L-methionine forms produced 100% mortality of
the larvae within 2 days after treatment.
Greater than 40% mortality was observed for the buffered 0.1% L-methionine
concentration with complete mortality for the remaining treatments within 5 days of


56
D S
considering Mexicos increasing demand for HFCS. Since HFCS and HFCS are not
estimated directly, HFCS demand (HFCS ) is forecasted based on the associated
scenarios with details provided in the following section.1 Once HFCS demand is
forecasted, the quantity of excess sugar destined to export is calculated by considering the
substitution between HFCS and indirect sugar consumption, which occurs at industry
A
(bulk users) level. Let Qlcsugar, ibe HFCS forecast-adjusted indirect sugar consumption,
then the sugar export forecast (SEt) is expressed with forecasted quantities of sugar
demanded and supplied (Qssugar, t, Q DCsugar, t ,and Q,csugar, ) as:
SE, = Q SSUGAR, f (Q DCSUGAR, i + Q 'CSUGAR, t ) (4.12)
In equation [4.34], it is also assumed that HFCS is consumed only to supplement sugar
consumption and its demand is met by readily available FtFCS from domestic production
and import from the United States. Accordingly, NAFTA provisions are defined,
depending on the Mexicos domestic sweetener balance; Mexico receives a larger quota
for the following year if is attained net surplus producer status for two successive years:
Q DCSUGAR, t + Q ICSUGAR, t + HFCS, < Q SSUGAR, t, (4.13)
otherwise, Mexico receives a smaller quota until 2008 when access to the U.S. market is
unlimited.
Model calibration
The model is calibrated by positioning the intercepts of inverse linear demand and
supply equations and by adjusting average transportation cost from the rest of the world
1 Alternatively HFCS demand can be forecasted balancing the total consumption of sweetener equation
with direct and indirect consumption sugar equations, yet it resulted in a poor forecast.


92
(Error bars @ 95%; F(00S)4> u= 0.98, F=3.33; /* =0.038)
1400
Control 0.1% 0.5% 1.0% Proline
Figure 7-5. Feeding rate of Neochetina eichhorniae on water hyacinth leaves treated
with L-methionine and Proline. No statistical differences were observed
between treatments (Tukeys MST, P=0.038).


71
Table 4-5. Strategies for the Sugar Trading Game
Country
Strategies
United States
Strategy 1
Maintains price support (status
quo)
Flexible quota
allocations among
Mexico and the
rest of the world
Strategy 2
Abandons price support and buys
up excess sugar in the market
Strategy 3
Introduces production control
with Mexico
Mexico
Strategy 1
Maintains the current policy (status quo)
Strategy 2
Higher sugar production
Strategy 3
Higher HFCS adoption
Strategy 4
Higher sugar production and higher HFCS adoption
Strategy 5
Introduces tax on HFCS


73
Table 4-8. Data Sources for U.S. Supply
Data
Unit
Source
Production of sugar, total
(dependent variable)
1000
short
tons
Sugar Statistical Compendium by (Stock
#91006, 1970-1990) and Sugar and
Sweetener Situation and Outlook
Yearbook (SSS-2002, 1980-2002) by
Economic Research Service, U.S.
Department of Agriculture
Production of cane sugar
(dependent variable)
Production of beet sugar
(dependent variable)
Retail price of refined sugar
Wholesale price of refined
cents/
pound
Sugar recovery rate, total
percent
Beet sugar recovery rate
Cane sugar recovery rate
Total farm production
expenses
million
US$
Database by Economic Research Service,
U.S. Department of Agriculture
Data length: 1960-2002
Table 4-9 Data Sources for Mexico Supply
Data
Unit
Source
Production of sugar
(dependent variable)
MT
Database by Comit de la Agroindustria
Azucarera (COAAZUCAR)
Wholesale price of standard
sugar
pesos /
kg
Cost of sugar production per
ton of sugar produced
pesos/
ton of
sugar
Downtime observed at mills
percent
Sugar loss during the process
percent
Duration of the harvest
days
Data length: 1988-2000


39
country, respectively; 1U¡, IUUi, IM¡, and IMM¡ represent the intercepts in each inverse
linear equation; and IU2, IUU2, 1M2, and IMM2 represent coefficients associated with
each quantity variable. The objective function of the U.S.-Mexico bilateral sugar trade
system is expressed with equations [3.19], [3.20], [3.21], and [3.22] with constraints such
as necessary conditions, i.e. demand-incoming shipment and supply-outgoing shipment
balance as well as those specific to the bilateral trade model such as Mexicos quota
allocation under NAFTA and U.S. price support:
Max I (IU, + IU2*QDus + Shifter?us) dQus
+ I (IMi + IM2*QICmx + Shifter?Mx)dQTCMX
- J (IUUi + IUU2*QSus + Shifter*us) dQ*US
- J (IMM, + IMM2*QSmx + Shifter*Mx)dQsmx
(TMX, US )*XMX. US
(Tus,mx)*X US, MX
(Tmx, us + OQTTmx, us )*XX mx, us
- (Trow, us + Prow)*Xrow, us
+ Prow *Xmx, row (3.23)
where US, MX, ROW represent Mexico, the United States, and the rest of the world,
respectively; T¡j is per unit transportation cost from i to j; OQTarMx. us is per unit per unit
over-quota tariff imposed on Mexican sugar shipped to the United States; and Prow is
world price of sugar. Note that the transportation cost within the country is assumed to be
zero. The over-quota import from the rest of the world to the United States (XXrow, c/s) is
ignored because of the high tariff rate imposed on the sugar from the rest of the world,


Table 5-11. Indexed Pay-off Matrix for the Trade Policy Game Played by Two Coalitions of Countries with the U.S. HFCS industry
[Baseline=100]
Mexico's strategies
Maintain the current policy
(status quo)
High production
High HFCS adoption
High production -
High HFCS adoption
U.S.
Mexico
U.S.
Mexico
U.S.
Mexico
U.S.
Mexico
U.S.
strategies
Maintain
price
support
(status
quo)
too
100
99.84
102.01
100.92
SX\\'\VV\S\V\'VVV\VS\V'\\\\\V
XXXxXXxXXN>
"N\"V
XXXXXXVVXXX
WWWAWWWW 'Xv- XX -XX
XXXXXXXXXxxXxxxXXXX- ----XX-
\\\\x\\\\\\\\s\\\\\\v\x\\v\\
100.96
97.33
Buying up
excess
sugar in
the market
100.03
95.57
100.01
102.01
101.10
S\\\\\\\\\\\XX\\\X\\\\X\\\V\V
SX>VXX'x
X X -X'
XxXxxxxxxxXXX
XX -X- x-
XXXXXXXXXXXXXXxXXXXXX
XXXXX- -XXXX X XXX' XXX XV
XXXXXXXXXXXXXXXXXXX
VXXXXXxXxVxVV
Xxxxxxxxxxxxxxxxxxx
ESrexsiS
XXXXXXv£. -- XXX XXX
XXXXX-XXXXXXXXX
VXX-XXXXXX
XXXXXXXXXXXXXX
xXX-XXXXXXXXXXXXX
xXXXXXXXXXXXXXxXXXXxXXX
xXXXXXXXxXXXXXXXXXXXX
.XXXXXXXXXXXXXXXXXXXXXXX
xxxxxx XXXX----XX- XV
XXXXXXXXxXXXXXXXXXX
XXXXXXXXXXXXXXXXXXXXXXXXXXXXX
101.12
97.33
Production
control
with
Mexico
100.06
95.57
I
1
if
101.16
XXXXXXXXXXXXXXXXXX.
xXXXXXXXxXXXXxXXXXXxxxXxXxXXX
-XXXXXXXXXXXXXxXXXXXV'
XXXXXXXXXXXXXXXXXXXXXXXXXXXXX
XX -XXXXXX-XX- -XXXXXX
XXXXXXXXXXXXXXXXxXX
.XXXXXXXXXXXXXXXXXXXXXXXXXXXXX
xxxxxxxxxxxxxxxxxxxxxxxxxxxxx
XXxXXXXXXXXXXXXXXXXXX
XXXXXXXXXXXXXXXXXXXXXXXXXXXXX
-xxxxxxxxxxxxxxxxxxxxxxxxxxxxx
XXXXx-XXXXXXXXX
XXXxXXXXXXXXXXXXXXXXXXX
XXXXXXXXXXXXXXXXXXXXXXXXX
XXXXXXXXXXXXXVXXXXXXV
,XXXXXXXXX-X-
XXXXXXxxxxxxxxxxxxxxxxv
XXXxxxxxXXxxxx
XXXXXXXXXXXXXXXxxxxxxxv
-XXXXXXXXXXXXxxxxxxxxx
XXXXXXXXXXX
XXXXXXXXXXX
100.94
-XXXXXXXXXXXXXXX xxx-
XXXXXXXXXXXXXXXXXXXXXX'
XXXXXXXXXXXXXxxxvx
XXXXXXXXXXXXXXXXXXX'
XXXXXXXXXXXXXXxxxX
XxXXXXXXXV
XXxxxxxxXXXXXXXXXXXX
XXXXXXXXXXXXXXXXXXXXX
.XXXXXXXXXXXXXXXXXXXXXXXXXXXXXx
XXXXXXXXXXXxXXXXXXXXXXXX
90.74
XXXXXXVXXXXXxx
XXXXXXXXXXXXXXxXXXXXXXXXXV
XxxxxXXXXXXx
XXXVXXXXXXXXXxXXXXXXXX
.XXXXX XXXX XXX xxxXXXXXXXXXXXX
XXVXXXXxxXXxXXXXXXXXXV
XXXXX X XXX XXXXXX VXX x XXX
XVXXXVXV
XXXXXXXvVxXX
XXXXXXXXXXXXXV
XXXXXXXXxxxXVXXXXXX' '


53
former regression, statistically significant estimates were 0.0286 (lag=3) and 0.0294
(lag=4) and from the latter, the estimate was not statistically significant.
U.S.-Mexico Bilateral Sugar Trade Model
Based on the theoretical framework presented in chapter 3, a bilateral trade model
of U.S.-Mexico sugar trade is developed and prepared for the empirical study. First,
linear inverse demand and supply equations (price endogenous) are formulated using
estimated elasticities from the previous analysis following the procedures by Spreen et al.
(2000) in order to formulate the objective function in the trade model. Detailed derivation
is noted in Appendix C. Although the double-log form is used in the demand and supply
analysis because of its statistical advantages in estimation, the linear form of demand and
supply equations are used in the trade analysis model for the following reasons: with the
linear form, welfare can be measured, whereas welfare cannot be measured using log
form equations due to the nature of the mathematical attributes; using the linear form
makes it possible for the demand or supply curves to shift with their slopes held constant;
and interpretation of Kuhn-Tucker conditions presented in chapter 3 is facilitated.
Sugar trade models have been developed by Koo and Taylor (2000) and Petrolia
and Kennedy (2000). The former model incorporates sugar production, consumption and
stock changes in seventeen sugar producing and consuming countries and the latter
encompasses the United States, Mexico, and Cuba. In these models, market equilibrium
is solved based on the market clearing condition, i.e. the sum of all countries excess
demand for sugar becomes zero as sugar price adjusts. These models are relatively simple
and may be suitable for macroscopic analysis since they are able to include many
economies without making the models large and complicated; however, the solutions
derived from these models pay little attention to countrys welfare or transfer costs of


32
Figure 3-9. Mortality of tobacco homworm larvae exposed to various concentrations of
L-methionine (njota 160) on excised eggplant leaves for feeding and
development trials. Proline (1.0%) and Btk were included for comparison
as positive and negative controls. Data were adjusted using Abbotts
formula for control mortality. Note the overlap in the 0.7% L-methionine,
1.0% L-methionine and Btk treatments at Day 1.


68
mechanisms and many are the result of a combination of different pesticide classes. The
CAATCH1 system is one that could be exploited in cases where the only alternative is
applying different or higher rates of pesticides to break resistance. Further research is
needed to determine compatibility of Bti and L-methionine for cases in which resistance
is observed in natural populations. Given the safety of L-methionine and the similar time
required for 100% mortality (when compared to Bti), this compound could represent a
viable alternative to traditional biorational compounds used in the management of the
YFM or other susceptible pest mosquito species.


105
Dimond, J.B., A.O. Lea and D.M. Delong. 1958. Nutritional requirements for
reproduction in insects. Proc. 10 Int Congr. Entomol. 2:135-137.
Droux, M, B.Gakire, L. Denis, S. Ravanel, L. Tabe, A.G. Lappartient, D. Job. 2000.
Methionine biosynthesis in plants: biochemical and regulatory aspects. Pp. 73-92.
/VBrunold, C., Rennenberg, H., De Kok, L.J., Stulen, L, Davidian, J.C. (eds.):
Sulfur Nutrition and Sulfur Assimilation in Higher Plants. Molecular,
Biochemical and Physiological Aspects. Paul Haupt Publishers. 447pp.
Durham, S. 2000. Hairy vetch thwarts Colorado potato beetle. Agricultural Research
Service, United States Department of Agriculture. Internet URL:
http://www.ars.usda.gov/is/pr/2000/000413.htm. Accessed April 2004.
Dwyer, J. 1999. Research Links 2000 Tobacco Homworm. Carleton College,
Department of Biology. Internet URL: http://www.acad.carieton.edu/curncular
/BIOL/resources/rlink. Accessed April 2004.
Ehler L.E. and D.G. Bottrell. 2000. The illusion of integrated pest management. Issues
in Science and Technology Online. National Academies and the University of
Texas (Dallas). Internet URL: http://www.nap.edU/issues/16.3/dder.htm.
Accessed April 2004.
Elliott, N.C., J.A. Webster, and S.D. Kindier. 1999. Developmental response of
Lysiphlebus testaceipes to temperature. Southwest Entomol. 24: 1-4.
Eymann, M. and W.G. Friend. 1985. Development of onion maggots (Dptera:
Anthomyiidae) on bacteria-free onion agar supplemented with vitamins and amino
acids. Ann. Entomol. Soc. Am. 78:182-185.
Feldman, D.H., W.R. Harvey and B.R. Stevens. 2000. A novel electrogenic amino acid
transporter is activated by K+ or Na+, is alkaline pH-dependent, and is CT-
independent. J. Biol. Chem. 275:24518-24526
Felton, G.W. and D.L. Dahlman. 1984. Allelochemical induces stress: Effects of L-
canavanine on the pathogenicity of Bacillus tkuringiensis in Manduca sexta. J.
Invert Path. 44: 187-191.
Ferro, D.N. 1985. Pest status and control strategies of the Colorado potato beetle. IN
Ferro, D.N. and R.H. Voss (eds.) Proceedings of the Symposium on the Colorado
potato beetle, XVII Inemational Congress of Entomology.
Fisher Scientific Internationa]. 2004. Online Catalog. Fisher Science International
Internet URL: https://wwwl.fishersci.com/index.jsp. Accessed April 2004.
Florida FIRST. 1999. Putting Florida FIRST: Focusing LFAS resources on solutions for
tomorrow. University of Florida, Institute of Food and Agricultural Sciences.
16pp.


51
equations, respectively; P d represents the real retail refined sugar price in the previous
year (deflated by CPI), real raw sugar price at NY sugar exchange in the previous year
(deflated by CPI), and real wholesale refined beet sugar price in the previous year
(deflated by CPI [cents / pound]), respectively; COST is real total farm production
expenses deflated by CPI (used as a proxy of sugar production cost [US$]); RCVs are
sugar recovery rates during sugar refining process [%]; TREND is a trend variable that
represents technology advancement; and ee,, uut, vv,are error terms. Prices in the
previous year are used assuming that decision-making on sugar production relies on sugar
crop production, recognizing growers decide their production plan with the price realized
in the previous year. Retail refined sugar price is used for total sugar production
estimation due to lack of wholesale price data. Total sugar recovery rate is expressed as
average of recovery rate of cane sugar and beet sugar computed by weighing each
production onto each recovery rate. Autoregressive term (production in the previous year)
is added in the regression to calculate long-run elasticities. Elasticities associated with
price production cost ((/(/?, UU9, and (7(7/5) are expected to be negative and the others to
be positive. Price elasticities are expected to be inelastic. Previously reported own-price
elasticities for the U.S. sugar supply at industry level are 0.14 for cane sugar and 0.34 for
beet sugar by Petrolia and Kennedy (2002), using the U.S. wholesale refined beet sugar
price reported at midwest markets. Price elasticities for land allowed to sugar production
estimated by Lopez (1990) were 0.103 for cane and 0.246 for beets. Both authors
reported higher elactisities for beet sugar.


86
compared with a One-way ANOVA and mean separation was performed using Tukeys
Multiple Comparison test (Zar, 1999).
Lvsiphlebus testaceipes
To test the effects of methionine on the GBP, cotton plants (Gossypium sp.;
Family: Malvacae) were grown and maintained at the University of Florida, Department
of Entomology and Nematology green and shade houses from 07 October 2002 to 25
November 2002. Aphids (A. gossypii Glover) were supplied from other experiments
using this organism and kept on plants within a sealed greenhouse to prevent unwanted
parasitism. Plants were maintained in the sealed greenhouse, infested with aphids and
then placed in the open shadehouse area to encourage parasitation. In total, 20 plants
were used for 2 treatments, 1.0% L-methionine and 0% L-methionine (Control) mixed
with deionized H20. Plants were sprayed weekly (12 October 2002 through 17
November 2002) with approximately 10 ml of solution using a hand-held spray bottle.
Counts of parasitized aphids began approximately two weeks after placing plants outside
to ensure adequate time for parasitism (Royer et al. 2001). Counts were made using a
hand lens and counter; mummies with exit holes were enumerated and removed. A
few parasitized aphids were removed and held in glass vials to ensure correct
identification of the parasitoid.
Data Analysis
Data from the parasitoid experiments were analyzed using Minitab Version 12
(Minitab, Inc.; State College, PA). Control and experimental plants were compared
against one another with a One-way ANOVA and separation of significant means was
performed with Tukeys Multiple Comparison test (Zar, 1999).


LC50 (% L-methionine Concentration)
30
24h 48h 72h Overall (216h)
Figure 3-8. Concentrations (%) of L-methionine required for the mortality of 50%
of test population of tobacco homworm after 9 days exposure
(nTota)~L540; n=180 for 3.0% L-methionine 10.0% L-methionine,
n=200 for remainder). Number range following value is the 95%
confidence limits. Determination of LC50 was performed using
PROBIT Version 1.5 (Ecological Monitoring Research Division,


14
Pr = a Pn + (1 a) Pex (2.3)
where Pr is wholesale price per kilogram of standard sugar to be used as the reference for
cane payment during the harvest; a is expected portion of harvest to be consumed
nationally (a equals one if expected consumption is greater than expected production);
Pn is reference price for standard domestic sugar calculated by comparing the October-
September average price of the previous year with the current year; (1-a) is expected
surplus as a portion of production; and Pex is expected export price of sugar, which is
calculated by a weighed average of the U.S. price (Contract No. 14) and the world price
(Contract No. 11) with corresponding export quantities.
The Instituto Medical y Seguro Social (IMSS), the Mexican Social Security
Institute, provides both pension and medical services to all the employees in Mexico as
well as to small farmers who grow sugarcane (Greene, 1998). It is often the case that
IMSS clinics are located to the next to the mills (Greene, 1998). Borrel (1991) as well as
Buzzanell and Lord (1995) have pointed out this special relationship as a source of
inefficiency in the Mexican sugar industry.
Mexicos Sugar Consumption
Since Mexico is neither an importer of sugar nor producer of sugar beets, sugar
consumed in the country is derived solely from sugarcane grown domestically. The main
use of sugarcane derivatives are shown in Figure 2.5. Sugarcane requires two steps in the
refining process to obtain the refined sugar used by households and industries. Raw
sugar, which is the product of the extraction process, is either stored or exported to other
countries where refinery facilities are available. Mexico consumes two kinds of refined
sugar, called standard sugar and white sugar. Standard sugar has a slight impurity,


APPENDIX A
MAJOR EVENTS IN THE SUGAR INDUSTRY HISTORY IN MEXICO AND THE
U.S.
Mexico
United States
1910
The Mexican Revolution
1917
Farmers ownership of Ejido land
started (Article 27, Mexican
Constitution)
1972
HFCS-42 production started.
1977
HFCS-55 production started.
1980
Coca Cola and PepsiCo replaced 50%
of sugar use with HFCS-55.
1988
Government started privatizing sugar
mills and dismantling AZUCAR S.A.
1989
Crystalline fructose production started.
1990
A minimum import quota of 1.256
million MT (raw value) of sugar was
established in agreement with the
implementation of GATT (October).
1990 Farm bill
All major brands of soft drinks utilized
100% HFCS as the nutritive sweetener
ingredient.
1991
COAAZUCAR was formed
(amendments in Decreto Caero).
Sugarcane growers started to be paid
by sucrose content instead of by the
weight of sugar cane (54 % of the
wholesale price of standard sugar
based on KARBE system).
1992
Agrarian reform (Article 27) allowed
to sell/rent Ejido land.
1994
NAFTA agreement (January)
Devaluation (December)
Sugarcane growers started to be paid
57% of the wholesale price of
standard sugar based on KARBE
system.
123


Diets and Survivorship 24
Feeding and Development 31
Choice Tests 31
Discussion 36
4 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF
THE COLORADO POTATO BEETLE, Leptinotarsa decemlineata, UNDER
LABORATORY CONDITIONS 39
Introduction 39
Materials and Methods 40
Survivorship 40
Feeding and Development 41
Preference Tests 41
Data Analysis 42
Results 43
Survivorship 43
Feeding and Development 43
Preference Tests 47
Discussion 47
5 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE
YELLOW FEVER MOSQUITO, Aedes aegypti, UNDER LABORATORY
CONDITIONS 52
Introduction 52
Materials and Methods 53
Bioassay 53
Growth and Development 54
Data Analysis 56
Results 56
Bioassay 56
Growth and Development 59
Discussion 66
6 EVALUATION OF L-METHIONINE UNDER NATURAL FIELD CONDITIONS69
Introduction 69
Materials and Methods 70
Preliminary Investigation of Silwet L-77 and L-methionine 70
Plot Design 70
Fruit Yield 71
Pest Introduction 71
Data Analysis 74
vi


30
Figure 2-10. U.S. HFCS Export, 1992-2001
Source: USDA, 2002a
HFCS
- Consumption of sugar per capita
- Consumption of sweetener per capita
Figure 2-11. Consumption of Sugar and HFCS per Capita in Mexico
Source: Garcia Chaves et al., 2004


54
temperature (23C) to permit the amino acid to fully dissolve before the addition of the
larvae. An additional trial of L-methionine buffered with Tris to a pH of 7.0 using a
Fisher Scientific Accumet pH 900 was conducted to determine if mortality was attributed
to a change in pH or exposure to the L-methionine.
Larvae of YFM (third instar) were obtained from the mosquito colony maintained
at the Department of Entomology and Nematology, University of Florida. Larvae were
transferred to the treatment jars using a camel hair, with 10 larvae per replicate for a total
of 40 Iarvae/treatment and niotai=240 for each amino acid bioassay experiment (Figure
5-1). Approximately 0.5g of finely ground fish food was added to serve as a larval food
source and nylon window screen was used to cover the tops of the jar to prevent the
escape of any emerged adults. Jars were held at 23C on a dedicated laboratory bench
top for approximately one week. The numbers of larvae surviving were recorded each
day.
Growth and Development
This experiment used the same Materials and Methods as the bioassay portion
with the exception of neonate larvae instead of 3rd instars. Eggs were placed in a tray of
water and held at 23 C for 2 days after eclosin. Neonates were removed using a camel
hair paintbrush and placed into each jar, with 10 larvae per replicate for a total of 40
larvae/treatment (nTOtai~240). Larval exuviae or dead larvae were removed and used to
examine growth rates by measuring the head capsules. Larvae head capsule widths were
measured (using an Olympus Tokyo Model 213598 stereomicroscope with an ocular
micrometer) as an evaluation of larval development.


117
Mexicos Export Potential
The opportunity for Mexico to export sugar to reach the 250,000 MT expanded
quota (which would be equivalent to approximately 20 percent of the U.S. minimum
import requirement) seems unlikely to be enjoyed due to Mexicos restricted production
compared to its sweetener consumption. Yet, Mexico possesses a large potential to export
over-quota as well as quota-free when all restrictions are lifted. The magnitude of over
quota and quota-free export depends on the expansion rate of Mexicos sugar production
and HFCS adoption. Specifically, if Mexico adopts HFCS at an increasing rate, a
considerable amount of surplus sugar destined to export will be generated, which
consequently poses a direct impact on the Mexican sugar industry and an extended
impact on the U.S. market.
The Impact of Changes in Mexican Market Situation
Mexicos sugar production, HFCS adoption, and a tax on HFCS are examined as
factors that change the Mexican sweetener market situation. The impact from these three
factors on Mexico is eminent. The Mexican sugar industry and welfare become better off
from higher sugar production which can be achieved through reduction of production
costs and mill downtime; however, this positive impact dissipates once HFCS is adopted
at higher rates. The negative impact from adopting HFCS on Mexico originates with the
imbalance between domestic sugar production and sweetener consumption, thus making
Mexico unable to attain a net sweetener surplus producer. Mexico then faces a large
sugar surplus that cannot be sold anywhere but the U.S. market with tariff (over-quota) or
the world market. Simulations showed that the sugar industry can tolerate a higher rate of
HFCS adoption if the gains from high production exceed the losses from adopting HFCS,
whereas the nations welfare cannot. This implies that the Mexican government faces


49
consumption into: sugar consumed by households, referred to as direct consumption;
and sugar consumed by bulk sugar users, referred to as indirect consumption. Only
indirect consumption of sugar is expected to be influenced by HFCS consumption as they
are the only consumers of HFCS. In addition to estimating sugar demand, it would have
been preferable to directly estimate HFCS consumption, but this was precluded because
of the relatively short time series available. Instead, total consumption of sweeteners (the
sum of sugar and HFCS consumption) is analyzed by regressing on sugar price and other
variables. A dummy variable that represents the availability of HFCS in the market is
also added to the regression forms of indirect consumption of sugar and total
consumption of sweeteners where substitution between sugar and HFCS occurs in the
market.
In(QDCsuGAR, t) = Mi + M2*In(Psugar, i) + M¡*ln(GDP,) + M4*In(POP,)+ u,
(4.2)
In(Q!Csugar, ,) = Ms + M6*In(Psugar, ,) + M7*In(GDPt) + M8*In(POP,)
+ U9*DHFCS + v, (4.3)
In(Q,Csugar, i) = M¡o + Mu*In(PDsugar, t) + Mi2*In(GDPt) + Mi3*In(POPt)
+ U,4*DHFCS + w, (4.4)
where QDC is direct consumption of sugar [MT], QIC is indirect consumption of sugar
[MT], Qr( is total consumption of sweeteners [MT], P is real retail price of standard
sugar (deflated by CPI [pesos / Kg]), GDP is real per capita GDP (deflated by CPI
[pesos]), POP is population, DHFCS is dummy variable for availability of high fructose
com syrup (HFCS) in the sweetener market (DHFCS= 1 after t=1992), and u,,vt,wtare
error terms. It is expected that the elasticities associated with price (M2, and M¡¡) and


10
The total number of sugarcane growers is reported as approximately 158,000
(COAAZICAR, 2003b), which is equivalent to roughly 2 percent of the total labor force
in the agricultural sector (INEGI, 2003). If related workers such as sugarcane cutters,
cane-transport employees, factory workers and administrative, and technical and
management personnel are included, total employment in the sugar sector exceeds
1,000,000 (Garcia Chaves et al., 2002) and accounts for more than 14 percent of
agricultural labor.
Land area per grower ranges from less than 1 ha, which accounts for 3.6 percent of
the total sugarcane area, to over 15 ha, which accounts for 17.5 percent of the land,
averaging 3.9 ha per grower (COAAZUCAR, 2003b). When the number of growers is
allotted to each land size category, a skewed distribution is revealed along the land scale
spectrum with many small-land holders and a few large-land holders (Figure 2-1). A
large number of small-scale sugarcane growers were created as a result of the Mexican
revolution and the sugar program that evolved after the revolution: the communal land
(ejido) has been divided and distributed among farmers since the revolution and the
Mexican sugar program offers social security and medical services to each grower-
proving to be a large incentive for farmers to grow sugarcane.
Mexicos Sugar Production
There are 60 operating sugar mills located across 15 states in the nation (Figure 2-
2). Sugar mills are responsible not only for milling sugarcane, but also supervising
sugarcane cultivation and organizing the harvest. This includes inspecting and advising
on cultivation, scheduling harvest dates, pooling and arranging laborers, and providing
trucks and drivers for the harvest.


