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The bionomics and life cycle of Trichodorus christiei

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Title:
The bionomics and life cycle of Trichodorus christiei
Creator:
Morton, Henry Vintcent Newton, 1936-
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English
Physical Description:
79 leaves : ill. ; 28 cm.

Subjects

Subjects / Keywords:
Corn ( jstor )
Crops ( jstor )
Fumigation ( jstor )
Plant roots ( jstor )
Roundworms ( jstor )
Soil science ( jstor )
Soils ( jstor )
Specimens ( jstor )
Tillage ( jstor )
Tomatoes ( jstor )
Dissertations, Academic -- Entomology and Nematology -- UF
Entomology and Nematology thesis Ph. D
Nematodes ( lcsh )
Trichodorus ( lcsh )
City of Sanford ( local )
Genre:
bibliography ( marcgt )
non-fiction ( marcgt )

Notes

Thesis:
Thesis (Ph. D.)--University of Florida, 1967.
Bibliography:
Includes bibliographical references (leaves 74-76).
General Note:
Typescript.
General Note:
Vita.
Statement of Responsibility:
by Henry Vintcent Newton Morton.

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University of Florida
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This item is presumed in the public domain according to the terms of the Retrospective Dissertation Scanning (RDS) policy, which may be viewed at http://ufdc.ufl.edu/AA00007596/00001. The University of Florida George A. Smathers Libraries respect the intellectual property rights of others and do not claim any copyright interest in this item. Users of this work have responsibility for determining copyright status prior to reusing, publishing or reproducing this item for purposes other than what is allowed by fair use or other copyright exemptions. Any reuse of this item in excess of fair use or other copyright exemptions requires permission of the copyright holder. The Smathers Libraries would like to learn more about this item and invite individuals or organizations to contact the RDS coordinator(ufdissertations@uflib.ufl.edu) with any additional information they can provide.
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THE BIONOMICS AND LIFE CYCLE OF

TricAodorus cAristiei










By
HENRY VINTCENT MORTON














A DISSERTATION PRESENTED TO THE GRADUATE COUNCIL OF
THE UNIVERSITY OF FLORIDA
IN PARTIAL FULFILLMENT OF THE REQUIBEMENTS FOR THE
DEGREE OF DOCTOR OF PHILOSOPHY











UNIVERSITY OF FLORIDA June, 1967















ACKNOWLEDGMENTS


The author wishes to thank all those who helped in making this dissertation a realization. In particular, supervisory committee co-chairman Professor V. G. Perry of the Department of Entomology for his guidance and encouragement in this work and for making available a teaching assistantship under visiting Professor G. Thorne, through the auspices of the Tropical Nematology Program, and to Dr. H. L. Rhoades, Assistant Nematologist, at the Central Florida Experiment Station, for the generous gift of his time, knowledge and facilities during the sixteen months the author spent at Sanford, made possible through the financial support of the Station.

Professor A. H. Krezdorn, Head of the Department of Fruit Crops, and Dr. G. C. Smart, Jr., Assistant Nematologist, Agricultural Experiment Station, for their helpful suggestions and criticisms in the writing of this dissertation.

Dr. H. N. Miller, Plant Pathologist, Agricultural Experiment Station, for the liberal use of his laboratory facilities and photomicrographic equipment.

To his wife, Virginia, sincerest appreciation for her patience and understanding. Lastly, to his parents, Commander and Mrs. E. B. Martino, who were always ready with financial support when needed.















TABLE OF CONTENTS


Page

ACKNOWLEDGMENTS . . . . . . . . . . . . . . . . . . . . . . . . ii

LIST OF TABLES . . . . . . . . . . . . . . . . . . . . vi

LIST OF FIGURES . . . . . . . . . . . . . . . . . . . . . . . . ix

INTRODUCTION . . . . . . . . . . . . . . . . . . ... . . . . . I1

REVIEW OF LITERATURE . . . . . . . . . . . . . . . . . . . . . 2

MATERIALS AND METHODS . . . . . . . . . . . . . . . . . . . . . 6

FIELD EXPERIMENTS . . . . . . . . . . . . . . . . . . . . 6

Field Experiment 1. Comparison of the effects of 1, 3dichloropropene 1, 2-dichloropropane (D-D soil fumigant)
fumigation with no fumigation on the population of Trichodorus christiei on fall, spring and summer crops . . . 6

Field Experiment 2. A split-plot experiment designed to
study the residual effect of three nematicides in the
control of Trichodorus christiei . . . . . . . . . . . 7

Field Experiment 3. The survival of Trichodorus
christiei in the field in the absence of a host . . . . 9

GREENHOUSE EXPERIMENTS . . . . . . . . . . . . . . . . . . 9

Greenhouse Experiment 1. Effects of four temperatures
on the reproductive rate of Trichodorus christiei . . . 9

Greenhouse Experiment 2. A test designed to determine
the host status of pangolagrass, Digitaria decumbens
Stent., to the ectoparasitic nematodes Trichodorus christiei
and Belonolaimus longicaudatus. . . . . . . . . . . . . 10

Greenhouse Experiment 3. The interrelationship of
population levels of Trichodorus christiei and Belonolaimus longicaudatus in the greenhouse utilizing
different levels of initial inocula . . . . . . . . . . 12










Page


Greenhouse Experiment 4. A comparison of the reproductive rate of 25 Trichodorus christiei in D-D fumigated soil with that in unfumigated soil . . . . . . . . . 12

Greenhouse Experiment 5. Investigation into the possible predaceous habit of three free-living nematodes and an oligochaete on Trichodorus christiei . . . . . . . . . . 13

Greenhouse Experiment 6. The effect of different kinds of potting containers on the reproduction of Trichodorus
christiei . . . . . . . . . . . . . . . . . . . . . . . 14

LABORATORY EXPERIMENTS . . . . . . . . . . . . . . . . . . 15

Laboratory Experiment 1. The life cycle of Trichodorus christiei in a constant temperature chamber set at 80OuF
with a 12-hour photoperiod . . . . . . . . . . . . . . . 15

Laboratory Experiment 2. Mode of molting by Trichodorus
christiei . . . . . . . . . . . . . . . . . . . . . . . 17

Laboratory Experiment 3. Determination of the reproductive potential of Trichodorus christiei . . . . . . . 17

Laboratory Experiment 4. Population dynamics of
Trichodorus christiei as affected by original numbers. . 17

Laboratory Experiment 5. An investigation into the
possibility of a stage resistant to D-D fumigation in
the life cycle of Trichodorus christiei . . . . . . . . 18

Laboratory Experiment 6. Survival of Trichodorus
christiei in the absence of a host at 80F . . . . . . 19

Laboratory Experiment 7. Morphological studies on
Trichodorus christiei . . . . . . . . . . . . . . . . . 20

RESULTS AND DISCUSSION . . . . . . . . . . . . . . . . . . 21

FIELD EXPERIMENTS . . . . . . . . . . . . . . . . . . . . 21

Field Experiment 1 . . . . . . . . . . . . . . . . . . . 21

Field Experiment 2 . . . . . . . . . . . . . . . . . . . 30

Field Experiment 3 . . . . . . . . . . . . . . . . . . . 36

GREENHOUSE EXPERIMENTS . . . . . . . . . . . . . . . . . . 36














Greenhouse Experiment 1 Greenhouse Experiment 2 Greenhouse Experiment 3 Greenhouse Experiment 4 Greenhouse Experiment 5 Greenhouse Experiment 6

LABORATORY EXPERIMENTS . .

Laboratory Experiment 1 Laboratory Experiment 2 Laboratory Experiment 3 Laboratory Experiment 4 Laboratory Experiment 5 Laboratory Experiment 6 Laboratory Experiment 7

THE BUILD-UP OF TRICHODORUS SUMMARY.. ..........

LITERATURE CITED . . . . . . . BIOGRAPHICAL SKETCH . . . . . .


Page

. . . . . . . . . . . . . . . . 37

. . . . . . . . . . . . . . . . 38

. . . . . . . . . . . . . . . . 41

. . . . . . . . . . . . . . . . 46

. . . . . . . . . . . . . . . . 46

. . . . . . . . . . . . . . . . 49

. . . . . . . . . . . . . . . . 51

S . . . . . . . . . . . . . . . 51

S . . . . . . . . . . . . . . . 55

S . . . . . . . . . . . . . . . 58

S . . . . . . . . . . . . . . . 58

. . . . . . . . . . . . . . . 61

S . . . . . . . . . . . . . . . 66

S . . . . . . . . . . . . . . . 67

CHRISTIEI . . . . . . . . ...69

. . . . . . . . . . . . . . . . 71

S . . . . . . . . . . . . . . . 74

S . . . . . . . . . . . . . . . 77















LIST OF TABLES


Table Page


1. Crops Used in Comparing D-D Fumigation with No Fumigation . . . . . . . . . . . . . . . . . .. . . . . . . . . 7

2. Nature and Source of the Three Soils Used in the Study of
the Host Status of Pangolagrass to Trichodorus christiei
and Belonolaimus longicaudatus . . . . . . . . . . . . . . 10

3a. Numbers of Trichodorus christiei Found in D-D Fumigated
Plots Compared to Those in Unfumigated Plots. (a) Fall
Application . . . . . . . . . . . . . . . . . . . . . . . 23

3b. Numbers of Trichodorus christiei Found in D-D Fumigated
Plots Compared to Those in Unfumigated Plots. (b) Spring
Application . . . . . . . . . . . . . . . . . . . . . . . 24

3c. Numbers of Trichodorus christiei Found in D-D Fumigated
Plots Compared to Those in Unfumigated Plots. (c) Summer
Application . . . . . . . . . . . . . . . . . . . . . . . 25

4. The Effect of Sorghum Root-depth on Populations of Trichodorus christiei at Harvest . . . . . . . . . . . . . . . . 27

5. Increase in Cabbage Yield and Numbers of Trichodorus christiei Following an Application of D-D Fumigant at 30 Gal per
Acre . . . . . . . . . . . . . . . . . . . . . . . . . . . 29

6a. A Comparison of the Effects Certain Nematicides Have On the
Build-up of Trichodorus christiei on 'Funks' Hybrid Field
Corn . . . . . . . . . . . . . . . . . . . . . . . . . . . 32

6b. A Comparison of the Residual Effects of a Split Application of Certain Nematicides on the Build-up of Trichodorus
christiei on Cabbage . . . . . . . . . . . . . . . . . . . 33

6c. Effect of Nematicide Treatments on Total Number and Weight
of Cabbage Heads on Three Harvest Dates. . . . . . . . . . 34

6d. A Comparison of the Residual Effects Certain Nematicides
Have on the Build-up of Trichodorus christiei On Unfumigated
'Gold Cup' Sweet Corn. . . . . . . . . . . . . . . . . . . 35












Table Page

7. A Comparison of the Survival of Trichodorus christiei
in the Presence and Absence of Organic Matter (O.M.)
Under Clean Cultivation in the Field . . . . . . . . . . . 36

8. The Effect of Temperature on the Reproductive Rate of 25
Trichodorus christiei on Sweet Corn and Tomato After 8
Weeks . . . . . . . . . . . . . . . . . . . . . . . . 37

9. Plant Parasitic Nematode Populations in 100 ml of Soil
From Under Pangolagrass Followed by Sweet Corn and Vice
Versa Maintained in the Greenhouse for 3 Months . . . . . 39

10. Total Dry Weight (Grams) of Roots From 5 Replicates of
Pangolagrass and Sweet Corn After 3 Months in Association
With Trichodorus christiei and Belonolaimus longicaudatus. 40

11. The Interrelationship of Two Ectoparasitic Nematodes At
Different Rates of Initial Inoculum While Feeding on Sweet
Corn in 6-inch Plastic Pots . . . . . . . . . . . .... . 42

12. Effect of Trichodorus christiei and Belonolaimus longicaudatus on the Dry Weights (Grams) of the Second Crop of
Sweet Corn. The Corn was Harvested 45 Days After Replanting in Pots in Which the Nematodes had Previously Been
Established . . . . . . . . . . . . . . . . . . . . . . . 43

13. The Reproductive Rate of Trichodorus christiei in D-D
Fumigated Soil and Unfumigated Soil in Greenhouse Pots . . 46

14. Effects of Oligochaetes on Number of Trichodorus christiei
After 8 Weeks . . . . . . . . . . . . . . . . . . . . . . 47

15. Effects of Three Possible Predaceous Nematodes on Populations of Trichodorus christiei . . . . . . . . . . . . . . 48

16. Effect of Aporcelaimellus obscurus on Established Populations of Trichodorus christiei . . . . . . . . . . . . . . 48

17. Numbers of Trichodorus christiei in 100 ml of Soil From
Four Different Containers Using Sweet Corn as a Host After
A Period of 8 Weeks . . . . . . . . . . . . . . . . . . . 49

18. Study of the Life Cycle of the Offspring of 10 Gravid Female
Trichodorus christiei at 80oF While Feeding on 'Homestead 24'
Tomato . . . . . . . . . . . . . . . . . . . . . . . . . . 52

19. Reproductive Rate of Individual Specimens of Trichodorus
christiei While Feeding on 'Homestead 24' Tomato Seedlings
Growing at A Constant Temperature of 80OF . . . . . . . . 59











Table Page


20a. Reproductive Rates of Trichodorus christiei at Different
Rates of Infestation Maintained at 800F. . . . . . . . . . 60

20b. Average Weights of 'Homestead 24' Tomatoes Inoculated with
Trichodorus christiei. . . . . . . . . . . . . . . . . . . 60

21a. Effects of D-D Fumigation on Trichodorus christiei, Belonolaimus longicaudatus and Hoplolaimus galeatus in Soil
Contained in Pint Jars . . . . . . . . . . ' . . . . . . . 64

21b. Effects of D-D Fumigation on a Pure Population of Trichodorus christiei in Soil Contained in Pint Jars . . . . . . 65

22. Survival of Trichodorus christiei in the Absence of a
Host in a 6-inch Pot at 800F . . . . . . . . . . . . . . . 67


viii














LIST OF FIGURES


Figure Page

1. Roots of a sweet corn plant showing stubby-root symptoms
caused by Trichodorus christiei . . . . . . . . . . . . . 26

2. Comparison of sweet corn seedling growth on unfumigated
Leon fine sand soil with that fumigated with D-D soil fumigant applied at 30 gal per acre . . . . . . . . . . . . . 28

3. The effect of Belonolaimus longicaudatus on the growth of
sweet corn 45 days after replanting in pots initially
inoculated with (left to right) 250, 100, 50 and 25 specimens . . . . . . . . . . . . .......................... 44

4. Growth of sweet corn 45 days after replanting in pots
initially inoculated with (left to right) 250, 100, 50
and 25 Trichodorus christiei . . . . . . . . . . . . . . . 45

5. Root growth of sweet corn after 8 weeks in the greenhouse
in a clay pot, glazed crock, and plastic pot . . . . . . . 50

6. Stages in the life cycle of Trichodorus christiei at (O10X);
(a) egg, (b) first larval stage, (c) second larval stage,
(d) third larval stage, (e) fourth larval stage, and (f)
adult female . . . . . . . . . . . . . . . . . . . . . . . 53

7. Variation in size between adults of Trichodorus christiei
collected from two locations near Sanford, Florida . . . . 54

8. Break in the old cuticle prior to molting of its "hood" by
Trichodorus christiei . . . . . . . . . . . . . . . . . . 56

9. A molting specimen of Trichodorus christiei. Note the
absence of the onchiostyle in the "hood" . . . . . . . . . 57

10. Characteristic position of Trichodorus christiei while feeding; i.e., the head at right angles to the cell upon which
it is feeding . . . . . . . . . . . . . . . . . . . . . . 62

11. Trichodorus christiei feeding upon the root-cap of tomato
in agar . . . . . . . . . . . . . . . . . . . . . . . . . 63

12. Water mount of a live Trichodorus christiei recovered from
soil with a moisture level of 1.7 per cent . . . . . . . . 68















INTRODUCTION


The nematode genus Trichodorus was erected in 1913 by Cobb (11) when he described the species Trichodorus obtusus. Micoletzky (22) transferred Dorylaimus primitivus de Man, 1880, to the genus Trichodorus and placed T. obtusus in synonomy. The genus received little further mention in the literature until 1951 when Christie and Perry (8) reported that an undescribed species of Trichodorus was parasitic to several crop plants in Florida. Subsequently this species, described by Allen (2) as Trichodorus christiei, has been recognized as a severe pathogen in many areas of the United States. To date, most of the research on this species deals with host-parasite relationships and control, with little consideration of its biology.

Extremely high population levels of T. christiei are frequently found following the application of soil fumigant nematicides. This unusual population build-up has led to considerable speculations as to the factors responsible for such a phenomenon.

This work on the bionomics and life cycle of T. christiei was

initiated as part of a program to attempt an explanation of the population increases of this nematode resulting from soil fumigation.














REVIEW OF LITERATURE


Thorne (33) placed the genus Trichodorus into his family Diphtherophoridae, subfamily Trichodorinae of the superfamily Dorylaimoidea and named T. primitivus (de Man, 1880) Micoletzky, 1922, the type species. Allen (2) published a monograph of the genus Trichodorus and described T. christiei, the species to which Christie and Perry (8) had given the common name of stubby-root nematode. Other species have since been added until now some 32 are recognized.

The feeding habits of T. christiei have been studied by a number of workers (5, 10, 27, 29 and 35). Russell and Perry (29) reported on three general types of feeding on wheat seedlings growing in agar; externally on roots, externally on root hairs, and within root caps. This latter method explains the measurable symptoms commonly found in association with low populations of T. christiei (1 and 29). Christie and Perry (8) proposed that the disease be termed stubby root because through the cessation of longitudinal growth, short, multibranched rootlets are produced.

The effects of temperature on the population numbers of T. christiei have been studied by Malek et al. (20). The greatest numbers after 60 days and 90 days were found at temperatures very close to or at 250C. The lowest rate of reproduction was at 150C. At the upper limit, Rhode and Jenkins (26) found that no reproduction took place at 350C.











Perry (24) observed a rapid build-up of T. christiei following soil fumigation with methyl bromide and other nematicides. Haasis et al.

(13), studying 14 species of nematodes associated with decline of woody ornamentals, found that they could control all but T. christiei with 1, 2-dibromo-3-chloropropane. This unique behavior of T. christiei to soil fumigation also has been observed by Martin (21) in Africa. Perry (24) suggested that this nematode has a high reproductive potential. Alhassan and Hollis (1) found that after 3 weeks the per cent increase in numbers of T. christiei was related inversely to the initial inoculum level and to plant damage.

The life cycle of T. christiei has been studied in part. Russell

(28) found that eggs hatched 66 to 68 hours after oviposition in vitro with apparently the first larval stage emerging. Rhode and Jenkins

(26) reported that T. christiei completed its life cycle in 16 to 17 days at 300C and 21 to 22 days at 220C. Four distinct groups were recovered; in each case specimens were larger at the lower temperature.

Bird (4) demonstrated that both host and geographic origin of T.

christiei could influence population densityindicating the possibility of races. The host also had an effect on the size of the nematode; for example, those specimens obtained from lettuce were larger than those from tomato and celery. Rhode and Jenkins (27) reported jimsonweed, asparagus, poinsettia and crotalaria to be the only non-hosts of T. christiei among the 42 species of plants they tested. Rhoades (25) in Florida found the numbers of T. christiei declined greatly during the summer months in plots planted to Crotalaria spectabilis Roth., but persisted in those with weeds.











Population numbers have also been reported to be affected by

seasons. Perry (24) found that damage was apparently most severe in a spring crop following a fumigated fall crop. A similar situation was found by Barker and Worf (3) on azaleas in Wisconsin, where no T. christiei were observed during the summer. Hoff and Mai (15) recorded the numbers of T. christiei recovered from onion fields on peat soil at eight depths at inervals of 6 inches down to 4 feet throughout the year in the state of New York. The maximum number was recorded at a depth of 0-6 inches around June 1. T. christiei was not found below 12 inches at any time. Hoff and Mai (15) concluded that "the natural population of T. christiei was maintained in a specific soil and root zone."

Comparing silt clay loam, loam and sandy loam, Thomason (31) found

that after twelve weeks the greatest reproduction of T. christiei occurred in the sandy loam. After 19 weeks, however, the sandy loam population was only 1/6 that of the loam. Survival in the absence of a host by T. christiei was also reported as being substantially greater in the loam. According to Christie (7), it seems unlikely that the distribution of T. christiei is greatly influenced by soil type.

Recent agricultural research has stressed biological control. A possible explanation for the rapid return of T. christiei following fumigation may be due to the reduction of predator populations. Working in vitro, Russell (28) found a number of predators which preyed upon T. christiei, the most aggressive of which were the nematode Aphelenchoides winchesi Christie, 1939 and the mite Protolaclaps











bickleyi Bram. Thorne (32) found the remains of Trichodorus primitivus in the gut of Mononchus acutus Cobb, 1917. Chitwood and Oteifa (6) observed oligochaetes feeding upon nematode eggs, while Schaerffenberg

(30) demonstrated that enchytraeids controlled Heterodera schachtii Schmidt, 1871 in greenhouse pots. Tardigrades were reported feeding upon Trichodorus aequalis Allen, 1957 by Hutchinson and Streu (16). Esser and Sobers (12) expressed doubt that tardigrades can be utilized in biological control as they are rarely found in abundance. Little is known of the presence of nematicidal fungi in Florida soils and there have been no studies on the effects fumigation have on these fungi.

A survey of the literature revealed no work on the competitive status of Trichodorus when in association with other plant parasitic nematodes. Krusberg and Sasser (18) observed that only small numbers of Meloidogyne sp. and Pratylenchus sp. were present when Hoplolaimus coronatus Cobb, 1923 was abundant. Thorne (34) noted that Xiphinema americanum Cobb, 1913 frequently dominates Pratylenchus penetrans (Cobb, 1917) Filipjev and Stekhoven, 1914 and Pratylenchus minus Sher and Allen, 1953 when in association with these smaller, slower moving pratylenchs.














MATERIALS AND METHODS


Field Experiments

Field plot experiments involving the use of nematicides were conducted at the Central Florida Experiment Station for the purpose of studying the population build-up of T. christiei following fumigation. Using a randomized block design, plots were laid out on Leon fine sand soil which had been fallow for the past ten years. The dominant plant species prior to plowing was bermudagrass (Cynodon dactylon (L.) Pers.). The area lacked tile which is normally used for drainage and subsurface irrigation; thus, when required, plots were irrigated by an overhead sprinkler system.

Soil samples were taken with a Hoffer soil sampler; eight-in-therow samples were taken from each plot. With one exception, samples were taken to a depth of 6 inches and the soil placed in polyethylene bags. Individual samples were thoroughly mixed and a 100 ml subsample processed by the centrifugal-flotation method (17). Samples were read within 48 hours of sampling. Nematodes were counted in a known area of a syracuse dish under a binocular microscope, and the number per 100 ml of soil was calculated. Field Experiment 1. --Comparison of the effects of 1,3-dichloropropene 1, 2-dichloropropane (D-D soil fumigant) fumigation with no fumigation on the population of Trichodorus christiei on fall, spring and summer crops.