119
flexible manner; if not Mexicos export will be dampened because of its comparative
disadvantage to the rest of the world. For the United States, accommodating Mexican
sugar can serve as an alternative form of price support because of the higher price of
Mexican sugar compared to that from the rest of the world; otherwise the U.S. price will
likely dip below the U.S. support price level caused by a large quantity of sugar from the
rest of the world flowing into the U.S. market at a lower price. In this sense, sugar trade
agreements were mutually beneficial for both the United States and Mexico.
Two alternative price stabilization policies are examined in the study: buying up
excess sugar in the market and production controls. Simulation results show some
improvement in both U.S. sugar industry revenues and cost-adjusted U.S. welfare as a
result of reduction in sugar program costs as well as an increase in tariff revenue from
Mexican sugar, yet overall positive effects are minimal. It is noteworthy, however, that
maintaining the price support may become extremely costly when combined with the
situation in where the U.S. government promises to accept a large amount of sugar from
the rest of the world. If the U.S. government switched policies from the price support to
buying up excess sugar, the timing to do so would be important so as to minimize the cost
incurred by the government: the cost of buying up excess sugar will rise immediately
after policies are switched whereas the cost of the price support will not because the U.S.
sugar price will be maintained relatively high in the early stage of the forecast horizon.
When buying up excess sugar policy and production control policy are compared,
losses in sugar industry revenues in both countries and in Mexicos welfare are larger for
the latter policy, given the similar magnitude of gains in U.S. welfare. In general, these
alternative policies still would be more effective than any price-led sugar policy because


Results 74
Effects of L-methionine and Silwett L-77 on Colorado Potato Beetle Adults
Under Laboratory Conditions 74
Effects of L-methionine and Silwett L-77 on yield 74
Survival of CPB larvae 74
Discussion 78
7 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE
NON-TARGET SPECIES 82
Introduction 82
Materials and Methods 84
Coleomegilla maculata 84
Neochetina eichhorniae 85
Lysiphlebus testaceipes 86
Data Analysis 86
Results 87
Coleomegilla maculata 87
Neochetina eichhorniae 87
Lysiphlebus testaceipes 87
Discussion 87
8 SUMMARY AND DISCUSSION 96
LIST OF REFERENCES 102
BIOGRAPHICAL SKETCH 114
Vll


65
P
Q
Mexican Market
P
Mexico-U.S. Market
P
Figure 4-1. Image of Model Calibration


18
(Congressional Research Service, 1999). Due to its liquid form, HFCS is considered as a
close substitute of sugar, though not a perfect one. Crystalline fructose, fine white
crystals of pure fructose is slightly sweeter than sugar and is another potential substitute
for sugar; however, crystalline fructose is more expensive than sugar and behaves
differently from sugar in most baking and other manufactured food uses thus limiting its
use as a sugar substitute (USDA, 1997).
HFCS is predominantly produced in the United States, which accounted for 74
percent of the world HFCS production in 2001(Buzzanel, 2002). HFCS is produced in
wet-milling facilities located in com growing regions in the U.S. HFCS producers outside
the United States and their recent production levels are: Japan (766,000 MT), Canada
(400,000 MT), Argentina (312,000 MT, estimated), Mexico (291,000 MT, estimated),
and European Union (293,000 MT) (Buzzanel, 2002). In the United States, most HFCS is
supplied and consumed domestically and only a small fraction is exported to Mexico and
Canada, the NAFTA member economies (Figure 2-8 and 2-9). The difference in trade
between these two countries is that Canada and the United States exchange a similar
amount of HFCS across the border while Mexico is a net buyer. Although the quantity
exported to Mexico, 122,800 MT in 2001, accounts for only 1.5 percent of total HFCS
demanded in the United States, the quantity accounts for 60 percent of total HFCS export
from the United States (Figure 2-10). This amount is equivalent to about 40 percent of
Mexicos domestic production ability (291,000 MT, Buzzanell, 2002).
The introduction of HFCS into the Mexican sugar market brought about significant
changes in the sugar consumption pattern in Mexico. Although HFCS is traded at a
higher price than sugar in Mexico, it has been gaining an increasing share of the Mexican


23
straightforward. According to the analysis by Moss and Schmitz (2002), the relationship
between sugar and HFCS prices has changed over time: HFCS price responded to
wholesale sugar price from 1983 to 1996, but not from 1997 to 2001. Another analysis by
Evans and Davis (2002) indicates that the estimated cross price elasticity of HFCS with
respect to sugar demand was found to be insignificant. This implies that the HFCS price
is set below the sugar price in order to attract bulk sweetener users but its behavior
remains ambiguous and arbitrary. Furthermore, other research reported that HFCS price
is not correlated with the price of com (Offenbach, 1995), but others found that the HFCS
price responded to the price of com from 1983 to 1996 but not from 1997 to 2001 (Moss
and Schmitz, 2002). Given the insignificant relationship between com price and HFCS
supply mentioned above, the indirect impact of changes in com production on HFCS
price would likely be small; however, any drastic changes in com program or large
changes in com export may eventually affect HFCS prices. Com statistics such as recent
com production for selected countries, industrial use of com in the U.S., and the com
price in the U.S. are illustrated in Appendix B.


Mean Head Capsule Width (mm)
33
(Errer Bars @ 95%; ^(o.os)7,i 52=2.37, F=18.2; P <0.001)
6
5
4
3
2
1
0
Control 0.1% 0.3% 0.5% 0.7% 1.0% Proline Btk
Figure 3-10. Mean head capsule widths of tobacco homworm larvae exposed
to excised eggplant leaves treated with various concentrations of
L-methionine (nrota]=320). Proline (1.0%) and Btk were included
for comparison as positive and negative controls. Error bars
denote 2 SE. Bars within treatments having the same letter are
not statistically different (Tukeys MST, P0.001).


LIST OF REFERENCES
Abbott, W.S. 1925. A method for computing the effectiveness of an insecticide. J.
Econ. Entomol. 18:265-267.
Aerts, M.J. and O.N. Neishiem. 1999. Florida Crop/Pest Management Profiles:
Tomatoes. CIR 1238. Pesticide Information Office. Food Science and Human
Nutrition Department, Florida Cooperative Extension Service, Institute of Food
and Agricultural Science, University of Florida. Internet URL:
http://www.edis.ifas.ufl.edu/ BODY_PI039.htm. Accessed April 2004.
Anand, R. and M. Anand. 1990. Nutritive effect of the D isomers of the essential amino
acids in casein diet on Dacus cucurbitae (Coquillett) maggots. Indian J. Entomol.
52(4): 525-528.
Andow, D.A., and S.J. Risch. 1985. Predation in diversified agroecosystems: Relations
between a coccinellid predator Coleomegilla maculata and its food. J. Appl.
Ecol. 22: 57-372.
Audsley, N., R.J. Weaver and J.P. Edwards. 1999. Juvenile hormone synthesis by
corpora allata of tomato moth, Lacanobia olercea (Lepidoptera: Noctuidae), and
the effects of allatostatins and allatotropin in vitro. Eur. J. Entomol. 96: 287-293.
Barfield, C.S. and M.E. Swisher. 1994. Integrated pest management: ready for export?
Historical context and internationalization of IPM. Food Reviews Intemat. 10(2):
215-267.
Baumhover, A.H., W.W. Cntelo, J.M. Hobgod, Jr., C.M. Knott, and J.J. Lam, Jr. 1977.
An improved method for mass rearing the tobacco horn worm. Agricultural
Research Service, Unites States Department of Agriculture ARS-S-167,13 pp.
Beck, S.D. and W. Hnec. 1958. Effects of amino acids on feeding behavior of the
European com borer, Pyraustra nubilialis (Hibn.). J. Insect Physiol. 2:85-96.
Bell, E.A. 1978. Toxins in seeds, pp. 143-161. IN J. Harboume (ed.), Biochemical
Aspects of Plant and Animal Coevolution. Academic Press, New York. 435pp.
Berge, M.A., G.A. Rosenthal and D.L. Dahlman. 1986. Tobacco budworm, Heliothis
virescens (Noctuidae) resistance to L-canavanine, a protective allelochemical.
Pest. Biochem and Physiol. 25: 319-326.
102


63
cents per pound) and the U.S. equilibrium price with the production quantity guaranteed
at the loan rate. This cost is captured by comparing between scenarios with and without
the price support, ceteris paribus. The cost of buying up excess sugar is calculated by
multiplying the U.S. net sugar import, i.e. total sugar import less 1,256,000 MT of
minimum import requirement with the U.S. equilibrium price. In the case of the policy to
buy excess sugar policy, sugar storage costs incurred by the government are ignored.
Note that since the values were converted in terms of U.S. dollars prior to simulations,
the exchange rate realized in the base year (2001) was implicitly used for calculating pay
offs.
All the games proposed are solved through a two-players pay-off matrix (two-
dimensions) with the two governments are the decision-makers. When coalitions are
formed, their individual pay-offs are pooled, assuming that the total pay-off is
redistributed among them (Morris, 1994). In doing so, it is also assumed that industry
revenue and each nations welfare can be added together. After pay-offs are calculated
for each party or coalition, these values are indexed as a relative gain or loss to the pay
offs in baseline scenario to facilitate the decision process. Mixed-strategy games are first
simplified by eliminating dominated strategies (strategies that played with zero
probability) and then solved through maximizing the expected pay-off to each player
from the game (Morris, 1994; Varan, 1992).
Sources of Data
Data for the Mexican sugar industry was obtained from the website of Comit de la
Agroindustria Azucarera (COAAZUCAR, Sugar Agro-Industry Committee). The
committee is in charge of monitoring sugar cane and sugar production at each mill as
well as determining the cane price paid to farmers in the country. The latter task was


58
Days of Exposure
Figure 5-3. Mortality of yellow fever mosquito larvae exposed to various
concentrations of D-methionine (nTOtai=240). Data were
adjusted using Abbotts formula for control mortality.


4
all risk of insect damage by using more applications and stronger pesticides (Schuster et
al. 1996).
Problems Associated with Pesticide Misuse
The use of pesticides is not completely ruled out under IPM strategies, but rather
IPM encourages responsible use to minimize environmental harm and to protect human
safety and health (Deedat, 1994). However, the misuse (both intentional, in terms of
more is better; and unintentional, as in agricultural runoff) also has resulted in
resistance in some of the target pests. For example, surveys in North Carolina have
shown that the Colorado potato beetle has become resistance to fenvalerate, carbofuran,
and azinphosmethyl as a result of control failures in the field (Heim et al. 1990).
Resistance to insecticides has also been observed in more than 450 arthropod pests
(Romoser and Stoffolano 1998). Bills et al. (2004) found a 38% increase in the number
of registered compounds used as pesticides from 1989-2000, and also a 16% increase in
pesticide resistance of arthropod species worldwide.
Losses are not limited to agricultural systems alone. Across Africa for example,
populations of insecticide-resistant mosquitoes are the result of a variety of mechanisms,
including exposure to pesticide residues in agricultural runoff, mutation of target sites,
and migration of resistant populations into areas where there were no previous problem
(FIC-NIH 2003). Parts of southwest Asia have seen a resurgence of malaria in some
areas where it was considered eradicated (due to a combination of resistance and the
economics associated with control of mosquito vectors) (Deedat 1994). The importance
of this example becomes even more relevant when one considers that one million
individuals die every year as a result of malaria, with upwards of 500 million cases per
year (Centers for Disease Control 2003). The existence of other mosquito-bome diseases


50
HFCS availability {Mg and Mm) to be negative and the others to be positive. Price
elasticities were expected to be inelastic. Serial correlation is anticipated and corrected by
the Yule-Walker Method with appropriate lags assigned.
U.S. Sweetener Supply Model
Regression analysis is used to estimate the U.S. sweetener supply utilizing time-
series data. Aggregate sugar supply is analyzed in three parts, i.e. total sugar supply,
sugar supply from sugarcane and sugar supply from sugar beets. Sugarcane and sugar
beet production are estimated separately as they have different production regions and a
different refining process. Sugar production is used as the dependent variable. It is
assumed that domestic sugar production is the primary source of sugar supply and
carried-over stock from the previous periods is considered constant over the estimated
time span. The model is specified as a double-log (natural) form and estimated
coefficients can be read as elasticities associated with each variable.
In(QStotal sugar, i) = UU ¡ + UU2*In(P dtotal sugar, t-i) + UU3*In(COST,)
+ U U4*Ih(RCVtoTAL SUGAR, t) + UU5*In(QSTOTALSUGAR,t-l)
+ UU6*TREND t + ee, (4.5)
In(QSCANE SUGAR, t) = UU7+ UU8*ln(P dCANE SUGAR, t-l) + UUg*ln(COSTt)
+ UU ¡o*ln(RCVcANE sugar, t) + UUu*ln(Q Scane sugar, t-i)
+ UU,2*TREND, + uu, (4.6)
I>l(QSBEET SUGAR, t) = UU¡3 + UU]4*Itl(P ^BEET SUGAR, t-l) + UU]5*Ill(COST,)
+ UUi6*Iti(RCVbeetsugar, t) + UUi7*In(Q SBEET SUGAR, t-l)
+ UUi8*TREND, + vv, (4.7)
where Q represents the total sugar quantity produced, sugar quantity produced from
sugarcane, and sugar quantity produced from sugar beets [1000 short tons] in the three



PAGE 1

EVALUATION OF THE AMINO ACID METHIONINE FOR BIORATIONAL CONTROL OF SELECTED INSECT PESTS OF ECONOMIC AND MEDICAL IMPORTANCE By LEWIS SCOTTY LONG A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2004

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Copyright 2004 by Lewis Scotty Long

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ACKNOWLEDGMENTS I thank Jim Cuda and Bruce Stevens for giving me the financial and intellectual freedom that made this work possible. I want to thank Jim for housing me in his lab and providing the facilities to perform this work, and Bruce for allowing me to take his initial work and elaborate on it as well as including me as a co-inventor of the research presented. Most of all, I would like to express my sincere appreciation to Judy Gillmore. Without her support and help this research would not have been completed. Judy was integral in every aspect of this endeavor and put up with more than her fair share of my "research". I extend heartfelt thanks to George Gerencser, James Maruniak, Simon Yu, and Susan Webb for serving as members of my supervisory committee. I would like to also thank Jim Lloyd, Jerry Butler, and Carl Barfield for all the experiences and knowledge shared. Finally, I want to express my deepest, eternal gratitude to my fellow graduate students Jim Dunford and Heather Smith, for providing support and guidance that only colleagues, intellectual equals, and close friends can give. I can only hope to repay them for their help by providing the same amount of support for their endeavors as they did mine. iii

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I dedicate this work to Karen, my wife and best friend. I thank her for putting up with living as a "graduate student" for the last 5 years in fulfillment of my childhood dream of being a "Doctor". She has been my pillar of support, and I would not have made it this far without her love and understanding.

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TABLE OF CONTENTS page ACKNOWLEDGMENTS iii LIST OF FIGURES viii ABSTRACT xi CHAPTERS 1 THE INTEGRATED PEST MANAGEMENT DILEMMA: ARE CONVENTIONAL PESTICIDES THE ONLY ANSWER? 1 Introduction 1 Importance of IPM in Florida and Surrounding States 2 Problems Associated with Pesticide Misuse 4 Biorational CompoundsAn Alternative to Chemical Pesticides 5 2 HISTORY OF THE USE OF AMINO ACIDS AS A MEANS TO CONTROL INSECT PESTS 7 Non-Protein Amino Acids 7 Essential Amino Acids 10 The Cation-Anion Modulated Amino Acid Transporter with Channel Properties (CAATCH1 ) System 9 Methionine 13 Research Objectives 16 3 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE TOBACCO HORNWORM, Manduca sexta, UNDER LABORATORY CONDITIONS 17 Introduction 17 Materials and Methods 18 Diets and Survivorship 18 Feeding and Development 20 Preference Tests 22 Data Analysis 24 Results 24 v

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Diets and Survivorship 24 Feeding and Development 31 Choice Tests 31 Discussion 36 4 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE COLORADO POTATO BEETLE, Leptinotarsa decemlineata, UNDER LABORATORY CONDITIONS 39 Introduction 39 Materials and Methods 40 Survivorship 40 Feeding and Development 41 Preference Tests 41 Data Analysis 42 Results 43 Survivorship 43 Feeding and Development 43 Preference Tests 47 Discussion 47 5 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE YELLOW FEVER MOSQUITO, Aedes aegypti, UNDER LABORATORY CONDITIONS 52 Introduction 52 Materials and Methods 53 Bioassay 53 Growth and Development 54 Data Analysis 56 Results 56 Bioassay 56 Growth and Development 59 Discussion 66 6 EVALUATION OF L-METHIONINE UNDER NATURAL FIELD CONDITIONS69 Introduction 69 Materials and Methods 70 Preliminary Investigation of Silwet L-77 and L-methionine 70 Plot Design 70 Fruit Yield 71 Pest Introduction 71 Data Analysis 74 vi

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Results 74 Effects of L-methionine and Silwett L-77 on Colorado Potato Beetle Adults Under Laboratory Conditions 74 Effects of L-methionine and Silwett L-77 on yield 74 Survival of CPB larvae 74 Discussion 78 7 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE NON-TARGET SPECIES 82 Introduction 82 Materials and Methods 84 Coleomegilla maculata 84 Neochetina eichhorniae 85 Lysiphlebus testaceipes 86 Data Analysis 86 Results 87 Coleomegilla maculata 87 Neochetina eichhorniae 87 Lysiphlebus testaceipes 87 Discussion 87 8 SUMMARY AND DISCUSSION 96 LIST OF REFERENCES 102 BIOGRAPHICAL SKETCH 114 vii

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LIST OF FIGURES Figure Page 21. The CAATCH1 system identified from the midgut of the tobacco hornworm 15 31. Rearing chamber for tobacco hornworm and Colorado potato beetle larvae used in the artificial and excised leaf diet tests 19 3-2. Setup for whole plant studies involving tobacco hornworm 21 3-3. Chambers used for tobacco hornworm and Colorado potato beetle preference tests 23 3-4. Amount of L-methionine present on leaf surface after treatment 25 3-5. Mortality of tobacco hornworm larvae exposed to various concentrations of L-methionine (nTotar^O) in artificial diet 26 3-6. Survivorship of THW larvae exposed to various concentrations of L-methionine (nTotai = 1,540) on excised eggplant leaves 28 3-7. Mortality of tobacco hornworm larvae exposed to various concentrations of L-methionine (nrotai=256) on whole plants 29 3-8. Concentrations (%) of L-methionine required for the mortality of 50% of test population of tobacco hornworm after 9 days exposure (nrotai^ 1,540; n=180 for 3.0% L-methionine 10.0% L-methionine, n=200 for remainder) 30 3-9. Mortality of tobacco hornworm larvae exposed to various concentrations of Lmethionine (nr 0 tai = 160) on excised eggplant leaves for feeding and development trials 32 3-10. Mean head capsule widths of tobacco hornworm larvae exposed to excised eggplant leaves treated with various concentrations of L-methionine (n To tai=320) 33 3-11. Total leaf area consumed by tobacco hornworm larvae exposed to excised eggplant leaves treated with various concentrations of L-methionine (nT O tai=320) 34 312. Mean leaf consumption by tobacco hornworm in the preference tests 35 41. Mortality of Colorado potato beetle larvae exposed to excised eggplant leaves treated with various concentrations of L-methionine (n To tai=560) 44 4-2. Concentrations (%) of L-methionine concentrations required for the mortality of 50% of the test population of Colorado potato beetle after 8 days exposure (n To tai=220) 45 vin

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4-3. Mean head capsule widths of Colorado potato beetle larvae exposed to excised eggplant leaves treated with various concentrations of L-methionine (n To tai = 320) 46 4-4. Total leaf area consumed by Colorado potato beetle larvae exposed to excised eggplant leaves treated with various concentrations of L-methionine (moai=320) 48 45. Mean leaf consumption by Colorado potato beetle in the preference tests 49 51. Bioassay setup for yellow fever mosquito larvae exposed to various concentrations of amino acids and Bti 55 5-2. Mortality of yellow fever mosquito larvae exposed to various concentrations of L-methionine (nr o tai = 240) 57 5-3. Mortality of yellow fever mosquito larvae exposed to various concentrations of D-methionine (nTotai = 240) 58 5-4. Mortality of yellow fever mosquito larvae exposed to various concentrations of Trisbuffered L-methionine (nT O taf = 240) 60 5-5. Mortality of YFM larvae exposed to various concentrations of Proline (n T otai = 240) 61 5-6. Mortality of yellow fever mosquito larvae exposed to various concentrations of L-leucine (nTotai=240) 62 5-7. Mortality of YFM larvae exposed to various concentrations of Beta-alanine (n To tai=240) 63 5-8. Mean head capsule widths of yellow fever mosquito larvae exposed to various Tris buffered (7.0 pH) concentrations of L-methionine (n To tai = 320) 64 59. Concentrations (%) resulting in 50% mortality (LC 50 ) of yellow fever mosquito larvae exposed to various amino acids after 10 days (nxotaf^O for each amino acid) 65 61. Overview of the design layout used to study the effects of L-methionine and Silwett L-77 solutions on yield of eggplant 72 6-2. Weed Systems, Inc. KQ 3L C0 2 backpack back sprayer used for application of L-methionine and Silwett L-77 solutions 73 6-3. Mortality of Colorado potato beetle adults exposed to excised eggplant leaves treated with L-methionine and the adjuvant Silwett L-77 (n To tai = 120) 75 6-4. Effects of L-methionine and Silwett L-77 on eggplant yield (A) and mean weight in grams of fruit (B) from 09 June to 3 1 August 2001 76 ix

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65. Mortality of Colorado potato beetle larvae on eggplants treated with L-methionine and Silwett L-77 87 71. Mortality of Coleomegilla maculata adults after exposure to L-methionine treated artificial diet 88 7-2. Mortality of Coleomegilla maculata adults after exposure to L-methionine treated cotton plant leaves infested with aphids 89 7-3. Feeding scars on water hyacinth (Eichhornia crassipes) leaf after exposure to Neochetina eichhorniae adults 90 7-4. Mortality of Neochetina eichhorniae on treated water hyacinth leaves 91 7-5. Feeding rate of Neochetina eichhorniae on water hyacinth leaves treated with L-methionine and Proline 92 7-6. Lysephlebius testiceipes parasitized aphids on cotton plants treated with L-methionine 93 x

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Abstract of Dissertation Presented to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy EVALUATION OF THE AMINO ACID METHIONINE FOR BIORATIONAL CONTROL OF SELECTED INSECT PESTS OF ECONOMIC AND MEDICAL IMPORTANCE By Lewis Scotty Long May, 2004 Chair: James P. Cuda Cochair: Bruce R. Stevens Major Department: Department of Entomology and Nematology Integrated pest management (IPM) strategies were developed in an effort to control pests with fewer pesticides. However, because of the misuse of pesticides and the failure to adopt IPM practices pesticide use is higher than ever. An alternative to conventional broad-spectrum pesticides is the use of biorational compounds; those that pose minimal risk to the environment and are specific to the target pests. The recent discovery of the CAATCH1 system in the midgut of the tobacco hornworm (THW), Manduca sexta, has revealed a novel means to control certain insect pests. This membrane protein works in alkaline conditions as both an amino acid transporter and also independently as a cation channel. However, the amino acid L-merhionine blocks amino acid transport thus altering the ion flow. xi

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Bioassays involving the tobacco hornworm, Colorado potato beetle (CPB), Leptinotarsa decemlineata, and the yellow fever mosquito (YFM), Aedes aegypti, were conducted to determine the insecticidal properties of this compound. L-methionine in artificial and natural diets resulted in the mortality of 50 to 100% in concentrations of 0.3% and higher for THW and CPB. Feeding rates and larval development also were affected with reduced levels (>0.1%) of L-methionine. Bioassay trials involving YFM larvae were similar, concentrations greater than 0.1% L-methionine produced mortality rates of 70 to 100%. All three species responded better to higher concentrations of Lmethionine than to Bacillus thuringiensis, the most commonly used and commercially available biorational pesticide. Field trials and non-target tests also were performed to determine L-methionine effectiveness under natural settings and safety to other organisms. Eggplant yield was not reduced by the application of L-methionine, which effectively controlled CPB larvae on the plants. Furthermore, several beneficial insects that were tested (a predator, a herbivore, and a parasitoid) were not affected by the addition of L-methionine to their diets. Based on these results, L-methionine was found to be effective in controlling selective agriculturally and medically important insect pest species, yet posed little threat to the crop plants applied to or to non-target organisms. The use of L-methionine as a pesticide, its relationship with insects and its possible use in delaying insecticide resistance were also examined. xu

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CHAPTER 1 THE INTEGRATED PEST MANAGEMENT DILEMMA: ARE CONVENTIONAL PESTICIDES THE ONLY ANSWER? Introduction Integrated Pest Management (IPM), the sustainable approach to the management of pest species using a combination of biological, chemical and cultural methods to reduce economic, environmental, and public health risk, was a result of economic losses associated with years of overuse of chemical control leading to resistance problems. The use of IPM strategies have certainly decreased pesticide usage and encouraged the use of methods that ensure a safer environment but many feel that it is not enough. After three decades of research efforts in the United States, IPM as it was envisioned in the 1970s was practiced on less than 8% of U.S. crop acreage based on Consumers Union estimates — well short of the national commitment to implement IPM on 75% of the total U.S. acreage by the end of the 1990s (Ehler and Bottrell 2000). This means that farm practices have changed little since the national IPM initiative was established in 1994 to implement biologically based alternatives to pesticides for controlling arthropod pests. It should be noted that the low percentage of IPM practices on commercial U.S. farmland may possibly be related to the lack of sufficient reporting means and actually may be higher than believed when the local growers and homeowners are included. However, the United States is considered the worlds' largest user of chemical pesticides, accounting for nearly 50% of total worldwide production and shows no sign of slowing (Deedat 1994). Pesticides remain the primary tool of pest consultants and farmers, because of the lack of economic incentives to adopt alternative strategies that require more effort to 1