After bringing the area to seed bed tilth, one half of each block was treated with D-D soil fumigant at the rate of 30 gal per acre











applied with a conventional shank injector at a depth of 6 inches. Two weeks later the plots were fertilized with a 5-5-8 mixture at a rate of 1000 lb per acre, disked and leveled. Two crops were planted, with 5 rows of each per replicate which were 35 ft long by 24 ft wide (Table 1). Data were taken from the 3 rows of each crop, within the border rows.

Table 1. Crops Used in Comparing D-D Fumigation With No Fumigation


Crop Cabbage Sorghum Cabbage Sweet corn Soybean Sorghum x sudangrass hybrid Cabbage Sorghum x sudangrass hybrid


Variety Marion market Beef builder Coppenhagen Gold cup Lee


Grazer A Market topper


Planting Date 10/14/64 3/25/65 7/19/65 11/12/65


Harvesting Date

2/3/65



6/10/65 10/20/65 2/25/66


Grazer A


Population counts of the nematodes present were made at the end of each month. Yields were taken on a fresh weight basis at harvest. Field Experiment 2. --A split-plot experiment designed to study the residual effect of three nematicides in the control ofTrichodorus christiei.

A comparison was made between a single nematicide application

prior to a summer crop of 'Funks' hybrid field corn and its effects on












the following fall crop of 'Greenback' cabbage with that of an application prior to both the summer and fall crops on plots 12.5 ft by 14 ft. The experiment was terminated by observing the residual effect of the nematicides on a crop of 'Gold-Cup' sweet corn the following spring.

The treatments included: (1) check, (2) 0-0 diethyl 0-2 pyrazinyl phosphorothioate (zinophos) at 3 lb per acre, (3) 1,2-dibromo-3chloropropane (Nemagon) at 2 gal per acre, (4) D-D soil fumigant at 15 gal per acre, and (5) D-D soil fumigant at 25 gal per acre.

Prior to fumigation 25 lb of soil containing 100 T. christiei per 100 ml was spread over each plot and disked in. The fumigants were applied by a hand applicator on a 12-inch lattice to a depth of 6 inches

2 weeks before planting. Zinophos was applied as a 10 per cent granular formulation to the soil surface just prior to planting and immediately forked into the soil to a depth of + 6 inches. The plots were then fertilized with a 5-5-8 fertilizer mixture at the rate of 1000 lb per acre, disked, leveled and planted.

The plots were cultivated to reduce weed growth and a side dressing of 5-5-8 fertilizer was applied at the rate of 500 lb per acre during the third and sixth week following planting. The insecticide Sevin was applied twice a week at 2 lb per 100 gal of water to control corn earworm. Cabbage looper control was obtained using 1/2 pint of parathion plus 1 quart of toxaphene in 100 gal of water.

Soil samples were taken prior to fumigation, during the growing season, and at harvest.










Field Experiment 3. --The survival of Trichodorus christiei in the field in the absence of a host.

A study was made on the effect of a green manure crop incorporated into the soil on the survival of T. christiei. Eight field plots 6 ft by 6 ft were planted to a fall crop of sorghum x sudangrass hybrid (variety Grazer A) a known host of T. christiei. After 7 weeks all the plots were harvested and the root systems removed. The green matter, at a rate of 17 tons per acre, then was incorporated in one plot of each of the 4 replicates. The experimental area was kept weed-free by hand for the following 16 weeks, during which time the survival of T. christiei was recorded in plots with and without organic matter.


Greenhouse Experiments

Greenhouse Experiment 1. --Effects of four temperatures on the reproductive rate of Trichordorus christiei.

The study was made in four constant temperature tanks designed by Harrison and Stall (14). Each of the tanks held 16 containers and the tanks were randomly alloted temperatures of 800, 85, 90* and 95*F + 1.5'F. Leon fine sand soil was fumigated with methyl bromide. After allowing the soil to aerate for a week a 5-5-8 fertilizer mixture was thoroughly mixed with the soil at the rate of 1000 lb per acre and 1100 ml of this soil was used to fill each of 64 plastic containers. Specimens of T. christiei were hand picked into distilled water; 25 females (males are very rare) were used to inoculate the soil in each container. At the time of inoculation half of the containers were planted to 'Gold Cup' sweet corn and half to 'Rutgers'tomato. The pots were watered with distilled water as needed. At the end of











8 weeks the experiment was terminated. The soil was thoroughly mixed

and a representative sample obtained. A 100 ml sub-sample was processed

using the sugar-flotation method to remove the nematodes.


Greenhouse Experiment 2. --A test designed to determine the host
status of pangolagrass, Digitaria decumbens Stent., to the ectoparasitic nematodes Trichodorus christiei and Belonolaimus longicaudatus.

Soil from the Immokolee fine sand series was obtained from three

sources (Table 2).


Table 2. Nature and Source of the Three Soils Used in the Study of the Host Status of Pangolagrass to Trichodorus christiei
and Belonolaimus longicaudatus.


Plant parasitic nematodes in 100 ml of soil
Origin pH O.M.a/ Stubby-root Stunt Ring "Sterile"b/ 5.3 4.48% -- -- -"Pangola" 7.0 4.32 14 -- -"Crop" 6.6 2.58 14 32 2 a/ Chromic acid determination of the organic matter. b "Sterile" = steam sterilized.

"Pangola" = from under a 3-year-old pangolagrass pasture.

"Crop" = from under sorghum, prior to which there had been 2 crops
of tomato.

Twenty 6-inch clay pots were filled with each of the three soils after incorporating the equivalent of 1000 lb per acre of 5-5-8 fertilizer mixture and 500 lb per acre of lime.

The pots were placed in the greenhouse with 5 replicates each of the following host-inoculation combinations in each soil:











Host Nematode

(1) Pangolagrass T. christiei

(2) Pangolagrass B. longicaudatus

(3) 'Gold Cup' sweet corn T. christiei

(4) 'Gold Cup' sweet corn B. longicaudatus

Individual sprigs of pangolagrass were planted in each of the

designated pots. These sprigs were obtained from one-month-old stem cuttings which had been grown in sterile soil. Three seeds of sweet corn were planted in each indicated pot.

The nematodes were inoculated in water suspensions around the seeds of corn and roots of pangolagrass. The respective treatments received 25 females of T. christiei and 20 females, plus 5 males, of B. longicaudatus. Both species of neamtodes were obtained from a field plot of sorghum.

The pots were maintained for 3 months. At the end of each month the grass was clipped to the first node and a side-dressing of 5-5-8 fertilizer mixture was applied at the rate of 1000 lb per acre to each replicate. On termination of the experiment, fresh weights of roots and tops were recorded, the soil in each replicate was thoroughly mixed, and three composite samples were obtained from each treatment. Nematode-free sprigs of pangolagrass then were planted in those replicates which had grown sweet corn, and sweet corn was planted in those replicates previously growing pangolagrass. These replants were maintained for an additional 3 months and processed in the same manner.











Greenhouse Experiment 3. --The interrelationship of population levels of Trichodorus christiei and Belonolaimus longicaudatus in the greenhouse utilizing different levels of initial inocula.

The rates of reproduction of pure populations of T. christiei and B. longicaudatus were compared with a third treatment having these 2 species in combination. The initial inoculum level of each nematode both in the pure and mixed treatments were 25,,50, 100 and 250. Inoculum of T. christiei was comprised of females only, while that of B. longicaudatus was a 4:1 ratio of females to males.

Plastic pots containing 1,400 ml of autoclaved Leon fine sand,

previously fertilized with an equivalent of 1000 lb per acre of 5-5-8 mixture, were planted to 'Gold Cup' sweet corn (3 seeds/pot). Suspensions of the nematode inocula were then washed into the planting holes, and the pots randomized on the greenhouse bench.

After 45 days the soil in each pot was thoroughly mixed and a 100 ml sample taken and processed. Both the corn tops and roots were collected and their dry weights recorded. Following the application of fertilizer the remaining soil was repotted and once again planted to 'Gold Cup' sweet corn. The experiment was terminated 45 days after replanting and similar data were obtained. Greenhouse Experiment 4. --A comparison of the reproductive rate of 25_Trichodorus christiei in D-D fumigated soil with that in unfumigated
soil.

The soil used in this experiment was a Leon fine sand which had been fumigated the previous spring with the nematicide D-D (27 gal per acre) prior to planting cantaloupe (Cucumis melo L. var reticulatus Naud.). A late summer cover crop of Crotalaria spectabilis was












planted following the cantaloupe. After the C. spectabilis had been harvested soil was removed from the field, sifted and placed in six 3-gal porcelain crocks. Three of these crocks were fumigated at a depth of 8 inches with 1.21 ml D-D (=30 gal per acre) by means of a pipette fitted with a propippette. The soil temperature at the time of fumigation was 660F. After waiting 2 weeks, six 6-inch clay pots were filled with fumigated soil and six with the unfumigated soil. At the time of potting a 100 ml sample of soil was processed and the number of nematodes recorded. The pots were seeded with oats (Avena sativa L.), and 25 specimens of T. christiei added to each. At the end of 2 months, the pots were removed and the numbers of T. christiei per 100 ml of soil recorded. The pots were then replanted to 'Gold Cup' sweet corn and allowed to grow for a further 2 months, before once again recording the numbers of nematodes present.

Greenhouse Experiment 5. --Investigation into the possible predaceous habit of three free-living nematodes and an oligochaete on Trichodorus christiei.

The nematode inocula were obtained from the test conducted on the host status of pangolagrass to T. christiei. The specimens were hand picked into distilled water and then poured into 6-inch clay pots containing autoclaved Leon fine sand soil which was then seeded with 'Gold Cup' sweet corn. The design of the treatments applied to each of the three possible predaceous nematodes being tested was as follows with 5 replicates of each:

(1) 100 T. christiei

(2) 100 T. christiei + 25 free-living specimens











(3) 100 T. christiei + 100 free-living specimens

(4) 25 free-living specimens

The free-living nematodes included Aporcelaimellus obscurus (Thorne and Swanger, 1936) Heyns, 1965; Eudorylaimus simplex (Thorne and Swanger, 1936) Andrassy, 1959 and Nylonchulus parabrachyurus (Thorne, 1924) Andrassy, 1958.

Since oligochaetes are commonly found in unfumigated soil, a test was designed to investigate the effect they might have. on the reproduction of T. christiei. The treatments consisted of the following three rates of hand picked inocula replicated five times:

(1) 100 T. christiei

(2) 100 T. christiei + 100 oligochaetes (3) 100 T. christiei + 500 oligochaetes

The inocula was poured into 6-inch plastic pots containing autoclaved Leon fine sand soil which had been fertilized at the equivalent rate of 1000 lb per acre of 5-5-8 mixture. The host plant used in this experiment was 'Homestead 24' tomato. After planting, the treatments were randomly placed in a growth chamber set at 80oF with a 12hour photoperiod. Top and root weights were recorded after 8 weeks, and the soil in each pot was thoroughly mixed and a 100 ml sample processed from each.

Greenhouse Experiment 6. --The effect of different kinds of potting containers on the reproduction of Trichodorus christiei.

Clay pots have conventionally been used for determining nematode pathogenicity and reproduction on a host. Frequently seedling roots











which escape nematode damage grow out to the walls of the pot, where they produce a mat. Owing to the porous nature of clay, there are greater moisture fluctuations along the pot walls, producing an environment which is less favorable to nematodes than the center of the pot.

Previous indications were that roots did not mat along the walls of glazed crocks and plastic pots. Experiments were conducted in a greenhouse and growth chamber set at 800F to determine the environmental effects in four different 6-inch containers on the reproduction of T. christiei. The containers used were as follows:

(1) Red clay pot; surface irrigated.

(2) Red clay pot; sub-surface irrigated.

(3) Glazed crock.

(4) Plastic pot.

Five containers of each type were filled with 1,350 ml of autoclaved Leon fine sand. Each of the pots was seeded with 3 'Gold Cup' sweet corn seeds, and an inoculum suspension of 25 T. christiei poured over the seeds. In addition to the inoculated pots, 5 replicates of uninoculated pots were set up in the growth chamber as checks.

After 8 weeks the pots were removed and top and root weights recorded. The soil in each pot was thoroughly mixed and a 100 ml sample processed to determine the nematode numbers.

Laboratory Experiments

Laboratory Experiment 1. --The life cycle of Trichodorus christiei in a constant temperature chamber set at 80oF with a 12-hour photoperiod.











Styrofoam cups were cut in half and 50 ml of autoclaved Leon fine sand placed in each. 'Homestead 24' variety of tomato was then sown in the cups and after germination thinned to 4 plants per cup. Two weeks after planting 10 gravid females of T. christiei obtained from a tomato host were poured into a hole adjacent to one of the tomato seedlings in each of 48 cups. From the third to the eighteenth day, 4 cups were removed per day.

The soil was processed using an unpublished technique devised by the late M. B. Linford of the University of Illinois. The soil and nematodes collected from the washings on the 325-mesh screen were poured onto a folded Kimwipe tissue held between two 2-ounce plastic funnels. The funnels were placed on the walls of a syracuse dish in such a manner as to suspend the tissue above the bottom of the dish. After allowing the washings to settle on the tissue (1-2 minutes) the funnels were transferred to a second syracuse dish. The water level in this dish was raised to a.level just above that of the tissue. After 6 hours the funnels were transferred to a third syracuse dish and the water level was adjusted as described above. The funnels were removed from this third dish after 2 hours, and the numbers and stages of development of the nematode specimens were recorded.

Specimens of the oldest stage collected each day were placed in a specimen vial filled 1/4 full of distilled water, and the remainder of the offspring were placed in another similar vial. The vials were then transferred to a hot water bath at 500C for 15 minutes,at which time the level in the vials was brought to 1/2,using 2 per cent formalin solution for fixation and preservation.











Laboratory Experiment 2. --Mode of molting by Trichodorus christiei.

Specimens of T. christiei were studied while molting in water

and in 3/4 per cent water agar contained in the top half of 60 x 15 mm plastic culture dishes. After picking the nemas into the agar, the surface of the agar was covered with a disk of plastic sheet to prevent drying and facilitate studies with the aid of a compound microscope. Other specimens were fixed at various stages throughout molting by placing them in 1 per cent formalin at 50C. These were then mounted on Cobb aluminum slide holders to facilitate study from both sides with the aid of oil immersion objectives. Laboratory Experiment 3. --Determination of the reproductive potential of Trichodorus christiei.

Styrofoam cups were prepared as above, but in this experiment 48 cups were inoculated with individual fourth stage larvae and placed in a growth chamber set at 800F with a 12-hour photoperiod. At the end of 16, 17, 18, 19 and 38 days, 8 cups were processed using Linford's funnels. Counts were made of the numbers of each life cycle stage of T. christiei present in each cup. This experiment was duplicated. Laboratory Experiment 4. --Population dynamics of Trichodorus christiei as affected by original numbers.

The effects of initial inocula numbers on the reproductive rate of T. christiei were studied in a constant temperature chamber set at 80oF with a photoperiod of 12 hours. The treatments included hand picked inocula of 25, 100 and 400 females of T. christiei inoculated into 4-inch plastic pots. Each pot contained 375 ml of autoclaved Leon











fine sand soil, and were planted to 'Homestead 24' variety of tomato. Following germination the tomato seedlings were thinned to 6 plants per pot. At the end of 21 days and 42 days, respectively, 6 replicates of each treatment were removed. Top and root weights were recorded and all the soil in each pot was processed using Christie and Perry's modified Baermann funnel technique (9). Counts were recorded after 12 hours.


Laboratory Experiment 5. --An investigation into the possibility of a stage resistant to D-D fumigation in the life cycle of Trichodorus christiei.

One pint wide-mouth fruit jars were filled with field soil and each of the following 10 treatments was replicated 5 times: Rate per acre Rate per jar

(1) Check

(2) 690 gal of mineral spirits 1.0 ml of mineral spirits

(3) 10 gal of D-D per acre 0.2 ml basic solution

(4) 15 " " " " 0.3 " " "

(5) 30 " " " " 0.6 " " "

(6) 50 " " " " 1.0 " " "

(7). 10 gal of water soluble D-D per acre

(8) 15 " " " " " "

(9) 30 " " " " " "

(10) 50 " " " " " "

The basic solution in treatments 3 through 6 was 92.1 ml mineral spirits + 7.2 ml D-D. The basic solution of the water soluble D-D contained 91.8 ml water + 7.2 ml D-D + 1.0 ml Triton X 100 surfactant. This latter solution was mixed by adding the surfactant to the D-D and then adding water.











The D-D was applied, using a 1 ml pipette filled with a propipette, to a depth of 4 inches within each jar. After the pipette was removed, each injector hole was carefully sealed.

The jars were placed in a laboratory cabinet (+ 720F) for 2 weeks. Water was added as needed to prevent drying out of the soil. A 100 ml sample of soil was then obtained from each jar and processed using the centrifugal-flotation method. Counts were made of living nematodes; i.e., those which were moving. The remainder of the soil from each jar was placed in a 4-inch clay pot, seeded with 'Homestead 24' variety of tomato and placed in the greenhouse. Three weeks later another 100 ml soil sample was processed as before.

This experiment was repeated using a wider range of water soluble D-D rates and a soil containing a pure population of T. christiei. The treatments included:


Rate per acre
(1) Check

(2) 25.0 gals of water-soluble D-D per acre

(3) 45.0 " " " " " "i

(4) 56.25 " " " " " "

(5) 75.0 " " " " " "

The basic solution was made up of 88.2 ml

1.0 ml Triton X 100 surfactant.


Laboratory Experiment 6.
absence of a host at 800F.


Rate per jar


0.33 ml of basic solution

0.60 " " " 0.75 " " " 1.00 " " " water + 10.8 ml D-D +


--Survival of Trichodorus christiei in the











Six clay pots were filled with Leon fine sand in which a pure population of T. christiei had been established, and 4 clay pots with the same type of field soil containing a mixed population of nematodes. The tops of the pots were covered with polyethylene secured by elastic bands, in order to limit the loss of moisture from the soil. The pots were then placed in a constant temperature chamber and watered as needed. Individual pots of each soil type were processed after 0, 2, 4, and 8 weeks. The remaining 2 pots containing the pure populations were taken down after 12 and 16 weeks. Two samples of a 100 ml size were processed from each pot, using the centrifugal-flotation method, and the number of surviving T. christiei were recorded. Laboratory Experiment 7. --Morphological studies on Trichodorus christiei.

Morphological studies were made on specimens of T. christiei

which had been killed and fixed as in Laboratory Experiment 1. However, better results were obtained by staining. The stain was prepared by adding 0.05 ml of green food color (water and propylene glycol) to 2.00 ml of water. Living specimens of T. christiei were placed in this stain which killed them within 24 hours to 48 hours. The killed specimens were then fixed in 1 per cent formalin on Cobb aluminum slides.














RESULTS AND DISCUSSION


Field Experiments

Field Experiment 1. Tables 3 a, b and c show that the build-up of T. christiei following soil fumigation described by Christie and Perry (8) may take place during any season of the year, provided a host is present. However, the more rapid increase obtained on the summer crop of sorghum indicates that the rates of build-up vary with differing levels of temperature.

The depth at which the young feeder roots of the host crop are located is of importance when studying the population of T. christiei (Fig. 1). Tables 3 a, b and c show that the major build-up in the

6 weeks following planting is in the top 6 inches of soil. Thereafter, until harvesting, the population of T. christiei increases very rapidly below the 6-inch depth. The extent to which this deeper population develops is not only dependent upon the nature of the host root system, but also on whether or not the soil has been fumigated (Table 4). The reason for this is apparently due to the fact that soil fumigation permits the young seedlings to develop a root-system capable of outgrowing the ensuing nematode infestation (Fig. 2). Thus, it was possible for the fumigated plots to out-yield those not fumigated (Table 5), while supporting a larger population of T. christiei from about the seventh week until harvesting. However, the control of Belonolaimus longicaudatus by fumigation should not be overlooked since this nema-











tode is far more pathogenic and destructive to vegetable crops in this area than T. christiei is.

When comparing the yield of cabbage from fumigated plots with

that from unfumigated plots, it should be noted that there is a significant different in yield at the 120-day harvest, whereas the yields obtained from the second harvest at 140 days were comparable (Table 5). The crops on fumigated plots of this experiment reached maturity as much as 2 weeks before those on unfumigated plots.









Table 3. Numbers of Trichodorus christiei Found in
Compared to Those in Unfumigated Plots.


D-D Fumigated Plots


(a) Fall Application

Treatment Sampling Crop Soil Temp. Sample No. of T. christiei/100 ml of soil Date Age at 6 in. depth Cabbage Sorgnum


Check 9/30/64a/ 0 890F 0-6 in 25 6-12 in 12
Fumigated 0-6 in 30 6-12 in 14

Check 10/31/64 16 days 780 0-6 in i/ 13b/ 6-12 in 1 0
Fumigated 0-6 in 0 0 6-12 in 0 0

Check 11/25/64 42 days 760 0-6 in 40 85 6-12 in 5 19
Fumigated 0-6 in 28 22 6-12 in 13 8

Check 12/29/64 76 days 720 0-6 in 184 191 6-12 in 147 83
Fumigated 0-6 in 173 156 6-12 in 181 110

Check 1/28/65 106 days 630 0-6 in 225 166 6-12 in 203 188
Fumigated 0-6 in 337 272 6-12 in 278 244


a/ Samples b/ Average


obtained just prior to fumigation. of 4 replicates.


Harvested 2/3/65.












Table 3. Numbers of Trichodorus christiei Found in D-D Fumigated Plots Compared to Those in Unfumigated Plots.
(b) Spring Application


Treatment Sampling Crop Soil Temp. Sample No. of T. christiei/100 ml of soil Date Age at 6 in. depth Cabbage Sweet corn


Check 3/29/65- 4 days 740F 0-6 in 69 40 6-12 in 68 55 Fumigated 0-6 in 3 7 6-12 in 7 12

Check 4/26/65 32 days 830 0-6 in 103 219 6-12 in 105 89 Fumigated 0-6 in 117 394 6-12 in 33 112

Check 5/31/65 67 days 850 0-6 in 84 67 6-12 in 63 63 Fumigated 0-6 in 238 273 6-12 in 375 272 a/ Fumigated 18 days Drior to this date.


Harvested 6/15/65.












Table 3. Numbers of Trichodorus christiei Found in D-D Fumigated Plots
Compared to Those in Unfumigated Plots.