PAGE 14

implement, produce unpredictable results, and require new knowledge (Barfield and Swisher 1994; Ehler and Bottrell 2000). Importance of IPM in Florida and Surrounding States Considerable effort has been devoted to developing IPM programs in Florida because of its unique pest problems and crop production systems, sensitivity to chemical pollutants, and increased urbanization (Capinera et al. 1994; Rosen et al. 1996). The necessity for developing IPM protocols for Florida's major plant and animal pests was underscored in a new statewide initiative. In November 1999, the Institute of Food and Agricultural Sciences (IF AS) at the University of Florida launched Putting Florida FIRST — Focusing IF AS Resources on Solutions for Tomorrow (Florida FIRST 1999). The Florida FIRST initiative was created (with input from stakeholders) to define the role of IF AS in shaping Florida's future in the 21 st century. Increasing concerns (expressed repeatedly by Florida's scientific community and the general public) about environmental contamination, food safety issues, and human and animal health problems resulting from the indiscriminate use (and often misuse) of pesticides are making existing methods for pest management obsolete. Successful implementation of "true" IPM, as it was envisioned by those who envisioned the original concept, will have the added benefit of helping Florida ". . enhance natural resources, provide consumers with a wide variety of safe and affordable foods, . provide enhanced environments for homes, work places and vacations, maintain a sustainable food and fiber system, and improve the quality of life. ." (Florida FIRST 1999). This effort to promote IPM programs in the state of Florida also benefits the surrounding states. For example, solanaceous crops produced in the southeastern U.S. (such as tomato, tobacco, eggplant, peppers and potato) are subjected to the same

PAGE 15

3 defoliation and fruit damage from various lepidopteran and coleopteran pests that also threaten Florida. The tomato pinworm [Keiferia lycopersicella (Walshingham) (Lepidoptera: Gelechidae)], armyworms [Spodoptera spp. (Lepidoptera: Noctuidae)], the Colorado potato beetle [Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae)], and hornworms [Manduca spp. (Lepidoptera: Sphingidae)] are some examples of pests that threaten both conventional producers and homeowners alike. For example, the estimated loss from and the cost of control of the tobacco hornworm, the number-one pest in tobacco crops in Georgia, reached $1 .5 (and $2.3 million), respectively, for the years 1996-1997 (Jones and McPherson 1997). From 1992-1998, tomato, eggplant, and pepper producing areas in the Southeast had a total of 1 ,247,000 pounds of endosulfan applied over 270,000 acres (Aerts and Neshiem 1999; Neshiem and Vulinec 2001). The cost of insecticides applied in Florida tomato production alone for 1993-1994 amounted to approximately $l,052/hectare for a total of $2.1 million; and rose to $2550/acre, totaling $103M for the 1996-1997 season (Aerts and Neshiem 1999; Schuster et al. 1996). The use of pesticides in Florida tomato production is high because tomatoes account for 30% of the total vegetable-crop value and 13% of the total vegetable acreage for the state, with 99% of production aimed toward the fresh market (Schuster et al. 1 996). For Florida potato producers, the cost of applying pesticides from 1 9951 996 was $1 1 .5M, and 96% of total Florida eggplant-crop acreage was treated with chemical insecticides (mainly methomyl and endosulfan) (Neshiem and Vulinec 2001). In addition to the monetary cost of pesticide use, commonly used insecticides such as endosulfan and fenvalerate show a high degree of toxicity to parasitoids of the tomato pinworm, thus negating the benefits of predation by natural enemies (Schuster et al. 1996). These figures may be the result of the "more is better" attitude of producers who want to avoid

PAGE 16

all risk of insect damage by using more applications and stronger pesticides (Schuster et al. 1996). Problems Associated with Pesticide Misuse The use of pesticides is not completely ruled out under IPM strategies, but rather IPM encourages responsible use to minimize environmental harm and to protect human safety and health (Deedat, 1994). However, the misuse (both intentional, in terms of "more is better;" and unintentional, as in agricultural runoff) also has resulted in resistance in some of the target pests. For example, surveys in North Carolina have shown that the Colorado potato beetle has become resistance to fenvalerate, carbofuran, and azinphosmethyl as a result of control failures in the field (Heim et al. 1990). Resistance to insecticides has also been observed in more than 450 arthropod pests (Romoser and Stoffolano 1998). Bills et al. (2004) found a 38% increase in the number of registered compounds used as pesticides from 1989-2000, and also a 16% increase in pesticide resistance of arthropod species worldwide. Losses are not limited to agricultural systems alone. Across Africa for example, populations of insecticide-resistant mosquitoes are the result of a variety of mechanisms, including exposure to pesticide residues in agricultural runoff, mutation of target sites, and migration of resistant populations into areas where there were no previous problem (FIC-NIH 2003). Parts of southwest Asia have seen a resurgence of malaria in some areas where it was considered eradicated (due to a combination of resistance and the economics associated with control of mosquito vectors) (Deedat 1994). The importance of this example becomes even more relevant when one considers that one million individuals die every year as a result of malaria, with upwards of 500 million cases per year (Centers for Disease Control 2003). The existence of other mosquito-borne diseases

PAGE 17

5 such as Dengue fever, yellow fever, and West Nile virus to name just a few, put countless millions more at risk. It would be dangerous to think that these diseases only occur in underdeveloped countries and not the United States. Integrated Pest Management practices also should be adopted for controlling the medical and veterinarian important insect vectors of these and other diseases. Biorational Compounds: An Alternative to Traditional Chemical Insecticides One way to reduce this reliance on traditional chemical pesticides and delay resistance is by increasing the variety and use biorational compounds. Biorational compounds are effective against selected pest species but are innocuous to nontarget or beneficial organisms; and have limited affect (if any) on biological control agents (Stansly et al. 1996). Biorational compounds include detergents, oils, pheromones, botanical products, microbes, and systemic and insect growth regulators (Perfect 1992; Wienzierl et al. 1998). Their safety lies in the low toxicity of the compound to nontarget organisms and the compound's short residual activity in the field. For example, Bacillus thuringiensis isrealensis (Bti) currently is one of the most widely used microbial pesticides for controlling aquatic dipteran pests (i.e., mosquitoes and black flies) because of its selectivity to this group and minimal nontarget effects (Glare and O'Callaghan 1998). However, resistance to Bt products has occurred in many species of lepidoptera from overuse of Bacillus thuringiensis kurstaki, and in some mosquito species to Bti, thus showing the need for alternatives to these compounds that are still effective (Brogdon and McAllister 1998; Marrone and Macintosh 1993). In addition to resistance, other problems are associated with the use of microbial control agents. Cook et al. (1996) discussed potential hazards, not properly identified in the planning stages, of displacement of native microorganisms, allergic responses in susceptible humans and

PAGE 18

6 animals, and eventual toxicity to nontarget organisms. Because of these problems, alternatives are needed to prevent another crisis like the one from which IPM originally arose.

PAGE 19

CHAPTER 2 HISTORY OF THE USE OF AMINO ACIDS AS A MEANS TO CONTROL INSECT PESTS Non-Protein Amino Acids One avenue of pest management explored in the field of biorational pesticides is the use nonprotein amino acids. Secondary plant materials such as these serve many functions in insect-plant relationships from attractants and repellents to crude insecticides (Dahlman 1980). Only a few nonprotein amino acids have been examined as a potential means to control insect pests. L-canavanine and its by-product of detoxification, Lcanaline, have been studied extensively, with a variety of effects ranging from developmental deformities to aberrant adult behavior (Dahlman and Rosenthal 1975; 1976; Rosenthal et al. 1995). L-canavanine is found mainly in leguminous plants, including several economic species (Bell 1978; Felton and Dahlman 1984). It is believed that plants produce this allelochemical for protection against feeding by phytophagous insects and herbivores (Rosenthal 1977). The mode of action for canavanine can be traced to several metabolic processes, including disruption of DNA/RNA and protein synthesis, arginine metabolism, uptake, anomalous canavanyl protein formation, and the reduction of active transport of K + in the midgut epithelium (Kammer et al. 1978; Racioppi and Dahlman 1980; Rosenthal 1977; Rosenthal et al. 1977; Rosenthal and Dahlman 1991). In contrast, canaline possesses neurotoxic characteristics with an unknown mode of action (Kammer et al. 1978). The species of choice for studies involving nonprotein amino acids has been the tobacco hornworm (THW), Manduca sexta (L.) (Lepidoptera: Sphingidae). 7

PAGE 20

8 L-canavanine exhibits a range of insecticidal effects in artificial diets when exposed to the THW. Dahlman (1977) demonstrated a reduction in consumption of artificial diet containing less than 1% canavanine (w/v) which resulted in a lower body mass and increased developmental time to the adult stage. Fecundity and fertility also was affected by L-canavanine. Rosenthal and Dahlman (1975) showed that concentrations as low as 0.5 mM L-canavanine in the diets of the THW resulted in the reduction of ovarial mass of adults, while Palumbo and Dahlman (1978) showed that concentrations of L-canavanine in agar-based diets resulted in the reduction of chorionated oocyte production in concentrations between 1 .0 and 2.0 mM. Under natural conditions, L-canavanine was found to retard development, and increased the susceptibility of exposed larvae to biotic and abiotic mortality factors (Dahlman 1980). However, field applications of L-canavanine were shown to be impractical because of the expense involved in synthesizing L-canavanine from its source, the jack bean (Canavalia ensiformis (L.) DC. (Family: Fabaceae)). Other sources of L-canavanine (i.e., analogues and homologues) were sought in an attempt to find a more practical source of the amino acid. Structural homologues of canavanine were examined and found to contribute to pupal deformities (and to a lesser degree, to mortality) (Rosenthal et al. 1998). Long-chain esters of L-canavanine were found to be more toxic than the parent compound when injected or added to an artificial diet exposed to last instar of THW specimens (Rosenthal et al 1998). Adding amino acids other than arginine (the parent compound to L-canavanine) to diets containing Lcanavanine increased deformities and mortality of THW larvae and was attributed to the structure and position of the functional groups on the added compounds (Dahlman and Rosenthal 1982). Although the THW has an effective means of degrading aberrant

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9 proteins (produced by the assimilation of L-canavanine) into newly synthesized proteins; the proteases involved do not efficiently degrade enough to prevent some damage from occurring in the insect (Rosenthal and Dahlman 1986; 1988). Surprisingly, L-canavanine also was shown to increase the effectiveness of Bacillus thuringiensis in vivo by altering membrane properties, mainly gut permeability, and active transport in the midgut of the THW (Felton and Dahlman 1984). However, despite the possible synergistic relationship between the relatively safe Bt product and this amino acid, no further research has been conducted to evaluate the combination for future commercial use. Other species of insects have also been tested for susceptibility to canavanine with a variety of results. Larvae of Drosophilia melanogaster Meigen (Diptera: Drosophilidae) showed no deleterious response to lower concentrations of canavanine, but showed mortality increased at concentrations over 1,000 ppm (Harrison and Holiday 1967). Lower concentrations also were ineffective against adult Pseudosarcophaga affinis (Fallen) (Diptera: Calliphoridae), with no effect on oocyte development (Hegdekar 1970). Dahlman et al. (1979) examined four species of muscoid flies and found greater than 70% mortality at the higher concentration (800 ppm) and decreased pupal weights as concentrations of canavanine increased. Despite the toxicity of canavanine to some insects, others have evolved detoxifying mechanisms to deal with high concentrations of this compound. Rosenthal et al. (1978) attributed the detoxification of canavanine in the bruchid Caryedes brasiliensis Thunberg (Coleoptera: Bruchidae) to the beetle's ability to convert canavanine to canaline, another toxic amino acid. Canaline is metabolized through reductive deamination to homoserine and ammonia, with the overall result being the detoxification

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10 of the two antimetabolites. This process actually increases the nitrogen intake from the foodstuff (from the increase of ammonia) (Rosenthal et al. 1976; Rosenthal et al. 1977). Another insect, the tobacco budworm {Heliothis virescens (Fab.) (Lepidoptera: Noctuidae)) was able to metabolize far more canavanine then the bruchid beetle larva ever takes in during its development, suggesting that generalists may have more than a single detoxification mechanism for compounds they may encounter (Berge et al. 1986). Metabolism of L-canavanine by the tobacco budworm was attributed to the gut enzyme canavanine hydrolase, and may have been the result of feeding on canavanine-containing plants of the Fabaceae (Melangeli et al. 1997). Essential Amino Acids In despite of the extensive toxicological research conducted on nonprotein amino acids, another group of amino acids, the essential ones, has been overlooked. One reason this avenue for research has not been pursued is that we do not want to give pests convenient access to an integral part of their diet. The fear of creating a "super" insect (that has been provided with compounds that actually aid in its development) is a rational one. Mittler (1967a; 1967b) found an increase in gustation in Myzus persicae (Sulzer) (Hemiptera: Aphididae), with amino acid levels as low as 0.2% concentration in a sucrose solution. Likewise, Sugarman and Jakinovich (1986) found increased gustatory response to both D-and L-methionine by Periplaneta americana (L) (Blattodea: Blattidae) adults. Another reason that essential amino acids have not been examined for use as a pesticide is the knowledge regarding the limited mode of action these compounds could be involved with (i.e., an active site or systemic response). Recent studies on the membrane proteins of insects show the possibility of a biophysiological system that can be exploited for insect control with certain essential amino acids.

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The CationAnion Modulated Amino Acid Transporter With Channel Properties (CAATCHH System Cation-Anion modulated Aminoacid Transporter with Channel properties (CAATCH1) is a recently cloned insect-membrane protein isolated from larval midgut/hindgut nutritive absorptive epithelium. This membrane protein exhibits a unique polypeptide and nucleotide sequence related to, but different from, mammalian Na + -, Clcoupled neurotransmitter transporters (Feldman et al. 2000). Using a unique PCR-based strategy, the gene encoding CAATCH1 was cloned from the digestive midgut of THW larvae. The unique biochemical, physiological, and molecular properties of CAATCH1 indicate that it is a multifunction protein that mediates thermodynamically uncoupled amino acid uptake, functions as an amino acid-modulated gated alkali cation channel, and is likely a key protein in electrolyte and organic-solute homeostasis of pest insects (Quick and Stevens 2001). In the presence of no amino acids, the cations K + and Na + are transported through the membrane via the channel (Figure 2A). When exposed to proline, the amino acid is transported through the membrane with an increase in cation flow, especially Na + (Figure 2B). However, when exposed to methionine, the amino acid transport is stopped and cation flow is altered, mainly the increased flow of K + and the decreased flow of Na + (Figure 2C). The CAATCH1 system works in alkaline conditions, at a pH optimum ~ 9.5. This alkaline condition is found in the midgut of several species (Nation 2001) and has been attributed to a variety of causes, from the detoxification of plant allelochemicals to amino acid uptake (Giordana et al., 2002; Leonardi et al. 2001).

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12 No Amino Acid Proline Methionine Na Na K* Figure 2. The CAATCH1 system identified from the midgut of the tobacco hornworm (modified from Quick and Stevens 2001). In the presence of no amino acids, ion flow is similar for both K + and Na + (A). With the addition of an amino acid, flows are changed. When proline is added (B), the transport occurs but the binding of the amino acid increases the ion flow, notably Na + However, when methionine is added (C) transport occurs and the binding of the amino acid greatly decreases the flow of Na + while K + is increased

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13 Several amino acids were found to initiate the blocking action of ion flow through the CAATCH1 protein, including threonine, leucine, and methionine, with the latter producing the greater response, based on CAATCH1 research (Feldman et al. 2000; Stevens et al. 2002; Quick and Stevens 2001). Methionine The amino acid methionine is considered essential in the diets of many organisms. Methionine is considered an indispensable amino acid in humans. Because the body does not synthesize it, uptake of methionine must occur in the diet. The recommended daily allowance of methionine for a healthy lifestyle ranges from 13 to 27 mg/kg/day for infants to full-grown adults (Young and El-Khoury 1996). This amino acid is linked to a decrease in histamine levels, increased brain function, and is found in a variety of sources; with the highest concentration in various seeds, greens, beef, eggs, chicken, and fish (Dietary Supplement Information Bureau 2000). Recently, research has centered on the genetic modification of crop plants to overproduce methionine to increase its nutritional quality (Zeh et al. 2001). Wadsworth (1995) discussed using methionine as a feed supplement, as an aid in the therapy of ketosis in livestock, and as a treatment for urinary infections in domestic pets. Onifade et al. (2001) examined the use of housefly larvae as protein foodstuffs, and found an increase in body weight gain and erythrocyte counts in rats whose diets were supplemented with fly larvae and methionine. Likewise, Koo et al. (1980) suggested dry face fly pupae could be used as a dietary supplement and foodstuff extender for poultry because of the high concentration of methionine. The environmental safety of methionine is well known, as it poses no risk to vertebrates due to a rather high oral LD 50 of 36g/kg _1 observed in rats (Mallinckrodt Baker 2001) and also

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in its use as a feed supplement for livestock under the trade name of Alimet (Novus, Inc., St. Louis, MO). In addition to vertebrates, methionine also is considered an essential amino acid for insects (Nation 2001). Based on research on nutritional requirements for insects, the amount of methionine needed in a diet for survival ranged from as little as 0.0007 mg/mL (for Aedes aegypti (L.) (Diptera: Culicidae) to as high as 100 mg/mL (for Heliothis zea (Broddie) (Lepidoptera: Noctuidae)) (Dadd and Krieger 1968; Eymann and Friend 1985; Friend et al. 1957; Kaldy and Harper 1979; Kasting et al. 1962; Koyama 1985; Koyama and Mitsuhashi 1975; Rock and Hodgson 1971; Singh and Brown 1957). Methionine occurs naturally as the L-isomer while the D-isomer (an optical enantiomer) is toxic to many insects and is not found in nature (Anand and Anand 1990). A few exceptions are known, (mainly Diptera and Lepidoptera) that actually are capable of metabolizing the normally unusable D-isomer (Dimond et al. 1958; Geer 1966; Rock 1971; Rock et al. 1973; Rock et al. 1975). The requirement for small amounts of this amino acid (as compared to other amino acids) may be a result of the ability for some insects to synthesize methionine from cysteine (a common sulfur containing amino acid) thus reducing the need to take in exogenous sources of methionine. Jaffe and Chrin (1979) found that A. aegypti adults were able to synthesize methionine from homocysteine with the aid of a methionine synthetase. They found this enzyme similar to those common in other metazoans, and found that the levels of methionine synthetase increased with the presence of filarial parasites. They hypothesized that this increase in methionine synthetase was a result of the parasite depleting the host of methionine. Fertility and fecundity also have been associated with methionine in some insects (mainly D. melanogaster,) with the possibility if it being a limiting factor during egg

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15 production (Sang and King 1961). Lack of methionine in the diet of the female may also explain the transfer of methionine in the ejaculate of the male during fertilization (Bownes and Partridge 1987). Methionine plays another role in insect biochemistry, especially in juvenile hormone biosynthesis, inhibitory allatostatins, and storage proteins known as hexamerins. Audsley et al. (1999) found that in vitro rates of juvenile hormone synthesis in females of the tomato moth {Mamestra oleracea (L.) (Lepidoptera: Noctuidae)) were dependent on the concentration of methionine present in the incubation medium. Tobe and Clarke (1985) found a direct relationship between methionine concentration and juvenile hormone biosynthesis in the cockroach, Diploptera punctata (Eschscholtz) (Blattodea: Blaberidae), further supporting the idea that methionine plays an important role in insect biochemistry. Storage proteins, or hexamerins, act as a storehouse for amino acids that can be sequestered for later use in the developmental cycle (Pan and Telfer 1996). Many Lepidoptera have been identified with hexamerins containing high concentrations of methionine and are metabolized during the last larval stage, and presumably used for egg production (Wheeler et al. 2000). Methionine as a potential pesticide has not been overlooked entirely. Tzeng (1988) tested a methionine and riboflavin mixture and found it successful in controlling various pests, including the larvae of Culex spp. (Diptera: Culicidae). The mode of action for this mixture was attributed to a photodynamic reaction and the production of oxygen rich radicals (Tzeng et al. 1990). Their research led to the use of this methionine compound as a control agent for sooty mold of strawberry (Tzeng and Devay 1989; Tzeng et al. 1990) but not as an insecticide.

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16 Discovery of novel means for controlling various insect pests is one tenant of IPM. The amino acid methionine, an environmentally safe organic compound, appears to be a candidate for further study. Before it can be considered for use in controlling insects pests, several issues must be addressed, including the determination of concentrations needed to provide effective control, compatibility with current application systems, safety to nontarget organisms (i.e., beneficial or biological-control agents), and to phytotoxicity. Research Objectives Our overall goal was to evaluate the effects of L-methionine, and its amino acid analogues, on the CAATCH1 system putatively in the midgut/hindgut as a means to control different insect pests. The working hypothesis is that the L-methionine only affects the CAATCH1 system and no other system, especially those involving Na+ channels or pumps (i.e., nervous tissue). The L-isomer of methionine was chosen because of the inability of most insect species to utilize the D-isomer. Ideal targets for this research are those pests that cause severe damage to agricultural systems and to human health. Specific objectives were to • Examine the effects of L-methionine as an insecticide on the larvae of M. sexta (Tobacco hornworm), L. decemlineata (Colorado potato beetle) and A. aegypti (Yellow-fever mosquito) under various conditions • Determine any adverse effects of L-methionine on plant health to ensure its safe use in a cropping system • Examine the effects of L-methionine on various nontarget insect species to ensure the environmental safety of L-methionine and thus its compatibility with natural enemies in the context of IPM.

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CHAPTER 3 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE TOBACCO HORNWORM, Manduca sexta, UNDER LABORATORY CONDITIONS Introduction Manduca sexta (L.) (Lepidoptera: Sphingidae), the tobacco hornworm (THW), is a widespread species considered an economic pest throughout North and South America. The caterpillar is known for its voracious appetite. In Georgia, the THW was responsible for between approximately $1 .2 to $1 .5 million in losses and costs for control annually in tobacco from 1997 to 2001 (Jones and McPherson 1997; McPherson and Jones 2002). In addition to its well-earned reputation as an agricultural pest of solanaceous crops, the THW has shown to be resistant to common pesticides (such as endrin and endosulfan), with the possibility of cross-resistance (Bills et al. 2004). The THW also is very important to scientific research outside the arena of economic entomology, with studies ranging from molecular-based research to ecological and physiological research, mainly because of its availability and ease in culturing (Dwyer 1999). One research area of interest to scientists involves the chemistry and physiology of the midgut Insect control (or the development of new insecticides) was probably not the main purpose of the research that resulted in identifying the CAATCH1 protein, yet it became the basis of our research project. Because little information is available on the insecticidal properties of methionine, several baseline experiments were necessary to determine that concentrations of this amino acid to test It also was necessary to test L-methionine and THW 17

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interaction in a variety of ways, including artificial diet, natural diet (excised leaves, whole plant, and choice tests. The purpose of this portion of this study was to determine whether L-methionine was detrimental to the survival and development of the THW and to determine if L-methionine could be used to control this species. Materials and Methods Eggs of THW were obtained from the insectary of North Carolina State University, and were held in 26.4L x 19.2W x 9.5H (cm) clear plastic rearing chambers with a hardware cloth (to facilitate cleaning) (Figure 3-1). Florida Reach-In Units (FRIUs) were used to control the environment for the rearing containers (Walker et al. 1993) Containers were held at 27 C, 60% relative humidity, and a 16L.8D photoperiod in FRR7s with either artificial or natural diet (excised eggplant leaves or whole plants) depending on the pending experiment. Neonates were allowed to feed for 2 days after eclosion before being transferred to treatment groups. A camel hair brush was used for transferring larvae, to minimize the risk of damage. Diets and Survivorship The artificial diet was prepared using the procedures outlined in Baumhover et al. (1977) with the inclusion of L-methionine for the treatment concentrations of 0.1%, 0.3%, 0.5%, 1.0%, 3.0%, 5.0% and 10.0% (wt/wt). The artificial diet was changed on a regular basis to prevent desiccation and fungal growth. Larvae were exposed to the artificial diet in the clear plastic rearing chambers with a hardware cloth, and kept in the FRKJs programmed with the aforementioned environmental constants. Natural diets consisted of excised eggplant leaves (Solarium melongena L.,"Classic" variety) of potted plants grown and maintained at the University of Florida,

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Figure 3-1. Rearing chamber for tobacco hornworm and Colorado potato beetle larvae used in the artificial and excised leaf diet tests. Hardware cloth stage supporting the leaf allowed for easy clean up and minimized disease problems by preventing larvae from coming in contact with fecal material (paper liner not shown).

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20 Department of Entomology and Nematology green and shade houses. Excised leaves were dipped in solutions of deionized H2O containing different concentrations of methionine; depending on the experiment and exposed to larvae in the same rearing chambers as the artificial diet trials under the same conditions. Survivorship data were pooled from several different trials for data analysis. In total, 64 potted eggplants were used for the whole-plant portion of the study. Plants were held in FRIUs under the same conditions as the artificial and excised leaf trials, in 38H x 15D (cm) plexiglas cylinders (Figure 3-2). Four THW neonates were placed on each plant for a total of 64 larvae (16 replicates) per treatment (nT 0 tai = 256 larvae). The treatment of L-methionine was applied to the test plants (using a hand-held sprayer calibrated to deliver approximately 10 mL of solution to each plant) before the addition of larvae. Feeding and Development To test L-methionine on the developmental rates of THW, larvae were exposed to excised eggplant leaves dipped in solutions containing the same concentrations of Lmethionine used in the artificial diet trials. Additional treatments of proline (1 .0%) and Bt-kurstaki (Dipel 86% WP at 3.5 grams/liter; Bonide, Oriskany, NY) were included as positive and negative controls, respectively. Leaves were scanned photometrically using the CI 203 Area Meter with conveyor attachment (CID, Inc.; Camas, WA) to measure leaf consumption before and after exposure to larvae. The difference in leaf areas resulting from the missing leaf tissue was assumed to be the amount eaten by the developing larvae. Larval head capsule widths were measured at the time of death or the

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Figure 3-2. Setup for whole plant studies involving tobacco hornworm. Top and portions of the sides were replaced with fine mesh to allow for airflow and to reduce condensation.

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22 end of the trial (using an Olympus Tokyo Model 213598 stereomicroscope with a optical micrometer) to monitor larval development Trials to determine the total amount of L-methionine applied to excised leaves also were included to quantify how much of the amino acid was physically present on leaves at the different concentration levels. Leaves were weighed before dipping into the control (0%) and L-methionine solutions (0.1%10%), allowed to air dry for 30 min and weighed again. The difference was assumed to be the actual amount of L-methionine residue on the leaf. This value then was used to determine the total amount of L-methionine on the leaf surface of the excised leaves and the amount of L-methionine consumed per gram of leaf material, based on calculations of the physical amount of the compound for each % concentration. Preference Tests It was unknown if the additional methionine acted to attract or repel larvae. Neonate larvae were used in the choice tests to determine if there was a preference between the control (deionized H 2 0) and the Treatments (1 .0% L-methionine). Leaves were obtained from potted plants maintained in the outdoor shade house. The tests consisted of 4 leaf disks (30 mm diameter) dipped into the control solution and placed into the chamber alternately with four leaf disks (30 mm diameter) dipped into the treatment solution and replicated with a total of 1 0 chambers. Each chamber consisted of a large petri dish (25.0 cm diameter x 9.0 cm depth) lined with a Seitz filter disk. The filter disk was moistened routinely with deionized H 2 0 to prevent the leaf disks from desiccation (Figure 3-3). Chambers were held in FRIUs at the same environmental constants described previously. The leaf disks also were scanned photometrically and

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Figure 3-3. Chambers used for tobacco hornworm and Colorado potato beetle preference tests. Two treatments (control and 1.0% Lmethionine) were used to determine if any larvae exhibited any preference or avoidance to L-methionine. Treatments were alternated in the chamber and neonates were released in the center of the dish and allowed to search for food. The filter paper in the bottom of the dish was moistened to prevent desiccation of the leaf disks and the test specimens.

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24 larval head capsule measurements made using the same procedures described in the Feeding and Development section. Data Analysis Sample sizes of all experiments were chosen according to the guidelines recommended by Robertson and Preisler (1991) for optimal sample size and data analysis. Probit analysis and determination of mean Lethal Concentration (LC50) were performed using PROBIT Version 1 .5 (Ecological Monitoring Research Division, USEPA) after Abbott's correction for control mortality (Abbott 1925). Survival data were normalized to the previous value when control mortality was greater than the treatment mortality, to produce a smoother trend line. Statistical analysis was performed on the data using Minitab Version 14 (Minitab, Inc.; State College, PA). Analysis of the data included One-way ANOVA and separation of significant means using Tukey's Multiple Comparison and Pearson Correlation was performed on the choice trial data to examine possible relationships between development and consumption of treated leaf material (Zar 1999). Regression analysis using lest squares were performed on the leaf weights before and after the L-methionine treatment for the equation used to convert % concentration to mg/g plant material (Figure 3-4). Results Diets and Survivorship The artificial diet resulted in 100% mortality of THW larvae for the 3.0% L-methionine to 10.0% L-methionine treatment after only one day of exposure (Figure 3-5). Approximately 80% mortality was observed in the 1.0% L-methionine treatment after 4 days, and 50% mortality for both the 0.3% L-methionine and 0.5% L-methionine

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L-methionine Concentration (%) Figure 3-4. Amount of L-methionine present on leaf surface after treatment. Excised leaves were weighed, dipped into various concentrations of L-methionine, allowed to dry, and then re-weighed. Difference assumed to be the amount of L-methionine remaining on leaf surface (T=22.43, and PO.001).

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26 Days of Exposure Figure 3-5. Mortality of tobacco horn worm larvae exposed to various concentrations of L-methionine (njotai^SO) in artificial diet. Data were adjusted using Abbott's formula to account for control mortality. Note the overlap in trend lines for the 3.0% L-methionine10.0% L-methionine concentrations after Day 1 and the 0.3% L-methionine and 0.5% L-methionine treatments from Day 1 to Day 10.

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27 treatment after 10 days of exposure. The 0.1% L-methionine concentration had lowest larval mortality with approximately 30% observed for the triaL The excised leaf trials exhibited higher mortality rates associated with the treatments than did the artificial diet trials. Again, complete mortality was observed with the 3.0% L-methionine thru 10.0% L-methionine concentrations after 1 day of exposure (Figure 3-6). Greater than 90% in the 0.5% L-methionine and 1 .0% L-methionine treatments, followed by 80% mortality in the 0.3% L-methionine treatment, and greater than 60% mortality occurred in the 0.1% L-methionine treatment after 8 days. Whole plant trials produced results similar to the excised leaf trials with greater than 90% larval mortality observed with the 1 .0% L-methionine treated plants after 14 days (Figure 3-7). Mortalities exceeding 20% and 60% were observed for the 0.1% L-methionine and 0.5% L-methionine treatments, respectively, during the same time interval. PROBIT analysis of a sample size of n To tai= 1,540 for 7 treatments (0.1% L-methionine, 0.3% L-methionine, 0.5% L-methionine, 1.0% L-methionine, 3.0% L-methionine, 5.0% L-methionine and 10.0% L-methionine) revealed an overall LC 50 of 0.66% (32.3 mg/g leaf material) concentration for the artificial diet and 0.074% (4.39 mg/g leaf material) concentration for the natural diet after 9 days of exposure (Figure 3-8). The LC 50 for the THW exposed to artificial diet was approximately half the value of that for the natural diet for the 24 to 72 hour exposure period. The LC 50 for the artificial diet of 1.08% (52.3 mg/g leaf material) for 24 h dropped to 1.0% (48.5 mg/g leaf material) after 48 h and to 0.57% (28.0 mg/g leaf material) after 72 h. As for the natural diet, the LC 50 of 0.53% (26.1 mg/g leaf material) was found to be lower than the artificial

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28 Days of Exposure Figure 3-6. Mortality of tobacco hornworm larvae exposed to various concentrations of L-methionine (niotaP 1,540) on excised eggplant leaves. Data were adjusted using Abbott's formula for control mortality. Note the overlap in trend lines for the 3.0% L-methionine10.0% L-methionine concentrations after Day 1

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29 Figure 3-7. Survivorship of tobacco hornworm larvae exposed to various concentrations of L-methionine (nT 0 tai = 256) on whole plants. Lmethionine was applied using a hand-held sprayer in the amount of 10 mL/treatment. Data were adjusted using Abbott's formula for control mortality.

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30 Figure 3-8. Concentrations (%) of L-methionine required for the mortality of 50% of test population of tobacco hornworm after 9 days exposure (nTotai = 1,540; n=180 for 3.0% L-methionine 10.0% L-methionine, n=200 for remainder). Number range following value is the 95% confidence limits. Determination of LC50 was performed using PROBIT Version 1.5 (Ecological Monitoring Research Division,

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31 diet at 24 h and dropped to 0.4% (19.9 mg/g leaf material) at 48 h and 0.25% (12.8 mg/g leaf material) after 72 h exposure. Overall, the LC50 at the end of the experiment for the natural diet was well below the value for the artificial diet, with close to a 90% reduction. Feeding and Development Mortality of THW for the developmental tests ranged from approximately 30% for the 0.1% L-methionine treatment and over 40% for the proline treatment (Figure 3-9). Complete mortality for the 0.3% L-methionine occurred after 7 days while the 0.5% L-methionine treatment took only 5 days. The Btk treatment mortality was similar to the 0.7% L-methionine and 1 .0-%L-methionine treatment, resulting in 100% mortality after 1 day of exposure to the amino acid. Both the mean head capsule width and amount of leaf material consumed showed significant differences between treatments, with the control, 0.1% L-methionine and proline treatments being different that the remaining treatments (Figures 3-10 and 3-1 1). Preference Tests The amount of control and 1.0% L-methionine leaf tissue consumed during the preference tests were found not to be statistically different (Figure 3-12). In addition to the amount of leaf material consumed between treatments not being different, the mean head capsule width {i.e., development) showed a correlation with the amount of control diet consumed (Pearson Correlation Coefficient 0.885, PO.001) while no correlation to the Treatment diet consumed (Pearson Correlation Coefficient 0.630, P=0.051) (Figure 3-11).

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32 Days of Exposure Figure 3-9. Mortality of tobacco hornworm larvae exposed to various concentrations of L-methionine (nTotai = 160) on excised eggplant leaves for feeding and development trials. Proline (1.0%) and Btk were included for comparison as positive and negative controls. Data were adjusted using Abbott's formula for control mortality. Note the overlap in the 0.7% L-methionine, 1 .0% L-methionine and Btk treatments at Day 1

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33 (Error Bars @ 95%; F (0 0 5)7,i 52=2.37, F=18.2; P<0.001) Control 0.1% 0.3% 0.5% 0.7% 1.0% Proline Btk Figure 3-10. Mean head capsule widths of tobacco hornworm larvae exposed to excised eggplant leaves treated with various concentrations of L-methionine (n To ta] = 320). Proline (1.0%) and Btk were included for comparison as positive and negative controls. Error bars denote 2 SE. Bars within treatments having the same letter are not statistically different (Tukey's MST, PO.001).

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34 (Error Bars @ 95% ; F(0.05)7,i52^2 37, F=18J; P<0.001) 300 -i Figure 3-11. Total leaf area consumed by tobacco homworm larvae exposed to excised eggplant leaves treated with various concentrations of Lmethionine (n To ta] = 320). Proline (1.0%) and Btk were included for comparison as positive and negative controls. Error bars denote 2 SE. Bars within treatments having the same letter are not statistically different (Tukey's MSTP, P<0.001).

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35 (&ror Bars @ 95% ; F(0.os)u 8=5.98, F=1.64; P=0.217) Control 1.0% Treatment Figure 3-12. Mean leaf consumption by tobacco hornworm in the preference tests. Error bars denote 95% SE, and treatments were found not to be statistically different. However, there was correlation between the control diet consumed and mean head capsule width (Pearson Correlation Coefficient 0.885, P=0.001) while no correlation was found between the Treatment diet consumed and mean head capsule width (Pearson Correlation Coefficient 0.630, P=0.05).

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36 Discussion The initial studies involving the high concentrations of L-methionine (i.e., 3.0-10.0%, which are outside the range normally encountered in nature) showed that a concentration of 1 .0% L-methionine was sufficient enough to provide good control of THW larvae reared on both artificial and natural diets. The 0.1%L-methionine concentration remained similar to that of the control for developmental and feeding trials (Figure 3-9), indicating a level of methionine that can be tolerated to some extent, as seen in the low mortality of this treatment. This is in stark contrast to the mortality seen in the excised leaf trials in which the same concentration had over 60% mortality (Figure 3-6). One possible explanation could be the amount of L-methionine present on the leaf disk being low enough and ingested at a slower rate than that of the whole leaf, which was left in the chamber with the larvae until the leaf was either completely consumed or too wilted for the larvae to ingest. The preference tests did show some preference towards control leaf disks over the 1 .0%L-methionine treated disks as seen in the correlation analysis of the diet consumed and the mean head capsule width of the larvae. Despite the lack of a statistical difference between the amount consumed, the larvae could have fed on the treated disks and then switched to the control disks based on a physiological cue. It is unclear if THW larvae possess specialized sensory structures to detect amino acids like those found in other Lepidoptera (Beck and Henec 1958; Dethier and Kuch 1971; Schoonhoven 1972), but the possible switch from the methionine rich treatment to the control leaf disks does indicate some sort of mechanism for detection. Del Campo and Renwick (2000) found THW larvae were induced to feeding on plants outside of their normal diet when the plants

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were treated with an extract from potato foliage suggesting induced host preference, attraction, and dependence on this compound in the extent of sustained feeding and development. A combination of sensory structures may be involved for the detection of specific amino acids and host plant compounds, which may explain the selection of methionine depleted host plants to avoid problems with the CAATCH1 system present in the midgut of the THW. The difference in the LC50 for the artificial and natural diets was striking considering the concentrations were the same. One possible explanation is the L-methionine on the natural diet was more readily available than that found in the artificial diet. With the artificial diet, the L-methionine is presumably spread throughout the diet and would therefore take longer for the THW to ingest enough to adversely affect the CAATCH1 system. In contrast, the L-methionine was found on the surface of the leaf in higher concentrations than that of the artificial diet and was also freely available once ingested. Thus, larvae were exposed to a higher concentration of L-methionine with less work to digest, resulting in lower survivorship in the same period of time. The 1.0%L-methionine concentration had the same mortality, feeding and developmental rates for THW, as did the Btk treatments (Figure 3-9). The 0.3% L-methionine, 0.5% L-methionine and 0.7% L-methionine treatments were virtually the same for mortality (Figure 3-9), developmental rate (Figure 3-10) and total leaf material consumed (Figure 3-1 1) and statistically the same as the 1 .0% L-methionine concentration and the Btk treatment. The similar mortality rate observed for the higher concentrations of L-methionine and Btk is encouraging considering the resistance to Bt seen in many insect species because of reduced receptor activity and binding (Bills et al.

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2004; Nester et al. 2002). Resistance in insects involves a variety of mechanisms and many are the result of a combination of different pesticide classes. The CAATCH1 system is one that could be used in cases where the only alternative is by adding more pesticides or at higher rates to break resistance. Further research is needed to determine compatibility of the different Bt insecticides and L-methionine with each other for cases in which Bt resistance is observed in natural populations. Given the safety of L-methionine and the shorter time required for 100% mortality (when compared to Btk results of this study), this compound could represent a viable alternative for pesticides currently used in the management of the THW.

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CHAPTER 4 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE COLORADO POTATO BEETLE, Leptinotarsa decemlineata, UNDER LABORATORY CONDITIONS Introduction Leptinotarsa decemlineta (Say) (Coleoptera: Chrysomelidae), the Colorado potato beetle (CPB), is considered an economic pest throughout North America. The larvae and adults of the CPB feed on a wide variety of solanaceous crop plants and are responsible for $150 million in losses and control related costs (Durham 2000). To further complicate matters, the CPB is resistant to numerous pesticides, including various pyrethroids and carbamates (Bills et al. 2004). Historically, CPB management relied heavily on chemical control methods that led to the development of resistance to different pesticides in several areas of the eastern United States (Forgash 1985; Gauthier et al. 1 98 1 ). Control of CPB without the use of chemicals is further complicated given the species ability to develop resistance and the limitations on the use of resistant varieties of potato (Ragsdale and Radcliffe 1999). The use of plant varieties that are resistant to CPB and other pests also run the risk of developing tolerance to chemical pesticides in other pest species (Sorenson et al. 1989). Despite the success of Bacillus thuringiensistenebrionis (Btt) and the biocontrol agents Podisus maculiventris Say (Hemiptera: Pentatomidae) and Edovum puttleri Grissel (Hymenoptera: Eulophidae), more biorational alternatives are necessary for controlling CPB to prevent yet another devastating threat to the potato industry because of this insect's ability develop resistance and overcome control methods (Boucher 1999; Ferro 1985; Tipping et al. 1999). This makes the CPB 39

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40 an excellent candidate for the evaluation of L-methionine as a possible means of controlling this devastating pest. Because little information is available on the insecticidal properties of L-methionine, several baseline experiments were necessary to determine what concentrations of this amino acid to test. Therefore, it was necessary to test L-methionine and CPB interaction in a variety of ways including survivorship of both larvae and adults, development of larvae when exposed to different concentrations of the amino acid, and preference tests. The purpose of this portion of this study was to conduct bioassays to determine if exposure to L-methionine was detrimental to the survival and development of the CPB and to determine if L-methionine could be used to control this species. Materials and Methods Eggs of CPB were obtained under UDSA permit from the insectary of the New Jersey Department of Agriculture and held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a hardware cloth (to facilitate cleaning) and held at 27 C, 60% relative humidity and 16L/8D photoperiod in FRIUs (Figure 3-1). Excised eggplant leafs were placed in the chambers with the neonates and they were allowed to feed for 2 days after eclosion before being transferred to experiments. A camel hair brush was used for transferring the neonates to minimize the risk of damaging the larvae. Survivorship Larvae and adults of the CPB were tested in preliminary experiments with the highest concentration (1 .0% L-methionine (wt/wt)) observed in tests done on the THW in the previous chapter. The diet for the larvae and adults consisted of excised eggplant leaves {Solarium melongena L.,"Classic" variety (Family: Solanaceae)) from plants grown and maintained at the University of Florida, Department of Entomology and

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41 Nematology green and shade houses. Excised leaves were dipped in solutions of deionized H2O containing different concentrations of methionine and held in the clear plastic boxes and held at the aforementioned environmental conditions (Figure 3-1). Additional treatments of proline (1.0%) and Bt-tenebrionis (Novodor FC @12.4 mL/L; Valent Biosciences, Libertyville, IL) were included as positive and negative controls, respectively. Survivorship data were pooled from several different trials for data analysis. Feeding and Development To test L-methionine on the developmental rates of CPB, larvae were exposed to excised eggplant leaves dipped in different concentrations of L-methionine under the same conditions as the survivorship trials. Additional treatments of proline (1.0%) and Btt were included as positive and negative controls, respectively. Leaves were scanned photometrically using the CI 203 Area Meter with conveyor attachment (CID, Inc., Camas, WA) before exposure to the larvae and measuring after leaf consumption. The difference in leaf areas resulting from the missing leaf tissue was assumed to be the amount eaten by the developing larvae. Larval head capsule widths were measured at the time of death or the end of the trial (using an Olympus Tokyo Model 213598 stereomicroscope with an ocular micrometer) as an evaluation of larval development. Preference Tests It was unknown if the additional methionine acted to attract or repel larvae. Neonate larvae were used in the choice tests to determine if there was a preference between the Control (deionized H 2 0) and the treatments (1.0% L-methionine). Leaves were obtained from potted plants maintained in the outdoor shade house. The tests

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42 consisted of 4 leaf disks (30 mm diameter) dipped into the Control solution and placed into the chamber alternately with four leaf disks (30 mm diameter) dipped into the treatment solution and replicated with a total of 1 0 chambers. Each chamber consisted of a large petri dish (25.0 cm diameter x 9.0 cm depth) lined with a Seitz filter disk. The filter disk was moistened routinely with deionized H 2 0 to prevent the leaf disks from desiccation (Figure 3-3). Chambers were held in FRIUs at the same environmental constants described previously. The leaf disks also were scanned photometrically and larval head capsule measurements made using the same procedures described in the Feeding and Development section. Data Analysis Sample sizes of all experiments were chosen according to the guidelines recommended by Robertson and Preisler (1991) for optimal sample size and data analysis. Probit analysis and determination of mean Lethal Concentration (LC 50 ) were performed using PROBIT Version 1.5 (Ecological Monitoring Research Division, USEPA) after Abbott's correction for control mortality (Abbott 1925). Survival data were normalized to the previous value when control mortality was greater than the treatment mortality, to produce a smoother trend line. Statistical analysis was performed on the data using Minitab Version 14 (Minitab, Inc.; State College, PA). Analysis of the data included One-way ANOVA and separation of significant means using Tukey's Multiple Comparison and Pearson Correlation was performed on the choice trial data to examine possible relationships between development and consumption of treated leaf material (Zar 1999).

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43 Results Survivorship Mortality of CPB larvae on treated excised eggplant leaves ranged from approximately 20% for the 0.1% L-methionine treatment after 4 days, 80% mortality for the 0.3% L-methionine treatment after 8 days of exposure and 100% for the remaining concentrations with the highest dose of 1 .0% L-methionine exhibiting complete control of CPB in 3 days post treatment (Figure 4-1). Some mortality (50%) was observed for the proline (1 .0%) treatment while the Bit larval treatment mortality was similar to the 1.0% L-methionine treatment, resulting in 100% mortality after 5 days. PROBIT analysis of a sample size of n to tai = l,320 for 6 treatments (Control), 0.1% L-methionine, 0.3% L-methionine, 0.5% L-methionine, 0.7% L-methionine and 1.0% L-methionine) revealed an overall LC50 of 0.21 8% concentration for the CPB after 8 days of exposure (Figure 4-2). The LC50 of 2.9% for 24 hours dropped to 1.1% after 48 hours and to 0.22% after 72 hours. Feeding and Development Mean head capsule widths between treatments were found to be statistically different (Figure 4-3). Four distinct groups were observed, with the Control, 0.1% L-methionine and proline treatments forming the first group. The second group of proline and 0.5% L-methionine were statistically the same and likewise the third group of the 0.3% L-methionine, 0.5% L-methionine, and 0.7% L-methionine treatments. The final group of Btt and 1.0% L-methionine treatments was statistically different from all other treatments.

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44 Days of Exposure Figure 4-1. Mortality of Colorado potato beetle larvae exposed to excised eggplant leaves treated with various concentrations of L-methionine (n T otai =: 560). Proline (1.0%) and Btt were included for comparison as positive and negative controls. Data were adjusted using Abbott's formula for control mortality.

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45 Figure 4-2. Concentrations (%) of L-methionine concentrations required for the mortality of 50% of the test population of Colorado potato beetle after 8 days exposure (nT O tai = 220). Number range following value is the 95% confidence limits. Determination of LC50 was performed using PROBIT Version 1.5 (Ecological Monitoring Research Division, USEPA), including Abbott's

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46 (Error Bars @ 95%; F(o.o5)7,3i2=1.14;F=576.71; P<0.001) Figure 4-3. Mean head capsule widths of Colorado potato beetle larvae exposed to excised eggplant leaves treated with various concentrations of Lmethionine (n To tai = 320). Proline (1.0%) and Bt were included for comparison as positive and negative controls. Error bars denote 2 SE. Bars within treatments having the same letter are not statistically different (Tukey's MST, PO.001).

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47 Feeding rates of CPB also were found to be statistically different among treatments (Figure 4-4). Three distinct groups were observed with the first group containing the Control and 0.1% L-methionine treatments while the second group of the 0.1% Lmethionine and 0.3% L-methionine, treatments were found to be statistically the same. The 0.5% L-methionine, 0.7% L-methionine, 1.0% L-methionine and Btt treatments were statistically different from the other groups. Overlap occurred with the proline treatment across all groups indicating no statistical difference with the rest of the treatments. Preference Tests The amount of Control and 1 .0% L-methionine leaf tissue consumed during the preference tests was found not to be statistically different (Figure 4-5). In addition, the mean head capsule width (/'.e., development) showed no relationship with either treatment based upon the low correlation coefficients. The 1.0% L-methionine concentration produced the same larval mortality, feeding and developmental rates for CPB, as did the Btt treatments (Figures 4-1, 4-3, and 4-4). The 0.3% L-methionine, 0.5% L-methionine and 0.7% L-methionine treatments took 4 days longer for complete control (Figure 4-1), but were statistically different for the developmental rates for the same treatments (Figure 4-3). As was the case with the THW survivorship, the 0.1% L-methionine concentration was not different from that of the Control. This may indicate a threshold of methionine that can be tolerated by the THW, and CPB to some extent, evidenced by the low mortality observed for this treatment. The Preference tests did not indicate any preference of leaf disks with or without L-methionine. The high mortality (90%) of the CPB larvae could be explained by a

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48 Figure 4-4. Total leaf area consumed by Colorado potato beetle larvae exposed to excised eggplant leaves treated with various concentrations of Lmethionine (n To tai=320). Proline (1.0%) and Btt were included for comparison as positive and negative controls. Error bars denote 2 SE. Bars within treatments having the same letter are not statistically different (Tukey'sMST,P<0.001).

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49 (Error Bars @ 95%; F (0 .o 5 )U8=5.98, F=1.64; P =0.217) Control 1 .0% L-methionine Figure 4-5. Mean leaf consumption by Colorado potato beetle in the preference tests. Error bars denote 95% SE, and treatments were found not to be statistically different. No correlation between either Control or Treatment Diet consumed and mean head capsule width was found (Pearson Correlation Coefficient 0.466, P=0.175 and 0.665, P=0.036, respectively).

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50 combination of the early consumption of the treated disks and mortality occurring after 48 hours, when a lower concentration is required for mortality. The larvae could have fed on the treated disks and then switched to the Control based on a physiological cue. Mitchell (1974) and Mitchell and Schoonhoven (1974) examined the taste receptors of CPB and found physiological and behavioral responses to some amino acids, mainly gamma aminobutyric acid (GABA) and alanine. They discussed the possibility that host selection in solanaceous plants may have been the result of these chemosensory structures and the concentration of amino acids in the leaves. It should be noted that both studies excluded methionine and no electrophysiological data were collected on the response of CPB to this amino acid. This is not surprising considering the fact that the diet of the CPB is low in methionine and therefore would not be a candidate for the inclusion in feeding stimulatory studies (Cibula et al. 1967). It is unknown if these sensory structures can detect methionine and possibly act as a means to avoid plant material high in this amino acid. This appears to be contradicted by the data in Figure 4-5, in which there was no difference between the treatments. The larvae feeding on the Control treatment, consuming the majority and then moving to the 1 .0% L-methionine treatment, could explain the lack of difference. There are some differences between some of the Feeding and Development treatments should be noted. The mean head capsule of the larvae in the 0.5% L-methionine treatment was higher than the 0.3% L-methionine treatment while the amount of leaf material consumed for the same treatment were the same indicating another factor involved with the greater head capsule width. The differences could be the result of the larger size of females and possibly could have included more females.

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51 The higher concentrations of L-methionine that produced mortality similar to the Btt is encouraging considering the occurrence of resistance to this compound seen in many pest insect species because of reduced receptor activity and binding (Bills et al. 2004; Nester et al. 2002). Resistance in insects involves a variety of mechanisms and many are the result of exposure to a combination of different pesticide classes. The Methionine-CAATCHl system could be exploited in cases where the only alternative is applying different pesticides or using higher rates to break resistance. Further research is needed to determine compatibility with Bt and L-methionine for cases in which resistance is observed in natural populations. Given the safety of L-methionine and the shorter time required for 100% mortality (when compared to Btt), this compound could represent a new biorational tool for the management of the CPB.

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CHAPTER 5 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE YELLOW FEVER MOSQUITO, Aedes aegypti, UNDER LABORATORY CONDITIONS Introduction Integrated Pest Management practices are not restricted to agricultural pests. Medically important insect pests are responsible for epidemics that have changed the course of human existence, from bubonic plague spread by the Oriental rat flea (Xenopsylla cheops Rothschild (Siphonaptera: Pulicidae)), to malaria carried by anopheline mosquitoes. One medically important species that has had a significant impact on human existence is the yellow fever mosquito (YFM), Aedes aegypti (L.) (Diptera: Culicide). This cosmopolitan species is found worldwide and is the primary vector for human dengue and yellow fever despite concerted efforts at eradication in the United States (Womack, 1993). In the United States alone, upwards of 150,000 lives were lost to yellow fever in the period starting in the late 1 8 th century and into the early 20 th century (Patterson, 1992). However, because of the development of a vaccine, yellow fever has been replaced by Dengue which is now second only to malaria as a worldwide threat (Gubler, 1998). Because Dengue fever is also vectored by the YFM, it poses a risk by affecting tens of millions of people worldwide (Gubler and Clark, 1 995). The inclusion of the YFM in this study was an effort borne of curiosity because of the lack of knowledge of the CAATCH1 system in other insects and the availability of specimens for study. Mosquito larvae are particulate feeders and have dietary 52

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53 requirements of methionine in the amounts of 0.0007mg/ml for the YFM. This amino acid also is considered essential for other species of mosquito in untraceable (in those studies) amounts (Chen, 1958; Singh and Brown, 1957). Given the high alkalinity found in the midgut of the YFM as well as other mosquito species, this physiological condition indicates the possibility of the presence of the CAATCH1 system in larval mosquitoes (Dadd, 1975). The purpose of this portion of the study was to examine the survival and development of YFM larvae exposed to water treated with excess L-methionine (adults were not tested given the feeding nature). In addition to L-methionine, other amino acids were tested in an effort to see if their response (i.e., survivorship) was similar CAATCH1 responses to methionine found by Feldman et al. (2000). Materials and Methods Bioassay The bioassay experiments consisted of six treatments (control, 0.1%, 0.3%, 0.5%, 0.7% and 1 .0%) each with four replicates. Both L-methionine and D-methionine were tested along with proline, Beta-alanine and L-leucine to examine the other amino acids that were found to be reactive to the CAATCH-1 system (Feldman et al., 2000). Bt-isrealiensis (Aquabac @ a rate of 2.3 mL/m 2 ; Biocontrol Network, Brentwood, TN) and proline also were included in some trials of L-methionine to allow for comparison of both positive and negative effects. Amino acids were weighed using a Denver Instruments Co. XD2-2KD digital scale and added to glass quart jars containing 500ml of deionized H 2 0. Concentrations were based on the proportion of lg/ 100ml for a 1% solution and for corresponding concentrations. Solutions were allowed to sit at room

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54 temperature (23C) to permit the amino acid to fully dissolve before the addition of the larvae. An additional trial of L-methionine buffered with Tris to a pH of 7.0 using a Fisher Scientific Accumet pH 900 was conducted to determine if mortality was attributed to a change in pH or exposure to the L-methionine. Larvae of YFM (third instar) were obtained from the mosquito colony maintained at the Department of Entomology and Nematology, University of Florida. Larvae were transferred to the treatment jars using a camel hair, with 10 larvae per replicate for a total of 40 larvae/treatment and nT O tai = 240 for each amino acid bioassay experiment (Figure 5-1). Approximately 0.5g of finely ground fish food was added to serve as a larval food source and nylon window screen was used to cover the tops of the jar to prevent the escape of any emerged adults. Jars were held at 23 C on a dedicated laboratory bench top for approximately one week. The numbers of larvae surviving were recorded each day. Growth and Development This experiment used the same Materials and Methods as the bioassay portion with the exception of neonate larvae instead of 3 rd instars. Eggs were placed in a tray of water and held at 23 C for 2 days after eclosion. Neonates were removed using a camel hair paintbrush and placed into each jar, with 10 larvae per replicate for a total of 40 larvae/treatment (nTot a r = 240). Larval exuviae or dead larvae were removed and used to examine growth rates by measuring the head capsules. Larvae head capsule widths were measured (using an Olympus Tokyo Model 213598 stereomicroscope with an ocular micrometer) as an evaluation of larval development.