(c) Summer Application

Treatment Sampling Crop Soil Temp. Sample No. of T. christiei/100 ml of soil Date Age at 6 in. depth Soybean2/ Sorghum


Check 6/30/65b/ 0 840F 0-6 in 28 9 6-12 in 28 11
Fumigated 0-6 in 0 0 6-12 in 0 0

Check 8/2/65 25 days 840 0-6 in 65 42 6-12 in 7 7
Fumigated 0-6 in 11 47 6-12 in 2 25

Check 8/26/65 49 days 860 0-6 in 42 147 6-12 in 21 56
Fumigated 0-6 in 137 312 6-12 in 56 459

Check 10/8/65 92 days 820 0-6 in 39 165 6-12 in 35 186
Fumigated 0-6 in 112 221 6-12 in 137 571
a/ Due to poor germination, the soybean plots were replanted 10 days after the initial planting.


V/ Fumigated 7 days prior to this date.


Harvested 10/20/65.
















































Fig. 1. - Roots of a sweet corn plant showing stubby-root
symptoms caused by Trichodorus christiei.












Table 4. The Effect of Sorghum Root-depth on Populations
of Trichodorus christiei at Harvest.

Depth Average No. of Dry-weight of No. of T. christiei/100 ml Root-tips roots (grams) of soil

Check Fumigated Check Fumigated Check Fumigated 0-2 in ?2/ 3/ 10.1 16.7 182 315 2-4 in 4 6 6.2 6.9 287 350 4-6 in 8 7 2.7 3.8 385 235 6-8 in 4 10 0.9 2.1 228 676 8-10 in 1 4 0.2 0.8 161 392 10-12 in - 2 0.05 0.4 53 239

12 in - 1 0.0 0.25 32 189 a/ Average of 6 samples.
b/ Average of 12 samples.


Below a depth of 6 inches, the plant parasitic nematode population

became increasingly dominated by T. christiei. In this respect very

few B. longicaudatus were found below the 8-inch depth.
















































Fig. 2. - Comparison of sweet
Leon fine sand soil fumigant applied at


corn seedling growth on unfumigated with that fumigated with D-D soil 30 gal per acre.









Table 5. Increase in Cabbage Yield and Numbers of Trichodorus christiei Following an
Application of D-D Fumigant at 30 Gal per Acre.


Replicate Yield of cabbage heads No. of T. christiei/100 ml of soil after 130 days.
After 120 Days After 140 Days 0-6 in 6-12 in


No. Weight (lb)


100.9 142.7 68.4 49.4


Weight (lb)


215.0 170.1 117.8 105.4


CHECK
No. Weight (lb)


50.0

39.3 48.4 58.6


No.


D-D
Weight (lb)


25.1

41.4 40.2 56.6


Average 64.25 152.1b/ 27.25 a/ Yield taken from three rows 35 ft long and 2.5 b/ The differences in yields of the D-D plots and
(FA = 11.088) LSD05 58.83 c/ Not significantly different.


40.8c/ 553 96 ft wide.
Check plots were significant at the 5% level


A

B C D

Average


54a/ 65 44 34

49.25


441 161 375 252

308


749

637 536 290











Field Experiment 2. When studying the data on the split-plots experiment in Tables 6a, b, c and d, designed to study the residual action of certain nematicides on the build-up of T. christiei, consideration should be given to the following:

a) The plots treated with the organo-phosphate nematicide,

zinophos, restricted the build-up of T. christiei. The residual action of this material also can be seen in the limitednumbers of T. christiei which were found on the second crop following a single application (Table 6b). This residual action was not seen in the final crop of sweet corn, though the yields on the plots receiving 2 applications of zinophos proved statistically greater than all other treatments.

b) The build-up of T. christiei was substantially greater following an application of D-D soil fumigant at 25 gal per acre than at 15 gal per acre. In addition, the numbers of T. christiei present at harvest were greater with a double application of D-D at both the 25 and 15 gal rates.

c) There were no significant differences in control of T. christiei between single and double applications of Nemagon at 2 gal per acre, and in both cases the build-up surpassed that of the control.

d) In Table 6a it can be seen that although the plots that

received 25 gal of D-D per acre had the highest numbers of T. christiei at harvest, they produced significantly greater yields of corn than did all other treatments. The same was found to be true on cabbages (Tables 6b and c) with the exception that the yield was not statistically different from those plots receiving 2 applications of zinophos at 3 lb per acre.











e) Table 6a shows that the application of each nematicide

resulted in significant yield increases of field corn. These yield increases were partly due to early control of T. christiei and partly due to control of other parasitic nematodes.

In addition, the number of T. christiei after 58 days was greater than that at 72 days. The reason for this is apparently due to the fact that at harvest the samples were taken to a 6-inch depth. As shown in the previous experiment, by crop maturity the majority of T. christiei are below this depth.

When cabbage was planted on the same plots following the corn and half the original number of plots retreated, none produced significant yield increases except those retreated with zinophos at 3 lb per acre and D-D at 25 gal per acre (Table 6c). The fact that the check plots yielded more than some of the treated plots suggests that cabbage, by virtue of their numerous rootlets, can withstand nematode damage better than corn. In addition, the larger initial populations of T. christiei may have influenced yields.

f) The counts of T. christiei on the final untreated crop of sweet corn are given in Table 6d. With the exception of Nemagon, the counts of all treatments are very similar at harvest, although the numbers 3 weeks after planting show considerable variation. The only statistically outstanding yield was obtained from the plots which had received 2 applications of zinophos at 3 lb per acre.










Table 6a.


A Comparison of the Effects Certain the Build-up of Trichodorus christi


Plot Numbers of T. christiei per 100 ml
No. Treatment After 58 days After 72 day


1 + 2 Check lOla/ 172

3 + 4 Zinophos
3 lb/acre 11 54

5 + 6 Nemagon
2 gal/acre 393 392

7 +8 D-D
15 gal/acre 721 391

9 + 10 D-D
25 gal/acre 833 462 a/ Average of 8 replicates. SYield data F 7 = 12.34**; i.e., significant at the 1% level.


Duncan Multiple Range Test on yield at Plot No.
2 1 5 6 7 3

13.833 17.375 27.438 31.438 33.250 33.625


Nematicides Have On ei on'Funks' Hybrid Field Corn. of soil Weight of 60 corn s (Harvest) plants 15.60 lb 34.82 lb 29.44 lb 34.57 lb 43.38 lb


the 5% level

8

35.875


4

36.000


10 9

41.563 45.188









A Comparison of the Residual Effects of a Split Application of Certain Nematicides on the Build-up of Trichodorus christiei on Cabbage.


No. Applicationsa


/ Numbers of T. christiei in 100
various intervals after At Fumigation 21 Days 50 Days


ml of soil at planting. 80 Days 112 Days(Harvest)


Check

Check

Zinophos

3 lb/acre

Nemagon

2 gal/acre

D-D

15 gal/acre

D-D

25 gal/acre Application -- made Application -- made


28

33

2 28

1 35 2 70 1 117 2 100 1 220 2 238 1 247
prior to the previous crop of prior to planting the cabbage.


Table 6b.


Plot No.


Treatment


1

2 3

4 5 6 7 8 9 10
8/ 1st
2nd


25 28 8 28

84 105 35

46 14 105

corn.


74

63 11 39 63 95 172 60 98

46


49 105 11

60 84 95

168 95

154 60


193 217 25

119 249 280

441 235 529 371








Effect of Nematicide Treatments on Total Number and Weight of Cabbage Heads on Three Harvest Dates.


No. Applications


1/27/66
No. Wt.(lb)


Harvest Date
2/18/66
No. Wt.(lb)


2/25/66
No. Wt. (lb)


Total Harvest
Harvest
No. Wt.(ib)


Check Check


Zinophos

3 lb/acre


5 Nemagon 2 10 6 2 gal/acre 1 11 7 D-D 2 26 8 15 gal/acre 1 15 9 D-D 2 58 10 25 gal/acre 1 30


Yield data F7 = 2.3096 *; i.e., significant


Plot No. 6


at the 5% level.


Duncan Multiple Range Test on yield at the 5% level.
5 8 2 1 4 10


31.125 31.425 32.025 35.425 35.525 36.475 37.050 39.125


Table 6c.


Plot


Treatment


21.2 29.5 58.7

27.3 12.6 13.0

39.2 21.3 85.9 38.0


83.9 84.4

116.6 79.1 64.6

72.6 71.8 61.4 91.0 86.7


36.7 27.8 33.9

39.5 48.5 38.9 45.5 45.4 47.4 23.5


141.8 141.7 209.2 145.9 125.7 124.5

156.5 128.1 224.3

148.2


52.3 56.075








Table 6d.


A Comparison of the Residual Effects Certain Nematicides Have on the Build-up of Trichodorus christiei On Unfumigated 'Gold Cup' Sweet Corn.


Plot Previous No. Applications No. of T. christiei Yield of Corn Ears No. Treatment per 100 ml of soil.
After 24 Days After 84 Days Wt. (lb)


Check Check


Zinophos

3 lb/acre


Nemagon


2 gal/acre D-D

15 gal/acre D-D

25 gal/acre


a/ Average of 4 replicates.
Yield data F 7 = 2.77*; i.e.,


Plot No.


1
33.8


significant at the 5% level.


Duncan Multiple Range Test on yield
4 7 6 5 8 34.5 34.8 35.0 39.3 40.0


at the
10
40.3


5% level.
2
40.5


14 63 200

119 112 42 105 53


33.8 40.5 49.1 34.5 39.3 35.0 34.8 40.0 40.9 40.3


9 3
40.9 49.1











Field Experiment 3. Table 7 shows that in the absence of a host there was a gradual mortality of T. christiei during the first 8 weeks. Between the eighth and twelfth weeks the mortality doubled that of the first 8 weeks. After 16 weeks, when the experiment was terminated, the T. christiei population had been reduced to 1/5 of the original number. A comparison of the survival of T. christiei under bare plots with bare plots in which organic matter had been incorporated (at the rate of 17 tons per acre) revealed no differences (Table 7). By contrast, Patrick et al. (23) obtained nematicidal substances from decomposing rye. It is possible that under summer conditions a more rapid decomposition of the organic matter might prove detrimental to T. christiei.


Table 7.


A Comparison of the Survival of Trichodorus christiei in the Presence and Absence of Organic Matter (0.M.) Under Clean Cultivation in the Field.


Time in Weeks
Treatment 0 3 1/2 8 12 16
Check 0.M. Check 0.M. Check 0.M. Check 0.M. Check 0.M.


No. of T. 414a/ 390 christiei/
100 ml of soil.

% reduction 0 0 from original population.

Soil Temp.(OF) 74


340 350


296 288


120 133


94 74


18 10 29 26 71 71 77 81 61


a/ Data from 4 replicates.








Greenhouse Experiments


Greenhouse Experiment 1. Of the four temperatures investigated, 80oF proved most favorable for reproduction by T. christiei (Table 8). There was, however, no statistical significance between the reproductive rates at 800 and 850F. Malek et al. (20) in New Jersey found 770F to be the optimum for T. christiei reproduction. Thus, the New Jersey and Florida populations of T. christiei appear to have similar optimum temperature requirements for reproduction.


Table 8. The Effect of Temperature on the Reproductive Rate of
25 Trichodorus christiei on Sweet Corn and Tomato After
8 Weeks.



Rep. I/ Rep. 2 Average
Temperature Corn Tomato Corn Tomato Corn Tomato

950F 5b/ 53b/ 583 2,844 294 1,449 90oF 2,253 10,500 5,103 7,489 3,678 8,995 850F 8,394 13,828 6,899 11,567 7,647 12,698 800F 9,891 15,402 10,626 12,788 10,259 14,095 a/ Replicate 1 ran from 8/1/65 to 9/25/65; and Replicate 2, from 11/5/65


to 1/2/66. Average count of 8 pots.
3
Corn F3 = 24.40
LSD
05 3,991.34


3 2** Tomato F3 = 63.2
3
LSD
05 2,591.45


The populations of T. christiei produced on Rutgers'tomato were greater in number than those produced on 'Gold Cup' sweet corn. As noted by Bird (4), the morphology of T. christiei was affected by the host upon which the specimens had been feeding. Of interest was the











presence of several males at 950F. Under field conditions males of this nematode are extremely rare.


Greenhouse Experiment 2. The data presented in Table 9 show that T. christiei and B. longicaudatus each parasitize and reproduce on pangolagrass, giving rise to as much as an eightfold increase in their numbers over a three-month period in sterile soil. In unsterilized soil taken from two different field locations, the populations did little more than maintain their numbers, which suggested the presence of a biological interaction. Consideration should be given to two factors in unsterilized soil, viz., (1) the presence of other plant parasitic nematodes and (2) the presence of certain nematode predators.

T. christiei proved more pathogenic to the pangolagrass root system than did B. longicaudatus (Table 10), but the latter was more pathogenic to sweet corn. The mixed population of plant parasitic nematodes present in "Pangola" and "Crop" soils undoubtedly contributed to the smaller root systems in these soils when compared to those in "Sterile" soil. Albeit, the terminal numbers of T. christiei and B. longicaudatus were larger in the "Sterile" soil.











Table 9. Plant Parasitic Nematode Populations in 100 ml of Soil From
Under Pangolagrass Followed by Sweet Corn and Vice Versa
Maintained in the Greenhouse for 3 Months.

Initially inoculated with 25 T. christiei Soil Host Nematode Stubby-root Lesion Stunt Sting Ring

"Sterile" Pangolagrass 14a/ - -
then Sweet Corn 907 - - -

"Pangola" Pangolagrass 11 39 - -
then Sweet Corn 151 5 - -

"Crop" Pangolagrass 14 42 2 - 83
then Sweet Corn 109 1 1 -"Sterile" Sweet Corn 54 - - -
then Pangolagrass 459 - - -

"Pangola" Sweet Corn 94 188 - -
then Pangolagrass 119 32 - -

"Crop" Sweet Corn 88 130 219 - 5
then Pangolagrass 105 25 70 -


Initially inoculated with 25 Belonolaimus Soil Host longicaudatus Nematode Stubby-root Lesion Stunt Sting Ring

"Sterile" Pangolagrass - - - 53
then Sweet Corn - - - 315

"Pangola" Pangolagrass - 46 - 7 4
then Sweet Corn 21 25 - 11

"Crop" Pangolagrass - 27 1 7 130
then Sweet Corn - 4 - - 67
"Sterile" Sweet Corn - - - 112
then Pangolagrass - - - 872

"Pangola" Sweet Corn 28 147 - 32
then Pangolagrass 60 49 - 42

"Crop" Sweet Corn 16 210 172 18 58
then Pangolagrass 112 25 144 6 14


/ Average of three samples.
each of the five 6-inch pot
cessed from each sample.


Samples collected by taking one probe from replicates. A 100 ml sub-sample was pro-












Key to Table 9:


Stubby-root Lesion

Stunt Sting Ring


= Trichodorus christiei = Pratylenchus brachyurus and
P. zeae
= Tylenchorhynchus spp. = Belonolaimus longicaudatus = Criconemoides spp.


Table 10.


Total Dry Weight (Grams) of Roots From 5 Replicates of Pangolagrass and Sweet Corn After 3 Months in Association with Trichodorus christiei and Belonolaimus longicaudatus.


Soil Crop Nematode Stubby-root Stubby-root Sting Sting
on 11/29/65 on 3/7/66 on 11/29/65 on 3/7/66

"Sterile" Sweet Corn 32.2a/ 21.4b/ 17.5a/ 18.7b/
Pangolagrass 14.8b/ 14.9a/ 25.8b/ 18.2a/

"Pangola" Sweet Corn 16.6 18.9 9.5 17.9
Pangolagrass 6.6 10.5 9.9 12.3

"Crop" Sweet Corn 17.1 16.2 12.2 15.1
Pangolagrass 12.6 9.6 10.7 18.1

a/ Same pots; planted first to sweet corn, then pangolagrass. b/ Same pots; planted first to pangolagrass, then sweet corn.











Greenhouse Experiment 3. The results of the experiment conducted to determine the effect of the competition by B. longicaudatus on the reproduction of T. christiei are presented in Table 11. After 45 days virtually no discernible effects of the competition were detected. In all probability, this was due to the low levels of initial inoculum. The data recorded 45 days after replanting these same pots suggest that T. christiei in the control pots had attained maximum levels of of infestation; i.e., + 1,000 individuals per 100 ml of soil, irrespective of the initial level of inoculum. Nevertheless, the presence of B. longicaudatus resulted in a marked reduction of the numbers of T. christiei. This reduction was more marked the greater the initial inoculum numbers, averaging up to 50 per cent. These differences were not statistically, significant, owing to wide variation in numbers and too few replicates. By contrast, the reproduction of B. longicaudatus was less effected the larger the initial numbers of T. christiei.

The average dry weights of the second sweet corn planting; i.e., from the 45th day to the 90th day, are given in Table 12. These data indicate that B. longicaudatus is much more pathogenic to sweet corn than is T. christiei (Fig. 3 and 4). In general the corn was damaged more by the combination of the species than by B. longicaudatus alone, though there were fewer numbers of B. longicaudatus in the combination.






Table 11.


The Interrelationship of Two Ectoparasitic Nematodes At Different Rates of Initial Inoculum While Feeding on Sweet Corn in 6-inch Plastic Pots.


AFTER 45 DAYS IN 100 ML OF SOIL Nematode Initial Rate of Inoculum 1 2 3 4 Average


Stubb -root a/ Sting_/ Stubby-root+Sting Stubby-root Sting
Stubby-root+Sting Stubby-root Sting
Stubby-root+Sting Stubby-root Sting
Stubby-root+Sting


25 25
25+25 50 50 50+50 100 100
100+100 250 250
250+250


18 28 39+25 46 91
53+56 112 105 81+133 312 375 287+392


53 35
21+11 63
119 46+21 112
196 49+196 207 469
186+161


60 39 60+70 60 53
84+116 112 179 60+119 224
413 382+291


95 38 63+28 77 67 133+140 196 196 158+242
210 256 130+102


57 35
46+34 60 83 79+83 133 169 87+173 238 378
246+239


AFTER 90 DAYS IN 100 ML OF SOIL


Stubby-root 25 1,
Sting 25
Stubby-root+Sting 25+25
Stubby-root 50 1,
Sting 50
Stubby-root+Sting 50+50
Stubby-root 100 1,
Sting 100
Stubby-root+Sting 100+100
Stubby-root 250 1,
Sting 250
Stubby-root+Sting 250+250
a/ Stubby-root = Trichodorus christiei. E/ Sting = Belonolaimus longicaudatus.


106 315 412+210 582 651 721+396 897 546 966+574 512 991 308+448


1,407 574
1,421+322 1,001 501 805+329 665 1,117
546+917 1,456 438
585+756


525 578
518+392 896
602 284+305 1,194 1,001
140+718 399
564 273+819


1,162
217 480+144 987 378
991+284 770
1,099
536+473 599
1,236 851+515


1,050 421
708+267 1,116.5 533
700+328.5 1,131.5
941
547+670.5
991.5
807
504+634.5


AFTER 90 DAYS IN i00 ML OF SOIL
























Table 12.


Effect of Trichodorus christiei and Belonolaimus longicaudatus on the Dry Weights (Grams) of the Second Crop of Sweet Corn. The Corn Was Harvested 45 Days after Replanting in Pots in Which the Nematodes had Previously Been Established.


T. christiei
Initial T. christiei B. longicaudatus + Inoculum B. longicaudatus 25 Tops 4.0 a/ 3.6 3.4

Roots 2.9 2.3 2.0

50 Tops 3.9 3.5 3.7

Roots 3.2 2.7 2.3 100 Tops 3.4 2.8 3.4 Roots 2.7 2.2 1.9 250 Tops 4.0 1.7 2.5 Roots 2.3 1.2 1.6 Check Tops 4.1

Roots 3.9

a/ Average of 4 replicates with three plants per replicate.















































Fig. 3 - The effect of Belonolaimus longicaudatus on
the growth of sweet corn 45 days after replanting in pots initially inoculated with (left to
right) 250, 100, 50 and 25 specimens.















































Fig. 4 - Growth of sweet corn 45 days after replanting
in pots initially inoculated with (left to right)
250, 100, 50 and 25 Trichodorus christiei.











Greenhouse Experiment 4. Results of the comparison of the reproductive rates of T. christiei in D-D fumigated soil with unfumigated soil are given in Table 13. After 8 weeks the number in the unfumigated soil was 1.7 times that in fumigated soil, due to higher initial numbers of T. christiei in the unfumigated soil. However, during the following 8 weeks the situation was reversed with D-D fumigated soil having

1.6 times more T. christiei. This demonstrated that a build-up of T. christiei, similar to that seen in the field, can take place when this nematode is inoculated into D-D fumigated soil in greenhouse pots.


Table 13. The Reproductive Rate of Trichodorus christiei in
D-D Fumigated Soil and Unfumigated Soil in Greenhouse Pots.



Numbers of T. christiei in 100 ml of soil Treatment Initially After 8 weeks After 16 weeks Host: Oats Host: Sweet Corn
Unfumigated 10 72a/ 560 D-D at 30 gal
per acre 2 42 910


/ Average of 2 samples which consisted of a probe taken from each of
the 6 replicates.


Greenhouse Experiment 5. The data obtained in the experiment conducted to determine whether oligochaetes have an effect on the reproduction of T. christiei are presented in Table 14. It should be noted that, although initially inoculated into steam-sterilized soil, the oligochaetes were able to feed and reproduce. However, no statistical difference











could be detected on the reproduction of T. christiei, under the conditions of the experiment, as a result of the increase in oligochaetes.


Table 14. Effects of Oligochaetes on Number Of Trichodorus christiei After 8 Weeks.



No. T. christiei ' No. Oligochaetes Inoculum
Inoculum per 100 ml of soil per 100 ml of soil


100 T. christiei alone 1,419a/ 100 T. christiei + 100 oligochaetes 1,326 806 100 T. christiei + 500 oligochaetes 1,285 1,238 a/ Average of 5 replicates.


The results of the study conducted on the possible predaceous habits of Eudorylaimus simplex, Aporcelaimellus obscurus, and Mylonchulus parabrachyurus on T. christiei are shown in Table 15. Of these three free-living nematodes, only the presence of A. obscurus resulted in reduced numbers of T. christiei. However, in a second test when specimens ofA. obscurus were inoculated into a previously established population of T. christiei, no such differences were observed (Table 16). From these tests it was concluded that none of these nematodes affect population levels of T. christiei.











Table 15. Effects of Three Possible Predaceous Nematodes on
Populations of Trichodorus 'christiei.