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55 Figure 5-1. Bioassay setup for yellow fever mosquito larvae exposed to various concentrations of amino acids and Bti. Jars contained 500mL of solution and were covered with screen to prevent the escape of emerging adults.

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56 Data Analysis Sample sizes of all experiments were selected according to the guidelines of Robertson and Preisler (1991) for optimal sample size and data analysis. Probit analysis and determination of mean Lethal Concentration (LC50) were performed using PROBIT Version 1.5 (Ecological Monitoring Research Division, USEPA) after Abbott's correction for control mortality (Abbott 1925). Probit analysis was performed on different concentrations (0.1%, 0.3%, 0.5%, 0.7% and 1.0%) of L-methionine, Trisbuffered L-methionine, D-methionine, Beta-alanine, proline and L-leucine for 24, 48, 72 and 168 hours (the end of the trials). Survival data were normalized to the previous value when control mortality was greater than the treatment mortality, to produce a smoother trend line. Statistical analyses were performed on the data using Minitab Version 12. Analysis (Minitab, Inc; State College, PA) of the data included One-way ANOVA and separation of means using Tukey's Multiple Comparison test (Zar 1999). Results Bioassav Mortality of YFM larvae in both the unbuffered L-and D-methionine trials was similar with low or no mortality, at the 0.1% concentrations (Figures 5-2 and 5-3). The 0.3% concentration had lower mortality with D-methionine (45%) than L-methionine (75%) and greater than 80% mortality for the 0.5% concentration for both isomers. Higher concentrations of both D-and L-methionine forms produced 100% mortality of the larvae within 2 days after treatment. Greater than 40% mortality was observed for the buffered 0.1% L-methionine concentration with complete mortality for the remaining treatments within 5 days of

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57 Exposure (Days) Figure 5-2. Mortality of yellow fever mosquito larvae exposed to various concentrations of L-methionine (n To tai=240). Data were adjusted using Abbott's formula for control mortality.

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58 Days of Exposure Figure 5-3. Mortality of yellow fever mosquito larvae exposed to various concentrations of D-methionine (nx o tai = 240). Data were adjusted using Abbott's formula for control mortality.

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59 exposure (Figures 5-4). The 1 .0%L-methionine treatment caused 100% mortality after 2 days while the Bti treatment took 3 days to reach the same level of control. The proline treatment caused less than 10% mortality. In contrast to methionine, survival of YFM larvae exposed to proline and L-leucine was higher, with only approximately 20% mortality for the higher 0.7% proline and 1.0% proline concentrations (Figure 5-5) and less than 3% mortality with the highest L-leucine concentration (Figure 5-6). Beta-alanine mortality was similar to the L-methionine treatments with between 75% and 83% mortality for the 0.5% Beta-alanine thru 1.0% Beta-alanine concentrations, respectively, greater than 40% mortality with the 0.3% Beta-alanine, and less than 5% mortality for the 0.1% Beta-alanine concentrations (Figure 5-7). Growth and Development Developmental rates of YFM larvae resulted in three distinct groups, with the control and proline treatments, producing virtually identical results; both were statistically different from the 0.1%L-methionine treatment and the remaining L-methionine treatments (Figure 5-8). The Bti treatment was statistically the same as the 03% L-methionine to 1 .0% L-methionine treatments, with very little growth taking place. Probit analysis for unbuffered L-methionine (nT O tai = 40 for 5 treatments; 0.1%, 0.3%, 0.5%, 0.7% and 1.0%) revealed an overall LC 50 of 0.19% concentration for the YFM after 7 days of exposure (Figure 5-9). The LC 50 of 1 .2% for 24 hours dropped to 0.41% after 48 hours and to 024% after 72 hours. When the L-methionine treatments (same concentrations) were buffered to a pH Of 7.0, the values dropped to 0.64% for 24

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60 Days of Exposure Figure 5-4. Mortality of yellow fever mosquito larvae exposed to various concentrations of Tris-buffered L-methionine (nxotai = 240). Data were adjusted using Abbott's formula for control mortality. Note the longer exposure because of the bioassay involving neonates instead of 3 rd instars. Note the overlap in some of the trend lines on Day 1 with the 0.3% L-methionine and 0.5% Lmethionine treatments.

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61 100 n 80 1 60 Days of Exposure Figure 5-5. Mortality of yellow fever mosquito larvae exposed to various concentrations of Proline (nT O tai = 240). Data were adjusted using Abbott's formula for control mortality. Note the overlap of trend lines for all treatments except the 0.7% L-methionine and 1 .0% L-methionine treatments. ^hControl *-0.10% A 0.30% -—0.50% ^^0.70% •—1.00%

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62 100 80 i 60 o 40 20 Control 0.10% 0.30% 0.50% 0.70% 1.00% 0 1 2 3 4 5 6 7 Days of Exposure Figure 5-6. Mortality of yellow fever mosquito larvae exposed to various concentrations of L-leucine (nTotai = 240). Data were adjusted using Abbott's formula for control mortality. Note the overlap in trend lines for all treatments.

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63 Figure 5-7. Mortality of yellow fever mosquito larvae exposed to various concentrations of Beta-alanine (nTotai = 240). Data were adjusted using Abbott's formula for control mortality.

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64 (Error Bars @ 95% ; F
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65 1.4 n 24h 48h 72h Overall (240h) Figure 5-9. Concentrations (%) resulting in 50% mortality (LC 50 ) of yellow fever mosquito larvae exposed to various amino acids after 10 days (nToti = 240 for each amino acid). Number range following value is the 95% confidence limits. Proline and L-leucine were also tested but did not exhibit sufficient mortality to allow for Probit Analysis.

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66 hours, and to 0.1 1% for 48-168 hours and remained constant since the trial lasted longer because of the use of neonates instead of 3 rd instars. The D-methionine treatments were similar with 0.44% for 24 and 48 hours, 0.33% for 72 hours and 0.32% after 168 hours. While not as striking as the others, Beta-alanine had a LC50 concentration of 1 .1% after 24 hours, dropped to 0.5% after 48 hours and leveled off around at 0.35% after 72 and 168 hours. Probit analysis of the Proline and L-leucine treatments was not performed, as the mortality associated with those treatments was too low (Figures 5-5 and 5-6). Discussion Although not commonly encountered, the Dform of methionine had virtually the same effect as the Lform on larval mosquito mortality. The D-and L-methionine trials showed that the Dform had lower mortality associated with it than the more reactive L-counterpart. Insects do not commonly use the Dform of amino acids, although D-methionine is metabolized by some orders to a limited extent (Ito and Inokuchi, 1981). The YFM could be an example of this phenomenon. Because of the nature of the CAATCH1 system in the alkaline midgut, buffering may have acted to increase the effectiveness of the system. Buffering the solutions did result in an increase in mortality, with even the lowest concentration of 0.1% L-methionine exhibiting a two-fold increase with the buffered form (Figure 5-4). Complete mortality was reached sooner with the buffered forms even for concentrations that did not reach 100% in the unbuffered form. In a field setting, the addition of L-methionine would be buffered naturally by the chemical properties of the bodies of water to which it was applied and similar results would be expected.

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67 Jaffe and Chrin (1979) found the adults of YFM females infected with Brugia, a filiaral parasite, were depleted of free form methionine because of the infection and were able to make up the difference by converting homocysteine to methionine with a special synthetase. The ability of YFM adults to synthesize methionine from homocysteine may be present in the larvae as well. This could be the result of the lack of methionine in the diet and possible evidence of the CAATCH1 system being present in at least the adult stage. The susceptibility of the larvae to L-methionine also could be the result of overexposure to a compound that is normally not encountered in high concentrations (>0. 1 %). However, the alkalinity of the particulate feeding larvae and the high mortality to L-methionine suggests that the CAATCH1 system is present and could be exploited in other species with similar midgut characteristics (Dadd, 1975). The survival of YFM larvae exposed to both Beta-alanine and L-leucine was unusual in that they each had the opposite effect on the YFM larvae. L-leucine was expected to have similar blocking properties as L-methionine based on CAATCH1 research (Feldman et al, 2000). Instead, almost no mortality was observed indicating the possibility of another system involved with the transport of this amino acid. Conversely, beta-alanine was not found to be reactive with the CAATCH1 system based on the work of Feldman et al. (2000). The unusually high larval mortality associated with this amino acid may be the result of a yet to be discovered midgut property. The similar mortalities observed for the higher concentrations of L-methionine and Bti is encouraging considering the resistance to this compound that has been documented in many insect species because of reduced receptor activity and binding (Bills et al., 2004; Nester et al., 2002). Resistance in insects involves a variety of

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68 mechanisms and many are the result of a combination of different pesticide classes. The CAATCH1 system is one that could be exploited in cases where the only alternative is applying different or higher rates of pesticides to break resistance. Further research is needed to determine compatibility of Bti and L-methionine for cases in which resistance is observed in natural populations. Given the safety of L-methionine and the similar time required for 100% mortality (when compared to Bti\ this compound could represent a viable alternative to traditional biorational compounds used in the management of the YFM or other susceptible pest mosquito species.

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CHAPTER 6 FIELD EVALUATION OF L-METHIONINE AS AN INSECTICIDE Introduction The role of methionine in animal systems is well known and only recently understood in plants. Methionine is required for protein synthesis; it is a precursor to several important biochemical compounds including ethylene and polyamines, sulfate uptake and assimilation, and also acts as an activator of threonine-synthase (Giovanelli et al. 1980; Droux et at. 2000; Bourgis et al. 2000; Zeh et al. 2001). Recently, research has focused on the transgenic modification of crop plants to overproduce methionine in order to increase their nutritional quality without affecting other biochemical processes (Zeh et al 2001). However, little work has been conducted on the effects of exogenous methionine and it became important to understand the role of externally applied methionine on plant health. Furthermore, the application of L-methionine to plants exposed to natural conditions presents additional problems in terms of how long the residue remains on the plant. Observations of other experiments using L-methionine revealed the tendency of this compound to crystallize after the aqueous portion evaporated forming a brittle, crusty coating that is easily removed. This coating does not appear to interfere with respiration and transpiration at the concentrations studied (1% and lower). To prevent the loss of L-methionine from the plants in a natural setting, the adjuvant Silwett L-77 (Helena Chemical; Collierville, TN) was included in this portion of the study in an effort to increase residual activity on the plant. Silwet L-77 is a nonionic organosilicate 69

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70 surfactant that has wetting and spreading properties (Helena Chemicals 2002) and was found to be compatible with solutions of L-methionine. The objectives for this portion of the study were to examine the effects of a methionine and Silwet L-77 mixture on a crop plant (eggplant) in terms of yield (both fruit weight and total yield) and to evaluate this mixture as an insecticide under natural conditions. Materials and Methods Preliminary Investigation of Silwet L-77 and L-methionine Adult CPBs were obtained from the University of Florida Horticultural Unit, Gainesville and held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a hardware cloth (to facilitate cleaning) and held at 27C, 60% relative humidity and 16L/8D photoperiod in FRIUs. Twenty-four adults were exposed used in each of the 5 treatments, with 4 replicates per treatment (niotar^O). Adults were used because of the lack of sufficient numbers of larvae to test. Excised leaves were dipped in solutions of deionized H2O containing different concentrations of methionine and Silwett L-77 (0.5% concentration), 0.1% L-methionine, 0.5% L-methionine, 1.0% L-methionine and controls of deionized H 2 0 and deionized H 2 0 +Silwet L-77. The additional control was to determine the possible insecticidal properties of Silwet L-77 alone and to make sure the addition of this adjuvant did not affect mortality or deter feeding. Plot Design Eggplants {Solarium melongena L.,"Classic" variety) were grown and maintained at the University of Florida Horticultural Unit, Gainesville, from 18 June to 04 November 2001 Eight, one hundred ft. rows of plants were used for this study, with two rows on each side consisting of buffer rows and four rows in the middle used for the experiments.

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71 Each row contained the 4 treatment plots of 10 plants (control (0% L-methionine), 0.1% L-methionine, 0.5% L-methionine and 1 .0% L-methionine in deionized water solutions) in a Latin square design. Plants within treatment plots were spaced 3 feet apart while treatment plots were 9 feet apart. Figure 6-1 shows the diagrammatic representation of the field plot. Plant Yield Before beginning the experiment, all developing eggplants were removed from the plants in an effort to standardize the treatments and ensure all eggplant development occurred after the exposure of methionine. Treatments were administered using a KQ 3L CO2 (Weed Systems, Inc.; Hawthorne, FL) backpack sprayer charged to 30 lbs PSI and a 3-nozzle boom to ensure complete coverage of the plant (Figure 6-2). Each treatment consisted of a 3L application over the 4 representative groups. The adjuvant Silwett L-77 (0.5% concentration) was included to improve the residual effect of the methionine under the field conditions. Plants were sprayed a total of nine times at approximately two-week intervals. Fruits were harvested at various times during the study and were weighed in the field using a Tokyo Electronics hand-held digital scale. Pest Introduction Neonate CPB larvae were reared on excised eggplant leaves for two days at 27C, 60% relative humidity and 16L/8D photoperiod in FRIUs to ensure healthy individuals for the test. Larvae were transferred to the field plants using a camel hairbrush and the branch marked with flagging tape. Introduction was made after the last spray treatment in November. Ten larvae were placed on each plant for a total sample size of 1 ,600 individuals. Plants were inspected for the next 5 days and larvae encountered noted.

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Barrier Barrier Rows Rows Control (A), 0.1% (C), 0.5% (B) and 1.0% (D) Figure 6-1 Overview of the design layout used to study the effects of L-methionine and Silwett L-77 solutions on yield of eggplant. Rows were four feet apart with individual plants three feet apart and treatments nine feet apart. Each letter represents a group of ten eggplants.

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73 Figure 6-2. Weed Systems, Inc. KQ 3L CO2 backpack back sprayer used for application of L-methionine and Silwett L-77 solutions. Boom consisted of three nozzles (middle top and end of each arm). In total, 3L were applied per treatment every two weeks from 09 July to 3 1 August 2001

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74 Data Analysis Data from the fruit and the CPB experiments were analyzed with ANOVA using Mini tab Version 12. Survivorship of CPB was corrected using Abbott's formula (Abbott 1925) to account for control mortality, mean separation was performed using Tukey's multiple comparison procedure (Zar 1999). Data for both the eggplant weight mean per treatment and also mean number of eggplants per treatment were analyzed using paired t-test. Results Effects of L-methionine and Silwett L-77 on CPB Adults Under Laboratory Conditions Little mortality was observed with the adult CPB at the 1 .0% L-methionine concentration (Figure 6-3). The 0.5% L-methionine concentration had the highest mortality of all the treatments at approximately 20% with the other treatments showing no adverse effects after correction for control mortality. Effects of L-methionine and Silwett L-77 on yield In total, 735 eggplants were collected during the course of this study from 09 June to 3 1 August 2001 Mean weight and yield of eggplants between the treatments were not statistically different from each other (Figures 6-4). Control plants produced 195 fruits with a mean weight of 276.9 grams, followed by the 0.1% treatment with 191 fruits at 281.2 grams. The 0.5% and 1.0% treatments yielded 175 and 174 fruits with mean weights of 295.7 grams and 283.6 grams, respectively. Survival of CPB larvae No statistical difference in survivorship of CPB larvae was observed between the three treatments for the first day after exposure (Figure 6-5) but treatment differences

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75 100 80 | to r 5 5 40 ^•Control Water A 0.10% -—0.50% •-1.00% -—Control Silwet 3 4 5 6 Days of Exposure Figure 6-3. Mortality of Colorado potato beetle adults exposed to excised eggplant leaves treated with L-methionine and the adjuvant Silwett L-77 (protaT^O). Data corrected for control mortality using Abbott's formula. Note the overlap in trend lines for the Control treatments and 0.1% L -methk>nine treatment.

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76 (Error Bars @ 95%; F (I)3> 1S6 =2.6626, F =030963; F= 0.81840) B Control 0.1% 0.5% 1.0% (n=195) (n=191) (n=175) (n=174) Figure 6-4. Effects of L-methionine and Silwett L-77 on eggplant yield (A) and mean weight in grams of fruit (B) from 09 June to 31 August 2001. Error bars denote 2 SE. There was no statistical difference for either eggplant yield or mean eggplant weight (Tukey's MST, P=0.05).

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77 80 Days after treatment Figure 6-5. Mortality of Colorado potato beetle larvae on eggplants treated with L-methionine and Silwett L-77. Mortality of larvae corrected using Abbott's formula (Abbott, 1925). Analysis performed on arcsin transformed data. Error bars denote 2 SE. Data points having by the same letter are not statistically different (Tukey's MST, P=0.05)

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78 were observed thereafter. By Day 4 the 1 .0% and 0.5% treatment were the only treatments that were statistically different from the control. There was substantial unexplained attrition of CPB larvae in the field for all treatments, which leveled off by Day 3. Data from day 5 was discounted because of the onset of a severe cold front that made it difficult to separate the effects of the weather from the treatments affects. Discussion The results of the field studies show that, using conventional application techniques, a mixture of methionine and Silwett L-77 did not appear to affect eggplant yield. Furthermore, the same combination produced substantial control of CPB larvae under natural field conditions after four days. Dahlman (1980) found that L-canavanine, a non-protein amino acid, could be used in the same manner for control of THW on tobacco, but the widespread use of this compound was limited by the cost ($107.85 for lg L-canavanine versus $3.35 for lg of L-methionine (Fisher Scientific International 2004)), adverse effect on plant development (Nakajima et al. 2001), and toxicity to vertebrates (Rosenthal 1977). Although complete coverage of the plant was not feasible, approximately 2.5 grams to 7.5 grams of L-methionine was applied to the plants in each of the treatment plots. Each plant, based on the amount applied, received approximately 7.5xl0 6 ug for the 1.0% L-methionine treatment, 3.8xl0 5 ug for the 0.5% L-methionine treatment and 2.5x1 0 4 ug for the 0.1% L-methionine treatment. This compares to only 4ug of L-canavanine, which resulted in decreased size, fecundity, and mortality of THW under field conditions (Dahlman 1980). It should be noted that the toxicity of L-canavanine is well documented and has a different mode of action than L-methionine and cannot be

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79 compared directly. However, the cost for the amount of L-canavanine required for largescale application far exceeds that of the largest amount of L-methionine needed. Despite the lack of a statistical difference between treatments for both mean weight and mean yield of eggplant, there were some interesting disparities within the data. First, there was an observable difference in mean weight of the eggplants between the treatments and the control. All eggplant weights were greater for the treatments than the control, with the 0.5% L-methionine concentration treatment producing the highest mean eggplant weight. It would appear that excess methionine decreases the number of fruit produced, but those fewer eggplants weighed more. Further research is needed to better understand the differences observed during this study. The addition of Silwet L-77 did not appear to adversely affect survival of CPB as seen in the preliminary tests on the adults and on the larvae during the field release (Figures 6-3 and 6-5). The low adult mortality observed could be attributed to the ability of this species to stop feeding and fly to a more suitable food source. Because the adults were unable to move to an untreated leaf, they were observed sitting motionless on the underside of the leaves. This was not observed in either of the controls as they were seen actively feeding the majority of the time. One aspect of this research that was not examined is that of fertility and fecundity of adults exposed to excess amounts of L-methionine. Despite the fact that methionine is used for egg production in many insect species, excess concentrations may act as a deterrent to feeding causing the adults to stop feeding and to seek other food sources. The lag time from the cessation in feeding to finding another food source may be long

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enough to significantly lower the fecundity of the females and possibly interfere with other behaviors such as mating. During the course of this portion of the study, some anecdotal data were collected based on personal observations. Predators (mainly arachnids) were observed on the plants until the end of the experiment. Other insects also were observed feeding on plants after treatments including piercing-sucking insects (i.e., aphids, coreids and cicadellids) with foliage feeders such as caterpillars rarely encountered except found only on control plants. Attempts to control predators via manual removal were unsuccessful, and predation may have contributed to the observed decrease in CPB. Because predators were present on all treatments, loss from predation was corrected with the use of Abbott's formula. The presence of natural enemies indicates the selectivity of the L-methionine in the field. The amount of methionine ingested by the predators was probably very small because they fed on other insects not plant material. Another set of observations on the safety of L-methionine was the exposure of potted eggplants to high (1 .0% methionine in distilled H 2 0 solution). In total, five plants were sprayed daily with the methionine solution and compared to five plants sprayed with water alone for 14 days. The only difference in the plants was the browning of the leaf tips and edges of the methionine sprayed plants. This also was seen in the excised leaf experiments with THW and CPB. A possible reason for this occurrence was the excess sulfur in the methionine might have burned the leaves. As mentioned earlier, the concentration was very high and also applied daily. Applications of the same concentration did not affect the plants in the field plots, indicating that treatments conducted at 2-week intervals would be safe for the plant.

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81 Overall, it appears that L-methionine can be used in a natural setting to control CPB larvae without affecting crop production. The adjuvant Silwett L-77 worked well with L-methionine in controlling CPB larvae but not the adults. The lack of effectiveness on the adults may be attributed to their ability to stop feeding and living off of reserves acquired during the larval stage until suitable food sources can be found. It is unknown if L-methionine, alone or in combination with Silwett L-77 adversely affects fecundity of the adults.

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CHAPTER 7 EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF SELECTED NONTARGET SPECIES Introduction A biorational pesticide is defined as one that is effective against pest species but innocuous to non-target organisms and not disruptive to biological control agents and beneficial species (Stansly et al. 1996). To test L-methionine as a potential pesticide and determine if it could be considered biorational, it was necessary to examine the effects of this compound on selected nontarget species that could possibly come into contact with it, either directly while on the plant or indirectly via incidental contact or as a host that has come into direct contact with this compound. The species chosen reflect a variety of non-target organisms, mainly those that were shown to be important in controlling some pest species. The pink spotted ladybird beetle, Coleomegilla maculata (DeGeer), the mottled water hyacinth weevil, Neochetina eichhorniae Warner, and the greenbug parasitoid, Lysiphlebus testaceipes (Cresson) all are beneficial insects that have been effective against pests in the state of Florida and also are common and readily available. Each species also represents a different feeding guild (predator, herbivore and parasitoid, respectively) to ensure a thorough examination of the possible effects of methionine as it might be encountered in under natural conditions. The pink spotted ladybird beetle (PSLB) is an abundant polyphagus species that is known to feed on many lepidopteran and coleopteran pests, including the Colorado potato beetle, in which it was responsible for over 50% of the predation on eggs and early 82

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83 instars (Andow and Risch 1985; Giroux et al. 1995; Griffin and Yeargan 2002; Groden et al. 1990; Hazzard et al. 1991; Hilbeck and Kennedy 1996; Munyaneza and Obrycki 1998). This species is widespread throughout North America, and has been shown to provide effective biological control in several crop species, including corn, crucifers, tomato and potato (Hoffman and Frodsham 1993). However, the PSLB was found to be susceptible to carbaryl and menthamidophos, the same pesticides used for the control of many aphid species (Hoffman and Frodsham 1993), Since its introduction into the United States in 1884, water hyacinth {Eichhornia crassipes (Mart.) Solms-Laubach) has infested waterways of the southeast that has cost upwards of $2 million to control in Florida alone (Schardt 1987). The mottled water hyacinth weevil (MWHW), native to Argentina, was first released in Florida in 1972 and subsequently to other states and countries in an effort to control water hyacinth (Center 1994). The genus is restricted to feeding on members of Pontederiaceae, with the MWHW feeding mainly on the introduced water hyacinth; it can be found virtually everywhere the host plant is present (Haag and Habeck 1991 ; Center et al. 1998). The greenbug parasitoid (GBP) is an important natural enemy of many cereal aphids. This species is known for the production of "mummies", the bodies of parasitized aphids that act as a protective case for the developing wasp pupa, and is considered by many to be tolerant to cold temperatures (Elliott et al. 1999; Knutson et al. 1993; Wright 1995). However, this greenbug parasitoid is an insect and is just as susceptible to pesticides despite the protective case of the immature form (Knutson et al. 1993).

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84 The purpose of this portion of the study was to examine the effects of Lmethionine on selected nontarget species that are both important in terms of being beneficial in controlling other pest species and also represent different feeding guilds that would come into contact with this compound in different ways (e.g., on prey items, on plant surfaces, hosts of parasitoids). Materials and Methods Coleomegilla maculata Adults were obtained from ENTOMOS, LLC (Gainesville, Florida), and were held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a hardware cloth stage inserted (to facilitate cleaning) at 27C, 60% relative humidity and 16L/8D photoperiod in FRIUs. Natural diet consisted of excised cotton leafs infested with aphids (Aphis gossypii Glover (Hemiptera: Aphididae)). Leaves were then dipped into either a 1 .0% L-methionine solution or 0% L-methionine (control) mixed with deionized H 2 0. Five adults were used in each replicate for a total n=30 for each treatment. Leaves were replaced every other day from 27 October 2002 to 07 November 2002. Artificial diet was obtained from ENTOMOS and prepared according to their guidelines with the exception of the inclusion of methionine for the 1.0% L-methionine treatment (wt/wt). Diets were replaced every other day from 27 October 2002 to 07 November 2002. Ten adults were used for each replicate for a total n=60 for each treatment. Data was normalized to 0% mortality when the treatments were corrected for control mortality (i.e., when the control mortality was greater than that of the treatment).

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85 Neochetina eichhorniae Adults of the MWHW were used in this study since the larvae and pupae are buried deep in plant tissue and therefore not likely to come into contact with methionine that could be present in a body of water. Specimens were supplied by Hydromentia, Inc. (Ocala, FL), from areas around South Florida. Weevils were maintained following the procedures outlined by Haag and Boucias (1991), with small petri dishes fitted with moistened filter paper and freshly cut water hyacinth leaves. Water hyacinth plants were collected from Lake Alice on the campus of the University of Florida and maintained in the University of Florida, Department of Entomology and Nematology greenhouse. Treatments consisted of cut leaves dipped in deionized H2O (control) or solutions containing 0.1% L-methionine, 0.5% L-methionine, 1.0% L-methionine or 1.0% proline. Prior to weevil exposures, each leaf was inspected for feeding scars or damage and noted to ensure the counts were based on current feeding. Each treatment consisted of 4 replicates with n=5 per replicate (n=20 per treatment and total n=100). Weevils and hyacinth leaves were held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a hardware cloth (to facilitate cleaning) and maintained at 27 C, 60% relative humidity and 16L/8D photoperiod in FRIUs. Fresh leaves were provided every 4 days; exposed leaves were preserved in sealed plastic bags and placed in a refrigerator until scars could be counted. Feeding damage was determined (with the use of an Olympus Tokyo Model 213598 stereo microscope) by the total number of scars present with each counted scar marked with a fine tipped permanent marker (Figure 7-3). Statistical analyses of the weevil data were performed using Minitab Version 12 (Minitab, Inc.; State College, PA). Feeding scars on control and treatment leafs were

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86 compared with a One-way ANOVA and mean separation was performed using Tukey's Multiple Comparison test (Zar, 1999). Lysiphlebus testaceipes To test the effects of methionine on the GBP, cotton plants (Gossypium sp; Family: Malvacae) were grown and maintained at the University of Florida, Department of Entomology and Nematology green and shade houses from 07 October 2002 to 25 November 2002. Aphids (A. gossypii Glover) were supplied from other experiments using this organism and kept on plants within a sealed greenhouse to prevent unwanted parasitism. Plants were maintained in the sealed greenhouse, infested with aphids and then placed in the open shadehouse area to encourage parasitation. In total, 20 plants were used for 2 treatments, 1.0% L-methionine and 0% L-methionine (Control) mixed with deionized H 2 0. Plants were sprayed weekly (12 October 2002 through 17 November 2002) with approximately 10 ml of solution using a hand-held spray bottle. Counts of parasitized aphids began approximately two weeks after placing plants outside to ensure adequate time for parasitism (Royer et al. 2001). Counts were made using a hand lens and counter; "mummies" with exit holes were enumerated and removed. A few parasitized aphids were removed and held in glass vials to ensure correct identification of the parasitoid. Data Analysis Data from the parasitoid experiments were analyzed using Minitab Version 12 (Minitab, Inc.; State College, PA). Control and experimental plants were compared against one another with a One-way ANOVA and separation of significant means was performed with Tukey's Multiple Comparison test (Zar, 1999).

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87 Results Coleomegilla maculata There was virtually no difference between the control and treatment groups for either the artificial or natural diet tests after correction for control mortality. Mortality was slightly higher for the control groups than the 1.0% L-methionine treatment (Figures 7-1 and 7-2). Further analysis was not necessary because of the identical numbers. Neochetina eichhorniae Total mortality for the treatments was less than 20% for all treatments, with the individual treatments having similar results (Figure 7-4). Feeding damage ranged between 2,000 and 4,000 scars per treatment and an average of 10.7 to 16.9 scars per survivor during the course of the experiment (Figure 7-5). No statistical differences were observed between the treatment and control groups Lvsiphlebus testaceipes In total, 188 and 232 aphid mummies with exit holes were found on treatment and control plants, respectively. Means for each treatment were not statistically different for each collection period or overall based on One-way ANOVA (Figure 7-6) with the only exception being the second and last collection period Discussion In general, L-methionine did not have the same toxic effect on the non-target organisms tested when compared to the pest species exposed to the compound in previous chapters. The pink spotted ladybird beetle adults actually showed the least amount of susceptibility to L-methionine. Survival of the adult beetles was higher in the 1 .0% L-methionine treatments than the control for both the artificial and natural diet

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88 100 80 60 40 20 0 "ControlAD ^H 1 .0% L-methionineAD Survivorship of 1 .0%L-methionine Grp> Control Grp 23456789 Days After Exposure 10 11 12 Figure 7-1. Mortality of Coleomegilla maculata adults after exposure to L-methionine treated artificial diet. Data corrected for control mortality using Abbott's formula.

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89 C8 t 100 80 60 40 •ControlND • 1 .0% L-methionine ND Survivorship of 1 .0%L-methionine Grp> Control Grp 4 5 6 7 8 Days After Exposure 10 11 12 Figure 7-2. Mortality of Coleomegilla maculata adults after exposure to Lmethionine treated cotton plant leaves infested with aphids. Data corrected for control mortality using Abbott's formula.

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90 Figure 7-3. Feeding scars on water hyacinth (Eichhornia crassipes) leaf after exposure to Neochetina eichhorniae adults. Black marks represent feeding scars marked with a fine tip marker to aid in counting (other side counted but not shown).

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91 100 80 5? | 60 r o 40 6 8 Days of Exposure 10 Control 0.10% 0.50% 1.00% •Proline 12 14 Figure 7-4. Mortality of Neochetina eichhorniae on treated water hyacinth leaves. Data corrected for control mortality using Abbott's formula

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Figure 7-5. Feeding rate of Neochetina eichhorniae on water hyacinth leaves treated with L-methionine and Proline. No statistical differences were observed between treatments (Tukey's MST, P=0.038).

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(Error bars @ 95%; F (00S)I>18 =4.41; F=3.25; P=0.005) Figure 7-6. Lysephlebius testiceipes parasitized aphids on cotton plants treated with L-methionine. Ten plants were used for each treatment and held in the shade house at the University of Florida, Department of Entomology and Nematology from 22 October to 25 November 2002. No statistical differences were observed except for the second and final collection date.

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trials. One possible explanation for this observation could be that the excess L-methionine increased the dietary quality of the artificial and natural diets for the PSLB in the treatments. However, because only adults were available, further tests are needed to determine if the larvae, also predaceous on the same pests as the adults, are sensitive to this compound. It should be noted that the midgut properties (i.e., alkalinity) for this species are not well known and may not even have the CAATCH1 proteins present in the midgut. The mottled water hyacinth weevil also appears not to be adversely affected by exposure to excess amounts of L-methionine despite its herbivorous habit like the THW and CPB. Another weevil within the same family (Anthonomus grandis Boheman (Coleoptera: Curculionidae)) is known to have an acidic midgut and the same could apply to the MWHW based on these results (Nation 2001). Therefore, this species and possibly other weevils may not be affected by compounds like L-methionine because of the lack of an alkaline midgut needed for the CAATCH1 protein to operate (Feldman et al. 2000; Quick and Stevens 2001). Again, further research is necessary to determine if CAATCH1 proteins are present in this weevil species. The greenbug parasitoid also was unaffected by exposure to the excess L-methionine found on treated leaves infested with aphids. Dadd and Krieger (1968) found higher methionine requirements for the greenbug Myzus persicae Sulzer (Hemiptera: Aphididae) when cysteine is scarce because of its ability to transform excess methionine to much needed sulfur and could possibly explain the parasitoid's tolerance to high methionine concentrations. Because of the life cycle of the GBP, and many other parasitoids, direct contact with compounds such as L-methionine would occur inside the

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95 body of the host, and not through direct contact with the foliage where the compound was applied. There is a possibility for the parasitoid having higher methionine requirements; based on filarial worm infected Aedes aegypti (L.) (Diptera; Culicidae) females and the associated drop in methionine levels in the haemolymph (Jaffe and Chrin 1979). This makes alternatives such as L-methionine safe for use around beneficial insects like the greenbug parasitoid. Overall, the results indicate that the PSLB (C. maculata), the MWHW (N. eichhorniae) and the GBP, (L. testaceipes) were not adversely affected by exposure to L-methionine in excess concentrations in a variety of artificial and natural diets. Survivorship and feeding rates were not statistically different between control and treatment groups for each species. From these data, it can be concluded that L-methionine is safe for use with beneficial insects and could be considered "biorational" in that it showed no adverse effects on non-target species. It also should be stressed that additional testing on other beneficial insects would be, on a case by case basis, necessary to examine the safety and "biorational" qualities of L-methionine.

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CHAPTER 8 SUMMARY AND DISCUSSION The creation and implementation of Integrated Pest Management (IPM) strategies to combat pest species were developed as a response to the economic losses associated with the overuse of chemical control. However. IPM strategies are not widely used because of the lack of alternatives and the ease of use of pesticides. This has resulted in the resistance to pesticides in many insect species, including economic and medical pests. In an effort to provide alternatives to traditional chemical control, biorational methods have been investigated and one such avenue is the use of non-protein amino acids. Chapter 2 covered the history of the use of non-protein amino acids as a pesticide, and discussed the CAATCH1 system and the safety of L-methionine. Only a handful of these amino acids have been investigated as a means of controlling insect pests but still lack the practicality and cost effectiveness as current chemical control methods. Recent discovery of a new midgut membrane protein, CAATCH1, has revealed a new possibility in insect control. The CAATCH1 system works in alkaline conditions and responds to different amino acids, mainly the reduction in ion flow after exposure to methionine, an essential amino acid required for normal development and metabolism of many species including humans. The use of a compound such as methionine would be an excellent addition to the IPM arsenal because of its relative safety to vertebrates and warrants further study as a pesticide. Chapters 3,4, and 5 were dedicated to examining the effects of L-methionine, a common analog of methionine, on three different economic and medically important 96

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97 pests. The tobacco hom worm (THW), Colorado potato beetle (CPB) and the yellow fever mosquito (YFM) were tested and found to be susceptible to concentrations greater than 0.1%. Diets, both natural and artificial, containing this compound resulted in the complete mortality of THW and also in the natural diet for CPB. Development and feeding rates were also affected by the addition of L-methionine to diets for THW and CPB. Survivorship and developmental rates of YFM were also affected by the addition of this amino acid to the larval habitat. In Chapter 6 it was found that the field application of L-methionine under natural conditions was able to control CPB. It was also determined that L-methionine was compatible with Silwett L-77, a commonly used adjuvant, and showed no detrimental effects on crop yield of eggplant. Finally, the application of a compound such as L-methionine has to be able to control the pests that it is used against and not have an effect on beneficial organisms that may come into contact with this compound. Chapter 7 detailed the results of tests that involved various beneficial insects from different feeding guilds (herbivore, predator and parasitoid) showed that L-methionine does not appear to pose a threat to nontarget organisms. One aspect of the use of a compound like L-methionine that is very important is the relative safety. The health hazards related to the contamination of the environment with pesticides are well documented and in the recent years have resulted of the review and removal of several insecticides from commercial and private use. The use of L-methionine as an insecticide would alleviate the dangers associated with other pesticides. The approved use as a nutritional supplement for livestock feed is a testament

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98 to the safety of this compound and residual found on the plant does not pose the same risk to the human population. It is difficult to understand how a compound such as methionine can be considered essential and deadly within the same organism. To understand this dichotomy, an examination of the role of this compound and how it relates to metabolism, development and reproduction is necessary. Although the diet of the THW is lacking high concentrations of methionine, the use of hexamerins may account for the levels needed for the biosynthesis of JH. The larvae take in methionine, metabolizing what is needed and storing the rest for later on during metamorphosis. In contrast, the larvae of the diamondback moth (Plutella xylostella (Lepidoptera: Plutellidae)), feeding mainly on methionine-rich crucifers, lack hexamerins with high methionine concentrations (Wheeler et tiL 2000). The levels of methionine encountered in a normal diet are below what the CAATCH1 proteins are capable of processing and may also be affected by the presence of symbiotic bacteria that is responsible for methionine oxidation in some insects (Gasnier-Fauchet and Nardon 1986a; 1986b). It is when the concentration exceeds the handling capacity of the midgut that problems occur. The time it takes to digest material containing natural amounts of methionine could be long enough for the CAATCH1 system to recover from exposure. The difference between the artificial and natural diet LC 50 for the THW (Figure 3-8) appears to support the idea that bound methionine (i.e., incorporated into the diet and not applied topically) takes longer to cause problems for the organism (if any) versus the relatively quick kill associated with the free methionine present on the leaf surface. The

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99 target ingests the methionine first as it feeds ensuring the overload CAATCH1 system and eventually death. As for the stored methionine, it is released from the storage proteins as needed to synthesize juvenile hormone and allow for transformation in addition to other functions. The remaining methionine is then used for protein synthesis in the tissues around the ovaries to boost yolk production, as seen in the transfer of methionine from male to female Drosophila species (Bownes and Partridge 1987). In the THW, the presence of hexamerins with high methionine content may be an alternative to the male contribution possibly found in its ejaculate. Methionine-rich hexamerins are common in Lepidoptera and have been shown to provide the larvae a source of amino acids during the synthesis of these proteins during the last stage of larval development (Wheeler et al. 2000). In addition to the need for methionine for metabolism and reproduction, the release of methionine may also in part account for the decrease in ion transport of the posterior region of the midgut during larval molts and the wandering stage present before pupation. Currently, little is known regarding the mechanisms involved with the decrease of ion transport during these developmental stages (Lee et al. 1998). Clearly there appears to be more to the role methionine plays in the development of some insects other than the vague designation of "essential" amino acid. Insects have evolved to deal with limiting resources, such as methionine, and have successfully found effective strategies like hexamerin storage or alternate pathways to deal with such problems. No attempt to link together all the aspects of the role of methionine in a whole organism or system context It appears that methionine actually may play a role far more important than that of just an essential amino acid. From the

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100 synthesis of homocysteine to produce methionine to the presence of methionine rich hexamerins and allophorins and protein synthesis, the role of methionine in plant-insect interactions may be larger than originally theorized. The production of methionine overproducing plants could also be used in future IPM strategies. Preliminary results indicate that genetically modified plants do produce enough methionine to affect the survivorship of caterpillars feeding on the plant (unpublished data). This could be used in crops in which improved nutritional quality is important as well as the insecticidal properties of the additional methionine. However, there appears to be a sublethal level (0.1%) of L-methionine in which THW and CPB can "tolerate" and survive with little mortality (Figures 3-9 and 4-1). Any system that makes use of a crop that can overproduce compounds like L-methionine would have to be able to express levels greater than this level to avoid any resistance/tolerance. This research has also provided more possibilities for the use of compounds such as L-methionine in the YFM portion of this study. The amino acid Beta-alanine provided similar levels of control, as did the methionine trials (Figure 5-7). Although unexpected (as discussed in Chapter 5), it shows that there are several other systems that can possibly be exploited in controlling some insects. Further research is necessary to determine if the combination of a compound like methionine and a pesticide already in use would result in the increase in toxicity or the decrease in the concentration of pesticide used. If compatibility between methionine and Bacillus thuringiensis does exists, then it is possible that resistance could be broken in a given population. For example, if a population of THW started to show resistance to Bacillus thuringiensis kurstaki then methionine could be used to remove both susceptible

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101 and resistant alike because of the difference in mode of action. Once the population was reduced, and the corresponding resistant genotype, Btk could be used once more at a lower concentration, closer to that of the susceptible population. This system could also be used for the reduction of Bt toxin resistance in the CPB and YFM if the compounds are compatible. In conclusion, it appears that L-methionine can be used as an insecticide to control insect pests of economic and medical importance. The target site (CAATCH1) is known and found in the midgut/hindgut (presumed) in at least three pest species (tobacco hornworm, Colorado potato beetle and the yellow fever mosquito) and possibly more. The compound (L-methionine) is a safe compound that is already used for livestock feed supplements, has very low mammalian toxicity, and is compatible with insecticide application systems. Non-target organisms were not affected with the application of Lmethionine, further supporting its use as a biorational insecticide. With increasing resistance to current insecticides in the study organisms, alternatives such as Lmethionine are needed now more than ever to further support of Integrated Pest Management strategies.

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106 Fogarty International Center and the U.S. National Institutes of Health (FIC-NIH). 2003. Multilateral Initiative on Malaria. U.S. National Institutes of Health. Internet URL: http://mim.nih.gov/englishyindex.html. Accessed April 2004. Forgash, AJ. 1985. Insecticide resistance in the Colorado potato beetle, pp. 33-52. IN D.N. Fcrro and R.H. Voss (eds.) Proceedings of the Symposium on Colorado Potato Beetle. XVII International Congress of Entomology, Massachusetts Agricultural Experiment Station Bulletin 704. Amherst Massachusetts. Friend, W.G., R.H. Backs and L.M. Cass. 1957. Studies on amino acid requirements of larvae of the onion maggot, Hylema antiqua (MG.), under aseptic conditions Can. J. Zool. 35: 535-543. Gasnier-Fauchet, F. and P. Nardon. 1986a. Comparison of sarcosine and methionine sulfoxide levels in symbiotic and aposymbiotic larvae of two sibling species, Sitophilus oryzae and Sitophilus zeamais (Coleoptera: Curculionidae). Insect Biochemistry 17(1): 17-20. Gasnier-Fauchet, F. and P. Nardon. 1986b. Comparison of methionine metabolism in symbiotic and aposymbiotic larvae of Sitophilus oryzae L. (Coloeptera: Curculionidae)II. Involvement of the symbiotic bacteria in the oxidation of methionine. Comp. Biochem. Physiol. 58(1): 251-254. Gauthier, V.L., R.N. Hoffmaster and M. Semel. 1981 History of Colorado potato beetle control, pp. 13-34. IN J.H. Cashcomb and R. Casagrande (eds.), Advances in Potato Pest Management. Hutchinson and Ross, Stroudsburg, PA. 672pp. Geer.B.W. 1966. Utilization ofD-amino acids for growth by Drosopbila melanocaster larvae. J. Nutr. 90: 3 1 -39. Giordana, B, M. Forcella, M.G. Leonardi, M. Casartelli, L. Fiandra, G.M. Hanozet and P. Parenti. 2002. A novel regulatory mechanism for amino acid absorption in lepidopteran larval midgut. J. Insect Physiol. 48: 585-592. Giovanelli,J,S.H.MuddandA.H.Datko. 1980. Sulfur amino acids in plants. Z2V BJ Miflin (ed.) The Bioclieinistry of Plants VoL 5, Academic Press, New York, pp. Giroux, S., R.M. Duchesne and D. Coderre. 1995. Predation of Leptmotarsa decemlimata (Coleoptera: Coccinellidae) by Coleomegilla maculata (Coleoptera: Coccmelhdae): Comparative effectiveness of predator developmental stages and effect of temperature. Environ. Entomol. 24: 748-754. Glare, TJR, and M. O'Callaghan, 1 998. Environmental and Health Impacts of Bacillus thurmgiensis isrealensis. Report for the New Zealand Ministry of Health, 58pp.

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no Nester, E.W., L.S. Thomashow, M. Metz and M. Gordon. 2002. One Hundred Years of Bacillus thruingiensis: A Critica Scientific Assessment Report from the American Academy of Microbiology. 16p. Onifade, A.A., O.O. Oduguwa, A.O. Fanimo, A.O. Abu, T.O. Olutunde, A. Arije and G. M. Babatunde. 2001. Effects of supplemental methionine and lysine on the nutritional value of housefly larvae meal (Musca domestica) fed to rats. Biores. Technol. 78: 191-194. Palumbo, R.E. and D.L. Dahhnan. 1978. Reduction of Manduca sexta fecundity and fertility by L-canavanine. J. Econ. Entomol. 71 : 674-676. Pan, M.L. and W.H. Telfer. 1996. Methionine-rich hexamerin and arylphorin as precursor reservoirs for reproduction and metamorphosis in female Luna moths. Arch. Insect Biochem. Physiol. 32: 149-162. Patterson, K.D. 1992. Yellow fever epidemics and mortality in the United States, 16931905. Soc. Sci. Med. 34(8): 855-865. Perfect, T. J. 1992. IPM in 2000, pp.47-53. IN A.A.S A. Kadir and H.S. Barlow (eds.), Pest Management and the Environment in 2000. CAB International, Oxford, UK. 401pp. Quick, M. and B.R. Stevens. 2001. Amino acid transporter CAATCH1 is also an amino acid-gated cation channel. J. Bio. Chem. 276(36): 33143-33418. Racioppi, J.V. and D.L. Dahhnan. 1980. Effects of L-canavanine on Manduca sexta (Sphingidae: Lepidoptera) larval hemolymph solutes. Comp. Biochem. Physiol 67: 35-39. Ragsdale, D. and E.B. Radcliffe. 1999. Colorado potato beetle management. University of Minnesota Cooperative Extension Service. Internet URL: http://ipmworld.umn. edu/arjhidalert/CPB~DWR.html. Accessed April 2004. Robertson, J.L. and H.K. Priesler. 1991. Pesticide bioassays with arthropods. CRC Press, Inc. Boca Raton, 127pp. Rock, G.C. 1971. Utilization of D-isomers of the dietary, mdispensable armno acids by Argyrotaenia velutinana larvae. J. Insect Physiol. 17: 2157-2168. Rock, G.C. and E. Hodgson. 1 971 Dietary amino requirements for Heliothis zea aetermined by dietary deletion and radiometric techniques. J. Insect Phvsiol 171087-1097. Rock, G.C, BG.Ligon, and E. Hogson. 1973. Utilization of methionine analoges by Argyrotaenia velutinana larvae. Ann. Entol. Soc. Amer. 66(1): 177-179.

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Ill Rock, G.C., A. Khan, and E Hodgson. 1975. The nutritional value of seven D-amino acids and a-keto acids for Argyrotaenia velutinana, He Hot his zea and Phormia regina. J. Insect Physiol. 21:693-703. Romoser, W.S. and J. G Stoffolano, Jr. 1998. The Science of Entomology, 4* edition. McGraw-Hill. Newyork, 605pp. Rosen, D., F.D. Bennett, and J.L Capinera. 1996. Prefece, pp. V-vi. IN D. Rosen, F.D. Bennett and J.L. Capinaera (eds.), Pest Management in the Subtropics: Biological Controla Florida Perspective. Intercept Limited, Andover, UK. 737pp. Rosenthal, G. A. 1977. The biological effects and mode of action of L-canavanine, a structural analogue of L-arginine. Q. Rev. Biol. 52(2): 155-178. Rosenthal, G.A. and D.L. Dahlman. 1975. Non-protein amino acid-insect interactions, n. Effects of canaline-urea cycle amino acids growth and development of the tobacco hornworm, Manduca sexta (L.) (Sphingidae). Comp. Biochem. Physiol 52: 105-108. Rosenthal, G.A. and D.L. Dahlman. 1988. Degradation of aberrant proteins by larval tobacco hornworm, Manduca sexta (L) (Sphingidae). Arch. Insect Biochem Physiol. 8: 165-172. Rosenthal, G.A.andD.L. Dahlman. 1991. Incorporation of L-canavanine into proteins and the expression of its antimetabolic effects. J. Ag. and Food Chem 39987990. Rosenthal, G.A., D.L. Dahlman, P.A. Crooks. S.N. Phuket, and L.S. Trifonov. 1995. Insecticidal properties of some derivatives of L-canavanie. J. Agric. Food Chem 43: 2728-2734. Rosenthal, G.A., D.L. Dahlman and D.H. Janzen. 1 976. A novel means for dealing with L-canavanine, a toxic metabolite. Science 192: 256-258. Rosenthal, G.A..D.L. Dahlman and D.H. Janzen. 1977. Degradation and detoxification of canavanine by a specialized seed predator. Science 196: 658-660. Rosenthal, G.A., D.L. Dahlman and D.H. Janzen. 1978. L-canaline detoxification: A seed predator's biochemical defense. Science 202: 528-529. Rosenthal, G.A., P. Nkomo and D.L. Dahlman. 1998. Effect of long-chained esters on the insecticidal properties of L-canavanine. J. Agric. Food Chem. 46(1): 296-299. Royer, T.A., K.L. Giles, S.D. Kindler and N.C. Elliott. 2001. Developmental response of three geographic isolates of Lysiphlebus testaceipes (Hymenoptera: Aphididae) to temperature. Environ. Entomol. 30(4): 637-641.