Numbers per 100 ml of soil after: 8 Weeks 25 Weeks

100 T. christiei alone 270 147 100 T. christiei + 266 228 25 Eudorylaimus simplex 0 84 100 T. christiei + 238 144 100 Eudorylaimus simplex 4 133 25 Eudorylaimus simplex alone 4 109 Numbers per 100 ml of soil after: 8 Weeks 16 Weeks
100 T. christiei alone 203 591 100 T. christiei + 217 353 25 Aporcelaimellus obscurus 0 66 100 T. christiei + 154 273 100 Aporcelaimellus obscurus 4 96 Numbers per 100 ml of soil after:
8 Weeks 25 Weeks

100 T. christiei alone 270 147 100 T. christiei + 207 196 25 Mylonchulus parabrachyurus 0 91 100 T. christiei + 221 210 100 Mylonchulus parabrachyurus 4 203 25 Mylonchulus parabrachyurus alone 0 420


Table 16. Effect of Aporcelaimellus obscurus on
Established Populations of Trichodorus christiei. Numbers per 100 ml of soil after 12 weeks:
Inoculum 1 2 3 Average T. christiei alone 805 630 735 723 T. christiei + 833 578 704 705 100 A. obscurus 315 305 291 304











Greenhouse Experiment 6. Results of the experiment conducted to determine which of four potting containers provided the most ideal environment for the reproduction of T. christiei are given in Table 17. In the greenhouse it was found that the glazed crock was statistically superior to plastic, to sub-surface irrigated clay, and to surface irrigated clay pots; there being no statistically significant difference among these latter three. In the constant temperature chamber, however, the plastic container was superior. In this case there was no statistically significant difference between the glazed crock and two types of clay pots. Similar data were obtained in a repetition of the constant temperature experiment in a different kind of chamber. Figure 5 shows the root system of sweet corn in three of the containers. Note the preponderance of roots lining the walls of the clay pot.

These data suggest that, in conditions where the greatest variable is temperature, glazed crocks are desirable, while in those conditions in which moisture is the most important variable, the plastic pots are best suited for rearing T. christiei.


Table 17. Numbers of T. christiei in 100 ml of Soil from Four
Different Containers Using Sweet Corn as a Host After
a Period of 8 Weeks.
Numbers of T. christiei per 100 ml Average dry weight Container of soil of roots per pot Greenhouse Chamber Greenhouse Chamber Plastic Pot 9398/ 1,209b/ 1.7 2.6 Glazed Crock 1,303 751 2.0 2.4 Clay Pot(surface
irrigated) 608 783 3.5 1.9 Clay Pot(sub-surface
irrigated) 885 796 1.7 3.5 a/ Two plants per pot.
b/ Four plants per pot.


















































Fig. 5. - Root growth of sweet corn after 8 weeks in the
greenhouse in a clay pot, glazed crock, and
plastic pot.











Duncan's Multiple Range Test: Surface Sub-Surface Greenhouse = Irrigated Irrigated Clay Clay Plastic Glazed 608 885 939 1,303



Glazed Surface ,Sub-Surface Plastic Growth chamber = Irrigated Irrigated ....Clay Clay ...

751 786 796 1,209




Laboratory Experiments


Laboratory Experiment 1. The results of a study on the life cycle of T. christiei following inoculation of 10 gravid females onto tomato seedlings growing at 80oF are given in Table 18. The different stages in the life cycle (Fig. 6) were first recorded on the following days after inoculation:

1st Larval stage . . . . . . . 4 days 2nd Larval stage . . . . . . . 4-5 days 3rd Larval stage . . . . . . . 7 days 4th Larval stage . . . . . . . 10 days Adult . . . . . . . . . . . . 14 days

Gravid female . . . . . . . . 17-18 days The criterion used to determine the stage of development was the size of the gonadal primordium. The reason for this can be seen in Fig. 7, which shows the size variation in adult T. christiei from two different habitats. Measurements of 10 specimens were made at molting in 1 per cent formalin. The average sizes were:










Table 18.


Study of the Life Cycle of the Offspring of 10 Gravid Female Trichodorus christiei at 80oF While Feeding on 'Homestead 24' Tomato.


No. Days Original Life Cycle Stage Av. No.
After Adults of
Inoculation Test Surviving Adult 4th 3rd 2nd 1st Offspring


3.0 3.0

7.3 7.0

3.8
7.5

5.0 4.5


16.5
5.5a/


27.3 22.0
15.5 11.5


13.0 5.0


47.5a/ 28.3 27.5 18.8


1.5 1.5 1.0 1.0

3.3 7.1 3.0 3.0

1.8 11.3
2.5 4.0 2.3 16.1
- 6.5

4.3 28.6
- 28.5

1.8 24.4
- 10.5

3.0 43.0
- 35.5

4.8 48.1 0.5 35.0 2.8 61.4 2.0 49.5 3.3 54.4 0.5 33.5 1.0 40.3 0.5 76.0 2.3 37.6 0 88.5

1.0 51.9 0 32.0 2.5 81.3 1.5 39.0 2.8 125.0


females recovered.


3.8


9.5
1.5

1.5 12.3
1.0 5.5 7.8 16.5
7.0 21.5 8.8 13.8 3.5 7.0

3.0 15.0 22.0
- 22.5 13.0

7.3 17.5 18.5 10.5 8.5 15.5 17.3 26.8 14.5
21.0 10.0 16.5 14.8 21.8 14.5 19.5 5.5 8.0 16.0 15.8 7.5
44.0 25.5 6.0 2.3 13.5 9.5 10.0 7.5 50.0 22.0 9.0 10.3 15.8 17.3 7.5 11.0 12.5 6.5 2.0


5.5
4.5


3.3 3.5

8.0 5.5

4.0 3.5

4.5 4.5


3.0 7.0 3.5 3.0 3.8 0.5 3.0


18


a/ Gravid



























(a) (b) (c) (d)






















0 0.7mm
(e) (f)






Fig. 6 - Stages in the life cycle of Trichodorus christiei at (IOX);
(a) egg, (b) first larval stage, (c) second larval stage;
(d) third larval stage, (e) fourth larval stage, and (f)
adult female.














































Fig. 7 - Variation in size between adults of Trichodorus
christiei collected from two locations near
Sanford, Florida.











Gonad
Length a b c Length lst-2nd 0.311mm 20.7 2.9 124.3 .008mm 2nd-3rd 0.41mm 16.2 3.7 98.3 0.03mm 3rd-4th 0.51mm 16.3 4.3 126.6 0.04mm 4th-Adult 0.70mm 16.0 5.6 100.0 0.23mm


Laboratory Experiment 2. The molting of T. christiei proved different than that of any other plant parasitic nematode. Prior to molting the specimens of T. christiei became quiescent. The posterior intestinal contents were ejected,while the remainder became granular with the denser material collecting around the perimeter of the intestine. This collection was most pronounced ventrally. Quiescence was terminated by a movement of the neck region from side to side, with the inner cuticle of the neck region becoming markedly wrinkled on the side to which the head was tilted. Concurrently the cuticle was, at times, stretched over the head as seen in Fig. 12. During these movements, esophageal palpitations were observed and the gland nuclei were clearly defined.

The old cuticle loosened,giving rise to a small overlap at the

head and tail. This was followed by a break in the old cuticle circumventing the body in the region of the base of the onchiostyle (Fig. 8). By contraction on the head a "hood" was formed on the dorsum which usually remained attached to the old cuticle at a single point (Fig. 9). This formation of a hood was somewhat similar to that recorded by Lapage

(19) for infective larvae of Trichostrongylus, Haemonchus and Ostertagia.
















w 0













N,
















Fig. 8 - Break in the old cuticle prior to molting of
its "hood" by Trichodorus christiei.














































Fig. 9 - A molting specimen of Trichodorus christiei.
Note the absence of the onchiostyle in the
"hood."










The head of T. christiei was then withdrawn from the hood.

Note in Fig. 9 that the portion of the body which had emerged from the old cuticle was greater in diameter than the portion still contained within both cuticles. The old cuticle was then shed within a few minutes by movements of the animal.

Detailed examinations of the head revealed the amphidial linings to be clearly visible, but no portion of the onchiostyle was found in any of the four molted cuticles. The tripartite onchiostyle of Trichodorus is unique among the plant parasitic nematodes as is the molting process.


Laboratory Experiment 3. The reproductive rates of individual specimens of T. christiei are given in Table 19. A marked increase (76%) in the numbers of offspring recovered occurred between the 16th and 17th days. Thereafter, until the 19th day, the increase was gradual. Between the 19th and 38th days, the population increased 12.2 times. Thus, although as many as 30 offspring were recovered from a single specimen in one generation, the average reproductive rate of this nematode is 12 offspring per 19 days at 80 0F using 'Homestead 24' tomato as the host plant. This figure was further substantiated by the data in Table 20a, prior to the influence of population pressures.


Laboratory Experiment 4. A study was made on the effect of varying initial inoculum numbers on the reproductive rate of T. christiei. The data from this study, using 6 'Homestead 24' tomato seedlings per 4-inch plastic pot of sterile soil,are presented in Table 20a. After









Table 19.


Reproductive Rate of Individual Specimens of T. christiei While Feeding on 'Homestead 24' Tomato Seedlings Growing at a Constant Temperature of 800-.


Days after Test Surviving original Numbers of the various stages inoculation number individuals divided in the life cycle recovered; mean by no. pots with of pots with offspring Dailya/ Daily Offspring Adult 4th 3rd 2nd 1st Total Max. Avg. 16 1 5/5 -- 2.2 2.6 0.8b/ -- 5.6 11 5.9

2 2/6 -- 0.7 2.8 2.0 0.7 6.2 12

17 1 4/4 0.9 3.2 4.9 3.9 -- 12.8 27 10.4

2 1/5 0.4 1.8 3.4 2.0 0.4 8.0 16

18 1 5/3 0.6 4.0 5.0 2.8 -- 12.4 29 11.0

2 2/6 0.2 3.2 4.5 1.8 0 9.7 17

19 1 4/6 2.7 2.8 2.8 2.0 -- 12.4 21 11.6

2 1/6 1.7 4.3 3.7 2.3 0.8 12.8 30

38 1 163 352
141
2 119 219 a/ Represents the maximum number of offspring from a single individual. / In the first test the figure given in the 2nd larval stage column represents the combined total
of 1st and 2nd larval stages.












Table 20a.


Reproductive Rates of Trichodorus christiei at Different Rates of Infestation Maintained at 800F.


Inoculum Number of T. christiei per 4-inch pot ( 400 ml)
level
At 21 Increase per At 42 Increase per Days individual Days individual 25 274a/ 10.9 4,092b/ 163.7 100 1,253 12.5 5,786 57.8 400 1,823 4.6 7,030 17.6 a/ Average of 6 replicates.
5/ Average of 12 replicates.






Table 20b. Average Weights of 'Homestead 24' Tomatoes Inoculated with Trichodorus christiei.


Fresh Weight (grams) Dry Weight (grams)
After 21 days After 42 days After 21 days After 42 days Tops 3.2a/ 8.1b/ 0.167 0.675 Roots 0.7 1.60 0.062 0.213 Tops 3.5 7.8 0.200 0.658 Roots 0.8 1.55 0.073 0.224 Tops 2.7 6.3 0.150 0.450 Roots 0.7 1.45 0.055 0.199 a/ Average of 6 pots; 6 plants per pot. '/ Average of 12 pots; 6 plants per pot.


25 100



400











21 days the greatest increases were found in those pots initially inoculated with 100 specimens. This was closely followed by those with 25 specimens. After 42 days the pots originally having 25 individuals produced increases of almost three times the rate of those with 100 and over nine times the rate of those with 400, thus demonstrating the importance of population pressures on the reproductive rate of T. christiei. Alhassan and Hollis (1) attributed the slower rise of higher initial inoculum numbers of T. christiei to an interaction with the host. Since there was an absence of necrosis, these authors have termed this relation one of "balanced parasitism." Figs. 10 and 11 show T. christiei feeding on tomato roots in agar.

Data relating to seedling weights are given in Table 20b. No

statistical difference was found between the fresh weights of the tops or the roots when the treatments were analyzed after 21 days and 42 days, respectively. Alhassan and Hollis (1) obtained similar results on cotton seedlings analyzed 21 days after inoculation with 0, 100, and 400 specimens of T. christiei. Nevertheless, their conclusion was that "seedling weights were related inversely to both initial and final populations of the nematode." The same conclusion may be drawn from the above experiment on 'Homestead 24' tomatoes after 42 days. Laboratory Experiment 5. Results from the investigation into the possibility of resistance in the eggs of T christiei to D-D fumigation are given in Tables 21a and b. Under the conditions of the experiment; i.e., recording the nematode population 2 weeks after fumigation, and 3 weeks after planting a susceptible host, it was found that whenever
















































Fig. 10 - Characteristic position of Trichodorus christiei
while feeding; i.e., the head at right angles to
the cell upon which it is feeding.





















































Fig. 11 - Trichodorus christiei feeding upon the
root-cap of tomato in agar.








Table 21a.


Effects of D-D Fumigation on Trichodorus christici, Belonolaimus longicaudatus and Hoplolaimus galeatus in Soil Contained in Pint Jars.


Average numbersa/ of nematodes per 100 ml of soil 2 weeks after fumigation (column 1), and 3 weeks after planting tomato (column 2) Trichodorus Belonolaimus Hoplolaimus Treatment christiei longicaudatus galeatus
1 2 1 2 1 2


1) Check 2) Mineral spirits
690 gal/acre


3) D-D
10 gal/acre

4) D-D
15 gal/acre

5) D-D
30 gal/acre

6) D-D
50 gal/acre

7) Emulsified D-D
10 gal/acre

8) Emulsified D-D
15 gal/acre

9) Emulsified D-D
30 gal/acre

10) Emulsified D-D
50 gal/acre
a/ Average of 5 replicates.


349 120 198 156 35 11 231 160 60


253


111i 141 109 32


24 143 152 117


233


154 68


49


5 0


84 44


232 88 37 35


4 4


99 50 28 36













Table 21b.


Effects of D-D Fumigation on a Pure Population of Trichodorus christiei in Soil Contained in Pint Jars.


Treatment Average numbersa/ of T. christiei per 100 ml of soil 2 weeks after fumigation, and 3 weeks after planting.

1) Check 354 420 2) D-D Emulsified
25 gal/acre 51 167 3) D-D Emulsified
45 gal/acre 23 37 4) D-D Emulsified
56.25 gal/acre 0 0 5) D-D Emulsified
75 gal/acre 0 0 a/ Average of 4 replicates.











the adults and larvae were killed no progeny of T. christiei could be found. This indicated that the eggs were killed.

Adults and larvae were found to survive following applications of D-D at 50 gal per acre to the jars, whether applied in mineral spirits or in a water-emulsified state. However, it would appear that many of these surviving specimens were incapable of infecting susceptible hosts. This phenomenon appeared more pronounced at the lower application rates of D-D.

The data in Table 21a indicate that T. christiei is more tolerant to D-D fumigation than is B. longicaudatus or Hoplolaimus galeatus. This experiment also demonstrates the marked susceptibility of B. longicaudatus to D-D fumigation.


Laboratory Experiment 6. The survival of T. christiei in the absence of a host was studied at a constant temperature of 80oF (Table 22). The rate of mortality of T. christiei was slightly greater in field soil than that of a pure population which previously had been established in sterile soil. This difference, however, was considered insufficient to indicate the presence of a successful predator.

A comparison of the data obtained for survival at 800F (Table 22) with those obtained in the field (Table 8) shows the rates of mortality at 800F to be slightly greater during the first 8 weeks. Thereafter, the field population declined much more rapidly until at the 16th week there was a difference of 20 per cent.

Often fallowing is associated with dry tillage. After allowing 6-inch clay pots of soil containing T. christiei to dry out in an air-conditioned laboratory to a moisture level as low as 1.7 per cent,











it was found that live specimens could still be recovered. Specimens of T. christiei recovered from this soil characteristically possessed a thickened cuticle (Fig. 12). This cuticular thickening suggests that


Table 22. Survival of Trichodorus christiei in the Absence of a Host in a 6-inch Pot at 800F.



Pure Population Field Soil
Numbers of T. Numbers of T.
Weeks christiei per Percentage christiei per Percentage elapsed 100 ml of soil reduction 100 ml of soil reduction


0 205a/ 0 % 69 0 %

2 196 3 66 3 4 157 23 47 32 8 140 32 42 40

12 115 44 16 88 57

a/ Average of two 100 ml samples. T. christiei can adapt itself to survive at low soil moisture levels. No living specimens were found in soil with a moisture level of 0.9 per cent.


Laboratory Experiment 7. The green food dye proved most satisfactory

for staining specimens of T. christiei, but the degree of staining

varied with different individuals. The dye dispersed evenly throughout the specimensonly after they were placed in a 1 per cent formalin solution. This stain was particularly successful in bringing out the

reproductive system and cuticular structures.






68






















































Fig. 12 - Water mount of a live Trichodorus christiei recovered
from soil with a moisture level of 1.7 per cent.











Among the cuticular structures not observed on unstained specimens were: (1) A series of dorsal and ventral pores spaced along the length of the body in the cuticle which appeared to be associated with a similar series of hypodermal cells; (2) two caudal pores which terminated within the cuticle; and (3) the posterior cephalids, completely encircling the cephalic region.


The Build-up of Trichodorus christiei

The data presented above do not directly answer the question of

what causes the build-up of T. christiei following fumigation; nevertheless, certain conclusions may be drawn. Indications are that predaceous nematodes play a minor role, as shown by the slow decline in high populations of T. christiei, in the absence of a host in the field. No statement can be made concerning possible effects of predaceous fungi.

There does not appear to be a stage in the life cycle of T.

christiei resistant to D-D fumigation. Complete eradication of nematodes has seldom been obtained in the field with soil fumigation. At Sanford, Florida, the control of T. christiei is made more difficult due to its presence at relatively greater depths than other plant parasitic nematodes.

Crops on fumigated soil have improved growth over those on unfumigated soil, particularly during the seedling stages. This is due, in part, to the control of B. longicaudatus by fumigation, and as a result competition between this nematode and T. christiei is absent in fumigated soil. Under these conditions, T. christiei can attain a higher reproductive rate as shown by the greenhouse experiment presented above.











It is interesting to note that T. christiei reaches its reproductive potential (i.e., 12 progeny per 19 days at 800F) at low levels of inoculum, but that population pressures are soon observed in 4-inch pots. The reason for this probably lies in the available food supply and its effect upon the reproductive rate. It is the considered opinion of the author that the crux of the problem of the build-up of T. christiei following fumigation is, in part, due to the increased number of available feeding sites.















SUMMARY


Population studies were made on the plant parasitic nematode,

Trichodorus christiei, following field applications of D-D soil fumigant on fall, spring, and summer vegetable crops. Results showed that T. christiei could be found in greater numbers in fumigated soil than in unfumigated soil within 7 weeks after planting. Nevertheless, due to better seedling growth, the fumigated areas out-yielded those unfumigated. This build-up of T. christiei was not seen on those plots treated with the organo phosphate nematicide zinophos. This latter nematicide was also found to have a significant residual effect on subsequent crop yields.

Indications are that T. christiei does not possess a resistant stage to D-D fumigation. However, consideration should be given to the fact that at harvest there is a greater population of T. christiei at soil depths below 6 inches than above 6 inches.

The rate of mortality of T. christiei in the absence of a host, whether or not organic matter was incorporated, did not suggest the presence of a predator. Nor were populations of T. christiei affected by association with three free-living nematodes or an oligochaete in sterile soil.

Both T. christiei and Belonolaimus longicaudatus were found to parasitize pangolagrass. The effect of these two nematodes upon one another was studied when in combination on sweet corn. The numbers of











T. christiei were greatly reduced. Belonolaimus longicaudatus was found to be more pathogenic to sweet corn than T. christiei. In general the corn was damaged more by the combination of the species than by B. 2longicaudatus alone, though there were fewer numbers of B. longicaudatus in the combination.

The type of potting container was found to significantly affect the population numbers of T. christiei. In the greenhouse a glazed crock provided the best environment of those studied, while in a growth chamber a plastic container provided the best conditions.

A study of the reproduction of T. christiei under four constant

temperatures, viz., 95*, 90*, 85* and 800, showed the latter to be the most suitable. The effect of initial inoculum numbers on the reproductive rate of T!. christiei was studied in sterile soil at 800F. After 42 days the pots originally having 25 individuals produced increases of almost 3 times the rate of those with 100, and over 9 times the rate of those with 400, thus demonstrating the importance of population pressures as it is affected by the available food supply.

Single T. christiei ,specimens feeding on tomato seedlings growing

at 80aF were found to give rise to an average of 12 progeny every 19 days. At this temperature the life cycle of T. christiei from egg to egg was found to be 17-18 days. Eclosion from the egg took place on the fourth day in the first larval stage which molted shortly thereafter. The third larval stage was first observed on the seventh day, after inoculating a gravid female onto tomato seedlings; the fourth larval stage, on the tenth day and the adult emerged after 2 weeks. The stage in the life cycle of T. christiei can best be judged by the size of the





73




gonadal primordium. Molting of T. christiei is described. This is the first record of a plant parasitic menatode not shedding its stylet, or any part of it, during molting.

Future work on the bionomics of T. christiei following fumigation should be directed towards the effects the improved host growth have on the reproduction of this nematode.















LITERATURE CITED


1. Alhassan, S. A., and J. P. Hollis. 1966. Parasitism of Trichodorus christiei on cotton seedlings. Phytopathology. 56: 573-574.

2. Allen, M. W. 1957. A review of the nematode genus Trichodorus with descriptions of ten new species. Nematologica. 2: 32-62.

3. Barker,K. R., and Gayle Worf. 1964. Parasitism of southern stock azaleas in Wisconsin by Tylenchorhynchusclaytoni, Trichodorus
christiei and Meloidogyne incognita. (Abstr.) Phytopathology.
54: 887.

4. Bird, G. W. 1966. Influence of host and geographic origin on populations of Trichodorus christiei. (Abstr.) Nematologica.
12: 88.

5. Chen, T. A., and W. F. Mai. 1965. The feeding of Trichodorus christiei on individually isolated corn root cells. (Abstr.)
Phytopathology. 55: 128.

6. Chitwood, B. G., and B. A. Oteifa. 1952. Nematodes parasitic on plants. Ann. Rev. Microbiol. 6: 175-177.

7. Christie, J. R. 1959. Plant Nematodes Their Bionomics and Control. Gainesville, Fla. Univ. Fla. Press. 1-256.

8. Christie, J. R., and V. G. Perry. 1951. A root disease of plants
caused by a nematode of the genus Trichodorus. Science. 113:
491-493.

9. Christie, J. R., and V. G. Perry. 1951a. Removing nematodes from
soil. Proc. Helminthol. Soc. Wash. 3: 69-72.

10. Christie, J. R., and V. G. Perry. 1959. Mechanism of nematode
injury to plants, p. 419-426. In C. S. Holton et al. (ed.), Plant Pathology Problems and Progress 1908-1958. Madison, Wisc.: Univ.
Wisconsin Press.

11. Cobb, N. A. 1913. New nematode genera found inhabiting fresh
water and non-brackish soils. J. Wash. Acad. Sci. 3: 432-444.

12. Esser, R. P., and E. K. Sobers. 1964. Natural enemies of nematodes. Soil and CropSci. Soc. of Fla. 24: 326-353.











13. Haasis, F. A., J. C. Wells, and C. J. Nusbaum. 1961. Plant
parasitic nematodes associated with decline of woody ornamentals
in North Carolina and their control by soil treatment. Plant
Disease Reptr. 45: 491-496.


14. Harrison, D. S.,and R. E. Stall. 1961.
tanks for studying soil-borne diseases.
1: 24-25.


Constant-temperature Trans. Am. Soc. Agr. Eng.


15. Hoff, J. K.,and W. F. Mai. 1964. Influence of soil depth and
sampling date on population levels of Trichodorus christiei.
Phytopathology. 54: 246.


16. Hutchinson, M. T.,and H. T. Streu.
nematodes. Nematologica. 5: 149.


1960. Tardigrades attacking


17. Jenkins, W. R. 1964. A rapid centrifugal-flotation technique for
separating nematodes from soil. Plant Disease Reptr. 48: 692.


18. Krusberg, L. R.,and J. N. Sasser. 1956.
ship of the lance nematode in cotton roots.
505-510.


19. Lapage, G. 1935.
Parasitology. 27:


Host-parasite relationPhytopathology. 46:


The second ecdysis of infective nematode larvae. 186-206.


20. Malek, R. B., W. R. Jenkins,and Ellen M. Powers. 1965. Effect
of temperature on growth and reproduction of Criconemoides curvatum
and Trichodorus christiei. (Abstr.) Nematologica. 11: 42.


21. Martin, G. C.


1966. Personal communication.


22. Micoletzky, H. 1922. Die freilebenden Erd-Nematoden. Archiv. f.
Naturgesch. 87: 1-650.

23. Patrick, Z. A., R. M. Sayre,and H. J. Thorpe. 1965. Nematocidal
substances selected for plant parasitic nematodes in extracts of
decomposing rye. Phytopathology. 55: 702-704.

24. Perry, V. G. 1953. Return of nematodes following fumigation of
Florida soils. Proc. Fla. Hort. Soc. 66: 112-114.

25. Rhoades, H. L. 1964. Effect of Crotalaria spectabilis and
Sesbania exaltata on nematode populations and subsequent yield of
snap beans and cabbage. Proc. Fla. State Hort. Soc. 77: 233237.

26. Rhode, R. A.,and W. R. Jenkins. 1957. Effect of temperature on
the life cycle of the stubby-root nematode. (Abstr.) Phytopathology. 47: 29.











27. Rhode, R. A.,and W. R. Jenkins. 1957a. Host range of Trichodorus
sp. and its host-parasite relationship on tomato. Phytopathology.
47: 295-298.

28. Russell, C. C. 1962. The embryology and parasitic habit of
Trichodorus christiei Allen with observations of predators.
Unpublished Masters Thesis, Univ. Florida. Gainesville, Florida.

29. Russell, C. C.,and V. G. Perry. 1966. Parasitic habit of
Trichodorus christiei on wheat. Phytopathology. 56: 357-358.

30. Schaerffenberg, B. 1950. Untersuchungen uber die Bedeutung der
Enchytraeiden als Humusbildner und Nematodenfeinde. Z. Pflanzenkrankh. Pflanzenschutz. 57: 183-191.

31. Thomason, I. J. 1959. Influence of soil texture on development
of stubby-root nematode. (Abstr.) Phytopathology. 49: 552.

32. Thorne, G. 1932. Specimensof Mononchus acutus found to contain
Trichodorus obtusus, Tylenchus robustus and Xiphinema americanum.
(Abstr.) J. Parasitol. 19: 90.

33. Thorne, G. 1939. A monograph of the nematodes of the superfamily
Dorylaimoidea. Capita Zool. 8: 1-190.

34. Thorne, G. 1961. Principles of Nematology. New York: McGraw
Hill. 1-553.

35. Zuckerman, B. M. 1961. Parasitism and pathogenesis of the
cultivated cranberry by some nematodes. Nematologica. 6: 135143.














BIOGRAPHICAL SKETCH


Henry Vintcent Newton Morton was born on September 15, 1936, in Johannesburg, S. Africa. After attending school at St. Johns College for 10 years, he assumed the job of farm-manager on a dairy farm in Natal for a year before enrolling, in 1955, in the University of Natal at Pietermaritzburg.

In 1959, he received a Bachelor of Science in Agriculture with a major in Horticulture.

In 1960, he travelled to England where he joined the staff of

Imperial Chemical Industries at Jealotts Hill Research Station, Berkshire, working with weedkillers.

In May, 1961, he immigrated to the United States by way of Canada and worked for Pulitzer Groves as grove foreman. In 1962, he became assistant to Dr. M. Cohen, Pathologist, at the Indian River Field laboratory, Fort Pierce, Florida.

In January, 1963, he enrolled in the Department of Fruit Crops at the University of Florida, where, with the aid of a Research Assistantship, he graduated with a degree of Master of Science in Agriculture in August, 1964.

In September, 1964, he enrolled in graduate work in Nematology. From May, 1965, through August, 1966, he conducted research at the Central Florida Experiment Station, Sanford, Florida.











He returned to Gainesville to aid Professor Thorne in teaching the first course offered in Tropical Nematology at the University of Florida.

He completed the requirements leading to the Doctor of Philosophy Degree in June, 1967.

The author is a member of Alpha Zeta, Gamma Sigma Delta and

Sigma Xi. In addition, he holds memberships in the.Society of Nematologists, American Society of Horticulture and Florida State Horticultural Society.

In January,1967, he married Virginia Ann Martin and adopted her two sons, Stephen and Scott.











This dissertation was prepared under the direction of the

chairman of the candidate's supervisory committee and has been

approved by all members of that committee. It was submitted to

the Dean of the College of Agriculture and to the Graduate Council, and was approved as partial fulfillment of the requirements for the

degree of Doctor of Philosophy.


June 20, 1967



Dean, College of Agriculture




Dean, Graduate School Supervisory Committee:




Co- C airman


Co-Ch TZ
c z. - e




Full Text

PAGE 1

BIONOMICS AND LIFE CYCLE OF Trichodorus christiei By HENRY VINTCENT MORTON A DISSERTATION PRESENTED TO THE GRADUATE COUNCIL OF THE UNIVERSITY OF FLORIDA JN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA June, 1967

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ACKNOWLEDGMENTS The author wishes to thank all those who helped in making this dissertation a realization. In particular, supervisory committee co-chairman Professor V. G. Perry of the Department of Entomology for his guidance and encouragement in this work and for making available a teaching assistantship under visiting Professor G. Thome, through the auspices of the Tropical Nematology Program, and to Dr. H. L. Rhoades, Assistant Nematologist , at the Central Florida Experiment Station, for the generous gift of his time, knowledge and facilities during the sixteen months the author spent at Sanford, made possible through the financial support of the Station. Professor A. H. Krezdorn, Head of the Department of Fruit Crops, and Dr. G. C. Smart, Jr., Assistant Nematologist, Agricultural Experiment Station, for their helpful suggestions and criticisms in the writing of this dissertation. Dr. H. N. Miller, Plant Pathologist, Agricultural Experiment Station, for the liberal use of his laboratory facilities and photomicrographic equipment. To his wife, Virginia, sincerest appreciation for her patience and understanding. Lastly, to his parents, Commander and Mrs. E. B. Martino who were always ready with financial support when needed. ii

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TABLE OF CONTENTS Page ACKNOWLEDGMENTS ii LIST OF TABLES vi « LIST OF FIGURES ix INTRODUCTION ' 1 REVIEW OF LITERATURE 2 MATERIALS AND METHODS 6 FIELD EXPERIMENTS 6 Field Experiment 1. Comparison of the effects of 1, 3dichloropropene 1, 2-dichloropropane (D-D soil fumigant) fumigation with no fumigation on the population of Tricho dorus christiei on fall, spring and summer crops ... 6 Field Experiment 2. A split-plot experiment designed to study the residual effect of three nematicides in the control of Trichodorus christiei. 7 Field Experiment 3. The survival of Trichodorus christiei in the field in the absence of a host .... 9 GREENHOUSE EXPERIMENTS 9 Greenhouse Experiment 1. Effects of four temperatures on the reproductive rate of Trichodorus christiei ... 9 Greenhouse Experiment 2. A test designed to determine the host status of pangolagrass , Digitaria decumbens Stent. , to the ectoparasitic nematodes Trichodorus christiei and Belonolaimus longicaudatus 10 Greenhouse Experiment 3. The interrelationship of population levels of Trichodorus christiei and Belono la imus longicaudatus in the greenhouse utilizing different levels of initial inocula 12 iii

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Page Greenhouse Experiment 4. A comparison of the reproductive rate of 25 Trichodorus christie i in D-D fumigated soil with that in unfumigated soil I 2 Greenhouse Experiment 5. Investigation into the possible predaceous habit of three free-living nematodes and an oligochaete on Trichodorus christiei " Greenhouse Experiment 6. The effect of different kinds of potting containers on the reproduction of Trichodorus christiei ^ LABORATORY EXPERIMENTS 15 Laboratory Experiment 1. The life cycle of Trichodorus christiei in a constant temperature chamber set at 80 F with a 12-hour photoperiod 15 Laboratory Experiment 2. Mode of molting by Trichodorus christiei ^ Laboratory Experiment 3. Determination of the reproductive potential of Trichodorus christiei 17 Laboratory Experiment 4. Population dynamics of Trichodorus christiei as affected by original numbers. . 17 Laboratory Experiment 5. An investigation into the possibility of a stage resistant to D-D fumigation in the life cycle of Trichodorus christiei 18 Laboratory Experiment 6. Survival of Trichodorus christiei in the absence of a host at 80 F 19 Laboratory Experiment 7. Morphological studies on Trichodorus christiei. 20 RESULTS AND DISCUSSION 21 FIELD EXPERIMENTS 21 Field Experiment 1 21 Field Experiment 2 30 Field Experiment 3 36 GREENHOUSE EXPERIMENTS 36 iv

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Page Greenhouse Experiment 1 37 O Q Greenhouse Experiment 2 Greenhouse Experiment 3 ^ Greenhouse Experiment 4 ^6 Greenhouse Experiment 5 > ^6 Greenhouse Experiment 6 • ^9 LABORATORY EXPERIMENTS 51 Laboratory Experiment 1 51 Laboratory Experiment 2 55 Laboratory Experiment 3 58 Laboratory Experiment 4 58 Laboratory Experiment 5 61 Laboratory Experiment 6 66 Laboratory Experiment 7 67 THE BUILD-UP OF TRICHODORUS CHRISTIEI 69 SUMMARY 11 LITERATURE CITED 74 BIOGRAPHICAL SKETCH 77 v

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Table LIST OF TABLES Page 1. Crops Used in Comparing D-D Fumigation with No Fumigation 2. Nature and Source of the Three Soils Used in the Study of the Host Status of Pangolagrass to Trichodorus christiei and Belonolaimus loneicaudatus 3a. Numbers of Trichodorus christiei Found in D-D Fumigated Plots Compared to Those in Unfumigated Plots. (a) Fall Application 3b. Numbers of Trichodorus christiei Found in D-D Fumigated Plots Compared to Those in Unfumigated Plots. (b) Spring Application 24 3c. Numbers of Trichodorus christiei Found in D-D Fumigated Plots Compared to Those in Unfumigated Plots. (c) Summer Application 25 A. The Effect of Sorghum Root-depth on Populations of Trichodorus christiei at Harvest 27 5. Increase in Cabbage Yield and Numbers of Trichodorus chris tiei Following an Application of D-D Fumigant at 30 Gal per Acre 29 6a. A Comparison of the Effects Certain Nematicides Have On the Build-up of Trichodorus christiei on 'Funks' Hybrid Field Corn 32 6b. A Comparison of the Residual Effects of a Split Application of Certain Nematicides on the Build-up of Trichodorus christiei on Cabbage 33 6c. Effect of Nematicide Treatments on Total Number and Weight of Cabbage Heads on Three Harvest Dates 34 6d. A Comparison of the Residual Effects Certain Nematicides Have on the Build-up of Trichodorus christiei On Unfumigated 'Gold Cup' Sweet Corn 35

PAGE 7

Table Pa § e 7. A Comparison of the Survival of Trichodorus christiei in the Presence and Absence of Organic Matter (O.M. ) Under Clean Cultivation in the Field 36 8. The Effect of Temperature on the Reproductive Rate of 25 Trichodorus christiei on Sweet Corn and Tomato After 8 Weeks 37 9. Plant Parasitic Nematode Populations in 100 ml of Soil From Under Pangolagrass Followed by Sweet Corn and Vice Versa Maintained in the Greenhouse for 3 Months 39 10. Total Dry Weight (Grams) of Roots From 5 Replicates of Pangolagrass and Sweet Corn After 3 Months in Association With Trichodorus christiei and Belonolaimus longicaudatus . 40 11. The Interrelationship of Two Ectoparasitic Nematodes At Different Rates of Initial Inoculum While Feeding on Sweet Corn in 6-inch Plastic Pots -. 42 12. Effect of Trichodorus christiei and Belonolaimus longicauda tus on the Dry Weights (Grams) of the Second Crop of Sweet Corn. The Corn was Harvested 45 Days After Replanting in Pots in Which the Nematodes had Previously Been Established 43 13. The Reproductive Rate of Trichodorus christiei in D-D Fumigated Soil and Unfumigated Soil in Greenhouse Pots . . 46 14. Effects of Oligochaetes on Number of Trichodorus christiei After 8 Weeks 47 15. Effects of Three Possible Predaceous Nematodes on Populations of Trichodorus christiei 48 16. Effect of Aporcelaimellus obscurus on Established Populations of Trichodorus christiei 48 17. Numbers o f Trichodorus christiei in 100 ml of Soil From Four Different Containers Using Sweet Corn as a Host After A Period of 8 Weeks 49 18. Study of the Life Cycle of the Offspring of 10 Gravid Female Trichodorus christiei at 80°F While Feeding on 'Homestead 24' Tomato . 52 19. Reproductive Rate of Individual Specimens of Trichodorus christiei While Feeding on 'Homestead 24' Tomato Seedlings Growing at A Constant Temperature of 80°F 59 vii

PAGE 8

Table Page 20a. Reproductive Rates of Trichodorus christiei at Different Rates of Infestation Maintained at 80°F 60 20b. Average Weights of 'Homestead 24' Tomatoes Inoculated with Trichodorus christiei. 60 21a. Effects of D-D Fumigation on Trichodorus christiei , Belono laimus longicaudatus and Hoplolaimus galeatus in Soil Contained in Pint Jars ' 64 21b. Effects of D-D Fumigation on a Pure Population of Tricho dorus christiei in Soil Contained in Pint Jars 65 22. Survival of Trichodorus christiei in the Absence of a Host in a 6-inch Pot at 80°F . . 67 viii

PAGE 9

LIST OF FIGURES Figure Pa S e 1. Roots of a sweet corn plant showing stubby-root symptoms caused by Trichodorus christiei 26 2. Comparison of sweet corn seedling growth on unfumigated Leon fine sand soil with that fumigated with D-D soil fumigant applied at 30 gal per acre 28 3. The effect of Belonolaimus loneicaudatus on the growth of sweet corn 45 days after replanting in pots initially inoculated with (left to right) 250, 100, 50 and 25 specimens **** 4. Growth of sweet corn 45 days after replanting in pots initially inoculated with (left to right) 250, 100, 50 and 25 Trichodorus christiei 45 5. Root growth of sweet corn after 8 weeks in the greenhouse in a clay pot, glazed crock, and plastic pot 50 6. Stages in the life cycle of Trichodorus christiei at (10X); (a) egg, (b) first larval stage, (c) second larval stage, (d) third larval stage, (e) fourth larval stage, and (f) adult female 53 7. Variation in size between adults of Trichodorus christiei collected from two locations near Sanford, Florida .... 54 8. Break in the old cuticle prior to molting of its "hood" by Trichodorus christiei 56 9. A molting specimen of Trichodorus christiei . Note the absence of the onchiostyle in the "hood" 57 10. Characteristic position of Trichodorus christiei while feeding; i.e., the head at right angles to the cell upon which it is feeding 62 11. Trichodorus christiei feeding upon the root-cap of tomato in agar 63 12. Water mount of a live Trichodorus christiei r ecovered from soil with a moisture level of 1.7 per cent 68 ix

PAGE 10

INTRODUCTION The nematode genus Trichodorus was erected in 1913 by Cobb (11) when he described the species Trichodorus obtusus . Micoletzky (22) transferred Dorylaimus primitivus de Man, 1880 J to the genus Trichodorus and placed T^ obtusus in synonomy. The genus received little further mention in the literature until 1951 when Christie and Perry (8) reported that an undescribed species of Trichodorus was parasitic to several crop plants in Florida. Subsequently this species, described by Allen (2) as Trichodorus christiei , has been recognized as a severe pathogen in many areas of the United States. To date, most of the research on this species deals with host-parasite relationships and control, with little consideration of its biology. Extremely high population levels of 1\ christiei are frequently found following the application of soil fumigant nematicides. This unusual population build-up has led to considerable speculations as to the factors responsible for such a phenomenon. This work on the bionomics and life cycle of T\. christiei was initiated as part of a program to attempt an explanation of the population increases of this nematode resulting from soil fumigation. 1

PAGE 11

REVIEW OF LITERATURE Thorne (33) placed the genus Trichodorus into his family Diphtherophoridae, subfamily Trichodorinae of the superfamily Dorylaimoidea and named T. primitivus (de Man, 1880) Micoletzky, 1922, the type species. Allen (2) published a monograph of the genus Trichodorus and described T. christiei , the species to which Christie and Perry (8) had given the common name of stubby-root nematode. Other species have since been added until now some 32 are recognized. The feeding habits of T. christiei have been studied by a number of workers (5, 10, 27, 29 and 35). Russell and Perry (29) reported on three general types of feeding on wheat seedlings growing in agar; externally on roots, externally on root hairs, and within root caps. This latter method explains the measurable symptoms commonly found in association with low populations of J\_ christiei (1 and 29). Christie and Perry (8) proposed that the disease be termed stubby root because through the cessation of longitudinal growth, short, multibranched rootlets are produced. The effects of temperature on the population numbers of T\_ christiei have been studied by Malek et al. (20). The greatest numbers after 60 days and 90 days were found at temperatures very close to or at 25°C. The lowest rate of reproduction was at 15°C. At the upper limit, Rhode and Jenkins (26) found that no reproduction took place at 35°C. 2

PAGE 12

3 Perry (24) observed a rapid build-up of christiei following soil fumigation with methyl bromide and other nematicides. Haasis et al^ (13), studying 14 species of nematodes associated with decline of woody ornamentals, found that they could control all but christiei with 1, 2-dibromo-3-chloropro P ane. This unique behavior of T\ christiei to soil fumigation also has been observed by Martin (21) in Africa. Perry (24) suggested that this nematode has a high reproductive potential. Alhassan and Hollis (1) found that after 3 weeks the per cent increase in numbers of T± christiei was related inversely to the initial inoculum level and to plant damage. The life cycle of T^ christiei has been studied in part. Russell (28) found that eggs hatched 66 to 68 hours after oviposition in vitro, with apparently the first larval stage emerging. Rhode and Jenkins (26) reported that T. christiei completed its life cycle in 16 to 17 days at 30°C and 21 to 22 days at 22°C. Four distinct groups were recovered; in each case specimens were larger at the lower temperature. Bird (4) demonstrated that both host and geographic origin of T\_ christiei could influence population density, indicating the possibility of races. The host also had an effect on the size of the nematode; for example, those specimens obtained from lettuce were larger than those from tomato and celery. Rhode and Jenkins (27) reported jimsonweed, asparagus, poinsettia and crotalaria to be the only non-hosts of T. christiei among the 42 species of plants they tested. Rhoades (25) in Florida found the numbers of T\ christiei declined greatly during the summer months in plots planted to Crotalaria spectabilis Roth. , but persisted in those with weeds.

PAGE 13

4 Population numbers have also been reported to be affected by seasons. Perry (24) found that damage was apparently most severe in a spring crop following a fumigated fall crop. A similar situation was found by Barker and Worf (3) on azaleas in Wisconsin, where no christiei were observed during the summer. Hoff and Mai (15) recorded the numbers of T\_ christiei recovered from onion fields on peat soil at eight depths at inervals of 6 inches down to 4 feet throughout the year in the state of New York. The maximum number was recorded at a depth of 0-6 inches around June 1. T. christiei was not found below 12 inches at any time. Hoff and Mai (15) concluded that "the natural population of christiei was maintained in a specific soil and root zone . " Comparing si/It clay loam, loam and sandy loam, Thomason (31) found that after twelve weeks the greatest reproduction of T. christiei occurred in the sandy loam. After 19 weeks, however, the sandy loam population was only 1/6 that of the loam. Survival in the absence of a host by T. christiei was also reported as being substantially greater in the loam. According to Christie (7) , it seems unlikely that the distribution of T. christiei is greatly influenced by soil type. Recent agricultural research has stressed biological control. A possible explanation for the rapid return of Tj_ christiei following fumigation may be due to the reduction of predator populations. Working in vitro , Russell (28) found a number of predators which preyed upon christiei , the most aggressive of which were the nematode Aphelenchoides winchesi Christie, 1939 and the mite Protolaclaps

PAGE 14

bickleyi Bram. Thome (32) found the remains of Trichodorus primitivus , in the gut of Mononchus acutus Cobb, 1917. Chitwood and Oteifa (6) observed oligochaetes feeding upon nematode eggs, while Schaerf fenberg (30) demonstrated that enchytraeids controlled Heterodera schachtii Schmidt, 1871 in greenhouse pots. Tardigrades were reported feeding upon Trichodorus aequalis Allen, 1957 by Hutchinson and Streu (16). Esser and Sobers (12) expressed doubt that tardigrades can be utilized in biological control as they are rarely found in abundance. Little is known of the presence of nematicidal fungi in Florida soils and there have been no studies on the effects fumigation have on these fungi. A survey of the literature revealed no work on the competitive status of Trichodorus when in association with other plant parasitic nematodes. Krusberg and Sasser (18) observed that only small numbers of Meloidogyne sp. and Pratvlenchus sp. were present when Hoplolaimus coronatus Cobb, 1923 was abundant. Thome (34) noted that Xiphinema americanum Cobb, 1913 frequently dominates Pratvlenchus penetrans (Cobb, 1917) Filipjev and Stekhoven, 1914 and Pratvlenchus minyus Sher and Allen, 1953 when in association with these smaller, slower moving pratylenchs.