PAGE 124

112 Schoonhoven, L.M. 1972. Plant recognition by lepidopterous larvae. Insect/Plant Relationships, Symposium of the Roy. Entomol. Soc. London 6:83-93. Schardt, J.D. 1987. 1987 Florida Aquatic Flora Survey Report. Florida. Department of Natural Resources, Bureau of Aquatic Plant Management. Tallahassee, FL. 49 pp. Schuster, D.J, J.E. Funderburk and P.A. Slansly. 1996. IPM in tomatoes, pp. 387-408. IN D. Rosen, F.D. Bennett and J.L. Capinaera (eds.), Pest Management in the Subtropics: Integrated Pest ManagementA Florida Perspective. Intercept Limited, Andover, UK. 737pp. Singh, K.RP. and A.W.A. Brown. 1957. Nutritional requirements of Aedes aegypti. J. Insect Physiol. 1(1): 199-220. Sorenson, C.E., RL. Fery and G.G.Kennedy. 1989. Relationship between Colorado potato beetle (Coleoptera: Chrysomelidae) amd tobacco hornworm (Lepidoptera:: Sphingidae) resistance in Lycopersicon hirsutism f. glagratum. J. Econ. Entomol 82(4): 1743-1748. Stansly, P.A., IX. Liu, D.J. Schuster and D.E. Dean. 19%. Role of biorational insecticides in management of Bemisia, pp. 605-615. IND. Gerling and R T. Mayer, Jr. (eds.) Bemisia: 1995 Taxonomy, Biology, Damage Control and Management. Andover, Hants, UKD. 702pp. Stevens, B.R., D.H. Feldman, Z. Liu and W.R Harvey. 2002. Conserved tyrosine147 plays a critical role in the ligand-gated current of the epithelial cation/anion acid transporter/channel CAATCH1. J. Exp. Biol. 205: 2545-2553. Sugarman, D. and W. Jakinovitch, Jr. 1986. Behavioral gustatory responses of adult cockroaches, Periplaneta americana to D and L amino acids. J Insect Phvsiol 32(1): 35-41. Tobe,S.S. and N.Clarke. 1985. The effect of L-methionine concentration on juvenile hormone biosynthesis by corpora allata of the cockroach Diploptera punctata Insect Biochem. 1 5(2): 1 751 79. Tipping, P.W., C.A. Holko, A.A. Abdul-Baki, and J.R Aldrich. 1999. Evaluating Edovum puttier i Grissell and Podiscus maculfventris (Say) for augmentative biological control of Colorado potato beetle in tomatoes. Biol. Control 16: 35-42. Tzeng,D.D. 1988. Photodynamic action of methionine-riboflavin mixture and its application in the control of plant diseases and other plant pests. Plant Protection Bulletin 30(2): 87-100. Tzeng,DD.andJ.E.Devay. 1989. Biocidal activity of mixtures of riboflavin against tlQ^SSSu 311(1 baCteria 311(1 P SSible m deS f aCti n MycoI 8 ia

PAGE 125

113 Tzeng,D.D., M.H. Lee, K.R. Chung and J.E.Devay. 1990. Products in light-mediated reactions of free methionine-riboflavin mixtures that are biocidal to microorganisms. Can. J. Microbiol. 36(7): 500-506. Wadsworth, D.J. 1995. Animal health products, pp. 257-284. Z7VC.R.A. Godfrey (ed.) Agrochemicals From Natural Products, Marcel Dekker, New York. 424pp. Walker, T.J., J J. Gaffhey, A.W. Kidder and A.B. ZifTer. 1993. Florida Reach-Ins: Environmental chambers for entomological research. American Entomologist 39(3): 177-182. Weinzierl, R., T. Henn and P.G. Koehler. 1998. Microbial insecticides. ENY-275 Cooperative Extension Service, Institute of Food and Agricultural Services University of Florida. 13pp. Wheeler, D.E., L Tuchinskaya, N.A. Buck and B.E. Tabahnik. 2000. Hexameric storage proteins during metamorphosis and egg production in the diamondback moth, Plutellaxylostella (Lepidoptera). J. Insect. Physiol. 46: 951-958. Womack, M. 1 993. The yellow fever mosquito, Aedes aegypti. Wing Beats 5(4): 4. Wright, R. 1995. Know Your Friends: Wasp Parasites of Greenbugs. Midwest Biological Control News Online, 11:9. Young, V.R. and A.E. El-Khoury. 1996. Human amino acid requirementsA reevaluation. Food and Nutrition Bulletin 17(3): 191-203. Zar,J.H. 1999. Biostansrical Analysis, 4th ed. Prentice Hall. New Jersey. 663pp. Zeh, M., A.P. Casazza, G Kreft U. Roessner, K. Bieberich, L. Willmitzer, R. Hoefgen and H. Hesse. 2001. Antisense inhibition of tlireonine synthase leads to high methionine content in transgenic potato plants. Plant Physiol. 127792-802

PAGE 126

BIOGRAPHICAL SKETCH Lewis Scotty Long was born in Calhoun, Georgia on August 20, 1971 He graduated from Madisonville High School (Madisonville, Tennessee) in May 1989. On a biology scholarship, Lewis attended Middle Tennessee State University (MTSU), where he earned his BS in May 1994. On graduation, he took a job as an aquatic biologist for Aquatic Resources Center (Franklin, Tennessee). Lewis worked there specializing in taxonomy of mayflies, stoneflies, caddisflies, and freshwater molluscs (snails and mussels). While still employed at Aquatic Resources Center, he started his graduate studies in 1996 at MTSU and continued the work he had started during his undergraduate years. In May of 1 999, Lewis graduated with his MS. After receiving his MS, Lewis moved to Florida and entered the PhD program at the University of Florida, Department of Entomology and Nematology. He worked with Dr. Bill Peters (Florida A&M University) on the worldwide taxonomic revision of an understudied group of mayflies. However, Dr. Peters unexpectedly passed away in 2000, and Lewis took this unfortunate event as a chance to broaden his expertise in entomology. In 2000, he took a part-time job with Drs. James Cuda and Bruce Stevens on research that was in the patent process. This was the research that Lewis undertook for his dissertation. Lewis also served as a teaching assistant for the department for classes such as Bugs and People, Life Sciences for Education Majors, Principles of Entomology, and Medical and Veterinary Entomology. He served as primary instructor for Insect Classification and Immature 114

PAGE 127

Insects. Lewis, along with fellow graduate student Jim Dunford, were awarded the Outstanding Teacher Award by the Entomology and Nematology Student Organization of the University of Florida for outstanding teaching accomplishments in the department. While at the University of Florida, Lewis joined the U.S. Army Reserve as a medical entomologist. He was assigned to the local Medical Detachments, and served there from 2000 to 2004. Originally he had planned on graduating in 2003, but was called to active duty with the 1469 th Medical Detachment as a part of Operation Enduring Freedom (OEF). Lewis was the OEF Theater entomologist, and served as the Executive Officer (responsible for the deployment of personnel and equipment to South West Asia). He was stationed at Kandahar Airfield, where he performed his duty and was awarded an Army Commendation Medal for his work in protecting soldiers from health hazards and diseases associated with the area. Lewis returned and continued his work toward graduation. Lewis was married in August 1992 to Karen Abbott, and is the father of Emilia Irene (1994) and Bryan Scott (1997). Lewis plans on having a career in the military as a medical entomologist, and all look forward to seeing the world and the rest of their future.

PAGE 128

I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. Jamas P. Cuda, Chair Assisrant Professor of Entomology and Nematology I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. j Bruce R. Stevens, Cochair Professor Physiology and Functional Genomics I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. George ASJ§erencser Professor of Physiology and Functional Genomics I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. Jarnies E. Maruniak Associate Professor of Entomology and Nematology I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. Simon S.J. Yu Professor of Entomology and Nematology

PAGE 129

I certify that I have read this study and that in my opinion it conforms to acceptable standards of scholarly presentation and is fully adequate, in scope and quality, as a dissertation for the degree of Doctor of Philosophy. Susan E. Webb Associate Professor of Entomology and Nematology This dissertation was submitted to the Graduate Faculty of the College of Agricultural and Life Sciences and to the Graduate School and was accepted as partial fulfillment of the requirements for the degree of Doctor of Philosophv. May 2004 Dean, College of Agricultural Sciences Dean, Graduate School


Table 5-4. Impact of Changes in U.S. Price Stabilization Policy on Pay-offs to the Industries and Nations Welfare [Billion US$]
- Flexible Quota Allocations -
United States
Mexico
Scenarios
HFCS
industry
revenue
Sugar
industry
revenue
Welfare
(A)
Adjusted
welfare
(A+B-C-D)
Tariff revenue
from Mexican
sugar
(B)
Cost of
price
support
(C)
Cost of
buying up
excess sugar
(D)
Net cost
(B-C-D)
Sugar
industry
revenue
Welfare
Baseline
(status quo)
5.684
(100)
35.713
(100)
354.486
(100)
354.370
(100)
0.093
(100)
0.208
(100)
0
( )
0.116
(100)
21.737
(100)
74.443
(100)
Scenario 4
("PA S F")
35.367
(99.03)
353.981
(99.86)
354.080
(99.92)
0.297
(320.15)
0.198
(95.11)
0
(-)
-0.099
(-85.43)
21.314
(98.05)
72.293
(97.11)
Scenario 8
("PA B F")
10.132
(178.27)
35.206
(98.58)
354.744
(100.07)
354.850
(100.14)
0.297
(320.15)
0
( )
0.191
( )
-0.106
(-91.62)
21.315
(98.06)
72.293
(97.11)
Scenario 12
("PA C F")
33.928
(95.00)
355.035
(100.15)
355.438
(100.30)
0.403
(434.64)
0
( )
0
( )
-0.403
(-348.70)
20.515
(94.38)
66.757
(89.68)
P = Mexico high production; A= Mexico high HFCS adoption; PA= Mexico high production-high HFCS adoption; T= Mexico tax on
HFCS; S= U.S. price support; B= U.S. buying up excess sugar; C= U.S. production control; F= U.S. flexible quota allocation; and M=
U.S. minimum quota allocations to the rest of the world.


CHAPTER 5
EFFECTS OF L-METHIONINE ON SURVIVAL AND DEVELOPMENT OF THE
YELLOW FEVER MOSQUITO, Aedes aegypti, UNDER LABORATORY
CONDITIONS
Introduction
Integrated Pest Management practices are not restricted to agricultural pests.
Medically important insect pests are responsible for epidemics that have changed the
course of human existence, from bubonic plague spread by the Oriental rat flea
(Xenopsylla cheops Rothschild (Siphonaptera: Pulicidae)), to malaria carried by
anopheline mosquitoes. One medically important species that has had a significant
impact on human existence is the yellow fever mosquito (YFM), Aedes aegypti (L.)
(Dptera: Culicide). This cosmopolitan species is found worldwide and is the primary
vector for human dengue and yellow fever despite concerted efforts at eradication in the
United States (Womack, 1993). In the United States alone, upwards of 150,000 lives
were lost to yellow fever in the period starting in the late 18th century and into the early
20th century (Patterson, 1992). However, because of the development of a vaccine,
yellow fever has been replaced by Dengue which is now second only to malaria as a
worldwide threat (Gubler, 1998). Because Dengue fever is also vectored by the YFM, it
poses a risk by affecting tens of millions of people worldwide (Gubler and Clark, 1995).
The inclusion of the YFM in this study was an effort borne of curiosity because of
the lack of knowledge of the CAATCH1 system in other insects and the availability of
specimens for study. Mosquito larvae are particulate feeders and have dietary
52


Table 5-5. Impact of Changes in U.S. Price Stabilization Policy on Pay-offs to the Industries and Nations Welfare [Billion US$]
- Minimum Quota Allocations -
United States
Mexico
Scenarios
HFCS
industry
revenue
Sugar
industry
revenue
Welfare
(A)
Adjusted
welfare
(A+B-C-D)
Tariff revenue
from Mexican
sugar
(B)
Cost of
price
support
(C)
Cost of
buying up
excess sugar
(D)
Net cost
(B-C-D)
Sugar
industry
revenue
Welfare
Baseline
(status quo)
5.684
(100)
35.713
(100)
354.486
(100)
354.370
(100)
0.093
(100)
0.208
(100)
0
( )
0.116
(100)
21.737
(100)
74.443
(100)
Scenario 13
("PA S M")
33.480
(93.75)
344.115
(97.07)
338.136
(95.42)
0.133
(142.87)
6.112
(2,931.51)
0
( )
5.979
(5,168.80)
12.159
(55.94)
69.095
(92.82)
Scenario 14
("PA B M")
10.132
(178.27)
27.886
(78.08)
356.639
(100.61)
354.819
(100.13)
0.180
(194.04)
0
( )
2.000
( )
1.820
(1,573.52)
14.532
(66.86)
69.882
(93.87)
Scenario 15
("PA-C-M")
26.516
(74.25)
357.077
(100.73)
357.277
(100.82)
0.200
(215.06)
0
( )
0
(-)
-0.200
(-172.54)
13.983
(64.33)
64.393
(86.50)
P = Mexico high production; A= Mexico high HFCS adoption; PA= Mexico high production-high HFCS adoption; T= Mexico tax on
HFCS; S= U.S. price support; B= U.S. buying up excess sugar; C= U.S. production control; F= U.S. flexible quota allocation; and M=
U.S. minimum quota allocations to the rest of the world.


98
cNcO''tm'Or--oo ONO^csm^tm
oooooSoooooooo
(N (N (N (N (N (N CN CN CN Year
- U.S. price A Mexico price
Figure 5-11. Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets
(Scenario 12 PA-C-F)
* U.S. price 6 Mexico price
Figure 5-12. Forecasted Equilibrium Sugar Prices in the U.S. and Mexican Markets
(Scenario 16 T-S-F)


112
Schoonhoven, L.M. 1972. Plant recognition by lepidopterous larvae. Insect/Plant
Relationships, Symposium of the Roy. Entomol. Soc. London 6:83-93.
Schardt,J.D. 1987. 1987 Florida Aquatic Flora Survey Report. Florida. Department of
Natural Resources, Bureau of Aquatic Plant Management. Tallahassee, FL. 49 pp.
Schuster, D.J, J.E. Funderburk and P.A. Slansly. 1996. IPM in tomatoes, pp. 387-408.
IN D. Rosen, F.D. Bennett and J.L. Capinaera (eds.), Pest Management in the
Subtropics: Integrated Pest Management- A Florida Perspective. Intercept
Limited, Andover, UK. 737pp.
Singh, K.R.P. and A.W.A. Brown. 1957. Nutritional requirements of Aedes aegypti. J.
Insect Physiol. 1(1): 199-220.
Sorenson, C.E., R.L. Fery and G.G. Kennedy. 1989. Relationship between Colorado
potato beetle (Coleptera: Chrysomelidae) amd tobacco homworm (Lepidoptera::
Sphingidae) resistance in Lycopersicon hirsutum f. glagratum. J. Econ. Entomol.
82(4): 1743-1748.
Stansly, P.A., T.X. Liu, D.J. Schuster and D.E. Dean. 1996. Role of biorational
insecticides in management of Bemisia, pp. 605-615. IN D. Gerling and R. T.
Mayer, Jr. (eds.) Bemisia: 1995 Taxonomy, Biology, Damage Control and
Management. Andover, Hants, UKD. 702pp.
Stevens, B.R., D.H. Feldman, Z. Liu and W.R. Harvey. 2002. Conserved tyrosine-147
plays a critical role in the ligand-gated current of the epithelial cation/anion acid
transporter/channel CAATCH1. J. Exp. Biol. 205: 2545-2553.
Sugarman, D. and W. Jakinovitch, Jr. 1986. Behavioral gustatory responses of adult
cockroaches, Periplaneta americana to D and L amino acids. J. Insect Physiol.
32(1): 35-41.
Tobe, S.S. and N. Clarke. 1985. The effect of L-methionine concentration on juvenile
hormone biosynthesis by corpora allata of the cockroach Diploptera punctata.
Insect Biochem. 15(2): 175-179.
Tipping, P.W., C.A. Holko, A.A. Abdul-Baki, and J.R. Aldrich. 1999. Evaluating
Edovum puttleri Grissell and Podiscus maculiventris (Say) for augmentative
biological control of Colorado potato beetle in tomatoes. Biol. Control 16: 35-42.
Tzeng, D.D. 1988. Photodynamic action of methionine-riboflavin mixture and its
application in the control of plant diseases and other plant pests. Plant Protection
Bulletin 30(2): 87-100.
Tzeng, D.D. and J.E. Devay. 1989. Biocidal activity of mixtures of riboflavin against
plant pathogenic fungi and bacteria and possible modes of action. Mycologia
81(3): 404-412.


5-6
Pay-off Matrix for the Trade Policy Game [Billion US$]
107
5-7 Indexed Pay-off Matrix for the Trade Policy Game without Coalitions [Baseline =
100] 108
5-8 Indexed Pay-off Matrix for the Trade Policy Game Played by Two Coalitions of
Countries without the U.S. HFCS industry [Baseline=100] 109
5-9 Indexed Pay-off Matrix for the Trade Policy Game Played by the Industry
Coalition and the Government Coalition without the U.S. HFCS industry
[Baseline=100] 110
5-10 Indexed Pay-off Matrix for the Trade Policy Game Played by the Grand Coalition
without the U.S. HFCS industry [Baseline=100] 111
5-11 Indexed Pay-off Matrix for the Trade Policy Game Played by Two Coalitions of
Countries with the U.S. HFCS industry [Baseline=100] 112
5-12 Indexed Pay-off Matrix for the Trade Policy Game Played by the Industry
Coalition and the Government Coalition with the U.S. HFCS industry
[Baseline=100] 113
5-13 Indexed Pay-off Matrix for the Trade Policy Game Played by the Grand Coalition
with the U.S. HFCS industry [Baseline=100] 114
C-l Coefficients for Inverse Linear Functions -U.S.- 129
C-2 Coefficients for Inverse Linear Functions -Mexico- 130
viii


I certify that I have read this study and that in my opinion it conforms to
acceptable standards of scholarly presentation and is fully adequate, in sqqpe and quality,
as a dissertation for the degree of Doctor of Philosophy.
Jamefe P. Cuda, Chair
Assistant Professor of Entomology and
Nematology
I certify that I have read this study and that in my opinion it conforms to
acceptable standards of scholarly presentation and is fully adequate, in scope and quality,
as a dissertation for the degree of Doctor of Philosophy.
Bruce R. Stevens, Cochair
Professor Physiology and Functional
Genomics
I certify that I have read this study and that in my opinion it conforms to
acceptable standards of scholarly presentation and is fully adequate, in scope and quality,
as a dissertation for the degree of Doctor of Philosophy.
Professor of Physiology and Functional
Genomics
I certify that I have read this study and that in my opinion it conforms to
acceptable standards of scholarly presentation and is fully adequate, in scope and quality,
as a dissertation for the degree of Doctor of Philosophy.
Jane^ E. Maruniak
Associate Professor of Entomology and
Nematology
I certify that I have read this study and that in my opinion it conforms to
acceptable standards of scholarly presentation and is fully adequate, in scope and quality,
as a dissertation for the degree of Doctor of Philosophy.
Simon S.J. Yu
Professor of Entomology and
Nematology


67
Jaffe and Chrin (1979) found the adults of YFM females infected with Brugia, a
filiaral parasite, were depleted of free form methionine because of the infection and were
able to make up the difference by converting homocysteine to methionine with a special
synthetase. The ability of YFM adults to synthesize methionine from homocysteine may
be present in the larvae as well. This could be the result of the lack of methionine in the
diet and possible evidence of the CAATCH1 system being present in at least the adult
stage. The susceptibility of the larvae to L-methionine also could be the result of
overexposure to a compound that is normally not encountered in high concentrations
(>0.1%). However, the alkalinity of the particulate feeding larvae and the high mortality
to L-methionine suggests that the CAATCH1 system is present and could be exploited in
other species with similar midgut characteristics (Dadd, 1975).
The survival of YFM larvae exposed to both Beta-alanine and L-leucine was
unusual in that they each had the opposite effect on the YFM larvae. L-leucine was
expected to have similar blocking properties as L-methionine based on CAATCH1
research (Feldman et al., 2000). Instead, almost no mortality was observed indicating the
possibility of another system involved with the transport of this amino acid. Conversely,
beta-alanine was not found to be reactive with the CAATCH1 system based on the work
of Feldman et al. (2000). The unusually high larval mortality associated with this amino
acid may be the result of a yet to be discovered midgut property.
The similar mortalities observed for the higher concentrations of L-methionine
and Bti is encouraging considering the resistance to this compound that has been
documented in many insect species because of reduced receptor activity and binding
(Bills et al., 2004; Nester et al., 2002). Resistance in insects involves a variety of


12
No Amino Acid
Proline
Methionine
Figure 2. The CAATCH1 system identified from the midgut of the tobacco
homworm (modified from Quick and Stevens 2001). In the
presence of no amino acids, ion flow is similar for both K+ and
Na+ (A). With the addition of an amino acid, flows are changed.
When proline is added (B), the transport occurs but the binding of
the amino acid increases the ion flow, notably Na+. However,
when methionine is added (C) transport occurs and the binding of
the amino acid greatly decreases the flow of Na+ while K+ is
increased


74
Table 4-10. Data Sources for Miscellaneous
Data
Source
U.S. Consumer Price Index (CPI) (1982-
1984=100)
Database by the Bureau of Labor
Statistics, U.S. Department of Labor
Mexico Consumer Price Index (CPI)
(1994=100)
Database by the Banco de Mexico
Exchange rate (US$- Mexican pesos)
Quota-tariff rates
Agricultural Outlook (1999) by Economic
Research Service, U.S. Department of
Agriculture
Sugar per unit transportation cost (bagged,
seaborne freight rate)
Personal communication with an exporter
of sugar
Loan rate for raw sugar in the U.S.
Haley and Suarez (2002)
Guarantee price for raw sugar in Mexico
Database by Comite de la Agroindustria
Azucarera (COAAZUCAR)


Table 5-13. Indexed Pay-off Matrix for the Trade Policy Game Played by the Grand Coalition with U.S. HFCS industry
[Baseline=100]
Maintain the current policy (status quo)
Grand coalition
High production
Grand coalition
Mexico's strategies
High HFCS adoption
Grand coalition
High production High HFCS adoption
Grand coalition
U.S.
strategies
Maintain
price
support
(status
quo)
Buying up
excess
sugar in
the market
Production
control
with
Mexico
100
99.16
99.18
99.74
98.88
99.04
99.08
100.25
100.38
98.95


APPENDIX B
CORN STATISTICS
Figure B-l shows the recent com production and consumption for selected
countries. Currently, the United States is the largest producer as well as consumer of
com, followed by China. However, in the 2002/2003 season, China became a net
producer. Trends show that production in China has been increasing for the last three
seasons while that of U.S. has been decreasing.
Figure B-2 shows the transition of food and industrial com use in the United States
during the past two decades. The largest industrial use for com is for fuel alcohol,
representing approximately 41 percent of total use in 2002. HFCS is the second largest
use, representing approximately 24 percent of total use. HFCS use has been level in the
last five years, while fuel alcohol use exhibits rapid growth.
Figure B-3 shows transition of the U.S. com price (No.2 Yellow) in Chicago
market during the past two decades. The price shows some fluctuation around 2.5 US
dollars per bushel.
Figure B-4 shows U.S. exports of products made from com in 2002 expressed in
million US dollars. Com gluten feed, com gluten meal, and com oil are the dominant
products made from com. When three kinds of fructose (fructose solids containing more
than 50% fructose, Chemically pure fructose, and fructose syrup with 50%+ fructose) are
aggregated, the total value becomes fourth largest, following com oil. If HFCS is already
saturated in the United States, it will have better marketing opportunities overseas;
however, the business size still will be small relative to that for the leading products.
125


88
relatively small, the choice by the Mexican government is made at the expense of its own
sugar industry. On the contrary, the industry coalition would lobby for the U.S.
production control strategy, assuming that the Mexican sugar industry promises to
compensate the loss of US $0.01 billion (US $10 million) bom by the U.S. sugar industry
with expected gain of US $1.27 billion (Table 5-6). With this contrast of the results
between a country coalition (Table 5-8) and a govemment/industry coalition (Table 5-9),
the U.S. government would likely form a coalition with its own industry while the
Mexican government would try to form a coalition with the U.S. government.
Results from the game played by the grand coalition that includes all players except
the U.S. HFCS industry are shown in Table 5-10. The solution is the Mexican
government high production strategy and the U.S. government buying excess sugar
strategy. Although this solution does not satisfy pareto optimality for all four payees, the
overall loss is minimal: the loss incurred by the U.S. sugar industry is US$ 0.31 billion
(Table 5-6) and is theoretically compensated by the total gain of US$ 2.29 billion (US$
0.36 billion from the U.S. government, US$ 0.13 billion from the Mexican sugar
industry, and US$ 1.80 billion from Mexicos welfare, Table 5-6) This solution
corresponds to the govemment/industry coalition seen in Table 5-9. In other words, if the
U.S. government sees the benefit from pooling gains with Mexico rather than with its
industry and if redistribution of gains is possible among governments and industries, no
one loses from the game. Such an arrangement and agreement in practice would be
expected to be difficult to reach and implement.
When the U.S. HFCS industry is included in coalitions, the same solutions are
reached: the combination of Mexican government high production strategy and U.S.


50
combination of the early consumption of the treated disks and mortality occurring after
48 hours, when a lower concentration is required for mortality. The larvae could have
fed on the treated disks and then switched to the Control based on a physiological cue.
Mitchell (1974) and Mitchell and Schoonhoven (1974) examined the taste receptors of
CPB and found physiological and behavioral responses to some amino acids, mainly
gamma aminobutyric acid (GABA) and alanine. They discussed the possibility that host
selection in solanaceous plants may have been the result of these chemosensory structures
and the concentration of amino acids in the leaves. It should be noted that both studies
excluded methionine and no electrophysiological data were collected on the response of
CPB to this amino acid. This is not surprising considering the fact that the diet of the
CPB is low in methionine and therefore would not be a candidate for the inclusion in
feeding stimulatory studies (Cibula et al. 1967). It is unknown if these sensory structures
can detect methionine and possibly act as a means to avoid plant material high in this
amino acid. This appears to be contradicted by the data in Figure 4-5, in which there was
no difference between the treatments. The larvae feeding on the Control treatment,
consuming the majority and then moving to the 1.0% L-methionine treatment, could
explain the lack of difference.
There are some differences between some of the Feeding and Development
treatments should be noted. The mean head capsule of the larvae in the 0.5%
L-methionine treatment was higher than the 0.3% L-methionine treatment while the
amount of leaf material consumed for the same treatment were the same indicating
another factor involved with the greater head capsule width. The differences could be the
result of the larger size of females and possibly could have included more females.


35
Theoretical Framework
Sweetener Market Analysis
The sweetener market in a country that consumes both sugar and HFCS can be
expressed as follows:
QdSUGAR =fl (PSUGARZ,)
(3.1)
Of1 HFCS -f2 (PhFCSZ2)
(3.2)
&SUGAR = hi (PSUGAR, Wj)
(3.3)
0sHFCS = h2 (PHFCS,, W2)
(3.4)
Q?SUGAR ~ Q'SUGAR
(3.5)
QhFCS = Q? HFCS
(3.6)
where Q? is aggregate quantity demanded, Qs is aggregate quantity supplied, P is a price,
Z and W are vectors of other factors that influence aggregate demand and supply of sugar
or HFCS, respectively. Equations [3.5] and [3.6] depict market clearing conditions for
each commodity.
Sweetener demand
Sweetener demand is defined based on consumer demand theory derived from
utility maximizing behavior. Following Varians demonstration (1992), aggregate sugar
demand is derived from maximizing utility of aggregated consumers, including industry
sugar users, by purchasing sugar:
max u ( Qsugar Qx)
s.t. Qsugar *Psugar + Qx*Px = m (3.7)
where Q is the quantity consumed, X represents all other goods consumed, Pisa price,
and m is national income. By solving maximization problem, the aggregate demand
function is expressed as:


6-5. Mortality of Colorado potato beetle larvae on eggplants treated with L-methionine
and Silwett L-77 87
7-1. Mortality of Coleomegilla maculata adults after exposure to L-methionine treated
artificial diet 88
7-2. Mortality of Coleomegilla maculata adults after exposure to L-methionine treated
cotton plant leaves infested with aphids 89
7-3. Feeding scars on water hyacinth {Eichhornia crassipes) leaf after exposure to
Neochetina eichhorniae adults 90
7-4. Mortality of Neochetina eichhorniae on treated water hyacinth leaves 91
7-5. Feeding rate of Neochetina eichhorniae on water hyacinth leaves treated with
L-methionine and Proline 92
7-6. Lysephlebius testiceipes parasitized aphids on cotton plants treated with
L-methionine 93
x


Figure 5-18. Absolute Effects of Production Improvement and HFCS Adoption on Pay-off to Mexicos Welfare


84
world. As a result of cheap sugar from the rest of the world flowing into the U.S. market,
the U.S. price will fall far below the support price. The loss bom by the Mexican sugar
industry particularly stands out: nearly a half of its expected revenue disappears due to
restricted access to the U.S. market. When fostering the sugar industry in its own country
and Mexico as a neighboring trade partner, the U.S. government needs to accept sugar
from Mexico more than from the rest of the world. This requires sensitive negotiations
among exporters.
Overall, the Mexican sugar industry and welfare are more prone to changes in the
Mexican sweetener market and U.S. sugar policy than the U.S. sugar industry and U.S.
welfare. Among tested changes, Mexicos HFCS adoption and U.S.s quota allocation
policy hold the greatest effects.
Effects of Mexicos production improvement and HFCS adoption on the Mexican
sugar industry and welfare are illustrated in Figures 5-17, 5-18, and 5-19. The rate of
production improvement ranges from the average rate realized between 1997 and 2001
(baseline) to additional 0.5, 1.0, and 1.5 percent to the average rate; the HFCS adoption
ranges from 25 percent share of indirect sweetener consumption (baseline) to 30,40, 45,
and 50 percent (HFCS adoption situation). The effects on the Mexican sugar industry are
shown in Figure 5-17. Expected accumulated revenue increases as sugar production
improves an additional 1 and 1.5 percent above the average rate. An increase in HFCS
adoption also brings about an increase in revenue until the share attained by HFCS climbs
up to 40 percent. When HFCS share reaches 50 percent, revenue shrinks compared to the
baseline. Increased revenue caused by HFCS adoption is due to higher domestic sugar
price and decreased revenue is due to decreased quantity of sugar demanded. The effects


69
Table 4-3. Assumptions for U.S. Sugar Policies
Categories
Policies
(code)
Assumptions
Stabilization
of the
demand price
Price support
(status quo)
(S)
The U.S. government maintains the price support
at 18 cents per pound (loan rate, 396.48 US$ per
MT).
Buying up excess
sugar in the market
(B)
The U.S. government abandons the price support
and buys up excess sugar in the market instead.
Production control
(C)
The U.S. government abandons the price. Instead,
the U.S. and Mexican governments collectively
control the sugar production in such a way that
the sum of quantities demanded in two countries
is primarily met by the sum of the quantities
supplied from two countries.
Quota
allocations
Flexible allocation
(status quo)
(F)
The U.S. government allocates quotas in a
flexible manner between Mexico and the rest of
the world.
Minimum quota
allocations for the
rest of the world
(M)
The U.S. government reserves the minimum
quotas (the remaining minimum import
requirement less allocated to Mexico) for the rest
of the world.