PAGE 15

MATERIALS AND METHODS Field Experiments Field plot experiments involving the use of nematicides were conducted at the Central Florida Experiment Station for the purpose of studying the population build-up of T^ christiei following fumigation. Using a randomized block design, plots were laid out on Leon fine sand soil which had been fallow for the past ten years. The dominant plant species prior to plowing was bermudagrass ( Cynodon dactylon (L.) Pers.). The area lacked tile which is normally used for drainage and subsurface irrigation; thus, when required, plots were irrigated by an overhead sprinkler system. Soil samples were taken with a Hoffer soil sampler; eight-in-therow samples were taken from each plot. With one exception, samples were taken to a depth of 6 inches and the soil placed in polyethylene bags. Individual samples were thoroughly mixed and a 100 ml subsample processed by the centrifugal-flotation method (17). Samples were read within 48 hours of sampling. Nematodes were counted in a known area of a Syracuse dish under a binocular microscope, and the number per 100 ml of soil was calculated. Field Experiment 1. -Comparison of the effects of 1 ,3-dichloropropene 1, 2-dichloropropane (D-D soil fumigant) fumigation with no fumigation on the population of Trichodorus christiei on fall, spring and summer crop s After bringing the area to seed bed tilth, one half of each block was treated with D-D soil fumigant at the rate of 30 gal per acre

PAGE 16

applied with a conventional shank injector at a depth of 6 inches. Two weeks later the plots were fertilized with a 5-5-8 mixture at a rate of 1000 lb per acre, disked and leveled. Two crops were planted, with 5 rows of each per replicate which were 35 ft long by 24 ft wide (Table 1). Data were taken from the 3 rows of each crop, within the border rows. Table 1. Crops Used in Comparing D-D Fumigation With No Fumigation Crop Variety Planting Date Harvesting Date Cabbage Marion market 10/14/64 2/3/65 Sorghum Beef builder Cabbage Coppenhagen 3/25/65 6/10/65 Sweet corn Gold cup Soybean Lee 7/19/65 10/20/65 Sorghum x sudangrass hybrid Grazer A Cabbage Market topper 11/12/65 2/25/66 Sorghum x sudangrass hybrid Grazer A Population counts of the nematodes present were made at the end of each month. Yields were taken on a fresh weight basis at harvest. Field Experiment 2. -A split-plot experiment designed to study the residual effect of three nematicides in the control of Trichodorus christiei A comparison was made between a single nematicide application prior to a summer crop of 'Funks' hybrid field corn and its effects on

PAGE 17

8 the following fall crop of 'Greenback' cabbage with that of an application prior to both the summer and fall crops on plots 12.5 ft by 14 ft. The experiment was terminated by observing the residual effect of the nematicides on a crop of 'Gold-Cup' sweet corn the following spring. The treatments included: (1) check, (2) 0-0 diethyl 0-2 pyrazinyl phosphorothioate (zinophos) at 3 lb per acre, (3) l,2-dibromo-3chloropropane (Nemagon) at 2 gal per acre, (4) D-D soil fumigant at 15 gal per acre, and (5) D-D soil fumigant at 25 gal per acre. Prior to fumigation 25 lb of soil containing 100 T. christiei per 100 ml was spread over each plot and disked in. The fumigants were applied by a hand applicator on a 12-inch lattice to a depth of 6 inches 2 weeks before planting. Zinophos was applied as a 10 per cent granular formulation to the soil surface just prior to planting and immediately forked into the soil to a depth of + 6 inches. The plots were then fertilized with a 5-5-8 fertilizer mixture at the rate of 1000 lb per acre, disked, leveled and planted. The plots were cultivated to reduce weed growth and a side dressing of 5-5-8 fertilizer was applied at the rate of 500 lb per acre during the third and sixth week following planting. The insecticide Sevin was applied twice a week at 2 lb per 100 gal of water to control corn earworm. Cabbage looper control was obtained using 1/2 pint of parathion plus 1 quart of toxaphene in 100 gal of water. Soil samples were taken prior to fumigation, during the growing season, and at harvest.

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9 Field Experiment 3 . — The survival of Trichodorus christiei in the field in the absence of a host . A study was made on the effect of a green manure crop incorporated into the soil on the survival of T\_ christiei . Eight field plots 6 ft by 6 ft were planted to a fall crop of sorghum x sudangrass hybrid (variety Grazer A) a known host of T\_ christiei. After 7 weeks all the plots were harvested and the root systems removed. The green matter, at a rate of 17 tons per acre, then was incorporated in one plot of each of the 4 replicates. The experimental area was kept weed-free by hand for the following 16 weeks, during which time the survival of T\_ christiei was recorded in plots with and without organic matter. , Greenhouse Experiments Greenhouse Experiment 1. — Effects of four temperatures on the reproductive rate of Trichordorus christiei . The study was made in four constant temperature tanks designed by Harrison and Stall (14) . Each of the tanks held 16 containers and the tanks were randomly alloted temperatures of 80° , 85° , 90° and 95°F + 1.5°F. Leon fine sand soil was fumigated with methyl bromide. After allowing the soil to aerate for a week a 5-5-8 fertilizer mixture was thoroughly mixed with the soil at the rate of 1000 lb per acre and 1100 ml of this soil was used to fill each of 64 plastic containers. Specimens of T_^ christiei were hand picked into distilled water; 25 females (males are very rare) were used to inoculate the soil in each container. At the time of inoculation half of the containers were planted to 'Gold Cup' sweet corn and half to 'Rutgers 'tomato. The pots were watered with distilled water as needed. At the end of

PAGE 19

10 8 weeks the experiment was terminated. The soil was thoroughly mixed and a representative sample obtained. A 100 ml sub-sample was processed using the sugar-flotation method to remove the nematodes. Greenhouse Experiment 2. -A test designed to determine the host status of pangolagrass, Digitaria decumbens Stent., to the ectonarasitic ne matodes Trichodorus christiei and Belonolaimus longi^ cauda tus. Soil from the Immokolee fine sand series was obtained from three sources (Table 2) . Table 2 Nature and Source of the Three Soils Used in the Study of the Host Status of Pangolagrass to Trichodorus christiei and Belonolaimus longicaudatus . ~ Plant parasitic nematodes in 100 ml of soil Origin pH P.M. 3 / Stubby-root Stunt Ring "Sterile ,,b _/ 5.3 4.48% "Pangola" 7.0 4.32 14 "Crop" 6.6 2.58 14 32 2 f/ Chromic acid determination of the organic matter. jV "Sterile" = steam sterilized. "Pangola" = from under a 3-year-old pangolagrass pasture. "Crop" = from under sorghum, prior to which there had been 2 crops of tomato. Twenty 6-inch clay pots were filled with each of the three soils after incorporating the equivalent of 1000 lb per acre of 5-5-8 fertilizer mixture and 500 lb per acre of lime. The pots were placed in the greenhouse with 5 replicates each of the following host -inoculation combinations in each soil:

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11 Host Nematode (1) Pangolagrass T. christiei (2) Pangolagrass B. longicaudatus (3) Gold Cup' sweet corn T. christiei (A) Gold Cup' sweet corn B. longicauda tus Individual sprigs of pangolagrass were planted in each of the designated pots. These sprigs were obtained from one-month-old stem cuttings which had been grown in sterile soil. Three seeds of sweet corn were planted in each indicated pot. The nematodes were inoculated in water suspensions around the seeds of corn and roots of pangolagrass. The respective treatments received 25 females of T. christiei and 20 females, plus 5 males, of B. longicaudatus . Both species of neamtodes were obtained from a field plot of sorghum. The pots were maintained for 3 months. At the end of each month the grass was clipped to the first node and a side-dressing of 5-5-8 fertilizer mixture was applied at the rate of 1000 lb per acre to each replicate. On termination of the experiment, fresh weights of roots and tops were recorded, the soil in each replicate was thoroughly mixed and three composite samples were obtained from each treatment. Nematode-free sprigs of pangolagrass then were planted in those replicates which had grown sweet corn, and sweet corn was planted in those replicates previously growing pangolagrass. These replants were maintained for an additional 3 months and processed in the same manner.

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12 flreenhouse Experiment 3 . The interrel ationship of population levels of Trichodorus christiei and B elonolaimus longicaudatus i n the greenhouse utilizing different level s of initial inocula. The rates of reproduction of pure populations of T^ christiei and B^ longicaudatus were compared with a third treatment having these 2 species in combination. The initial inoculum level of each nematode both in the pure and mixed treatments were 25,, 50, 100 and 250. Inoculum of Tj_ christiei was comprised of females only, while that of Ek longicaudatus was a 4:1 ratio of females to males. Plastic pots containing 1,400 ml of autoclaved Leon fine sand, previously fertilized with an equivalent of 1000 lb per acre of 5-5-8 mixture, were planted to 'Gold Cup' sweet corn (3 seeds/pot). Suspensions of the nematode inocula were then washed into the planting holes, and the pots randomized on the greenhouse bench. After 45 days the soil in each pot was thoroughly mixed and a 100 ml sample taken and processed. Both the corn tops and roots were collected and their dry weights recorded. Following the application of fertilizer the remaining soil was repotted and once again planted to 'Gold Cup' sweet corn. The experiment was terminated 45 days after replanting and similar data were obtained. Greenhouse Experiment 4. -A comparison of the reprod uctive rate of 25 Trichodorus christiei in D-D fumigated soil with that in unfumigated soil . The soil used in this experiment was a Leon fine sand which had been fumigated the previous spring with the nematicide D-D (27 gal per acre) prior to planting cantaloupe ( Cucumis melo L. var reticulatus Naud.). A late summer cover crop of Crotalaria spectabilis was

PAGE 22

13 planted following the cantaloupe. After the C. spectabilis had been harvested soil was removed from the field, sifted and placed in six 3-gal porcelain crocks. Three of these crocks were fumigated at a depth of 8 inches with 1.21 ml D-D (=30 gal per acre) by means of a pipette fitted with a propippette. The soil temperature at the time of fumigation was 66°F. After waiting 2 weeks, six 6-inch clay pots were filled with fumigated soil and six with the unfumigated soil. At the time of potting a 100 ml sample of soil was processed and the number of nematodes recorded. The pots were seeded with oats ( Avena sativa L.), and 25 specimens of T^ christiei added to each. At the end of 2 months, the pots were removed and the numbers of T^ christiei per 100 ml of soil recorded. The pots were then replanted to 'Gold Cup' sweet corn and allowed to grow for a further 2 months, before once again recording the numbers of nematodes present. Greenhouse Experiment 5. -Investigation into the possible predaceou s habit of three free-living nematodes and an oligochaete on Trichodorus christiei . The nematode inocula were obtained from the test conducted on the host status of pangolagrass to T_j_ christiei . The specimens were hand picked into distilled water and then poured into 6-inch clay pots containing autoclaved Leon fine sand soil which was then seeded with 'Gold Cup' sweet corn. The design of the treatments applied to each of the three possible predaceous nematodes being tested was as follows with 5 replicates of each: (1) 100 T^ christiei (2) 100 T. christiei + 25 freeliving specimens

PAGE 23

14 (3) 100 Tj_ christiei + 100 freeliving specimens (4) 25 freeliving specimens The freeliving nematodes included Aporcelaimellus obscurus (Thome and Swanger, 1936) Heyns, 1965; Eudorylaimus simplex (Thorne and Swanger, 1936) Andrassy, 1959 and Mylonchulus parabrachyurus (Thorne, 1924) Andrassy, 1958. Since oligochaetes are commonly found in unfumigated soil, a test was designed to investigate the effect they might have on the reproduction of T\ christiei . The treatments consisted of the following three rates of hand picked inocula replicated five times: (1) 100 T;_ christiei (2) 100 Tj_ christiei + 100 oligochaetes (3) 100 T_;_ christiei + 500 oligochaetes The inocula was poured into 6-inch plastic pots containing autoclaved Leon fine sand soil which had been fertilized at the equivalent rate of 1000 lb per acre of 5-5-8 mixture. The host plant used in this experiment was 'Homestead 24' tomato. After planting, the treatments were randomly placed in a growth chamber set at 80°F with a 12hour photoperiod. Top and root weights were recorded after 8 weeks, and the soil in each pot was thoroughly mixed and a 100 ml sample processed from each. Greenhouse Experiment 6. -The effect of different kinds of potting containers on the reproduction of Trichodorus christiei . Clay pots have conventionally been used for determining nematode pathogenicity and reproduction on a host. Frequently seedling roots

PAGE 24

15 which escape nematode damage grow out to the walls of the pot, where they produce a mat. Owing to the porous nature of clay, there are greater moisture fluctuations along the pot walls, producing an environment which is less favorable to nematodes than the center of the pot. Previous indications were that roots did not mat along the walls of glazed crocks and plastic pots. Experiments were conducted in a greenhouse and growth chamber set at 80°F to determine the environmental effects in four different 6-inch containers on the reproduction of T;_ christiei . The containers used were as follows: (1) Red clay pot; surface irrigated. (2) Red clay pot; sub-surface irrigated. (3) Glazed crock. (4) Plastic pot. Five containers of each type were filled with 1,350 ml of autoclaved Leon fine sand. Each of the pots was seeded with 3 'Gold Cup 1 sweet corn seeds, and an inoculum suspension of 25 T. christiei poured over the seeds. In addition to the inoculated pots, 5 replicates of uninoculated pots were set up in the growth chamber as checks. After 8 weeks the pots were removed and top and root weights recorded. The soil in each pot was thoroughly mixed and a 100 ml sample processed to determine the nematode numbers. Laboratory Experiments Laboratory Experiment 1. -The life cycle of Trichodorus christiei in a constant temperature chamber set at 80°F with a 12-hour photoperiod.

PAGE 25

16 Styrofoam cups were cut in half and 50 ml of autoc laved Leon fine sand placed in each. 'Homestead 24' variety of tomato was then sown in the cups and after germination thinned to 4 plants per cup. Two weeks after planting 10 gravid females of T_j_ christiei obtained from a tomato host were poured into a hole adjacent to one of the tomato seedlings in each of 48 cups. From the third to the eighteenth day, 4 cups were removed per day. The soil was processed using an unpublished technique devised by the late M. B. Linford of the University of Illinois. The soil and nematodes collected from the washings on the 325-mesh screen were poured onto a folded Kimwipe tissue held between two 2-ounce plastic funnels. The funnels were placed on the walls of a Syracuse dish in such a manner as to suspend the tissue above the bottom of the dish. After allowing the washings to settle on the tissue (1-2 minutes) the funnels were transferred to a second Syracuse dish. The water level in this dish was raised to a. level just above that of the tissue. After 6 hours the funnels were transferred to a third Syracuse dish and the water level was adjusted as described above. The funnels were removed from this third dish after 2 hours, and the numbers and stages of develop ment of the nematode specimens were recorded. Specimens of the oldest stage collected each day were placed in a specimen vial filled 1/4 full of distilled water, and the remainder of the offspring were placed in another similar vial. The vials were then transferred to a hot water bath at 50°C for 15 minutes, at which time the level in the vials was brought to 1/2, using 2 per cent formalin solution for fixation and preservation.

PAGE 26

17 Laboratory Experiment 2. -Mode of molting by Trichodorus christiei. Specimens of christiei were studied while molting in water and in 3/4 per cent water agar contained in the top half of 60 x 15 mm plastic culture dishes. After picking the nemas into the agar, the surface of the agar was covered with a disk of plastic sheet to prevent drying and facilitate studies with the aid of a compound microscope. Other specimens were fixed at various stages throughout molting by placing them in 1 per cent formalin at 5°C. These were then mounted on Cobb aluminum slide holders to facilitate study from both sides with the aid of oil immersion objectives. Laboratory Experiment 3. -Determination of the reprodu ctive potential of Trichodorus christiei . Styrofoam cups were prepared as above, but in this experiment 48 cups were inoculated with individual fourth stage larvae and placed in a growth chamber set at 80°F with a 12-hour photoperiod. At the end of 16, 17, 18, 19 and 38 days, 8 cups were processed using Linford's funnels. Counts were made of the numbers of each life cycle stage of T\ christiei present in each cup. This experiment was duplicated. Laboratory Experiment 4. — Population dynamics of Trichodorus christiei as affected by original numbers. The effects of initial inocula numbers on the reproductive rate of T. christiei were studied in a constant temperature chamber set at 80°F with a photoperiod of 12 hours. The treatments included hand picked inocula of 25, 100 and 400 females of Tj_ christiei inoculated into 4-inch plastic pots. Each pot contained 375 ml of autoc laved Leon

PAGE 27

18 fine sand soil, and were planted to 'Homestead 24' variety of tomato. Following germination the tomato seedlings were thinned to 6 plants per pot. At the end of 21 days and 42 days, respectively, 6 replicates of each treatment were removed. Top and root weights were recorded and all the soil in each pot was processed using Christie and Perry's modified Baermann funnel technique (9). Counts were recorded after 12 hours. Laboratory Experiment 5 . -An investigation into the possibility of a stage resistant to D-D fumigation in the life cycle of Trichodorus christiei . One pint wide-mouth fruit jars were filled with field soil and each of the following 10 treatments was replicated 5 times: Rate per acre Rate per jar (1) Check (2) 690 gal of mineral spirits 1.0 (3) 10 gal of D-D per acre 0.2 (4) 15 " ii ii ii 0.3 (5) 30 " ii ii ii 0.6 (6) 50 " ii ii ii 1.0 (7) 10 gal of water soluble D-D per acre (8) 15 " it it ii ii ii (9) 30 " it ii ii ii ii (10) 50 '»' it ii ii ii ii The basic solution in treatments 3 through II II II M spirits + 7.2 ml D-D. The basic solution of the water soluble D-D contained 91.8 ml water + 7.2 ml D-D + 1.0 ml Triton X 100 surfactant. This latter solution was mixed by adding the surfactant to the D-D and then adding water.

PAGE 28

19 The D-D was applied, using a 1 ml pipette filled with a propipette, to a depth of 4 inches within each jar. After the pipette was removed, each injector hole was carefully sealed. The jars were placed in a laboratory cabinet (+ 72°F) for 2 weeks. Water was added as needed to prevent drying out of the soil. A 100 ml sample of soil was then obtained from each jar and processed using the centrifugal-flotation method. Counts were made of living nematodes; i.e., those which were moving. The remainder of the soil from each jar was placed in a 4-inch clay pot, seeded with 'Homestead 24' variety of tomato and placed in the greenhouse. Three weeks later another 100 ml soil sample was processed as before. This experiment was repeated using a wider range of water soluble D-D rates and a soil containing a pure population of T;_ christiei . The treatments included: Rate per acre Rate per jar (1) Check (2) 25.0 gals of water-soluble D-D per acre 0.33 ml of basic solution (3) 45.0 " " " " " " 0.60 " " " (4) 56.25 " " " " " " 0.75 " " " (5) 75.0 " " " " " " 1.00 " " " The basic solution was made up of 88.2 ml water + 10.8 ml D-D + 1.0 ml Triton X 100 surfactant. Laboratory Experiment 6. -Survival of Trichodorus christiei in the absence of a host at 80°F.

PAGE 29

20 Six clay pots were filled with Leon fine sand in which a pure population of christiei had been established, and 4 clay pots with the same type of field soil containing a mixed population of nematodes. The tops of the pots were covered with polyethylene secured by elastic bands, in order to limit the loss of moisture from the soil. The pots were then placed in a constant temperature chamber and watered as needed. Individual pots of each soil type were processed after 0, 2, 4, and 8 weeks. The remaining 2 pots containing the pure populations were taken down after 12 and 16 weeks. Two samples of a 100 ml size were processed from each pot, using the centrifugal-flotation method, and the number of survivin g T. christiei were recorded. Laboratory Experiment 7. — Morpholog ical studies on Trichodorus christiei. Morphological studies were made on specimens of T^ christiei which had been killed and fixed as in Laboratory Experiment 1. However, better results were obtained by staining. The stain was prepared by adding 0.05 ml of green food color (water and propylene glycol) to 2.00 ml of water. Living specimens of T. christiei were placed in this stain which killed them within 24 hours to 48 hours. The killed specimens were then fixed in 1 per cent formalin on Cobb aluminum slides.

PAGE 30

RESULTS AND DISCUSSION Field Experiments Field Experiment 1. Tables 3 a, b and c show that the build-up of T^ christiei following soil fumigation described by Christie and Perry (8) may take place during any season of the year, provided a host is present. However, the more rapid increase obtained on the summer crop of sorghum indicates that the rates of build-up vary with differing levels of temperature. The depth at which the young feeder roots of the host crop are located is of importance when studying the population of T^ christiei (Fig. 1). Tables 3 a, b and c show that the major build-up in the 6 weeks following planting is in the top 6 inches of soil. Thereafter, until harvesting, the population of T. christiei increases very rapidly below the 6-inch depth. The extent to which this deeper population develops is not only dependent upon the nature of the host root system, but also on whether or not the soil has been fumigated (Table 4). The reason for this is apparently due to the fact that soil fumigation permits the young seedlings to develop a root-system capable of outgrowing the ensuing nematode infestation (Fig. 2). Thus, it was possible for the fumigated plots to out-yield those not fumigated (Table 5) , while supporting a larger population of T\ christiei from about the seventh week until harvesting. However, the control of Belonolaimus longicaudatus by fumigation should not be overlooked since this nema21

PAGE 31

22 tode is far more pathogenic and destructive to vegetable crops in this area than T_j_ christiei is. When comparing the yield of cabbage from fumigated plots with that from unfumigated plots, it should be noted that there is a signif cant different in yield at the 120-day harvest, whereas the yields obtained from the second harvest at 140 days were comparable (Table 5) The crops on fumigated plots of this experiment reached maturity as much as 2 weeks before those on unfumigated plots.

PAGE 32

23 w 5 -i-i o O r-l c Fx CM o •rl •H •a 4J 0) 0) 03 •H 4-> O 4-1 •H n bO r-l «H •H & Sj •C m -i M B oco cu CO T3 a s • H -H vO o u CO to Cu CU O M 00 C •H OJ r-l 4J Cu CO e q CO CO cu i r-l i l t O vO O * in o> cn oo 00 r-l CM o 00 r-co -3 vO CO O o a) X) OJ 4J CO o m oo co ^ TO X) CN oCN •* o A o CJ •U 03 00 •r-< i-l CO v£) O cti oo m h vO 00 CN > 03 T3 vO CM CN in co r-» oo cm o co CM CM CO CM •rl 1-1 1-1 -rl CM vO r-l vO I I I I QiOO* o CO vO CO vD o m co CM •o T3 CU co <

PAGE 33

24 c o •H 4-1 co O •H I— ( a a c H M a rH •H O Cfl MH Q o (J , — 1 4_) £ a; a) o CO rH •H O 1 •H 4J en H M rC a a) .i 60 ~ 1 CO n i4-< o CO O O 2; QJ rl-< rH 4J cx g o> TO X) B 3 d • H rH H o 4-1 CO ao> o 60 M << U DO •rH a) rH 4-1 a CO E Q a C/j 4-1 C a> B 4-1 CO 1) u H O r-. cm O 00 CO N C C! C C •H -H -H *H CM CM vD i— I vO i—i • I I I o * rH o o m 00 in CO m co u CD -C u X) 0) 4J (0 t>0 •H m x) 01 41 cn 0) 5l CO X CO x) rG 4-1 O 4-1 u o H u a. CO CO XI oo XI 0) CO 60 CO I

PAGE 34

25 43 CO H o H 4-1 CD u a a 4-l rO o >v o o 01 rC I— 1 a a § •H o 4J CO TO a o 0) 60 o <; 60 c •H CJ r-l 4J a co e q CO CO c a) 0 4J CO CJ r-l H OlHQO oo co o o CN CN c c c c •H -H -i-l *H CN CN VD H ^ r4 till fa o 00 oj in m in CN c d C c CN CN vO t-l v£> H I I I I OvOO* .a I in O tn O CJ -G o -a cj 4J CO 60 •H fa o 00 CO TO m CN vO CN 1 — 00 CJ 0) u CJ 4J TO 60 •r-l fa I — . vD CN CJ> v TO XI O m vo CN 00 r-l r-l 60 m vO 00 CN in c vD |— ( f— 4 CN in •H \ 4J o (-J CN CO r-l o a i-H <— i T3 CO CJ •H 4J 4-1 CO •rl CU > •rl OJ ,c On lO CN 4J CO CO t — 1 cn r-l r-l |_l OJ 4J CM CO CO c c (J CO •H -r-l •r-l •H •a CN CN o vO r— 1 vD r-l r-^ O vO | | VO T3 CN 0) u o\ XI CO >> X) o co CO •H CJ .c rC 4-1 4J in vO oo o X) X. OJ CJ 4J 4-1 CO CO 60 60 o •H CJ •i-l CJ cu rC rC CJ fa u fa

PAGE 35

Roots of a sweet corn plant showing stubby -root symptoms caused by Trichodorus christiei .

PAGE 36

Table 4. The Effect of Sorghum Root-depth on Populations of Trichodorus christiei at Harvest. Depth Average No. of Dry-weight of No. of T. christiei/ 100 ml Root-tips roots (grams,) of soiT~ VjilcL. in. Check Fuxniga ted Check Fumigated 0-2 in 2&/ 3b/ 10.1 16.7 182 315 2-4 in 4 6 6.2 6.9 287 350 4-6 in 8 7 2.7 3.8 385 235 6-8 in 4 10 0.9 2.1 228 676 8-10 in 1 4 0.2 0.8 161 392 10-12 in 2 0.05 0.4 53 239 12 in 1 0.0 0.25 32 189 a/ Average of 6 samples, b/ Average of 12 samples. Below a depth of 6 inches, the plant parasitic nematode population became increasingly dominated by T. christiei . In this respect very few B. longicaudatus w ere found below the 8-inch depth.

PAGE 37

28 2. Comparison of sweet corn seedling growth on unfumigat Leon fine sand soil with that fumigated with D-D soil fumigant applied at 30 gal per acre.

PAGE 38

29 d •H O O I •H 03 Q) >> •H CO 4J T3 CO •H O >-i co o H . Q) H -U IH IW CD o r-l . -i-l O O S3 W H I o CO X) CD M eg .O -Q CO U o •H CD O P. 01 05 CO n Q O M 01 4J 14-1 < CO >s n P o CN cu o bt H 0) o H o 2 UO CN CN CN m oo CN m rH rH in CN 00 vO m o rH co CN CO o CO o 00 oo in CO in o I CN Q i O CO m CM co CO o oCO CO u XI 00| •H vO m CN m rH CN vO oo in rH o vO o CN CD in o> rC u co a CO o jC 14-4 CJ in T3 CN CO ' co C 4J CO O 0) M| CO 0 > O oo tn oo" "O in CO V cj > C oo o) oo u o 0) • IH rH 14-1 rH •H l|-( P _ c/i 4-1 c CO o •H >4-l •H d M II CO Q) dJ rHJH 4J •rl^h O >4 h 0 a

PAGE 39

30 Field Experiment 2. When studying the data on the split-plots experiment in Tables 6a, b, c and d, designed to study the residual action of certain nematicides on the build-up of T_j_ christiei , consideration should be given to the following: a) The plots treated with the organo-phosphate nematicide, zinophos, restricted the build-up of T. christiei . The residual action of this material also can be seen in the limited numbers of T. christiei which were found on the second crop following a single application (Table 6b). This residual action was not seen in the final crop of sweet corn, though the yields on the plots receiving 2 applications of zinophos proved statistically greater than all other treatments. b) The build-up of T_j_ christiei was substantially greater following an application of D-D soil fumigant at 25 gal per acre than at 15 gal per acre. In addition, the numbers of T\_ christiei present at harvest were greater with a double application of D-D at both the 25 and 15' gal rates. c) There were no significant differences in control of T^ christiei between single and double applications of Nemagon at 2 gal per acre, and in both cases the build-up surpassed that of the control. d) In Table 6a it can be seen that although the plots that received 25 gal of D-D per acre had the highest numbers of J_j_ christiei at harvest, they produced significantly greater yields of corn than did all other treatments. The same was found to be true on cabbages (Tables 6b and c) with the exception that the yield was not statistically different from those plots receiving 2 applications of zinophos at 3 lb per acre.

PAGE 40

31 e) Table 6a shows that the application of each nematicide resulted in significant yield increases of field corn. These yield increases were partly due to early control of T^ christiei and partly due to control of other parasitic nematodes. In addition, the number of T^ christiei after 58 days was greater than that at 72 days. The reason for this is apparently due to the fact that at harvest the samples were taken to a 6-inch depth. As shown in the previous experiment, by crop maturity the majority of T\ christiei are below this depth. When cabbage was planted on the same plots following the corn and half the original number of plots retreated, none produced significant yield increases except those retreated with zinophos at 3 lb per acre and D-D at 25 gal per acre (Table 6c). The fact that the check plots yielded more than some of the treated plots suggests that cabbage, by virtue of their numerous rootlets, can withstand nematode damage better than corn. In addition, the larger initial populations of T. christiei may have influenced yields. f) The counts of 1\_ christiei on the final untreated crop of sweet corn are given in Table 6d. With the exception of Nemagon, the counts of all treatments are very similar at harvest, although the numbers 3 weeks after planting show considerable variation. The only statistically outstanding yield was obtained from the plots which had received 2 applications of zinophos at 3 lb per acre.

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PAGE 43

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36 Field Experiment 3. Table 7 shows that in the absence of a host there was a gradual mortality of christiei during the first 8 weeks. Between the eighth and twelfth weeks the mortality doubled that of the first 8 weeks. After 16 weeks, when the experiment was terminated, the T. christiei population had been reduced to 1/5 of the original number. A comparison of the survival of T\_ christiei under bare plots with bare plots in which organic matter had been incorporated (at the rate of 17 tons per acre) revealed no differences (Table 7). By contrast, Patrick ej: al. (23) obtained nematicidal substances from decomposing rye. It is possible that under summer conditions a more rapid decomposition of the organic matter might prove detrimental to T_;_ christiei . Table 7. A Comparison of the Survival of Trichodorus christiei in the Presence and Absence of Organic Matter (O.M. ) Under Clean Cultivation in the Field. Time in Weeks Treatment 0 3 1/2 8 12 16 Check O.M. Check O.M. Check O.M. Check O.M. Check O.M. No. of t 414f/ 390 340 350 296 288 120 133 94 74 christiei / 100 ml of soil. % reduction 0 0 18 10 29 26 71 71 77 81 from original population. Soil Temp. (°F) 74 61 f/ Data from 4 replicates.

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37 Greenhouse Experiments Greenhouse Experiment 1. Of the four temperatures investigated, 80°F proved most favorable for reproduction by T. christiei (Table 8). There was, however, no statistical significance between the reproductive rates at 80° and 85°F. Malek et al. (20) in New Jersey found 77°F to be the optimum for T\_ christiei reproduction. Thus, the New Jersey and Florida populations of T^ christiei appear to have similar optimum temperature requirements for reproduction. Table 8. The Effect of Temperature on the Reproductive Rate of 25 Trichodorus christiei on Sweet Corn and Tomato After 8 Weeks. Temperature Rep. Rep. 2 Avera Corn Toma to Corn Tomato Corn Toma to 95°F 53W 583 2,844 294 1,449 90°F 2,253 10,500 5,103 7,489 3,678 8,995 85°F 8,394 13,828 6,899 11,567 7,647 12,698 80°F 9,891 15,402 10,626 12,788 10,259 14,095 fy Replicate 1 ran from 8/1/65 to 9/25/65; and Replicate 2, from 11/5/65 to 1/2/66. b/ Average count of 8 pots. Com F^ = 24.40* Tomato F^ = 63.2** LSD LSD 05 3,991.34 05 2,591.45 The populations of T_j_ christiei produced on 'Rutgers' tomato were greater in number than those produced on 'Gold Cup' sweet corn. As noted by Bird (4), the morphology of T. christiei was affected by the host upon which the specimens had been feeding. Of interest was the

PAGE 47

38 presence of several males at 95°F. Under field conditions males of this nematode are extremely rare. Greenhouse Experiment 2. The data presented in Table 9 show that T. christiei and longicaudatus each parasitize and reproduce on pangolagrass, giving rise to as much as an eightfold increase in their numbers over a three-month period in sterile soil. In unsterilized soil taken from two different field locations, the populations did little more than maintain their numbers, which suggested the presence of a biological interaction. Consideration should be given to two factors in unsterilized soil, viz. , (1) the presence of other plant parasitic nematodes and (2) the presence of certain nematode predators. T. christiei proved more pathogenic to the pangolagrass root system than did longicaudatus (Table 10), but the latter was more pathogenic to sweet corn. The mixed population of plant parasitic nematodes present in "Pangola" and "Crop" soils undoubtedly contributed to the smaller root systems in these soils when compared to those in "Sterile" soil. Albeit, the terminal numbers of T. christiei and B. longicaudatus were larger in the "Sterile" soil.

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39 Table 9. Plant Parasitic Nematode Populations in 100 ml of Soil From Under Pangolagrass Followed by Sweet Corn and Vice Versa Maintained in the Greenhouse for 3 Months. Soil Host Initially inoculated with 25 Nematode T. christiei Stubby-root Lesion Stunt St ins Ring "Sterile" Pangolagrass then Sweet Corn 14 a / 907 "Pangola" Pangolagrass then Sweet Corn 11 151 39 5 4 _ _ "rrnn" Pangolagrass then Sweet Corn 14 109 42 1 2 • 1 83 "Sterile" Sweet Corn then Pangolagrass 54 459 "Pangola" Sweet Corn then Pangolagrass 94 119 188 32 "Crop" Sweet Corn then Pangolagrass 88 105 130 25 219 70 5 Soil Host Initially inoculated with 25 longicaudatus Nematode Belono laimus Stubby-root Lesion Stunt Sting Ring "Sterile" Pangolagrass then Sweet Corn 53 315 "Pangola" Pangolagrass then Sweet Corn 21 46 25 7 11 4 "Crop" Pangolagrass then Sweet Corn 27 4 1 7 130 67 "Sterile" Sweet Corn then Pangolagrass 112 872 "Pangola" Sweet Corn then Pangolagrass 28 60 147 49 32 42 "Crop" Sweet Corn then Pangolagrass 16 112 210 25 172 144 18 6 58 14 fj Average of three samples. Samples collected by taking one probe from each of the five 6-inch pot replicates. A 100 ml sub-sample was processed from each sample.

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40 Stubby-root Lesion Stunt Sting Ring Key to Table 9: = Trichodorus christiei Pratylenchus brachyurus and P. zeae Tylenchorhynchus spp. Belonolaimus longicaudatus Criconemoides spp. Table 10. Total Dry Weight (Grams) of Roots From 5 Replicates of Pangolagrass and Sweet Corn After 3 Months in Association with Trichodorus christiei and Belono laimus longicaudatus . Soil Crop Nematode Stubby-root Stubby-root Sting Sting on 11/29/65 on 3/7/66 on 11/29/65 on 3/7/6 "Sterile" Sweet Corn 32. 2 a / 21. 4 b / 17. 5 a / 18.7b/ Pangolagrass 14.8b/ 14.9a/ 25 . 8b/ 18.2a/ "Pangola" Sweet Corn 16.6 18.9 9.5 17.9 Pangolagrass 6.6 10.5 9.9 12.3 "Crop" Sweet Corn 17.1 16.2 12.2 15.1 Pangolagrass 12.6 9.6 10.7 18.1 £/ Same pots; planted first to sweet corn, then pangolagrass. b/ Same pots; planted first to pangolagrass, then sweet corn.

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41 Greenhouse Experiment 3. The results of the experiment conducted to determine the effect of the competition by longicaudatus on the reproduction of T. christiei are presented in Table 11. After 45 days virtually no discernible effects of the competition were detected. In all probability, this was due to the low levels of initial inoculum. The data recorded 45 days after replanting these same pots suggest that T. christiei in the control pots had attained maximum levels of of infestation; i.e., + 1,000 individuals per 100 ml of soil, irrespective of the initial level of inoculum. Nevertheless, the presence of B. longicaudatus resulted in a marked reduction of the numbers of T. christiei . This reduction was more marked the greater the initial inoculum numbers, averaging up to 50 per cent. These differences were not statistically, significant, owing to wide variation in numbers and too few replicates. By contrast, the reproduction of B. longicaudatus was less effected the larger the initial numbers of T_;_ christiei . The average dry weights of the second sweet corn planting; i.e., from the 45th day to the 90th day, are given in Table 12. These data indicate that B^ longicaudatus is much more pathogenic to sweet corn than is T. christiei (Fig. 3 and 4). In general the corn was damaged more by the combination of the species than by B^ longicaudatus alone, though there were fewer numbers of Bj_ longicaudatus in the combination.

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A3 Table 12. Effect of Trichodorus christiei and Belonolaimus longicaudatus on the Dry Weights (Grams) of the Second Crop of Sweet Corn. The Corn Was Harvested 45 Days after Replanting in Pots in Which the Nematodes had Previously Been Established. T. christiei Initial T^ christiei B. longicauda tus + Inoculum B^ longicauda tus 25 Tops 4.0 U 3.6 3.4 Roots 2.9 2.3 2.0 50 Tops 3.9 3.5 3.7 Roots 3.2 2.7 2.3 100 Tops 3.4 2.8 3.4 Roots 2.7 2.2 1.9 250 Tops 4.0 1.7 2.5 Roots 2.3 1.2 1.6 Check Tops Roots 4.1 3.9 a/ Average of 4 replicates with three plants per replicate.

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44 Fig. 3 The effect of Belonolaimus longicaudatus on the growth of sweet corn 45 days after replanting in pots initially inoculated with (left to right) 250, 100, 50 and 25 specimens.

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45 ig. 4 Growth of sweet corn 45 days after replanting in pots initially inoculated with (left to right) 250, 100, 50 and 25 Trichodoru s christiei.

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46 Greenhouse Experiment 4 . Results of the comparison of the reproductive rates of T\_ christiei in D-D fumigated soil with unfumigated soil are given in Table 13. After 8 weeks the number in the unfumigated soil was 1.7 times that in fumigated soil, due to higher initial numbers of T\_ christiei in the unfumigated soil. However, during the following 8 weeks the situation was reversed with D-D fumigated soil having 1.6 times more T. christiei . This demonstrated that a build-up of T. christiei. similar to that seen in the field, can take place when this nematode is inoculated into D-D fumigated soil in greenhouse pots. Table 13. The Reproductive Rate of Trichodorus christiei in D-D Fumigated Soil and Unfumigated Soil in Greenhouse Pots. Numbers of T. christiei in 100 ml of soil Treatment Initially After 8 weeks After 16 weeks Host: Oats Host: Sweet Corn Unfumigated 1Q 72£/ 560 D-D at 30 gal per acre 2 42 910 tl Average of 2 samples which consisted of a probe taken from each of the 6 replicates. Greenhous e Experiment 5 . The data obtained in the experiment conducted to determine whether oligochaetes have an effect on the reproduction of T. christiei are presented in Table 14. It should be noted that, although initially inoculated into steam-sterilized soil, the oligochaet< were able to feed and reproduce. However, no statistical difference

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47 could be detected on the reproduction of T_j_ christiei, under the conditions of the experiment, as a result of the increase in oligochaetes. Table 14. Effects of Oligochaetes on Number Of Trichodorus christiei After 8 Weeks. No. T. christiei No. Oligochaetes Inoculum per 100 ml of soil per 100 ml of soil 100 T. christiei alone 1,4191/ 100 T. christiei chaetes + 100 oligo1,326 806 100 T. christiei chaetes + 500 oligo1,285 1,238 / Average of 5 replicates. The results of the study conducted on the possible predaceous habits of Eudorylaimus simplex , Aporcelaimellus obscurus , and Mylonchulus parabrachyurus on T^ christiei are shown in Table 15. Of these three free-living nematodes, only the presence of A. obscurus resulted in reduced numbers of T^ christiei . However, in a second test when specimens o f A . obscurus were inoculated into a previously established population of T. christiei , no such differences were observed (Table 16). From these tests it was concluded that none of these nematodes affect population levels of T. christiei.

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48 Table 15. Effects of Three Possible Predaceous Nematodes on Populations of Trichodorus christiei . Numbers per 100 ml of soil after: 8 Weeks 25 Weeks 100 T. christiei alone 270 147 100 T. christiei + 25 Eudorylaimus simplex 266 0 228 84 100 T. christiei + 100 Eudorylaimus simplex 238 4 144 133 25 Eudorylaimus simplex alone 4 109 Numbers per 100 8 Weeks ml of soil after: 16 Weeks 100 T. christiei alone 203 591 100 T. christiei + 25 Aporcelaimellus obscurus 217 0 353 66 100 T. christiei + 100 Aporcelaimellus obscurus 154 4 273 96 Numbers per 100 8 Weeks ml of soil after: 25 Weeks 100 T. christiei alone 270 147 100 T. christiei + 25 Mylonchulus parabrachyurus 207 0 196 91 100 T. christiei + 100 Mylonchulus parabrachyurus 221 4 210 203 25 Mylonchulus parabrachyurus a lone 0 420 Table 16. Effect of Aporcelaimellus obscurus on Established Populations of Trichodorus christiei . Numbers per 100 ml of soil after 12 weeks: Inoculum 1 2 3 Average T. christiei alone 805 630 735 723 T. christiei + 833 578 704 705 100 A. obscurus 315 305 291 304

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49 Greenhouse Experiment 6. Results of the experiment conducted to determine which of four potting containers provided the most ideal environment for the reproduction of T_j_ christiei are given in Table 17. In the greenhouse it was found that the glazed crock was statistically superior to plastic, to sub-surface irrigated clay, and to surface irrigated clay pots; there being no statistically significant difference among these latter three. In the constant temperature chamber, however, the plastic container was superior. In this case there was no statistically significant difference between the glazed crock and two types of clay pots. Similar data were obtained in a repetition of the constant temperature experiment in a different kind of chamber. Figure 5 shows the root system of sweet corn in three of the containers. Note the preponderance of roots lining the walls of the clay pot. These data suggest that, in conditions where the greatest variable is temperature, glazed crocks are desirable, while in those conditions in which moisture is the most important variable, the plastic pots are best suited for rearing Tj_ christiei . Table 17. Numbers of T. christiei in 100 ml of Soil from Four Different Containers Using Sweet Corn as a Host After a Period of 8 Weeks. Numbers of T^ christiei per 100 ml Average dry weight Container of soil of roots per pot Greenhouse Chamber Greenhouse Chamber Plastic Pot 939£/ 1,209^/ 1.7 2.6 Glazed Crock 1,303 751 2.0 2. A Clay Pot (surface irrigated) 608 783 3.5 1.9 Clay Pot (sub-surface irrigated) 885 796 1.7 3.5 f/ Two plants per pot. W Four plants per pot.

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50 WW Fig. 5. Root growth of sweet corn after 8 weeks in the greenhouse in a clay pot, glazed crock, and plastic pot.

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51 Duncan's Multiple Range Test: Surface Sub-Surface Greenhouse = Irrigated Irrigated Clay Clay Plastic Glazed 608 885 939 1.303 Glazed Surface .Sub-Surface Plastic Growth chamber = Irrigated Irrigated Clay Clay 751 786 796 1,209 Laboratory Experiments Laboratory Experiment 1. The results of a study on the life cycle of T. christiei following inoculation of 10 gravid females onto tomato seedlings growing at 80°F are given in Table 18. The different stages in the life cycle (Fig. 6) were first recorded on the following days after inoculation: 1st Larval stage 4 days 2nd Larval stage 4-5 days 3rd Larval stage 7 days 4th Larval stage 10 days Adult 14 days Gravid female 17-18 days The criterion used to determine the stage of development was the size of the gonadal primordium. The reason for this can be seen in Fig. 7, which shows the size variation in adult T. christiei from two different habitats. Measurements of 10 specimens were made at molting in 1 per cent formalin. The average sizes were:

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52 Table 18. Study of the Life Cycle of the Offspring of Trichodorus christiei at 80°F While Feeding Tomato. 10 Gravid Female on 'Homestead 24' No. Days Original After Adults Inoculation Test Surviving Life Cycle Stage Adult 4th Av. No. of 3rd 2nd 1st Offspring 3 1 3.0 0 2 3.0 4 1 7.3 • • 1.5 1.5 2 7.0 x . \j i n 5 1 3.8 3.8 3.3 7.1 2 7.5 J . yJ J • u 6 1 5.0 9.5 1.8 11.3 2 4.5 9 5 t. . j a n H . u 7 1 6.8 1.5 12.3 2.3 16.1 2 7.5 1 0 j • j 0. J 8 1 5.5 7.8 16.5 4.3 28.6 2 4.5 7 O ^ X. J 98 CO . J 9 1 7.5 8.8 13.8 1.8 24.4 2 2.0 5 j • j 7 0 in ^ xU . j 10 1 3.3 3.0 15.0 22.0 3.0 43.0 2 3 5 99 " • J ij . U 35.5 11 1 8.0 7.3 17.5 18.5 4.8 48.1 2 j • j in 1U . j o . i> 1 c c 15 . 5 0.5 35.0 12 1 4.0 17.3 26.8 14.5 2.8 61.4 2 3 5 91 n in n iu. U 1 c c lb. 5 2.0 49.5 13 1 4.5 14.8 21.8 14.5 3.3 54.4 2 H.J in c 19.5 5.5 8.0 0.5 33.5 14 1 5.5 16.0 15.8 7.5 1.0 40.3 2 5.5 44.0 25.5 6.0 0.5 76.0 15 1 3.0 2.3 13.5 9.5 10.0 2.3 37.6 2 7.0 7.5 50.0 22.0 9.0 0 88.5 16 1 3.5 10.3 15.8 17.3 7.5 1.0 51.9 2 3.0 11.0 12.5 6.5 2.0 0 32.0 17 1 3.8 16.5 27.3 22.0 13.0 2.5 81.3 2 0.5 5.5:7 15.5 11.5 5.0 1.5 39.0 18 1 3.0 47. 5 a / 28.3 27.5 18.8 2.8 125.0 Gravid females recovered.