7
framework employed to analyze U.S.-Mexico sugar trade is presented. In chapter 4 and 5,
empirical procedures as well as data set used in the study and the results form the
empirical study are presented. Lastly, conclusions and implications for policy are
discussed in chapter 6.


Table 5-10. Indexed Pay-off Matrix for the Trade Policy Game Played by the Grand Coalitions without the U.S. HFCS industry
[Baseline=100]
Mexico's strategies
Maintain the current policy
(status quo)
High production
High HFCS adoption
High production -
High HFCS adoption
U.S.
HFCS
U.S.
adjusted
welfare and
Mexico's
welfare
U.S. and
Mexican
sugar
industries
U.S.
HFCS
U.S. L
adjusted ^
welfare and
Mexico's
welfare
'.S. and
fexican
sugar
dustries
U.S.
HFCS
U.S.
adjusted
welfare and
Mexico's
welfare
U.S. and
Mexican
sugar
industries
U.S.
HFCS
U.S.
adjusted
welfare and
Mexico's
welfare
U.S. and
Mexican
sugar
industries
U.S.
strategies
Maintain
price
support
(status
quo)
100
100
-
100.27
-
97.96
-
99.34
Buying up
excess
sugar in
the market
-
99.15
-
f
-
98.11
-
99.47
Production
control
with
Mexico
-
99.17
-
99.74
-
98.16
-
98.02


76
annual crop. These results reflect these differences and imply that sugar beet production
is more sensitive to price changes. Although cane sugar production is assumed to respond
to raw sugar price, the coefficient of the price variable is not significant. Estimated price
elasticities were both inelastic for total sugar supply and beet sugar supply as anticipated.
The long-run supply price elasticities were calculated dividing the estimated price
elasticities (short-run) by (1- A.) where A. represents the estimates for production in the
previous year (autoregressive term). Computation yielded all inelastic long-run price
elasticities: 0.3875 and 0.6764 for total sugar and beet sugar, respectively.
Results for the demand and supply analysis for Mexico are summarized in Table 5-
2. In the demand equation, signs of significant estimates associated with each variable
were consistent with a priori expectations. The only statistically significant estimate
among the three price elasticities was direct consumption, and it was inelastic. The
population variable accounted for most of the explanatory power of consumption in all
models. Significant estimates associated with GDP in indirect sugar and total sweetener
consumption indicate that consumers tend to consume more sugar through sugar-
containing products as their income increases.
In the supply equation for Mexico, the signs of estimates associated with each
variable corresponded with a priori expectations. The estimate associated with price was
inelastic. While reduction in production cost and factory downtime indicated an increase
in production, the length of sugarcane harvest duration was almost perfectly correlated to
sugar output from the mills. The coefficient for the variable representing sugar loss
during the process was not significantly related to sugar output implying that the degree
of sugar loss was not as critical as other factors such as production cost and factory


11
In the 2001/02 crop season, average sugarcane yield was 70.32 MT per hectare in
Mexico, yielding 4,872,388 MT of sugar (CO A AZUCAR, 2003a). Irrigation is one of the
factors which influences cane yield; however, irrigation systems are found only in the
area where less rainfall is expected (Figure 2-3): 30 percent of total sugarcane area has no
irrigation system and 25 percent has full irrigation system (COAAZUCAR, 2003c).
Harvest is the most labor-intensive part of sugarcane production; the harvest season
lasts for about six months starting between November and January and ending in June in
most regions, depending on weather and size of enterprise. Harvest competes for
growers labor with other winter crops since many growers are also engaged in
production of crops such as maize, vegetables and citrus. Most of the harvest is carried
out manually; only 9 percent of total sugarcane processed at mills is harvested by
machine; 27 out of 60 mills do not employ a machine harvester at all; however, a cane
loader is used in most cases (COAAZUCAR, 2003d). Since the mills own machine
harvesters, cane loaders, and trucks, growers do not need to own them; however, it means
growers have no means to harvest and sell their sugarcane without the mills assistance
and coordination. Similar situations regarding growers capacity in harvest are found in
other crops such as citrus.
Upon harvest, sugarcane is bought by mills from growers and processed into sugar.
Sugarcane quality is vital to the sugar production process; high sucrose content cane
leads to high sugar production. Yet, it is often the case in Mexico that trucks endure long
waiting times to unload cane due to limited milling capacities. The average wait time
observed in 2001 was almost 30 hours across mills (COAAZUCAR, 2003e). The longer
trucks wait, the lower the quality of sugarcane becomes. Scattered and fragmented


60
alternatives to the current price support. In one policy, instead of directly supporting
sugar price, the government indirectly supports a sugar price through buying up excess
sugar in the market to assure that sugar price will not fall due to excess supply from
overseas. In this assumption, the government buys up as much sugar as the net sugar
import, i.e. total sugar import less minimum import requirement. In the bilateral trade
model, the cost of buying up excess sugar is not included in the objective function,
assuming that the government does not spend ex ante cost. Also for simplicity, sugar
storage costs incurred by the government are ignored. The second policy option is to
introduce sugar production controls in both the United States and Mexico. This option
requires cooperation from the Mexican sugar industry: defection by either party will
result in an unsuccessful outcome. In this way, both the United States and Mexico are
assumed to control their sugar production according to forecasted demand, avoiding
excess supply of sugar. In other words, the idea implies that the United States is willing
to import sugar from Mexico as long as Mexico cooperates to reduce its sugar production
to meet the sugar demand in both countries and also that Mexico can avoid excess surplus
sugar which cannot be sold anywhere except in the world market. This sugar policy does
not involve financial support from the government: the cost of the program is zero.
U.S. policy levers related to quota allocations to exporters are treated with two
different approaches: the U.S. government allocates import quotas in a flexible manner
between Mexico and the rest of the world (status quo); and minimum quotas are
maintained (the remainder of the minimum import requirement less that allocated to
Mexico) to the rest of the world no matter how much Mexico exports to the United


52
Mexican Sweetener Supply Model
Analysis of the aggregate supply of sugar in Mexico is conducted in a similar
manner as the U.S. supply analysis except that Mexico produces sugar only from
sugarcane.
In(QS,) = MM] + MM2*In(P D,.¡) + MM3*ln(COSTt) + MM4*In(DTt)
+ MM5*In(SUGLOSS,) + MM6*In(DURTN,) +MM7*TREND,+ww,
(4.8)
where Q ss are total sugar quantity produced [MT], P d is real wholesale standard sugar
price in the previous year (deflated by CPI [pesos / Kg]), COST is real average
production cost per ton of sugar realized at sugar mill deflated by CPI [pesos], DT is
average mill operation downtime ratio observed at sugar mill [%], SUGLOSS is average
sugar loss ratio observed during sugar production process at mill [%], DURTN is average
duration of the harvest in each season [days], TREND is a trend variable that represents
technology advancement; and ww, is error term. Price in the previous year is used
assuming that decision-making on sugar production relies on sugar crop production,
recognizing growers decide their production plan with the price realized in the previous
year. An autoregressive term (production in the previous year) is not added because of
limited length of data available for regression. It is expected that the elasticities
associated with price production cost (MM3), downtime (MM4), and sugar loss (MM3) to
be negative and the others to be positive. Price elasticities are expected to be inelastic.
Previously reported own- price elasticities for Mexican sugar supply at industry level is
0.67 by Petrolia and Kennedy (2002). Borrel (1991) estimated two sugarcane price
elasticities by regressing sugarcane yield on sugar cane price in an Almon polynomial
distributed lag model and by regressing sugarcane acreage on sugarcane price. From the


22
manufacturers (Moss and Schmitz, 2002). The other entity in the U.S. sweetener market
is Coalition for Sugar Reform (CSR), the industrial sugar users coalition. CSR opposes
U.S. sugar policy, but has been unsuccessful at bringing a lower sugar price to the market
from which industrial sugar users as well as consumers would benefit. Many studies have
shown that producers clearly gain while consumers and industrial users lose in the U.S.
sugar program: an analysis conducted by the General Accounting Office (GAO, 2000)
indicates that food manufacturers could be substantial gainers from elimination of the
sugar program (Moss and Schmitz, 2002).
A history of sugar and HFCS prices are shown in Figure 2-12. Both sugar and
HFCS prices exhibit a continuing declining trend. The decline in both U.S. domestic and
export prices of the HFCS price is due to sophistication of wet-milling technology in
combination with decreases in the tariff schedule under NAFTA for the latter. HFCS has
been marketed at a lower price than raw sugar in the U.S. market. Although the U.S.
HFCS industry sells at a lower price than sugar, it still benefits from the sugar program
because the price of sugar is maintained higher than it would be without the program.
This structure confirms why the HFCS industry has been supporting the sugar program as
a member of ASA; however, an opportunity for the HFCS industry to increase marketing
overseas such as in Mexico may weaken the incentive to support the sugar program: if
the sugar price drops as a result of Mexican sugar flowing into the U.S. market through
the large import quota promised under NAFTA, the industry has to weigh both costs and
benefits to the industry (Moss and Schmitz, 2002).
Although the economic implications of the price differential between sugar and
HFCS are identified, estimation of a quantitative relationship for HFCS supply is not


37
max 71- PSUGAR Qsugar C (Qsugar,, V )
(3.11)
The first-order condition gives the supply relationship for the industry by equating output
price to marginal cost. Expressed in a general form with a vector of other factors that
influence aggregate supply:
QS SUGAR (PSUGAR, Wj ).
(3.12)
Similarly HFCS supply from industry (Q'hfcs ) is theoretically expressed as a
function of the HFCS price and the supply shifters; however, the quantitative relationship
is expected to be insignificant: HFCS pricing is ambiguous and arbitrary as discussed in
the previous chapter and thus the estimated relation does not likely represent the
associated marginal cost curve.
U.S.-Mexico Bilateral Sugar Trade System
A spatial equilibrium model is used to portray the U.S.-Mexico bilateral sugar
trade system, following the Takayama and Judge formulation (1964). The model provides
the optimal equilibrium price as well as quantities demanded and supplied at the
equilibrium through maximization of welfare in each region, i.e. the sum of the consumer
and producer surplus, given the demand and supply equations and the transportation cost
among regions. Let S¡(Y¡) represent the inverse supply function (price-dependent form)
for sugar in region i; Y¡ represents the quantity produced in region i; Dj{Qj) represents the
inverse demand function (price-dependent form) for sugar in region j; and Qj represents
the quantity consumed in region j.
j
S.t. X(x(y + xx,) (3.14)


64
taken over from the former government body after the privatization of the milling sector.
The committee carries an extensive data set regarding not only physical production and
price but also detailed productivity and efficiency indicators such as sugar and fiber
content in cane, mill downtime and sugar production loss during the process across 60
operating mills. Data for the U.S. sugar industry was obtained from the Sugar and
Sweetener Situation and Outlook Yearbook and other publications by the Economic
Research Service, USDA. Historic data for population were obtained form the website of
the U.S. Bureau of the Census; and those for GDP, exchange rates and consumer price
index were taken from OECD documents.
Data sources used in the study are summarized in Tables 4-6 through 4-10. For
demand analysis, aggregate time-series data at the national level are used in the
regressions for both the United States and Mexico. Missing data were found in U.S. sugar
consumption and related prices in 1991 and filled with average values of two adjacent
years in order to maintain the data continuity.
For the supply analysis, aggregate time-series data at industry level are used in the
regression for both the United States and Mexico. No missing data were found in the data
set. Nominal values of old Mexican pesos before devaluation found in the data are
converted into current pesos. Local units are preserved during the estimation of
elasticities in demand and supply analyses. In bilateral trade model, different local units
are converted into common units such as metric tons and US dollars so that the model can
achieve the equilibrium point in the system.


83
instars (Andow and Risch 1985; Giroux et al. 1995; Griffin and Yeargan 2002; Groden et
al. 1990; Hazzard et al. 1991; Hilbeck and Kennedy 1996; Munyaneza and Obrycki
1998). This species is widespread throughout North America, and has been shown to
provide effective biological control in several crop species, including com, crucifers,
tomato and potato (Hoffman and Frodsham 1993). However, the PSLB was found to be
susceptible to carbaryl and menthamidophos, the same pesticides used for the control of
many aphid species (Hoffman and Frodsham 1993).
Since its introduction into the United States in 1884, water hyacinth (Eichhornia
crassipes (Mart.) Solms-Laubach) has infested waterways of the southeast that has cost
upwards of $2 million to control in Florida alone (Schardt 1987). The mottled water
hyacinth weevil (MWHW), native to Argentina, was first released in Florida in 1972 and
subsequently to other states and countries in an effort to control water hyacinth (Center
1994). The genus is restricted to feeding on members of Pontederiaceae, with the
MWHW feeding mainly on the introduced water hyacinth; it can be found virtually
everywhere the host plant is present (Haag and Habeck 1991; Center et al. 1998).
The greenbug parasitoid (GBP) is an important natural enemy of many cereal
aphids. This species is known for the production of mummies, the bodies of
parasitized aphids that act as a protective case for the developing wasp pupa, and is
considered by many to be tolerant to cold temperatures (Elliott et al. 1999; Knutson et al.
1993; Wright 1995). However, this greenbug parasitoid is an insect and is just as
susceptible to pesticides despite the protective case of the immature form (Knutson et al.
1993).


19
Figure 3-1. Rearing chamber for tobacco homworm and Colorado potato
beetle larvae used in the artificial and excised leaf diet tests.
Hardware cloth stage supporting the leaf allowed for easy
clean up and minimized disease problems by preventing
larvae from coming in contact with fecal material (paper liner
not shown).


84
The purpose of this portion of the study was to examine the effects of L-
methionine on selected nontarget species that are both important in terms of being
beneficial in controlling other pest species and also represent different feeding guilds that
would come into contact with this compound in different ways (e.g., on prey items, on
plant surfaces, hosts of parasitoids).
Materials and Methods
Coleomeeilla maculata
Adults were obtained from ENTOMOS, LLC (Gainesville, Florida), and were
held in 26.4L x 19.2W x 9.5H (cm) clear plastic boxes with a hardware cloth stage
inserted (to facilitate cleaning) at 27C, 60% relative humidity and 16L/8D photoperiod
in FRIUs. Natural diet consisted of excised cotton leafs infested with aphids (Aphis
gossypii Glover (Hemiptera: Aphididae)). Leaves were then dipped into either a 1.0%
L-methionine solution or 0% L-methionine (control) mixed with deionized H2O. Five
adults were used in each replicate for a total n=30 for each treatment. Leaves were
replaced every other day from 27 October 2002 to 07 November 2002. Artificial diet was
obtained from ENTOMOS and prepared according to their guidelines with the exception
of the inclusion of methionine for the 1.0% L-methionine treatment (wt/wt). Diets were
replaced every other day from 27 October 2002 to 07 November 2002. Ten adults were
used for each replicate for a total n=60 for each treatment. Data was normalized to 0%
mortality when the treatments were corrected for control mortality (i.e., when the control
mortality was greater than that of the treatment).


47
Feeding rates of CPB also were found to be statistically different among treatments
(Figure 4-4). Three distinct groups were observed with the first group containing the
Control and 0.1% L-methionine treatments while the second group of the 0.1% L-
methionine and 0.3% L-methionine, treatments were found to be statistically the same.
The 0.5% L-methionine, 0.7% L-methionine, 1.0% L-methionine and Btt treatments were
statistically different from the other groups. Overlap occurred with the proline treatment
across all groups indicating no statistical difference with the rest of the treatments.
Preference Tests
The amount of Control and 1.0% L-methionine leaf tissue consumed during the
preference tests was found not to be statistically different (Figure 4-5). In addition, the
mean head capsule width (i,e development) showed no relationship with either treatment
based upon the low correlation coefficients.
Discussion
The 1.0% L-methionine concentration produced the same larval mortality, feeding
and developmental rates for CPB, as did the Btt treatments (Figures 4-1,4-3, and 4-4).
The 0.3% L-methionine, 0.5% L-methionine and 0.7% L-methionine treatments took 4
days longer for complete control (Figure 4-1), but were statistically different for the
developmental rates for the same treatments (Figure 4-3). As was the case with the THW
survivorship, the 0.1% L-methionine concentration was not different from that of the
Control. This may indicate a threshold of methionine that can be tolerated by the THW,
and CPB to some extent, evidenced by the low mortality observed for this treatment.
The Preference tests did not indicate any preference of leaf disks with or without
L-methionine. The high mortality (90%) of the CPB larvae could be explained by a


LIST OF REFERENCES
Alvarez, Jose and Leo Polopolus. The Florida Sugar Industry. EDIS document
SC042, Food and Resource Economics Department, University of Florida, Gainesville,
FL, June 2002a.
Alvarez, Jose and Leo Polopolus. The Sugar Program: Description and Debate.
EDIS document SC020, Food and Resource Economics Department, University of
Florida, Gainesville, FL, June 2002b.
Azcar S.A. de C.V. Estaditica Azucareras. Ventas de azcar el pas por clase,
destino y tipo de operacin 1970-1990. Mexico, DF, 1990.
Borrel, Brent. The Mexican Sugar Industry. International Economics
Department, Policy, Research and External Affairs Working Paper Series No.596.
Washington, DC: World Bank, February 1991.
Buzzanell, Peter. The U.S.-Mexico High Fructose Com Syrup (HFCS) Trade
Dispute. In Schmitz, Andrew, Thomas H. Spreen, William A. Messina, Jr., and Charles
B. Moss, Sugar and Related Sweetener Markets: International Perspectives (53-64). New
York: GABI Publishing, 2002.
Buzzanell, Peter, and Ron Lord. Mexico: Sugar and Com Sweetener, an Update.
Sugar and Sweetener S&O V 20(2), June 1995.
Comit de la Agroindustria Azucarera (COAAZUCAR). Desarrollo Operative
Campo-Fabrica 1996/2002. Internet site:
http://www.sagarpa.gob.mx/Coaazucar/menu2/nacional.htm (Accessed May 2003a).
Comit de la Agroindustria Azucarera (COAAZUCAR). Rangos de Superficie y
Nmero de Caeros a Nivel Nacional Zafra 2000/2001. Internet site:
http://www.sagarpa.gob.mx/Coaazucar/menu6/ann09.htm (Accessed May 2003b).
Comit de la Agroindustria Azucarera (COAAZUCAR). Superficie de Riego y
Temporal y su Produccin de Caa Zafra 2000/2001. Internet site:
http://www.sagarpa.gob.mx/Coaazucar/menu6/sprt03.htm (Accessed May 2003c).
Comit de la Agroindustria Azucarera (COAAZUCAR). Avance del Tipo de
Cosecha, Numero de Cortadores, Cosechadoras Integrales y Alzadoras al 17 de Febrero
del 2001 Zafra 2000/2001. Internet site:
http://www.sagarpa.gob.mx/Coaazucar/menu4/ult_estim_prod.htm (Accessed May
2003d).
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Figure 7-1. Mortality of Coleomegilla maculata adults after exposure to L-methionine
treated artificial diet. Data corrected for control mortality using Abbotts
formula.


20
approximately 48 and 45 percent of the total sugarcane production in 2001, respectively
(USDA, 2001a). Floridas sugar production from sugarcane made up 50 percent of total
cane sugar production in the United States in 2001 (USDA, 2002a), exceeding that of
Louisiana (40 percent) due to a higher sugar recovery rate from cane. Sugar beet
production regions are classified into four regions: Great Lakes, Upper Midwest, Great
Plains, and Far West. The Upper Midwest, which includes Minnesota and North Dakota,
produces approximately 47 percent of total sugar beet production; the Far West, which
includes California, Idaho, Oregon and, Washington, produces approximately 26 percent
of total sugar beet production in 2001 (USDA 2002a). Total sugar production from
sugarcane and sugar beets was 4,017,000 short tons (3,615,000 MT) and 4,000,000 short
tons (3,600,000 MT), respectively (USDA, 2002a). The proportion of sugar produced in
the United States from sugarcane and sugar beets is about equal.
Compared to Mexico, sugarcane production in the United States is regionally
concentrated and highly mechanized as well as vertically integrated. In the case of the
Floridas sugar industry, sugarcane is grown areas concentrated in flat land in south
Florida. Sugarcane growing activities such as planting, harvesting and transporting
harvested crop are fully mechanized, unlike Mexico. Six raw sugar mills, which are
located near the sugarcane fields, possess an average daily processing capacity of 20,750
tons of sugarcane (Alvarez and Polopolus, 2002a). Two sugar refineries are located
adjacent to two sugar mills. U.S. Sugar, the countrys largest sugar producer operating in
Florida, owns a fully integrated cane sugar refinery facility that manages not only sugar
refining but also packaging and warehousing. With the facility built next to the existing


implement, produce unpredictable results, and require new knowledge (Barfield and
Swisher 1994; Ehler and Bottrell 2000).
Importance of IPM in Florida and Surrounding States
Considerable effort has been devoted to developing IPM programs in Florida
because of its unique pest problems and crop production systems, sensitivity to chemical
pollutants, and increased urbanization (Capinera et al. 1994; Rosen et al. 1996). The
necessity for developing IPM protocols for Floridas major plant and animal pests was
underscored in a new statewide initiative. In November 1999, the Institute of Food and
Agricultural Sciences (IFAS) at the University of Florida launched Putting Florida FIRST
Focusing IFAS Resources on Solutions for Tomorrow (Florida FIRST 1999). The
Florida FIRST initiative was created (with input from stakeholders) to define the role of
IFAS in shaping Floridas future in the 21st century. Increasing concerns (expressed
repeatedly by Floridas scientific community and the general public) about environmental
contamination, food safety issues, and human and animal health problems resulting from
the indiscriminate use (and often misuse) of pesticides are making existing methods for
pest management obsolete. Successful implementation of true IPM, as it was
envisioned by those who envisioned the original concept, will have the added benefit of
helping Florida ... enhance natural resources, provide consumers with a wide variety of
safe and affordable foods,... provide enhanced environments for homes, work places
and vacations, maintain a sustainable food and fiber system, and improve the quality of
life... (Florida FIRST 1999).
This effort to promote IPM programs in the state of Florida also benefits the
surrounding states. For example, solanaceous crops produced in the southeastern U.S.
(such as tomato, tobacco, eggplant, peppers and potato) are subjected to the same


48
(Error Bars @ 95%; F(0.05)7,312=1.14;F=40.1; P<0.001)
400
Control 0.1% 0.3% 0.5% 0.7% 1.0% Proline Btt
Figure 4-4. Total leaf area consumed by Colorado potato beetle larvae exposed
to excised eggplant leaves treated with various concentrations of L-
methionine (nTOtai=:320). Proline (1.0%) and Btt were included for
comparison as positive and negative controls. Error bars denote 2
SE. Bars within treatments having the same letter are not statistically
different (Tukeys MST, P0.001).


Abstract of Dissertation Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Doctor of Philosophy
EVALUATION OF THE AMINO ACID METHIONINE FOR BIORATIONAL
CONTROL OF SELECTED INSECT PESTS OF ECONOMIC AND MEDICAL
IMPORTANCE
By
Lewis Scotty Long
May, 2004
Chair: James P. Cuda
Cochair: Bruce R. Stevens
Major Department: Department of Entomology and Nematology
Integrated pest management (IPM) strategies were developed in an effort to
control pests with fewer pesticides. However, because of the misuse of pesticides and the
failure to adopt IPM practices pesticide use is higher than ever. An alternative to
conventional broad-spectrum pesticides is the use of biorational compounds; those that
pose minimal risk to the environment and are specific to the target pests.
The recent discovery of the CAATCH1 system in the midgut of the tobacco
homworm (THW), Manduca sexta, has revealed a novel means to control certain insect
pests. This membrane protein works in alkaline conditions as both an amino acid
transporter and also independently as a cation channel. However, the amino acid
L-methionine blocks amino acid transport thus altering the ion flow.
xi


90
welfare and let its domestic sugar industry be taken care of by the U.S. sugar and HFCS
industry, there will be little reason for the Mexican government to cooperate with the
United States; however, Mexicos abandoning support to the sugar and its related
industries may trigger political and social instability.
Lastly, the results from the game played by the grand coalition are shown in Table
5-13. Although the solution of the game is the same as one without the U.S. HFCS
industry, the pay-offs from Mexican government high production-high HFCS adoption
strategy are almost as good. This result implies that adopting HFCS in the Mexican
market is not necessarily unbeneficial as long as Mexico increases sugar production,
under the assumption that gains to the grand coalition are redistributed among countries
and industries. The challenge is how to put this into practice.


43
Results
Survivorship
Mortality of CPB larvae on treated excised eggplant leaves ranged from
approximately 20% for the 0.1% L-methionine treatment after 4 days, 80% mortality for
the 0.3% L-methionine treatment after 8 days of exposure and 100% for the remaining
concentrations with the highest dose of 1.0% L-methionine exhibiting complete control
of CPB in 3 days post treatment (Figure 4-1). Some mortality (50%) was observed for
the proline (1.0%) treatment while the Bit larval treatment mortality was similar to the
1.0% L-methionine treatment, resulting in 100% mortality after 5 days.
PROBIT analysis of a sample size of ntotai=L320 for 6 treatments (Control), 0.1%
L-methionine, 0.3% L-methionine, 0.5% L-methionine, 0.7% L-methionine and 1.0%
L-methionine) revealed an overall LC50 of 0.218% concentration for the CPB after 8 days
of exposure (Figure 4-2). The LC50 of 2.9% for 24 hours dropped to 1.1% after 48 hours
and to 0.22% after 72 hours.
Feeding and Development
Mean head capsule widths between treatments were found to be statistically
different (Figure 4-3). Four distinct groups were observed, with the Control, 0.1%
L-methionine and proline treatments forming the first group. The second group of
proline and 0.5% L-methionine were statistically the same and likewise the third group of
the 0.3% L-methionine, 0.5% L-methionine, and 0.7% L-methionine treatments. The
final group of Bit and 1.0% L-methionine treatments was statistically different from all
other treatments.


29
Figure 3-7. Survivorship of tobacco homworm larvae exposed to various
concentrations of L-methionine (niotai=256) on whole plants. L-
methionine was applied using a hand-held sprayer in the amount of
10 mL/treatment. Data were adjusted using Abbotts formula for
control mortality.