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53 (b) (c) (d) 0.7 mm (f) 6 Stages in the life cycle of Trichodorus christiei at (10X); ( a ) egg, (b) first larval stage, (c) second larval stage; (d) third larval stage, (e) fourth larval stage, and (f) adult female.

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54 Fig. 7 Variation in size between adults of Trichodorus christiei collected from two locations near Sanford, Florida.

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55 Gonad Length a b c Length lst-2nd 0.311mm 20.7 2.9 124.3 .008mm 2nd-3rd 0 . 4 1mm 16.2 3.7 98.3 0.03mm 3rd-4th 0.51mm 16.3 4.3 126.6 0.04mm 4th-Adult 0.70mm 16.0 5.6 100.0 0. 23mm Laboratory Experiment 2. The molting of christiei proved different than that of any other plant parasitic nematode. Prior to molting the specimens of christiei became quiescent. The posterior intestinal contents were ejected, while the remainder became granular with the denser material collecting around the perimeter of the intestine. This collection was most pronounced ventrally. Quiescence was terminated by a movement of the neck region from side to side, with the inner cuticle of the neck region becoming markedly wrinkled on the side to which the head was tilted. Concurrently the cuticle was, at times, stretched over the head as seen in Fig. 12. During these movements, esophageal palpitations were observed and the gland nuclei were clearly defined. The old cuticle loosened, giving rise to a small overlap at the head and tail. This was followed by a break in the old cuticle circumventing the body in the region of the base of the onchiostyle (Fig. 8). By contraction on the head a "hood" was formed on the dorsum which usually remained attached to the old cuticle at a single point (Fig. 9). This formation of a hood was somewhat similar to that recorded by Lapage (19) for infective larvae_ of Trichostrongylus . Haemonchus and Ostertagia .

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Fig. 8 Break in the old cuticle prior to molting of its "hood" by Trichodorus christiei.

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58 The head of T_j_ christiei was then withdrawn from the hood. Note in Fig. 9 that the portion of the body which had emerged from the old cuticle was greater in diameter than the portion still contained within both cuticles. The old cuticle was then shed within a few minutes by movements of the animal. Detailed examinations of the head revealed the amphidial linings to be clearly visible, but no portion of the onchiostyle was found in any of the four molted cuticles. The tripartite onchiostyle of Trichodorus is unique among the plant parasitic nematodes as is the molting process. Laboratory Experiment 3. The reproductive rates of individual specimens of T^ christiei are given in Table 19. A marked increase (767<>) in the numbers of offspring recovered occurred between the 16th and 17th days. Thereafter, until the 19th day, the increase was gradual. Between the 19th and 38th days, the population increased 12.2 times. Thus, although as many as 30 offspring were recovered from a single specimen in one generation, the average reproductive rate of this nematode is 12 offspring per 19 days at 80°F using 'Homestead 24' tomato as the host plant. This figure was further substantiated by the data in Table 20a, prior to the influence of population pressures. Laboratory Experiment 4. A study was made on the effect of varying initial inoculum numbers on the reproductive rate of T. christiei . The data from this study, using 6 'Homestead 24' tomato seedlings per 4-inch plastic pot of sterile soil, are presented in Table 20a. After

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59 r-H •rH 60 • CO !> m O Q <| o 00 00 MH CD 1 3 o >, . •rH r-H X i— i 13 CD •rH co OJ M CD £ 01 3 Q Ph 4J CD 0) u r-H 1 — 1 1-1 r-l C CO o SO O CJ c CJ o O CD •H ^> •H M a, -3 00 H co CD CO a 4-1 > r-H UH CM o m CD CJ UH o a) >, o 60 x; CJ ta c 4J rC 3 •rH a) u •a vO '4H m •rH a o o •rH CO CM H i — 1 O O co co V u CD 4J a w a) x: 0 60 X) u a. CM a 4-1 i — i •rH ~j m CM CD r-H 2; •H o 3 T> "3 CD •H 01 > 00 4J •H i-H T) o 3 1 s 4J T3 1 H CO T3 4-1 OJ 60 •H •rH 60 •H 13 3 0 CD CM }_l •H pS o CO CO H T3 r-H 4J a. CO 60 CO o 10 in > 0) a 3 a mh •H 4J •rH T) >4H m U ta > •r-i o O ai •H > o 3 E > •rH a -a o S-i -3 o W 3 3 >^ to •rH xi a, CD 3 o M • a-) 0) CO XI r-H QJ i-H H a; 3 r-i x> CD H s r-l 0 CM CO in r-H CO CM vf oo CM CM co as t-H r-H vO r-H r-H i-H 00 o CO CM 00 CM 00 CO CM vO vD CN 00 CO i — i CD | | O 4-1 — — 1 r-H •H o T) 0 3 'rH CD JH CD S r-H O M -3 I4H 3 CM 60 3 CD •rH JZ U 4J (X CO 3 m •rH UH o CD MH > O •rH 60 (H • 0) CD CO JO U CD 3 60 60 CD C3 •H 4-1 p 4H CO CD r-H E x; CD rH 4J > u CO 4J CD e CO 1-1 CD 0) 4J T3 XI 3 4J 4J CM CO CO U •3 J-l •rH 3 3 UH CD CD CO CD 4-1 CD x; CO U 4J r-H a, CD 3 UH aJ H o

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60 Table 20a. Reproductive Rates of Trichodorus christiei at Different Rates of Infestation Maintained at 80°F. Inoculum level Number of T. christiei per 4-inch pot ( 400 ml) At 21 Days Increase per individual At 42 Days Increase per individua 1 25 274f/ 10.9 4,092V 163.7 100 1,253 12.5 5,786 57.8 400 1,823 4.6 7,030 17.6 aj Average of 6 replicates. ]V Average of 12 replicates. Table 20b. Average Weights Inoculated with of 'Homestead 24' Tomatoes Trichodorus christiei. Fresh Weight (grams) Dry Weight (grams) After 21 days After 42 days After 21 days After 42 days 25 Tops 3.2f/ 8.lW 0.167 0.675 Roots 0.7 1.60 0.062 0.213 100 Tops 3.5 7.8 0.200 0.658 Roots 0.8 1.55 0.073 0.224 400 Tops 2.7 6.3 0. 150 0.450 Roots 0.7 1.45 0.055 0.199 £/ Average of 6 pots; 6 plants per pot. °J Average of 12 pots; 6 plants per pot.

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61 21 days the greatest increases were found in those pots initially inoculated with 100 specimens. This was closely followed by those with 25 specimens. After 42 days the pots originally having 25 individuals produced increases of almost three times the rate of those with 100 and over nine times the rate of those with 400, thus demonstrating the importance of population pressures on the reproductive rate of T. christiei . Alhassan and Hollis (1) attributed the slower rise of higher initial inoculum numbers of 1\_ christiei to an interaction with the host. Since there was an absence of necrosis, these authors have termed this relation one of "balanced parasitism." Figs. 10 and 11 show T. christiei feeding on tomato roots in agar. Data relating to seedling weights are given in Table 20b. No statistical difference was found between the fresh weights of the tops or the roots when the treatments were analyzed after 21 days and 42 days, respectively. Alhassan and Hollis (1) obtained similar results on cotton seedlings analyzed 21 days after inoculation with 0, 100, and 400 specimens of T. christiei . Nevertheless, their conclusion was that "seedling weights were related inversely to both initial and final populations of the nematode." The same conclusion may be drawn from the above experiment on 'Homestead 24' tomatoes after 42 days. Laboratory Experiment 5 . Results from the investigation into the possibility of resistance in the eggs of L. christiei to D-D fumigation are given in Tables 21a and b. Under the conditions of the experiment; i.e., recording the nematode population 2 weeks after fumigation, and 3 weeks after planting a susceptible host, it was found that whenever

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62 Fig. 10 Characteristic position of Trichodorus christiei while feeding; i.e., the head at right angles to the cell upon which it is feeding.

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63 Fig. 11 Trichodorus chr istiei feeding upon the root-cap of tomato in agar.

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66 the adults and larvae were killed no progeny of Tj_ christiei could be found. This indicated that the eggs were killed. Adults and larvae were found to survive following applications of D-D at 50 gal per acre to the jars, whether applied in mineral spirits or in a water-emulsified state. However, it would appear that many of these surviving specimens were incapable of infecting susceptible hosts. This phenomenon appeared more pronounced at the lower application rates of D-D. The data in Table 21a indicate that T^ christiei is more tolerant to D-D fumigation than is longicaudatus or Hop lo la imus galeatus. This experiment also demonstrates the marked susceptibility of B. longicaudatus to D-D fumigation. Laboratory Experiment 6 . The survival of T. christiei in the absence of a host was studied at a constant temperature of 80°F (Table 22). The rate of mortality of T_j_ christiei was slightly greater in field soil than that of a pure population which previously had been established in sterile soil. This difference, however, was considered insufficient to indicate the presence of a successful predator. A comparison of the data obtained for survival at 80°F (Table 22) with those obtained in the field (Table 8) shows the rates of mortality at 80°F to be slightly greater during the first 8 weeks. Thereafter, the field population declined much more rapidly until at the 16th week there was a difference of 20 per cent. Often fallowing is associated with dry tillage. After allowing 6inch clay pots of soil containing T^ christiei to dry out in an air-conditioned laboratory to a moisture level as low as 1.7 per cent,

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67 it was found that live specimens could still be recovered. Specimens of T. christiei recovered from this soil characteristically possessed a thickened cuticle (Fig. 12). This cuticular thickening suggests that Table 22. Survival of Trichodorus christiei in the Absence of a Host in a 6-inch Pot at 80°F. Pure Population Field Soil Numbers of T\ Numbers of T\_ Weeks christiei per Percentage christiei per Percentage elapsed 100 ml of soil reduction 100 ml of soil reduction 0 205f/ 0 7, 69 0 7. 2 196 3 66 3 4 .157 23 47 32 8 140 32 42 40 12 115 44 16 88 57 a/ Average of two 100 ml samples. T. christiei can adapt itself to survive at low soil moisture levels. No living specimens were found in soil with a moisture level of 0.9 per cent. Laboratory Experiment 7 . The green food dye proved most satisfactory for staining specimens of J_j_ christiei , but the degree of staining varied with different individuals. The dye dispersed evenly throughout the specimens only after they were placed in a 1 per cent formalin solution. This stain was particularly successful in bringing out the reproductive system and cuticular structures.

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Fig. 12 Water mount of a live Trichodorus christiei recovered from soil with a moisture level of 1.7 per cent.

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69 Among the cuticular structures not observed on unstained specimens were: (1) A series of dorsal and ventral pores spaced along the length of the body in the cuticle which appeared to be associated with a similar series of hypodermal cells; (2) two caudal pores which terminated within the cuticle; and (3) the posterior cephalids, completely encircling the cephalic region. The Build-up of Trichodorus christiei • The data presented above do not directly answer the question of what causes the build-up of T\_ christiei following fumigation; nevertheless, certain conclusions may be drawn. Indications are that predaceous nematodes play a minor role, as shown by the slow decline in high populations of 1\_ christiei , in the absence of a host in the field. No statement can be made concerning possible effects of predaceous fungi. There does not appear to be a stage in the life cycle of T. christiei resistant to D-D fumigation. Complete eradication of nematodes has seldom been obtained in the field with soil fumigation. At Sanford, Florida, the control of T. christiei is made more difficult due to its presence at relatively greater depths than other plant parasitic nematodes. Crops on fumigated soil have improved growth over those on unfumigated soil, particularly during the seedling stages. This is due, in part, to the control of Bj_ longicaudatus by fumigation, and as a result competition between this nematode and christiei is absent in fumigated soil. Under these conditions, T. christiei can attain a higher reproductive rate as shown by the greenhouse experiment presented above.

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70 It is interesting to note that Tj_ christiei reaches its reproductive potential (i.e., 12 progeny per 19 days at 80°F) at low levels of inoculum, but that population pressures are soon observed in 4-inch pots. The reason for this probably lies in the available food supply and its effect upon the reproductive rate. It is the considered opinion of the author that the crux of the problem of the build-up of T. christiei following fumigation is, in part, due to the increased number of available feeding sites.

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SUMMARY Population studies were made on the plant parasitic nematode, Trichodorus christiei , following field applications of D-D soil fumigant on fall, spring, and summer vegetable crops. Results showed that T. christiei could be found in greater numbers in fumigated soil than in unfumigated soil within 7 weeks after planting. Nevertheless, due to better seedling growth, the fumigated areas out-yielded those unfumigated. This build-up of christiei was not seen on those plots treated with the organo phosphate nematicide zinophos. This latter nematicide was also found to have a significant residual effect on subsequent crop yields. Indications are that T_j_ christiei does not possess a resistant stage to D-D fumigation. However, consideration should be given to the fact that at harvest there is a greater population of T\_ christiei at soil depths below 6 inches than above 6 inches. The rate of mortality of T^ christiei in the absence of a host, whether or not organic matter was incorporated, did not suggest the presence of a predator. Nor were populations of T^_ christiei affected by association with three free-living nematodes or an oligochaete in sterile soil. Both christiei and Belonolaimus longicaudatus were found to parasitize pangolagrass . The effect of these two nematodes upon one another was studied when in combination on sweet corn. The numbers of 71

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72 T. christiei were greatly reduced. Belonolaimus longicaudatus was found to be more pathogenic to sweet corn than T\_ chrjstiei . In general the corn was damaged more by the combination of the species than by longicaudatus alone, though there were fewer numbers of B. longicaudatus in the combination. The type of potting container was found to significantly affect the population numbers of T\ christiei . In the greenhouse a glazed crock provided the best environment of those studied, while in a growth chamber a plastic container provided the best conditions. A study of the reproduction of T\ christiei under four constant temperatures, viz., 95°, 90°, 85° and 80°, showed the latter to be the most suitable. The effect of initial inoculum numbers on the reproductive rate of T.'. christiei was studied in sterile soil at 80°F. After 42 days the pots originally having 25 individuals produced increases of almost 3 times the rate of those with 100, and over 9 times the rate of those with 400, thus demonstrating the importance of population pressures as it is affected by the available food supply. Single T\ christiei ^specimens feeding on tomato seedlings growing at 80°F were found to give rise to an average of 12 progeny every 19 days. At this temperature the life cycle of T^ christiei from egg to egg was found to be 17-18 days. Eclosion from the egg took place on the fourth day in the first larval stage which molted shortly thereafter. The third larval stage was first observed on the seventh day, after inoculating a gravid female onto tomato seedlings; the fourth larval stage, on the tenth day and the adult emerged after 2 weeks. The stage in the life cycle of T^ christiei can best be judged by the size of the

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73 gonadal primordium. Molting of T. christiei is described. This is the first record of a plant parasitic menatode not shedding its stylet, or any part of it, during molting. Future work on the bionomics of T\_ christiei following fumigation should be directed towards the effects the improved host growth have on the reproduction of this nematode.

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LITERATURE CITED 1. Alha ssan, S. A. , and J. P. Hollis. 1966. Parasitism of Trichodorus christiei on cotton seedlings. Phytopathology. 56: 573-574. 2. Allen, M. W. 1957. A review of the nematode genus Trichodoru s with descriptions of ten new species. Nematologica . 2: 32-62. 3. Barker, K. R. , and Gayle Worf. 1964. Parasitism of southern stock azaleas in Wisconsin by Ty lenchorhynchus c laytoni . Trichodoru s christiei and Meloidogyne incognita . (Abstr.) Phytopathology. 54: 887. 4. Bird, G. W. 1966. Influence of host and geographic origin on populations of Trichodorus christiei . (Abstr.) Nematologica. 12: 88. 5. Chen, T. A., and W. F. Mai. 1965. The feeding of Trichodoru s christiei on individually isolated corn root cells. (Abstr.) Phytopathology. 55: 128. 6. Chitwood, B. G. , and B. A. Oteifa. 1952. Nematodes parasitic on plants. Ann. Rev. Microbiol. 6: 175-177. 7. Christie, J. R. 1959. Plant Nematodes Their Bionomics and Control . Gainesville, Fla. Univ. Fla. Press. 1-256. 8. Christie, J. R. , and V. G. Perry. 1951. A root disease of plants caused by a nematode of the genus Trichodorus . Science. 113: 491-493. 9. Christie, J. R. , and V. G. Perry. 1951a. Removing nematodes from soil. Proc. Helminthol. Soc. Wash. 3: 69-72. 10. Christie, J. R. , and V. G. Perry. 1959. Mechanism of nematode injury to plants, p. 419-426. In C. S. Holton et al. (ed.), Plan t Pathology Problems and Progress 1908-1958 . Madison, Wise . : Univ. Wisconsin Press. 11. Cobb, N. A. 1913. New nematode genera found inhabiting fresh water and non-brackish soils. J. Wash. Acad. Sci. 3: 432-444. 12. Esser, R. P., and E. K. Sobers. 1964. Natural enemies of nematodes. Soil and Crop. Sci. Soc. of Fla. 24: 326-353. 74

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75 13. Haasis, F. A., J. C. Wells, and C. J. Nusbaum. 1961. Plant parasitic nematodes associated with decline of woody ornamentals in North Carolina and their control by soil treatment. Plant Disease Reptr. 45: 491-496. 14. Harrison, D. S.,and R. E. Stall. 1961. Constant-temperature tanks for studying soil-borne diseases. Trans. Am. Soc. Agr. Eng. 1: 24-25. 15. Hoff, J. K.,and W. F. Mai. 1964. Influence of soil depth and sampling date on population levels of Trichodorus christiei . Phytopathology. 54: 246. 16. Hutchinson, M. T.,and H. T. Streu. 1960. Tardigrades attacking nematodes. Nematologica . 5: 149. 17. Jenkins, W. R. 1964. A rapid centrifugal-flotation technique for separating nematodes from soil. Plant Disease Reptr. 48: 692. 18. Krusberg, L. R.,and J. N. Sasser. 1956. Host-parasite relationship of the lance nematode in cotton roots. Phytopathology. 46: 505-510. 19. Lapage, G. 1935. The second ecdysis of infective nematode larvae. Parasitology. 27: 186-206. 20. Malek, R. B. , W. R. Jenkins, and Ellen M. Powers. 1965. Effect of temperature on growth and reproduction of Criconemoides curvatum and Trichodorus christiei . (Abstr.) Nematologica. 11: 42. 21. Martin, G. C. 1966. Personal communication. 22. Micoletzky, H. 1922. Die freilebenden Erd-Nematoden. Archiv. f. Naturgesch. 87: 1-650. 23. Patrick, Z. A., R. M. Sayre, and H. J. Thorpe. 1965. Nematocidal substances selected for plant parasitic nematodes in extracts of decomposing rye. Phytopathology. 55: 702-704. 24. Perry, V. G. 1953. Return of nematodes following fumigation of Florida soils. Proc. Fla. Hort. Soc. 66: 112-114. 25. Rhoades, H. L. 1964. Effect of Crotalaria spectabilis and Sesbania exa ltata on nematode populations and subsequent yield of snap beans and cabbage. Proc. Fla. State Hort. Soc. 77: 233237. 26. Rhode, R. A., and W. R. Jenkins. 1957. Effect of temperature on the life cycle of the stubby-root nematode. (Abstr.) Phytopathology. 47: 29.

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76 Rhode, R. A., and W. R. Jenkins. 1957a. Host range of Trichodoru s sp. and its host-parasite relationship on tomato. Phytopathology. 47: 295-298. Russell, C. C. 1962. The embryology and parasitic habit of Trichodorus christiei Allen with observations of predators. Unpublished Masters Thesis, Univ. Florida. Gainesville, Florida. Russell, C. C, and V. G. Perry. 1966. Parasitic habit of Trichodorus christiei on wheat. Phytopathology. 56: 357-358. Schaerff enberg, B. 1950. Untersuchungeri uber die Bedeutung der Enchytraeiden als Humusbildner und Nematodenf einde. Z. Pflanzenkrankh. Pf lanzenschutz. 57: 183-191. Thomason, I. J. 1959. Influence of soil texture on development of stubby-root nematode. (Abstr.) Phytopathology. 49: 552. Thorne, G. 1932. Specimens of Mononchus acutus found to contain Trichodorus obtusus , Tylenchus robustu s and Xiphinema americanum . (Abstr.) J. Parasitol. 19: 90. Thorne, G. 1939. A monograph of the nematodes of the superfamily Dorylaimoidea. Capita Zool. 8: 1-190. Thorne, G. 1961. Principles of Nematology . New York: McGraw Hill. 1-553. Zuckerman, B. M. 1961. Parasitism and pathogenesis of the cultivated cranberry by some nematodes. Nematologica. 6: 135143.

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BIOGRAPHICAL SKETCH Henry Vintcent Newton Morton was born on September 15, 1936, in Johannesburg, S. Africa. After attending school at St. Johns College for 10 years, he assumed the job of farm-manager on a dairy farm in Natal for a year before enrolling, in 1955, in the University of Natal at Pietermaritzburg. In 1959, he received a Bachelor of Science in Agriculture with a major in Horticulture. In 1960, he travelled to England where he joined the staff of Imperial Chemical Industries at Jealotts Hill Research Station, Berkshire, working with weedkillers. In May, 1961, he immigrated to the United States by way of Canada and worked for Pulitzer Groves as grove foreman. In 1962, he became assistant to Dr. M. Cohen, Pathologist, at the Indian River Field laboratory, Fort Pierce, Florida. In January, 1963, he enrolled in the Department of Fruit Crops at the University of Florida, where, with the aid of a Research Assistantship, he graduated with a degree of Master of Science in Agriculture in August, 1964. In September, 1964, he enrolled in graduate work in Nematology. From May, 1965, through August , 1966, he conducted research at the Central Florida Experiment Station, Sanford, Florida. 77

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78 He returned to Gainesville to aid Professor Thome in teaching the first course offered in Tropical Nematology at the University of Florida. He completed the requirements leading to the Doctor of Philosophy Degree in June, 1967. The author is a member of Alpha Zeta , Gamma Sigma Delta and Sigma Xi. In addition, he holds memberships in the .Society of Nematologists, American Society of Horticulture and Florida State Horticultural Society. In January, 1967, he married Virginia Ann Martin and adopted her two sons, Stephen and Scott.

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This dissertation was prepared under the direction of the chairman of the candidate's supervisory committee and has been approved by all members of that committee. It was submitted to the Dean of the College of Agriculture and to the Graduate Council, and was approved as partial fulfillment of the requirements for the degree of Doctor of Philosophy. . June 20, 1967 Dean, College of Agriculture Dean, Graduate School Supervisory Committee: