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Survival of Subterranean Termites (isoptera

Permanent Link: http://ufdc.ufl.edu/UFE0024743/00001

Material Information

Title: Survival of Subterranean Termites (isoptera Rhinotermitidae) Isolated in Wood of Various Moisture Contents
Physical Description: 1 online resource (58 p.)
Language: english
Creator: Chan, Wai-Han
Publisher: University of Florida
Place of Publication: Gainesville, Fla.
Publication Date: 2010

Subjects

Subjects / Keywords: body, moisture, oak, pine, relative, subterranean, termite, water, wood
Entomology and Nematology -- Dissertations, Academic -- UF
Genre: Entomology and Nematology thesis, M.S.
bibliography   ( marcgt )
theses   ( marcgt )
government publication (state, provincial, terriorial, dependent)   ( marcgt )
born-digital   ( sobekcm )
Electronic Thesis or Dissertation

Notes

Abstract: Subterranean termites nest in the soil, where they forage looking for a source of food, because they are dependent on its moisture for survival. However, some subterranean termite species are able to form aerial infestations, with no soil contact. This experiment looked at the mortality rate and water weight loss in the Formosan subterranean termite, Coptotermes formosanus Shiraki, and the Eastern subterranean termite, Reticulitermes flavipes (Kollar), at various levels of wood moisture content (WMC). Termites were isolated in northern red oak (NRO) and southern yellow pine (SYP) wood blocks ranging from 5 to 35% WMC. Using destructive sampling, termite mortality and body water content were measured over a two-week period. Separate bioassays were conducted to measure relative humidity in the air surrounding wood at 5 to 35% moisture content. Termite mortality decreased significantly with increase in WMC. Termites were unable to survive two weeks when placed in wood with < 20% WMC. Termites in 30% WMC and above is the point at which the wood is at or above full saturation, showed little to no mortality over the two week period in both NRO and SYP. Termite body water contents (BWC) decreased with a decrease in WMC. When BWC reached approximately 40%, rapid mortality occurred due to desiccation. Results showed that within air space surrounded by wood at WMC ?25%, relative humidity was measured at 98%. WMC ?25% resulted in significantly lower RH values. Termites stressed for moisture showed ability to conserve water by reabsorbing form their feces, producing fecal pellets. Our results suggest that, while termites do require their ambient environment to be close to 100% relative humidity, they depend on free water in wood for long term survival when not in contact with other moisture sources as in aerial infestations.
General Note: In the series University of Florida Digital Collections.
General Note: Includes vita.
Bibliography: Includes bibliographical references.
Source of Description: Description based on online resource; title from PDF title page.
Source of Description: This bibliographic record is available under the Creative Commons CC0 public domain dedication. The University of Florida Libraries, as creator of this bibliographic record, has waived all rights to it worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law.
Statement of Responsibility: by Wai-Han Chan.
Thesis: Thesis (M.S.)--University of Florida, 2010.
Local: Adviser: Koehler, Philip G.

Record Information

Source Institution: UFRGP
Rights Management: Applicable rights reserved.
Classification: lcc - LD1780 2010
System ID: UFE0024743:00001

Permanent Link: http://ufdc.ufl.edu/UFE0024743/00001

Material Information

Title: Survival of Subterranean Termites (isoptera Rhinotermitidae) Isolated in Wood of Various Moisture Contents
Physical Description: 1 online resource (58 p.)
Language: english
Creator: Chan, Wai-Han
Publisher: University of Florida
Place of Publication: Gainesville, Fla.
Publication Date: 2010

Subjects

Subjects / Keywords: body, moisture, oak, pine, relative, subterranean, termite, water, wood
Entomology and Nematology -- Dissertations, Academic -- UF
Genre: Entomology and Nematology thesis, M.S.
bibliography   ( marcgt )
theses   ( marcgt )
government publication (state, provincial, terriorial, dependent)   ( marcgt )
born-digital   ( sobekcm )
Electronic Thesis or Dissertation

Notes

Abstract: Subterranean termites nest in the soil, where they forage looking for a source of food, because they are dependent on its moisture for survival. However, some subterranean termite species are able to form aerial infestations, with no soil contact. This experiment looked at the mortality rate and water weight loss in the Formosan subterranean termite, Coptotermes formosanus Shiraki, and the Eastern subterranean termite, Reticulitermes flavipes (Kollar), at various levels of wood moisture content (WMC). Termites were isolated in northern red oak (NRO) and southern yellow pine (SYP) wood blocks ranging from 5 to 35% WMC. Using destructive sampling, termite mortality and body water content were measured over a two-week period. Separate bioassays were conducted to measure relative humidity in the air surrounding wood at 5 to 35% moisture content. Termite mortality decreased significantly with increase in WMC. Termites were unable to survive two weeks when placed in wood with < 20% WMC. Termites in 30% WMC and above is the point at which the wood is at or above full saturation, showed little to no mortality over the two week period in both NRO and SYP. Termite body water contents (BWC) decreased with a decrease in WMC. When BWC reached approximately 40%, rapid mortality occurred due to desiccation. Results showed that within air space surrounded by wood at WMC ?25%, relative humidity was measured at 98%. WMC ?25% resulted in significantly lower RH values. Termites stressed for moisture showed ability to conserve water by reabsorbing form their feces, producing fecal pellets. Our results suggest that, while termites do require their ambient environment to be close to 100% relative humidity, they depend on free water in wood for long term survival when not in contact with other moisture sources as in aerial infestations.
General Note: In the series University of Florida Digital Collections.
General Note: Includes vita.
Bibliography: Includes bibliographical references.
Source of Description: Description based on online resource; title from PDF title page.
Source of Description: This bibliographic record is available under the Creative Commons CC0 public domain dedication. The University of Florida Libraries, as creator of this bibliographic record, has waived all rights to it worldwide under copyright law, including all related and neighboring rights, to the extent allowed by law.
Statement of Responsibility: by Wai-Han Chan.
Thesis: Thesis (M.S.)--University of Florida, 2010.
Local: Adviser: Koehler, Philip G.

Record Information

Source Institution: UFRGP
Rights Management: Applicable rights reserved.
Classification: lcc - LD1780 2010
System ID: UFE0024743:00001


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SURVIVAL OF SUBTERRANEAN TERMITES (ISOPTERA: RHINOTERMITIDAE)
ISOLATED IN WOOD OF VARIOUS MOISTURE CONTENTS





















By

WAI-HAN CHAN


A THESIS PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT
OF THE REQUIREMENTS FOR THE DEGREE OF
MASTER OF SCIENCE

UNIVERSITY OF FLORIDA

2010



































2010 Wai-Han Chan



































To my family









ACKNOWLEDGMENTS

I thank my committee advisor Dr. Philip Koehler and committee members Dr. Roberto

Pereira and Dr. Nan Yao Su for their help and support throughout my graduate career. I would

also like to thank my parents for guiding me through this with all their love and support.









TABLE OF CONTENTS



A C K N O W L E D G M E N T S ..............................................................................................................4

L IST O F T A B L E S ...... .. ................ ............ ...... ...... .. ........................ ..............

LIST O F FIG U RE S ................................................................. 7

ABSTRAC T .........................................................................................

1 IN TRODU CTION ....................................... ..... ............................. 10

2 M A TER IA L S A N D M E TH O D S ........................................ ............................................18

In se cts .................... .................................................................. 1 8
W ood Blocks for Term ite Bioassay......................................................... ............... 18
R elative H um idity E xperim ent ................................................................ .. .....................19
M moisture Experim ents ......................... .... .................... .... .. .. .... ........ ......20
Termite Mortality at Various Wood Moisture Contents ....................................................21
Termite Body Water Content at Various Wood Moisture Contents.................. ............21
D ata A analysis ................................................... 22

3 R E SU L T S .............. ... ................................................................25

R elative H um idity .............................. ...............................................................25
Termite Mortality at Different Wood Moisture Content Levels ........................................25
Kinetics of Wood Moisture Effects on Termite Survival ....................................................28
Body W after Content ............................... ...................... ..........30

4 D IS C U S S IO N .....................................................................................................4 6

L IST O F R E FE R E N C E S ...................................................................................... ...................54

B IO G R A PH IC A L SK E T C H .............................................................................. .....................58









LIST OF TABLES

Table page

3-1 Effect of WMC, and estimated relative humidity, on the mortality of Reticulitermes
flavipes and Coptotermesformosanus ........................................ ......................... 43

3-2 Analysis of Variance results for termite mortality in 5-35% wood moisture content
for different treatm ents (a=0.05) .............................................. ............................. 44

3-3 Coefficients of determination (r2) used to determine N (kinetic orders) used for
survivalship curves .................................................... ................ 45









LIST OF FIGURES


Figure page

3-1 Percent relative humidity of space enclosed by wood blocks at various wood
m moisture contents. .......................................... ............................ 32

3-2 Effect of wood moisture on mortality of Reticulitermesflavipes and Coptotermes
f orm o sa n u s ...................................... ....................................................... 3 3

3-3 Survival curves of Reticulitermesflavipes placed in northern red oak moistened at
various w ood m oisture contents................................................ ............................. 34

3-4 Survival curves of Reticulitermesflavipes placed in southern yellow pine moistened
at various w ood m oisture contents...................... .... ......... ..................... ............... 35

3-5 Survival curves of Coptotermesformosanus placed in northern red oak moistened at
various w ood m oisture contents................................................ ............................. 36

3-6 Survival curves of Coptotermesformosanus placed in southern yellow pine
moistened at various wood moisture contents. ...................................... ............... 37

3-7 Death curve using the kinetic order m odel (n) ............... ............................... .......... 38

3-8 Death curves representing observed tim e ........................................ ....... ............... 39

3-9 Comparison of estimated (Fig. 3-7) and actual (Fig. 3-8) death curves ..........................40

3-10 Effect of wood moisture on body water content of Reticulitermesflavipes and
C op toterm esform osanus ......................................................................... ....................4 1

3-11 Relationship between classes of percent body water content (BWC) and survivorship
of Reticulitermes flavipes and Coptotermesformosanus................ ...............42

4-1 Dry fecal pellets obtained from Reticulitermesflavipes. ................................................52

4-2 Dessicated Reticulitermesflavipes worker with dried fecal pellet protruding from
a n u s .......................................................... ..................................... 5 3









Abstract of Thesis Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Master of Science

SURVIVAL OF SUBTERRANEAN TERMITES (ISOPTERA: RHINOTERMITIDAE)
ISOLATED IN WOOD OF VARIOUS MOISTURE CONTENTS

By

Wai-Han Chan

August 2010

Chair: Philip Gene Koehler
Major: Entomology and Nematology

Subterranean termites nest in the soil, where they forage looking for a source of food,

because they are dependent on its moisture for survival. However, some subterranean termite

species are able to form aerial infestations, with no soil contact. This experiment looked at the

mortality rate and water weight loss in the Formosan subterranean termite, Coptotermes

formosanus Shiraki, and the Eastern subterranean termite, Reticulitermesflavipes (Kollar), at

various levels of wood moisture content (WMC). Termites were isolated in northern red oak

(NRO) and southern yellow pine (SYP) wood blocks ranging from 5 to 35% WMC. Using

destructive sampling, termite mortality and body water content were measured over a two-week

period. Separate bioassays were conducted to measure relative humidity in the air surrounding

wood at 5 to 35% moisture content. Termite mortality decreased significantly with increase in

WMC. Termites were unable to survive two weeks when placed in wood with <20% WMC.

Termites in 30% WMC and above is the point at which the wood is at or above full saturation,

showed little to no mortality over the two week period in both NRO and SYP. Termite body

water contents (BWC) decreased with a decrease in WMC. When BWC reached approximately

40%, rapid mortality occurred due to desiccation. Results showed that within air space

surrounded by wood at WMC >25%, relative humidity was measured at 98%. WMC <25%









resulted in significantly lower RH values. Termites stressed for moisture showed ability to

conserve water by reabsorbing form their feces, producing fecal pellets. Our results suggest that,

while termites do require their ambient environment to be close to 100% relative humidity, they

depend on free water in wood for long term survival when not in contact with other moisture

sources as in aerial infestations.









CHAPTER 1
INTRODUCTION

Termites are structural pests that damage homes and buildings aesthetically and

physically. Of the 2,300 species of termites that exist worldwide, about 50 are found in the

United States, and about 18 are pests of structures (Su and Scheffrahn 1990). The Formosan

subterranean termite, Coptotermesformosanus Shiraki, is the most destructive and aggressive

subterranean termite species found in the United States, whereas the Eastern subterranean

termite, Reticulitermesflavipes (Kollar), is one of the most widespread and important economic

pests.

All termites are social insects living in colonies that can consist of over a million

individuals (Thorne 1998). In social colonies, each member completes certain tasks, dividing

their work among specific members. Termite colonies have specific pheromones which are used

to recognize nest mates of different castes and maintain separate existence from other colonies

(Potter 2004). Castes are groups of individuals of distinct morphological characteristics that

perform certain tasks within the colony. This division of labor results in an efficient, productive,

and cooperative society that can continue for years (Thorne 1998). Termite castes consist mainly

of reproductive, workers and soldiers. Colonies will also contain eggs and immatures. The

subterranean life cycle begins as an egg which hatches into larvae. These larvae later become

workers which can then turn into separate castes, including nymphs, secondary reproductive, or

soldiers.

Worker termites are considered the most important caste of the colony (Snyder 1948).

They consist of both males and females and are blind. Workers consume wood and other

cellulose based products, and are responsible to feeding and hydrating other caste members.









They are responsible for tending the nursery, tunneling through soil, building mud tubes, and

grooming the nest mates from mites and fungal infestations (Thorne 1998). These workers can

Soldier termites are the defensive caste. They act aggressively toward invaders but

sometimes can become passive and retreat from a confrontation (Thorne 1998). Their mandibles

are very effective against certain predators such as ants, and certain species that have a chemical

defense system (Lee and Wood 1971). Colonies of C. formosanus consist of -5-10% soldiers,

whereas in the species R. flavipes soldiers are only -1-2% of the colony (Haverty 1977, Haverty

et al. 2000). Proportion of soldiers in a colony is dependent on food availability, seasonal

changes, worker nurturing capacity, and other factors (Su and La Fage 1987). Because soldier

mandibles are developed for defense rather than for chewing food, soldiers are dependent on

workers for nourishment. Although soldiers do not eat directly from the food source, they do

forage and participate in food scouting (Potter 2004).

Workers can also molt into apterous neotenics, which are wingless secondary

reproductive. These are sexually mature and can reproduce if needed by the colony. Workers

can also molt into nymphs. Nymphs are not sexually mature but have a small set of wing buds.

These nymphs can lose their wing buds and molt back into a worker or can grow into an alate.

Nymphs also have the ability to molt into brachypterous neotenics, which are secondary

reproductive with wing buds. Like the apterous neotenics, brachypterous neotenics are also

sexually mature. The abdomen of these females becomes enlarged by physogastry, but they are

less mature than the queen and king of the colony.

Termites live underground in wood or in nests and will typically form tunnels in the soil

in search of food. Underground, termites are able to acquire resources and are protected from

predators, sunlight, air currents, and desiccation (Potter 2004). In moist, loose sand, the









subterranean termite will form tunnels by either pushing its head forward and then pressing the

grains of soil from side to side with its head, body, and mandibles (Ebeling and Pence 1957) or

by removing soil particles in compact soil. Su and Puche (2003) showed tunnel differences

within different moisture gradients in R. flavipes and C. formosanus. Termites tunnel more in

sand with higher moisture.

Termite species differ considerable in moisture requirements, some being able to live in

wood above ground and in dry and hot environments while others are restricted to mesic or

wetter regions (Collins 1969). Unlike other insects, which have a cuticle that forms an effective

barrier to protect them from water evaporation, subterranean termites are lightly sclerotized

(Wigglesworth 1945). They are soft-bodied insects and are prone to desiccation because of their

inability to retain water very well; therefore, subterranean termites are more dependent on

moisture for survival than other insects (Delaplane and La Fage 1989).

Because subterranean termites lose water more readily from their integument rather than

from respiration (Collins 1969), it is imperative that the moisture content of the ambient air is

very high. If the surrounding air is too dry, subterranean termites must leave that area in search

of water; otherwise they will die (Collins 1969). If the relative humidity is near 100%, then the

termites can survive and will continue to feed (Forschler 1999). In laboratory studies, Delaplane

and LaFage (1989) found that subterranean termites showed a distinct preference for moister

wood. To maintain this moisture, termites will bring moisture from their water source, such as

soil, or deposit fecal material into the galleries (Collins 1969).

Although subterranean termites typically obtain their moisture from the soil, they are

capable of forming aerial nests, with no ground contact. If the moisture content of wood above

ground level is high enough, subterranean termites can survive and multiply for an indefinite









period with no soil contact (Potter 2004). For example, an intact colony of R. flavipes was found

infesting a floating lake house and a houseboat in Kentucky, both completely suspended over

water with no soil connection (Potter et al. 2000 and 2001). Aerial infestations of C. formosanus

account for about 25% of structural infestations in Florida (Su and Scheffrahn 1986) and 50% of

high rise buildings in Honolulu, Hawaii (Tamashiro et al. 1987). Because these two species are

subterranean termites, they do require a source of moisture in order to survive. The moisture

source can be from faulty plumbing, leaky roofs, condensation, or others.

The relative humidity (RH) of the ambient air plays a large role in subterranean termite

survival. Other termite species, such as the drywood termite, Cryptotermes brevis (Walker),

obtain the majority of its moisture from the ingested wood. The rectal pads in these species are

more developed than those found in subterranean termites, allowing drywood termites to

reabsorb most of the water from their feces (Collins 1969). When exposed to highly humid

environments, however, they become progressively more bloated and die of "water poisoning"

(Buxton 1932). Subterranean termites do not have rectal pads as specialized as drywood termites

and depend on water from their food and environment.

Other insects depend on the relative humidity of their environment, as well. The stored

product beetles Sitophilus granary (Linnaeus), and S. zeamais (Motschulsky) both showed

highest survival at 100% RH, compared to <95% RH. Other beetles, such as the Tenebriodes

mauritanicus (Linnaeus), however, had peak survival at RH between 65 and 80% (Khan 1983).

The cat flea, Ctenophelidesfelis (Bouche), is unable to live as a larva when RH is <45 or >95%

(Bruce 1948). It has also been reported that in the desert fleas, Xenopsylla ramesis (Rothschild)

and X conformis (Wagner), humidity was the most important factor in determining survival.

Low humidities cause a lower lifespan in these species, decreased larval activity, and soften









cocoons. Subterranean termites are also dependent on RH and so have developed ways to

sustain the RH in aerial infestations.

Subterranean termites are able to utilize a nesting building material known as carton,

consisting of soil and masticated wood cemented together with saliva and excrement, which

allow them to sustain aerial nests. This nest carton, which helps retain and conserve water, can

be found along walls, attics, or roofs of termite-infested buildings. These nests provide termites

with moisture, food, and protection away from the soil with no ground contact. Flat roofs are

ideal locations for termites to start an aerial colony because of water pools in low areas, giving

termites the moisture they need to sustain a colony (Su et al. 1990). Just as soil tubes are used as

a guide to find active subterranean termite colonies, tubes can also be formed by termites living

in an aerial nest. Because termites found in aerial infestations have no soil contact, these tubes

will contain little to no soil and the majority of it will consist of saliva, feces, and masticated

wood (Blasingame 1987).

In urban southeastern Florida, 25% of all C. formosanus infestations are aerial. In

Honolulu, Hawaii, 50% of all C. formosanus infestations are found in high-rise buildings in

Honolulu, Hawaii (Su and Scheffrahn 1987). These infestations can be started by alates which

find a suitable area above ground to start a colony. Another way aerial infestations can be

formed is from a colony that is initially underground. Sometimes, worker termites will forage

into an above ground area and find it more suitable than the original nest. In this situation,

workers will transport the king and queen to the location above ground and the connection to the

ground is severed. The third way aerial infestations can form is from secondary reproductive

that get separated from the rest of the colony and are cut off from the ground. These secondary

reproductive are sexually mature males and females. When they are separated from their









original colony, sometimes from pesticide applications, male and female secondary

reproductive start new colonies by budding (Blasingame 1987; Su and Scheffrahn 1987).

Wood moisture content plays a large role in colony survival (Delaplane and La Fage

1989). When wood is of lower moisture content, termites will rely on external sources of

moisture. These external moisture sources allow subterranean termites to be able to survive in

wood with moisture content less than 14.8% (Blasingame 1988). However, too much water can

be lethal to termites resulting in reduction of foraging populations or cause termites to enter a

state of quiescence for several hours (Potter 2004). If the wood containing termites is above

15% WMC, the interstitial air space occupied by the termites is believed to be near 100%

relative humidity (Sponsler and Appel 1990). This relative humidity level should protect

termites from cuticular desiccation.

Depending on termite species, wood moisture content can be the factor determining

whether or not termites can sustain activity. The subterranean termites Odontotermes sp. are

known to survive best at 96% RH but can restore their moisture from the timber on which they

are feeding (Milner et al. 1997). In a choice test of wood moisture preference, R. flavipes did not

prefer moist wood over dry wood as long as they had a source of external moisture. However, C.

formosanus preferred moist wood over drier wood, even if there was an available external source

of moisture (Delaplane and La Fage 1989).

According to Forschler (1998), wood moisture readings above 15% could indicate

conditions suitable enough to support subterranean termite activity with no external water

sources. Wood moisture content below 15% would be unsuitable for the termites and would

result in death by cuticular water loss, if no external source of moisture was available. Because

termites can be found infesting wood at 15% moisture content, subterranean termites do not









necessarily have to be in fully saturated wood in order to survive and continue to feed on the

structure. Wood fiber saturation does not occur until the wood reaches 25-30% moisture content

(Carll and Highly 1999). At WMC above 25-30% wood is to be above saturation point and free

water is available for termites. Little research exists on wood moisture content required in order

sustain aerial infestations without an external source of moisture (Potter 2004).

Delaplane and La Fage (1989) showed that damp wood in buildings are more prone to

sustain aerial termite infestations and that bait blocks should have higher moisture content in

order to increase its effectiveness. McManamy et al. (2008) conducted a wood moisture

experiment and concluded that wood moisture content needed to be at least 30% WMC in order

for eastern subterranean termites to be able to survive more than six months. McManamy et al.

(2008) also stated that wood moisture content <24% was not enough to sustain an aerial

infestation, but did not measure whether relative humidity had an effect. These authors also

measured body weight and concluded that termite mortality drastically increased once termites

dropped 30-40% of their body weight. Sponsler and Appel (1990) obtained similar results and

found that water loss at time of death was an average of 50.5% for C. formosanus and 53.5% for

R. flavipes.

This study was conducted to determine effects of wood moisture content on subterranean

termites. Two types of wood, northern red oak (NRO) and southern yellow pine (SYP), and two

species of termites, C. fomosanus and R. flavipes, were used. My first objective was to

determine the air relative humidity of a void inside the wood moistened at 5-35% and compare

our results between a hard wood, NRO, and a soft wood, SYP. My second objective was to

determine the wood moisture content (WMC) needed for long-term subterranean termite survival

with no soil contact. For this experiment, two species of termites and the two species of wood









were compared and determine the wood-moisture death kinetics for subterranean termites.

Using these results, I attempted to correlate air relative humidity and subterranean termite

mortality. Finally, I wanted to determine what percent body water loss that resulted in

subterranean termite death, and to compare the results for the two termite species living on the

two wood species.









CHAPTER 2
MATERIALS AND METHODS

Insects

Three colonies ofReticulitermesflavipes (Koller), separated by more than 1.5 km, were

collected in Gainesville, FL as described by Tucker (2004). Briefly, termites were field-

collected in 6-L plastic buckets inserted in the ground and covered with a lid accessible just

above the soil surface. Two rolls of corrugated cardboard were inserted into the buckets and

were checked every 7-10 days. At those times, cardboard rolls infested with termites were

replaced with new rolls and infested rolls were taken back to the lab for termite removal. The

termites were removed from cardboard rolls and placed on moist corrugated cardboard sheets

and reared at room temperature (-230C) in plastic containers (27.0 by 19.0 by 9.5 cm) for <1 wk

before used in experiments.

Coptotermesformosanus Shiraki were collected from the Audubon Zoo in New Orleans,

LA. Termites were collected from monitoring traps stocked with cylindrical pieces of wood.

Infested wood was placed in 20-L plastic buckets and driven to the University of Florida in

Gainesville, FL. Termites in infested wood were then kept in 30.5 by 14.0 by 20.3 cm Sterilite"

Show OffsTM (Sterilite Corporation, Townsend, MA) containers with 0.95 L builder sand at

approximately 10% moisture. Termites collected from different collection sites in the New

Orleans Zoo were kept in separate containers and assumed to represent different colonies. These

containers were stored in a lit room at room temperature (-230C) for later use.

Wood Blocks for Termite Bioassay

Northern red oak (NRO) boards and southern yellow pine (SYP) stakes were cut into

small blocks (7 by 4 by 2 cm). Three adjoining holes (1 cm deep) were drilled using a 1.9 cm

Forstner bit on one side of each block to produce a void (5 by 1.9 by 1 cm). A separate hole, (0.5









cm diam. by 1 cm deep) was drilled into the corner of each block 0.5 cm from one of the corners

to allow moisture replacement. Blocks were oven-dried at 600C for 48 hr to remove any water

from the wood. Each block was then weighed and dry weights were recorded.

Relative Humidity Experiment

To measure the relative humidity in the air space surrounding moist wood, a device was

constructed to simulate the use of a sling psychrometer. A small fan, connected to either a 9V

battery or electrical outlet, was placed in the center of a five-sided 15.3 by 8.8 by 7.0 cm wooden

box. This fan moved the air and created the drying effect necessary to measure a differential of

temperature between a dry and a wet thermocouple. The wooden box was placed in a 1.9-L

Glad Gladware container (The Clorox Company, Oakland, CA).

To create the wooden boxes, NRO boards or SYP stakes were cut into five 11.3 by 3.5 by

2.0 cm (long blocks) and four 8.8 by 3.5 by 2.0 cm (short blocks). Each block had a 0.8 cm deep

and 2.5 cm wide drilled void to create blocks that were similar to the ones described above which

were used in moisture experiments with termites. The void was drilled so that it started 1.5-2 cm

from one end of the block and continued to the opposite end. Wood blocks were placed in a

600C oven for 48 hr to dry and were then moistened to 5-35% WMC by weight in increments of

5%. Wood blocks were then individually placed in 0.5-L Ziploc bags which were placed into

3.8-L Ziplocbags to prevent moisture loss. Blocks were kept in plastic bags for 36-48 hr to

allow water to completely disperse within the wood block before blocks were used for

experiments. One hour before the blocks were used for experiments, water was added to replace

any evaporated water and reestablish the desired WMC.

Nine wood blocks with the same WMC were used to create a five-sided 15.3 by 8.8 by

7.0 cm wooden box. Blocks were positioned so the side with the void faced inward in the box.

The length of the box consisted of two stacked long blocks and the width consisted for two

19









stacked short blocks. The bottom of the box was a single long block, with the void facing

upward. Size-16 rubber bands were used to secure the box. Formed and secured wooden boxes

were placed open side up in the center of the 1.9-L Glad Gladware plastic containers, which

were sealed with their plastic lids. Wooden boxes and containers remained in at room

temperature for the remainder of the experiment.

Two 3-mm holes, 1.0 cm apart, were created at the center of the lid of the plastic

container to allow two thermocouples to be inserted through the top of the lid. One of the two

thermocouples was wrapped with a wet sponge (-0.4 cm in diam. and 1.5 cm long). Size-4

cotton thread was used to secure the sponge on the thermocouple with several knots. An

additional hole was placed on the lid of the container to insert a disposable pipette. Tap water

was placed in the pipette and the pipette was placed so its tip was touching the sponge-wrapped

thermocouple to allow water replenishment of the sponge throughout the experiment.

Temperature measurements from both dry and wet thermocouples were taken with an

EasyView Dual Input Thermometer (Extech Instruments Corp., Waltham, MA) at 15-min

increments until temperatures stabilized (3-4 hr). Using an online psychometric calculator

(http://www.sugartech.co.za/psychro/index.php), wet bulb and dry bulb temperature values were

used to compute the relative humidity within the wooden boxes constructed with wood at

different WMC levels.

Moisture Experiments

Immediately after the wood blocks were oven-dried, water was added to the large void of

the wood blocks to achieve 5, 10, 15, 20, 25, 30, and 35% (wt/wt) WMC. Moistened blocks

were placed into individual 0.5-L Ziplocbags (S.C. Johnson and Son, Inc, Racine, WI), which

were kept in larger 3.8-L Ziplocbags to further help prevent moisture absorption or loss for 36-









48 hours until added water was evenly distributed throughout the wood block. After this period,

blocks were weighed and moist weight was recorded.

Termite Mortality at Various Wood Moisture Contents

Four separate experiments were conducted with R. flavipes or C. formosanus placed on

NRO or SYP using the following procedure. Termites were separated and counted to result in

101 R. flavipes or 110 C. formosanus individuals in order to mimic natural field ratios of 1 soldier

for 100 workers for R. flavipes and 10 soldiers for 100 workers for C. formosanus. After

separation from the colony, termites were chilled over ice for -1 min to slow the termites so they

could be placed into the large void in the wood blocks without escaping. Clear acetate paper (7.0

by 4.0 cm) was hot-glued over the opening of the block void to prevent termite escape and allow

for an enclosed air space. The total weight of the blocks, including termites, acetate paper and

hot glue, was recorded immediately after setup, and blocks were sealed back into individual 0.5-

L Ziplocbags. Bags with wood blocks were then placed in larger 3.8-L Ziplocbags and kept

at room temperature (-230C) for the duration of the experiment. Water was added every 7 days

throughout the experiments to the moisture reservoirs to maintain initial weight and WMC.

Termite Body Water Content at Various Wood Moisture Contents

Blocks were prepared for destructive sampling at 1, 2, 3, 5, 7, 10, and 14 days. At each

sampling day, total block weights were recorded to allow an estimation of the amount of WMC

that had been retained. Termite mortality was determined and all the surviving termites were

weighed together, then killed and dried at 600C for 30 minutes, and re-weighed to a 0.01-mg

precision. Percent body water content (BWC) was calculated by subtracting the termite dry mass

from the initial mass of the living termites and dividing the result by the initial mass. Mass loss

was assumed to be entirely from water loss.









Data Analysis

Relative humidity was not measured directly from the wood blocks in which the termites

were placed, but rather from a separate wooden box comprised of blocks of wood moistened at

WMC levels 5, 10, 15, 20, 25, 30, or 35% as described before. It was assumed that relative

humidity in the air space of the wood blocks to which termites were exposed was the same as

that within the wooden boxes used to measure relative humidity for each WMC. Death and

survivorship curves were calculated for WMC 5-25% because it was at those levels that

significant mortality in termites was observed, whereas no significant termite mortality occurred

at 30 and 35% WMC.

For the relative humidity experiment, a completely random design was conducted, using

two wood species and 7 moisture levels. Each experimental unit consisted of a wooden box built

with nine blocks of wood placed in a plastic Gladware container with a psychrometer. There

were three replicates totaling 42 experimental units. One-way ANOVA was conducted to

determine effects of wood species on ambient wood relative humidity.

Using statistical analytical software (SAS Institute 2007), termite mortality and percent

BWC were arcsine-square-root transformed. A randomized complete block design was used for

the termite mortality experiment. Three replications, from different termite colonies, with 100

termite workers plus soldiers for each termite species, C. formosanus and R. flavipes, were used.

Using destructive sampling, termites were exposed to seven different WMC levels (5, 10, 15, 20,

25, 30, and 35%) and mortality was observed every 1, 2, 3, 5, 7, 10, and 14th day. At each

sampling date for each WMC level, BWC for remaining living termites was measured.

Significant differences were determined by graphically comparing standard error bars

surrounding the means. One way analysis of variance was used to view significance differences

between treatments and significance of day and moisture within treatments. Separation of means

22









was conducted using student Neumann keuls. A t-test was conducted to measure significant

differences between termite species and wood species.

The fundamental empirical model used to estimate thermal mortality kinetics (Johnson et

al. 2009, Wang et al. 2002, Alderton and Snell 1970) was used to determine the effect of WMC

on termite mortality. Time was measured in days in order to estimate kinetics of termite

mortality when insects were exposed to different levels of WMC. Termite survival at each

WMC was plotted against exposure days, and survival curves were obtained for each WMC by

linear regression. A WMC death curve was obtained by plotting the observed minimum time in

days [Log (time) plotted on y-axis] needed to obtain 100% termite mortality at the different

WMC (x-axis). Plots were obtained with kinetic orders 0, 0.5, 1, 1.5, and 2 following the

methods described by Wang et al. (2002). Coefficients of determination (r2) were calculated for

each curve. Based on these coefficients, the kinetic order equations that best fitted the survival

data were selected. Parameters for kinetic orders 0 and 0.5 order equations were used to

determine the effect of WMC on termite mortality. These kinetic orders were chosen based on

the best combination of coefficient of determinations, comparing all kinetic orders.

Slope and intercept from the regression equations were used to estimate the number of

days to 100% mortality at different WMC for each of four treatments (C. formosanus on SYP, C.

formosanus on NRO, R. flavipes on SYP, and R. flavipes on NRO). Estimated time to cause

100% termite mortality according to these curves were plotted against WMC. Confidence

intervals (95%) of slope and intercept were used to compare significance in the regressions lines

for each treatment.

To determine the accuracy of these estimates, another WMC death curve was obtained

plotting WMC against the number of days (LOG) where 100% mortality was actually observed









in the wood blocks. Confidence intervals (95%) of slope and intercept were used to compare

regressions lines obtained for each termite species/wood type combination. Differences between

treatments were determined to be significant when confidence intervals did not overlap.

Termite BWC for each of the 7 sampling dates (1, 2, 3, 5, 7, 10, and 14 d) was graphed

for each WMC level (5, 10, -35%). Significant differences between BWC within treatments

were determined by comparing standard error values; means with overlapping standard errors

were considered not significantly different and means with standard errors that did not overlap

were considered significantly different. The percent BWC relationship to survival of the termites

was measured by comparing the amount of termites that survived at certain BWC levels. Termite

BWC was grouped into increments of 5%. Number of surviving termites measured at specific

BWC was graphed.









CHAPTER 3
RESULTS

Relative Humidity

Relative humidity (RH) in the air surrounding the wood box arenas showed a significant

increase with an increase in WMC, reaching at plateau at 25% WMC for both NRO and SYP

(Fig. 3-1). There was no significant difference in RH between the two types of wood (df-1;

F=0.10; P 0.7583). At WMC levels >25%, RH values were >90%, however no RH values

above 98% were measured using the methods in these studies. Relative humidities for 25, 30

and 35% WMC were not significantly different on NRO or SYP.

Termite Mortality at Different Wood Moisture Content Levels

An increase in WMC yielded a decrease in termite mortality and increase in longevity for

both R. flavipes and C. formosanus on NRO and SYP (Table 3-1). Termite mortality was

significantly lower at higher WMC for all treatments (Table 3-2). Analysis of variance showed a

significant relationship between WMC and termite mortality, representing a significant decrease

in mortality with increase in WMC. Neither species of termites were able to sustain activity

longer than 7 d in arenas where surrounding air was <90% RH.

Mortality of R. flavipes on NRO (Fig. 3-2A) decreased with an increase in WMC yet a

minimum of 25% WMC was required to yield some survival for the 14-day duration of the

experiment. Rapid mortality occurred at 5-20% WMC within the first 7 days. By 7 d, all termites

had died at up to 15% WMC. At 20% WMC, termite mortality was <10% within the first 5 d but

increased to 72% by 7 d. There was a slight increase in mortality at 25% WMC on 10 d but no

significant differences, with less than 10% mortality, occurred between 25, 30, and 35% WMC at

14 d for R. flavipes on NRO (df-2; F=2.00; P 0.2160).









Similarly, R. flavipes on SYP at 5-15% WMC (Fig. 3-2B) were not able to survive longer

than 7 d. At 20% WMC, rapid mortality occurred between 3 and 10 d with an average of 89%

population mortality. R. flavipes mortality at 25% was not observed until after 7 d, averages for

mortality were 0.3% at 7 d, 19.8% at 10 d, and 43.2% at 14d. There were no significant

differences in mortality between sampling days at 30% (df 6; F=0.35; P 0.8953) or 35% (df=6;

F=0.80; P=0.5890) WMC, both with <3 mortality throughout the experiment.

Coptotermesformosanus on NRO (Fig. 3-2C) were not able to survive 5-15% WMC

longer than 7 d. At 20% WMC, termites had <20% mortality for the first seven days but at 10 d,

mortality was 100%. At 25% WMC, mortality slowly increased from 8.5% at 7 d, to 11.2% at

10d, and rapidly increased to 41.2% at 14 d. At 30 and 35% WMC, mortality maintained below

10% and showed no significant differences between sampling days (30% WMC: df=6; F=0.43;

P=0.8447; 35% WMC: df=6; F=0.77; P=0.6029)

Coptotermesformosanus on SYP at 5-15% WMC (Fig. 3-2D) were also unable to survive

past 7 d. When termites were placed in SYP at 20% WMC, C. formosanus mortality began to

increase rapidly from 5 to 10 d to an average of 72%. On SYP at <20% WMC, C. formosanus

SYP were unable to survive 14 d. At 25% WMC, the majority of mortality for C. formosanus

was observed between 7 and 14 d with an average of 53% mortality. C. formosanus mortality on

SYP at 25% WMC slowly increased from 5.45% at 7 d to 10.91% at 10 d and increased to

61.82% at 14 d. There was significant mortality at 30% WMC and at 35% WMC at 14 d.

However, there was no significant difference between 30 and 35% WMC mortalities at 14 d.

One way analysis of variance showed significant differences in mortality between

treatments (df=3; F=4.64; P=0.0033). Separation of means showed no significant differences

between C. formosanus on SYP, R. flavipes on NRO, and R. flavipes on SYP. However, C.









formosanus on NRO did have significantly lower mortality. There were no significant

differences in mortality between the two termite species (df-559; T=-1.47; P=0.1416). However

mortality was significantly greater on SYP than NRO (df=559; T=-2.73; P=0.0064).

For R. flavipes at 5% WMC on NRO, the air RH was 54% which lead to 11% termite

mortality by 1 d, 96% by 2 d, and 100% mortality by 3 d (Table 3-1). A 5% increase in WMC

increased air RH to 75% at 10% WMC, which caused 98% termite mortality by 3 d and 100%

by 5 d. At 15% WMC, air RH in wooden boxes had increased to 85% though it was not enough

to allow termites to survive beyond 7 d. Mortality at 10 d for R. flavipes on NRO was 96% at

20% WMC and 1.3% at 25% WMC, though air RH for both test arenas were estimated to be

95% (Table 3-1). Wooden NRO boxes with 30 and 35% WMC were measured to have 97% air

RH and resulted in <2% termite mortality throughout the 2-week period (Table 3-1).

On SYP, RH's of the air surrounding wood at 5-15% WMC were not significantly

different from air RH's surrounding NRO at similar WMC (Table 3-1), yet R. flavipes mortalities

were significantly higher on SYP than on NRO. On SYP at 5% WMC, air RH was 52% and

resulted in 78% mortality at 1 d and 100% mortality by 2 d. On SYP at 10% WMC, air RH in

wood boxes was 73% which resulted in 25% R. flavipes mortality on 1 d, 64% at 2 d, 99% at 3 d,

and 100% by 5 d. At 25% WMC, RH was 95% and <2% termite mortality was observed until 14

d when there was 43% mortality (Table 3-1).

Coptotermesformosanus on NRO were not able to survive >10 d when the RH in the air

surrounding the wood was assumed to be <90% from the RH data at <20% WMC (Table 3-1).

When the RH was 95% (20% WMC), C. formosanus mortality reached 100% at 10 d. Relative

humidity was not significantly different between 20 and 25% WMC on NRO however, termite









mortality was significantly lower at 25% WMC than 20% WMC. At 98% RH (35% WMC on

NRO), C. formosanus mortality was <5% throughout the two-week period.

For C. formosanus on SYP at 25% WMC, though the air RH in the arenas was measured

in wooden boxes to be 95%, as was seen in NRO, 64% mortality occurred at 14 d (Table 3-1).

At 20 and 25% WMC (97% RH), mortality slowly increased to 13% by 14 d at 30% WMC, and

to 18% at 14 d at 35% WMC.

Kinetics of Wood Moisture Effects on Termite Survival

Daily mortality rates for both termite species on both wood species at 30 and 35% WMC

were not estimated because the estimated slopes for the mortality lines were not different from 0.

Coefficients of determinations (r2) were determined to choose the best kinetic model for

death curve estimates (Table 3-3). The 0.5-order kinetic model (Wang et al. 2002) were used for

R. flavipes on NRO (Fig. 3-3). At 5% WMC, mortality was estimated to begin to occur before 1

d and to reach 100% at 3 d, increasing at a rate of 45.4% per day. For 10% WMC, mortality was

also estimated to begin before 1 d and to reach 100% at 4.6 d. At 10% WMC, live populations

were estimated to decrease by 24.5% per day. When NRO had 15% WMC, mortality was

estimated to begin at 1.1 d and to reach 100% at 9.5 d, with populations decreasing by 11.9% a

day. At 20% WMC, mortality was estimated to begin at 4.4 d, and to reach 100% mortality by

10 d, with the population decreasing 16.4% per day. For wood at 25% WMC, mortality was

estimated to begin at 4.8 d. Populations were not estimated to reach 100% mortality until 52.3 d

and were estimated to reach 19% mortality at 14 d, increasing at a rate of at 2.1% per day.

The 0-order kinetic model was used for R. flavipes on SYP and C. formosanus on NRO

and SYP (Fig. 3-4, 3-5 and 3-6). Survivorship curves for all three treatments showed a decrease

in their rate of mortality with an increase in WMC. For R. flavipes on SYP (Fig. 3-4), at 5%

WMC, mortality was estimated to begin within the first day and reach 100% by 2 d. At 15%

28









WMC, mortality was estimated to begin before 1 d and to reach 100% by 7 d, with populations

decreasing at 16.8% per day. With 20% WMC, mortality was estimated to begin at 1.3 d and to

reach 100% mortality by 12 d, decreasing 8.9% a day. At 25%, mortality was estimated to begin

at 6.9 d and to reach 43% at 14 d. Using this death curve, it was estimated that R. flavipes at

25% WMC in SYP would reach 100% mortality by 23.3 d, with a mortality rate of 6.1 % per

day.

The wood-moisture death kinetics for C. formosanus on NRO (Fig. 3-5) estimated that

termites started dying before 1 d at 5% WMC and that mortality reached 100% mortality at 2 d,

with populations decreasing 68.7% per day. At 10% WMC, mortality was estimated to begin

before 1 d and reach 100% at 4.5 d. Populations of C. formosanus are estimated to decrease by

22.7% per day when placed on NRO at 10% WMC. At 15% WMC, C. formosanus were

estimated to start dying at 1.9 d and to reach 100% mortality by 6.9 d, with populations

decreasing by 19.7% per day. When C. formosanus termites were placed in NRO at 20% WMC,

mortality was estimated to begin at 4.7 d and reach 100% mortality 10.6 d, with populations

decreasing by 17.1% per day. At 25% WMC, mortality was estimated to begin at 6.1 d and

reach 38% mortality at 14 d. Using this curve, 100% mortality was estimated to occur at 26.8 d.

When C. formosanus was placed in SYP (Fig. 3-6) at 5% WMC, mortality was estimated

to begin before 1 d and 100% mortality was estimated to occur by 2.7 d. Populations were

estimated to decrease at a rate of 29.8% per day when placed in 5% WMC. At 10% WMC,

mortality was estimated to start before 1 d and reach 100% by 4.2 d, with populations decreasing

by 18.8% per day. At 15% WMC, mortality was estimated to start before 1 d and reach 100% at

6.8 d, with populations decreasing by 14.4% per day. For 20% WMC, the wood moisture death

curve estimated mortality to begin at 1.2 d and reach 100% by 11.0 d, with mortality rates of









10.2% per day. At 25% WMC, mortality was estimated to begin at 4.8 d and reach 62% at 14 d.

When the SYP was 25% WMC, mortality for C. formosanus was not estimated to reach 100% at

19.7 d.

The WMC death curves plotted using the slopes and intercepts from the model-estimated

survival curves (Fig. 3-7) showed that there were no significant differences in kinetics of WMC-

effects between R. flavipes and C. formosanus on NRO. On SYP, R. flavipes was estimated to

survive longer than C. formosanus for each respective WMC. The 95% CI for the wood

moisture death curves generated with the observed data on time to reach 100% mortality (Fig. 3-

8) showed that survival of both termite species on SYP and NRO were not significantly different.

A comparative graph was made combining death curves from the model-estimated survival

curves and death curves generated with observed data (Fig. 3-9). No differences were observed

between mortality estimates based on model-estimated kinetics and actual mortality data.

Body Water Content

Body water content (BWC) in termites started to decrease within the first day for all

treatments at all WMC levels (Fig. 3-10). Termites confined to blocks with higher WMC lost

BWC at a slower rate. The average initial BWC for R. flavipes was 77%. On NRO, termite

mortality for WMC 5-20% occurred rapidly after termite BWC decreased to an average of 40%

(Fig. 9A).

In SYP at 5% WMC (Fig. 3-10B), R.flavipes's lowest BWC measurement was 39% at 1

d. On SYP, termites decreased to a BWC average of 38% when placed on wood at 5-20% WMC

before reaching 100% mortality. There were only two remaining live termites after 3 d from one

rep at 10% WMC which were measured to have reduced their BWC to 22% before reaching

100% mortality. At 30 and 35% WMC, termite BWC reached levels -70% before recovering

previously lost BWC.









Coptotermesformosanus initial average BWC was 71%. The lowest BWC measurement

of live termites on NRO between 5, 10, and 15% WMC was between 40 and 50% (Fig. 3-10C)

before all termites in the treatment died. Termites reached an average of 39% BWC before

reaching 100% mortality at 5-20% WMC. Live termites on wood at 25-35% WMC did not

replenish all the BWC lost and were measured to have an average BWC of 54% at 14 d.

On SYP, C. formosanus BWC decreased on the first day for all WMC levels (Fig. 3-

10D). The lowest BWC recorded for termites at 5, 10, and 20% WMC was -45%. Four

remaining surviving termites subjected to 15% at 5 d were measured to have 31.63%. C.

formosanus BWC had decreased to an average of 41% for insects on wood at 5-20% WMC

before reaching 100% mortality. After 14 d, termites in 25-35% WMC had an average of 52%

BWC. Standard error differentiation showed these values to be significantly lower than initial

BWC.

There was a positive correlation between termite BWC and survival for all treatments

(Fig. 11). Survival of R.flavipes on NRO (Fig. 3-11A) was highest when BWC was 51-55%.

For this treatment, there was an increase of 49% in survival between 36-40% and 41-45% BWC.

For R. flavipes on SYP, (Fig. 3-11B), termite survival increased more gradually with increase in

BWC, however, <50% of termites were able to survive at BWC lower than 41-45%. Similarly,

with C. formosanus on NRO (Fig. 3-11C), an average of 44% of termites were able to survive

when BWC was 41-45% or less. C. formosanus survival on NRO was highest when BWC was

51-55% and greater. For C. formosanus on SYP (Fig. 3-11D), survival also showed a gradual

increase with an increase in termite BWC. C. fomosanus survival on SYP was >50% when BWC

was 36-40% and 68% survival was observed in termites with 41-45% BWC. When BWC was

above 50%, termite mortality became minimal (89% survival or higher).











100

90
80






-- SSYP
30

20 -
0NR 0
10 -
30
20
10


0 5 10 15 20 25 30 35 40
Wood Moisture Content

Figure 3-1. Percent relative humidity (RH) of space enclosed by Southern Yellow Pine (SYP)
and Northern Red Oak (NRO) at 5, 10, 15, 20, 25, 30, and 35% wood moisture
content (WMC).





























0 2 4 6 8 10 12 14


100
90
80
S70
S60
I 50
S40
S30
^ 20
10
0


0 2 4 6 8
Day


10 12 14


0 2 4 6 8
Day


10 12 14


Figure 3-2. Effect of wood moisture on mortality of Reticulitermesflavipes and Coptotermes
formosanus placed in southern yellow pine (SYP) and northern red oak (NRO) wood
blocks with 5, 10, 15, 20, 25, 30 and 35% moisture contents: A) R.flavipes on NRO;
B) R. flavipes on SYP; C) C. formosanus on NRO; D) C. formosanus on SYP.


0 2 4 6 8 10 12 14
















0.9


0.8 y = -0.4539x+ 1.2828
S0.05 R2 = 0.8488

S0.7 \ y = -0.2453x+1.1242
S0.10 R2= 0.8167

0.6 -
A y = -0.1193x + 1.1358
A 0.15 R2= 0.9344
S0.5
y = -0.164x + 1.7187
04 0.20 R2= 0.9152
o 0.4


SR'= 0.9872

0.2

0.1 A


0.0
0 2 4 6 8 10 12 14

Day

Figure 3-3. Survival curves of Reticulitermesflavipes placed in northern red oak moistened at 5,
10, 15, 20, and 25% moisture contents.

























*0.05


10.10


A0.15


X0.20


H,0.25


= -0.4''"" i\ +
R, = *| "i-t,

-0.3726x + 1.1173
R- = 0.9987

0.16"8x+ 1.1152
R- = 0.9096

S-0.089x+ 1.1123
R' = 0.9376
-0.0611x+ 1.4206
R2 = 0.9992


o ,, \ "x
0 2 4 6 Day 8 10 12 14
Figure 3-4. Survival curves of Reticulitermesflavipes placed in southern yellow pine moistened
at 5, 10, 15, 20, and 25% moisture contents.














0.9 w\

0.8 -

2 0.7

S0.6 -
S0.05 y -0.6869x+ 1.3748
S0.5 R2 = 1

04 0.10 y = -0.2272x + 1.0318
S0R2= 0.8743

6 0.3 0.15 y =-0.1974x+ 1.3687
S\\ R2R'= 0.9656
0.2
02 0.20 y = -0.1708x + 1.8077
\ R'= 0.8011
0.1
\ 0.25 y = -0.0483x + 1.2959
0.0 R2= 0.8718
0.0
0 2 4 6 8 10 12 14 16
Day
Figure 3-5. Survival curves of Coptotermesformosanus placed in northern red oak moistened at
5, 10, 15, 20, and 25% moisture contents.











1.0
X y= -0.298x+0.8001
0.9- 0.05 R2= 0.7667

A m0.10 Y =-0.1875x+0.7942
0.8 R = 0.6665

0.7 0.15 y= -0.1439x + 0.9844
X R2= 0.8509

0.6 0.20 y=-0.1019x+1.1238
1 \ R2= 0.8828
4 0.5 0.25 Y=-0.0674x+1.3258
R2= 0.9791
S0.4
A \\0
r 0.3

S0.2

S0.1

0.0 -
0 2 4 6 8 10 12 14 16
Day
Figure 3-6. Survival curves of Coptotermesformosanus placed in southern yellow pine
moistened at 5, 10, 15, 20, and 25% moisture contents.










2
1.8
1.6
1.4
1.2
1
0.8
0.6
0.4
'0.2

0


I 2
. 1.8

S1.6
1.4
H 1i


2
y= 0.058x+0.0967 1.8 y 0.055x- 0.0117
R2= 0.9024 / 1. R2= 0.9929
s 1.6 -
s/ 1.4 -
s/ 1.2 -

/ U 1 /I

R 0.9771 .8 R0 .9 0.

-0 10.6 0






1.2 -
R 0. 9771 10. 1.8 R 0.9979




-'n=0 0.6
S0.4 -
S 0 -










0 5 10 15 20 25C 0 10 15 20 2.D
Percent Wood Moisture Content
0 --0.6

_0.4


Figure 3-7. Death curve using the kinetic order model (n) representing LOG of days to reach
100% mortality of Reticulitermesflavipes and Coptotermesformosanus placed in
southern yellow pine (SYP) and northern red oak (NRO) blocks with 5, 10, 15, 20,
and 25% wood moisture contents (WMC), dotted lines represent 95% confidence
interval of regression line: A) R. flavipes on NRO, B) R. flavipes on SYP, C) C.
formosanus on NRO, D) C. formosanus on SYP.













y= 0.0523x+0.2025 s
R = 0.9924

/0





JI
-S




0 S

S
'D








5 10 15 20 2


y = 0.0449x+0.1505 /
R 0. 93 7,
) 5 10 15 20 2


y =0.0449x+0.1505 s
R= 0.9327 s








-
-


2

1.8 -

1.6

1.4

1.2

1

0.8

0.6

0.4

0.2
a


k


y= 0.0567x+0.0775
R = 0.9617










'S7;


4,
4,


'V


0 5 10 15 20 25


y =0.0462x+0.2535 ,-
R 0.9846 ,

r
j,
SL'
z,


jJ o, '


r
a.
4,


0 5 10 15 20 25 0 5 10 15 20 25
C D
Percent Wood Moisture Content
Figure 3-8. Death curves representing observed time (Log[days]) and wood moisture content
(WMC) combinations in Reticulitermesflavipes and Coptotermesformosanus
yielding 100% mortality, dotted lines represent 95% confidence interval of regression
line: A) R. flavipes on NRO; B) R. flavipes on SYP; C) C. formosanus on NRO; D) C.
formosanus on SYP.


0.8

0.6

0.4

0.2

0


2


1.8 -

1.6

1.4

1.2

1

0.8

0.6

0.4

0.2

0


.


S
S
S
S
S









2
1.8
1.6
1.4
1.2
1
0.8
0.6
0.4
,0.2
0


2
- 1.8
1.6
1.4
E 1.2
1
0.8
0.6
0.4
0.2
0


0 5 10 15 20 25
B


0 5 10 15 20 25 0 5 10 15 20 25
C D
Percent Wood Moisture Content
Figure 3-9. Comparison of estimated (Fig. 3-7) and actual (Fig. 3-8) death curves representing
time (Log[days]) and wood moisture content (WMC) combinations in Reticulitermes
flavipes and Coptotermesformosanus yielding 100% mortality, dotted lines represent
95% confidence interval of regression line. A) R. flavipes on NRO; B) R. flavipes on
SYP; C) C. formosanus on NRO; D) C. formosanus on SYP


10 15 20 25
A


0 5








--0-5 --W10 -~15 "-220 --25 --4030 -35


80
70
60
50
40
30
20
0

80
70
60
50
40
30
20 -
0

80
70
60
50
40
30


4 6 8


Day
Figure 3-10. Effect of wood moisture on body water content of Reticulitermesflavipes and
Coptotermesformosanus placed in southern yellow pine (SYP) and northern red oak
(NRO) wood blocks with 5, 10, 15, 20, 25, 30 and 35% moisture contents: A) R.
flavipes on NRO, B) R. flavipes on SYP, C) C. formosanus on NRO, D) C.
formosanus on SYP.


14
A
























S "63o 3LS *'o 1 '. 0s S o


-' o 6 /-6 I 'l I S S6


S 1630 36*S 0lS 1 S6, 50 -


^-x' =?63J /'3'6'o i' 3> s ^ 6D


Percent Body Water Content
Figure 3-11. Relationship between classes of percent body water content (BWC) and
survivorship of Reticulitermesflavipes and Coptotermesformosanus confined to
wood at different WMC. A) R. flavipes on NRO; B) R. flavipes on SYP; C) C.
formosanus on NRO; D) C. formosanus on SYP.









Table 3-1. Effect of WMC, and estimated relative humidity, on the mortality of Reticulitermesflavipes and Coptotermesformosanus
over 14 d in northern red oak (NRO) and southern yellow pine (SYP)
Reticulitermes flavipes Coptotermesformosanus
Days Days
Estimated
wood % % Relative
species WMC Humidity 1 2 3 5 7 10 14 1 2 3 5 7 10 14
NRO 5 54 11 96 100 31 100 -
10 75 2 45 98 100 08 44 85 100 -
15 86 0 23 23 64 98 100 05 09 11 68 100
20 95 0 06 02 03 72 96 100 02 04 1 19 14 100
25 96 0 03 04 02 08 19 09 03 02 03 08 08 11 41
30 95 0 0 02 0 01 01 04 01 04 02 03 05 07 04
35 98 0 0 02 0 01 02 0 01 01 0 03 02 02 05
SYP 5 52 78 100 4 99 100 -
10 73 25 64 99 100 15 78 96 100 -
15 86 02 08 58 77 99 100 01 16 63 72 99 100
20 90 01 03 06 3 6 95 100 0 05 34 27 49 98 100
25 95 0 01 02 01 0 2 43 0 04 07 05 11 31 64
30 97 01 01 01 0 0 0 0 0 06 06 08 08 13 13
35 97 0 01 01 01 0 02 01 0 03 06 07 05 11 18









Table 3-2. Analysis of Variance results for termite mortality in 5-35% wood moisture content
for different treatments (a=0.05)
Treatment DF MS F-value P-value
R. flavipes on NRO 6 6.51 33.45 <0.0001
R. flavipes on SYP 6 7.77 50.39 <0.0001
C. formosanus on NRO 6 1.79 12.96 <0.0001
C. formosanus on SYP 6 6.38 32.82 <0.0001











Table 3-3. Coefficients of determination (r2) used to determine N (kinetic orders) used for
survivorship curves to estimate termites mortality at any given wood moisture
content. Highlighted values represent those used to develop death curves
Wood Moisture Content
Treatment N 5 10 15 20 25 30 35
R. flavipes
NRO 0 1 0.8743 0.9656 0.8011 0.8718 0.2763 1
0.5 1 0.979 0.9505 0.8204 0.8306 0.3027 1
1 1 0.933 0.8067 0.8357 0.8521 0.3312 1
1.5 1 0.8066 0.7035 0.8413 1 0.3621 1
2 1 0.7755 0.6854 0.842 0.8379 0.3951 1

R. flavipes
SYP 0 0.7667 0.6665 0.8509 0.8828 0.9791 0.843 0.9992
0.5 0.8089 0.7984 0.9388 0.8193 0.9808 0.8398 0.9998
1 0.8909 0.9201 0.9646 0.7151 0.7158 0.8366 1
1.5 0.946 0.9469 0.8972 0.6542 0.1016 0.8335 0.9998
2 0.9608 0.9493 0.8571 0.6432 0.0342 0.8303 0.999

C. formosanus
NRO 0 0.7829 0.7753 0.9122 0.8678 0.9944 0.9301 0.1263
0.5 0.8488 0.8167 0.9344 0.9152 0.9872 0.9265 0.1241
1 0.9684 0.8701 0.9010 0.9665 0.9778 0.9229 0.1219
1.5 0.9970 0.8872 0.8549 0.9846 0.9666 0.9192 0.1197
2 0.9777 0.8893 0.8444 0.9867 0.9543 0.9155 0.1175

C. formosanus
SYP 0 1.0000 0.9987 0.9096 0.9376 0.9992 1.0000 0.2500
0.5 1.0000 0.9728 0.9629 0.9520 1.0000 1.0000 0.2421
1 0.7500 0.8426 0.8755 0.8980 0.9995 1.0000 0.2344
1.5 0.7500 0.8706 0.6943 0.8371 0.9975 0.8176 0.2267
2 0.7500 0.8842 0.6346 0.9268 0.7784 0.8176 0.2192









CHAPTER 4
DISCUSSION

Though studies have been conducted demonstrating that wood moisture content needs to be

high in order for termites to be able to sustain activity, the exact wood moisture content needed,

without external moisture sources was unknown. My study aimed at determining the wood moisture

content effects on termite survival and body water content. Previous studies have shown that

termites prefer moist wood over dry wood (Delaplane and La Fage 1989), even if given an additional

source of water. However, these authors did not examine the effects the lower moisture wood had

on subterranean termites.

WMC affects the relative humidity in the ambient environment around the block of wood.

Though it has been noted that termites are able to survive in wood as long as the moisture content is

>16% (Forschler 1999), my data suggests otherwise. It has been suggested that when wood has

>16% WMC, the relative humidity in the interstitial spaces is at or near 100% and, therefore,

termites should not die from water loss through the cuticle. However, my results show that even in

cases where the wood is >16% WMC, and even at 25% WMC, termites still lose body water.

I estimated the relative humidity of the air within 15% WMC blocks to be <90%, which

explains why termites were unable to survive for a week at 15% WMC or below. At 20% WMC,

neither R. flavipes nor C. formosanus were able to survive longer than 10 days, where relative

humidity was lower than that found at 25% WMC. This suggests that relative humidity needs to be

>95% in order for termites to sustain activity as shown before. McManamy et al. (2008) found that

WMC had to be >24% in order to sustain subterranean termite aerial infestations with no soil

contact. However, the authors assumed that the relative humidity within the wood was at or near

100% based on Forchler's review (1999). Our results suggest that relative humidity may be lower









than previously expected when the wood has >24% WMC, therefore, termites can lose cuticular

body water.

Relative humidity results peaked to 98% relative humidity at 25% WMC and there was no

significant difference between the relative humidity in the area surrounding wood boxes moistened

to 30 and 35% WMC. I was unable to obtain a relative humidity value of 100% when wood was

moistened at a point higher than saturation. This may have occurred because tap water was used to

wet the wood blocks. Wood saturation with distilled water may have given a close relative humidity

of 100% (Almeida and Hernandez 2005). Tap water may have contained dissolved salts which

prevented measurements of ambient environment from reaching 100%. Studies have shown that tap

water contains higher fluoride content than distilled water (Lalumandier et al. 2000).

Sponsler and Appel (1990) exposed termites to conditions of very low relative humidity (0-

2%). Our results show that termites were able to lose more BWC throughout time than previous

studies before death occurred. The longer survival observed in my studies may have been a result of

moisture from wood maintaining a higher ambient relative humidity for a longer period of time. A

higher relative humidity would have allowed termites to retain their BWC for a longer time,

maintaining BWC above a critical level at which death occurs.

Relative humidity and the temperature to which termites are exposed cause changes in

termite BWC which determine survivorship of the insect. Previous studies have also shown that R.

flavipes will live 5.1 hr at 300C and 0-2% RH, with 53.5% BWC at time of death, and C. formosanus

will live 7.1 hr, with 50.5% BWC at time of death (Sponsler and Appel 1990). Collins (1969)

showed that R. flavipes workers were able to live 3.0-6.2 hr and drop to 56.5% BWC before 100%

mortality in 34-35C and 0-4% relative humidity conditions.









Although a surviving R. flavipes termite on SYP at 10% WMC was measured as having

21.7% BWC, termites began to die rapidly at approximately 35-49% body weight loss in my

experiment, which is lower than the BWC observed at time of death by Collins (1969) and Sponsler

and Appel (1990). McManamy et al. (2008) found similar results where 20% WMC caused termites

to drop approximately 40% body weight which led to rapid mortality.

In addition to ambient relative humidity, temperature also has an effect on termite mortality.

Our experiment were maintained at room temperature (23 C), contrary to those in previous

mentioned experiments (Collins 1969; Sponsler and Appel 1990) where termites were held in

>300C. Lower temperatures may have had an effect on the length of time it took to reach 100%

termite mortality, allowing termites to survive longer.

Unlike Collins (1969) who exposed termites to high temperatures (30C) and low relative

humidities (0-2%), my experiment maintained termites at room temperature and relative humidities

>60%. Lower temperatures and higher relative humidities may have allowed termites to decrease

their BWC over a longer time before reaching 100% mortality. My experiment showed that even

though BWC decreased, termites were able to survive longer than 24 hr with ambient relative

humidity <60%. As shown by Mellanby (1939), termites were able to lower their BWC and still be

able to sustain activity.

My results showed that even though there was no significant difference between termite

mortality at 30, and 35% WMC, mortality was significantly greater at 25% WMC for termites in

SYP. McManamy et al. (2008) reported that wood moisture content needed to be at least 30% in

order for subterranean termites to sustain an aerial infestation. Wood fiber saturation does not occur

until the wood reaches 25-30% moisture content (Carll and Highly 1999); any moisture content

above that is free water in the wood. This suggests that termite mortality is not only dependent on









ambient relative humidity but also require access to free water. Both R. flavipes and C. formosanus

on NRO showed no significant difference in mortalities between 25 and 30% WMC at the end of the

experimental period.

Insects are able to draw upon their hemolymph as a liquid reserve without impairing their

locomotion or respiration (Mellanby 1939). Termites are able to do this and have been seen walking

after substantial hemolymphic fluid loss following flattening of the abdomen (Collins 1969). By

flattening their abdomens, termites were able to sustain more body water weight loss while being

exposed to lower relative humidities. This explains why termites in my experiments were able to

live for days at lower relative humidities.

Another source of moisture when stressed for water may be the feces of the subterranean

termite. Subterranean termite feces is usually a muddy texture, with no solid shape, unlike drywood

termite feces. Nakayama et al (2004) suggested that when subterranean termites become stressed for

moisture, they will excrete liquid feces to increase ambient relative humidity. Contrary to these

reports, solid pellets of feces were found in blocks with WMC <25% in my experiments (Fig. 4-1).

Similar observations were made where species of Kalotermitidae showed a plug of hard material

protruding from the anus following experimental drying (Collins 1969). This suggests that, when

stressed for moisture, subterranean termites may be able to absorb extra moisture from their feces.

McManamy et al. (2008) suspected termites to be losing water through their feces, which would be

minimized if termites were excreting solid feces, as I observed. Dead termites exposed to low

relative humidities died from desiccation. A closer observation to those termites showed pellets of

feces protruding from their anus (Fig. 4-2). This proves that when stressed for moisture,

subterranean termites will try to retain as much moisture as possible from their feces.









Additionally, termites may have better access to moisture depending on the type of wood

they are feeding on. C. formosanus prefers hard wood over southern yellow SYP (Morales-Ramos

and Rojas 2001), however R. flavipes has not shown to have any preference. Feeding preference in

previous studies may have been affected by the way water is retained in wood. Though hardwoods

have tighter wood fibers than soft woods, hard woods contain vesicles which are able to retain more

isolated amounts of water, allowing termite access to more water (Manchester 1996). This fact

would further explain why both termite species were able to survive in NRO at lower wood moisture

contents than SYP.

Using the thermal kinetic method was an effective way to estimate termite mortality. Slopes

for all estimated survival curves using the kinetic equations decreased with an increase in wood

moisture content for all treatments. This shows that, with an increase in wood moisture content,

termites are able to survive longer. This occurs because, with an increase in wood moisture content,

there is an increase in water uptake as termites consume the wood. The more water they are able to

consume, the longer they are able to survive, decreasing the slope of the survival curve. At 30-35

WMC, termite life span is not limited by lack of moisture and termite colonies should be able to

sustain an aerial infestation without other water sources.

Another factor that affects termite survival is group size. Studies have shown that termites

exposed to dry conditions will live longer if tested in groups rather than individually (Collins 1969).

Different amounts of total insects were used per wood block per species; 101 R. flavipes and 110 C.

formosanus. This could explain why C. formosanus on NRO showed lower mortality rates than all

other treatments. In our experiments, because 101-110 termites were used in each individual block,

mortality in block with WMC at 30 and 35% may have been caused by cannibalism, mold (which

was formed in the wood), or other natural causes. In field settings, termites have the ability to move









from an area heavily infested with mold. However in my experiments, termites were confined to an

area where mold may have affected the longevity of the termites. Also, because both termite species

were field collected and age was not determined, that may have had an effect on termite mortality.

Wood moisture content is a very important factor in determining whether or not a termite

colony is able to survive in aerial infestations without soil contact or other water sources. Because

these infestations have no soil contact, it is imperative that that moisture content of the nest and

relative humidity within the nest are high enough to sustain the colonies. My research shows that

relative humidity needs to be <95% in order to sustain long term termite survival in wood with no

external source of moisture. Furthermore, termites seem to need free water in the wood to be able to

survive, requiring WMC to be >25%. Termites in wood with low WMC were probably not able to

obtain enough water to replace water they were losing through the cuticle due to low ambient

relative humidities. My experiments showed that when subterranean termites are stressed for

moisture, they are capable of conserving water by reabsorbing it from their feces, forming pellets.

However, once their BWC lowers to -35-45%, subterranean termites cannot survive and rapid

mortality occurs. For this reason, termites rely greatly on external sources of moisture such as leaky

roofs, faulty plumbing, or excess rain water to live long term in aerial infestations.





































Figure 4-1. Dry fecal pellets obtained from Reticulitermes flavipes.













































Figure 4-2. Desiccated Reticulitermesflavipes worker with dried fecal pellet protruding from
anus









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BIOGRAPHICAL SKETCH

Wai-Han Chan grew up in Miami, FL. She is the second of four daughters to Man Tat

Chan and Leyon Chan. She attended Gulliver Preparatory for high school. Upon graduation in

2003, she attended Purdue University where she obtained her Bachelor of Science in entomology

in 2007. While studying at Purdue University, she was chosen for the Pat Baker/pow Pest

Control, Inc. Memorial Scholarship in 2005 and the Oser Family Scholarship for entomology in

2007. She then earned her Master of Science in agricultural and life sciences in urban

entomology Summer of 2010 from the University of Florida under the mentorship of Dr. Philip

Gene Koehler and Dr. Roberto Manoel Pereira. While obtaining her research degree studying

the wood moisture effects on subterranean termites, she had the opportunity to give six

professional presentations at annual meetings including the Entomological Society of America,

Florida Pest Management Association, Southeastern pest management conference, and the

Florida Entomological Society where she won 2nd place in a student paper competition. In 2008,

she was elected president of Urban Entomological Society while being a member of the

Entomological Society of America and Florida Entomological Society. Upon graduating from

the University of Florida, she has completed six publications in Florida Pest Pro magazine,

University of Florida Featured Creatures, and Pests in and Around the home.

Wai-Han Chan currently lives in Melbourne, FL. She is currently working at TyraTech

Laboratories as a Product Development Staff Scientist. She plans and executes scientific studies

and interprets data for the development of insecticidal products made from plant essential oils in

applied entomology testing on urban and agricultural test species.





PAGE 1

1 SURVIVAL OF SUBTERRANEAN TERMITES (ISOPTERA: RHINOTERMITIDAE) ISOLATED IN WOOD OF VARIOUS MOISTURE CONTENTS By WAI HAN CHAN A THESIS PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF T HE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE UNIVERSITY OF FLORIDA 2010

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2 2010 Wai Han Chan

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3 To my family

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4 ACKNOWLEDGMENTS I thank my committee advisor D r. Philip Koehler and committee members Dr. Roberto Pereira and Dr. Nan Yao Su for their help and support throughout my graduate career. I would also like to thank my parents for guiding me through this with all their love and support

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5 TABLE OF CONTENTS page ACKNOWLEDGMENTS ................................ ................................ ................................ ............... 4 LIST OF TABLES ................................ ................................ ................................ ........................... 6 LIST OF FIGURES ................................ ................................ ................................ ......................... 7 ABSTRACT ................................ ................................ ................................ ................................ ..... 8 1 INTRODUCTION ................................ ................................ ................................ .................. 10 2 MATERIALS AND METHODS ................................ ................................ ........................... 18 Insects ................................ ................................ ................................ ................................ ..... 18 Wood Blocks for Termite Bioassay ................................ ................................ ........................ 18 Relative Humidity Experiment ................................ ................................ ............................... 19 Moi sture Experiments ................................ ................................ ................................ ............. 20 Termite Mortality at Various Wood Moisture Contents ................................ ......................... 21 Termite Body Water Content at Various Wood Moisture Contents ................................ ....... 21 Data Analysis ................................ ................................ ................................ .......................... 22 3 RESULTS ................................ ................................ ................................ ............................... 25 Relative Humidity ................................ ................................ ................................ ................... 25 Termite Mortality at Different Wood Moisture Content Levels ................................ ............ 25 Kinetics of Wood Moisture Effects on Termite Survival ................................ ....................... 28 Body Water Content ................................ ................................ ................................ ............... 30 4 DISCUSSION ................................ ................................ ................................ ......................... 46 LIST OF REFERENCES ................................ ................................ ................................ ............... 54 BIOGRAPHICAL SKETCH ................................ ................................ ................................ ......... 58

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6 LIST OF TABLES Table page 3 1 Effect of WMC, and estimated relative humi dity, on the mortality of Reticulitermes flavipes and Coptotermes formosanus ................................ ................................ ............... 43 3 2 Analysis of Variance results for termite mortality in 5 35% wood moisture content for different treatments ................................ ................................ ....................... 44 3 3 Coefficients of determination (r) used to determine N (kinetic orders) used for survivalship curves ................................ ................................ ................................ ............. 45

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7 LIST OF FIGURES Figure page 3 1 Percent relative humidity of space enclosed by wood blocks at various wood moisture contents ................................ ................................ ................................ .............. 32 3 2 Effect of wood moisture on mortality of Reticulitermes flavipes and Coptotermes formosanus ................................ ................................ ................................ ........................ 33 3 3 Survival curves of Reticulitermes flavipes placed in northern red oak moistened at various wood moisture contents ................................ ................................ ........................ 34 3 4 Survival curves of Reticulitermes flavipes placed in southern yellow pine moistened at various wood moisture contents ................................ ................................ ..................... 35 3 5 Survival curves of Coptotermes formosanus placed in northern red oak moistened at various wood moisture contents ................................ ................................ ........................ 36 3 6 Survival curves of Coptotermes formosanus placed in southern yellow pine moistened at various wood moisture contents ................................ ................................ .. 37 3 7 Death curve using the kinetic order model (n) ................................ ................................ ... 38 3 8 Death curves representing observed time ................................ ................................ .......... 39 3 9 Comparison of estimated (Fig. 3 7) and actual (Fig. 3 8) death curves ............................ 40 3 10 Effect of wood moisture on body water content of Reticulitermes flavipes and Coptotermes formosanus ................................ ................................ ................................ ... 41 3 11 Relationship between classes of percent body water content (BWC) and survi vorship of Reticulitermes flavipes and Coptotermes formosanus ................................ ................... 42 4 1 Dry fecal pellets obtained from Reticulitermes flavipes. ................................ ................... 52 4 2 Dessicated Reticulitermes flavipes worker with dried fecal pellet protruding from anus ................................ ................................ ................................ ................................ .... 53

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8 Abstract of Thesis Presented to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Master of Science SURVIVAL OF SUBTERRANEAN TERMITES (ISOPTERA: RHINOTERMITIDAE) ISOLATED IN WOOD OF VARIOUS MOISTURE CONTENTS By Wai Han Chan August 2010 Chair: Philip Gene Koehler Major: Entomology and Nematology S ubterranean termites nest in the soil, where they forage looking for a source of food, because they are dependent on its moisture for survival. However, some subterranean termite species are able to form aerial infestations, with no soil contact. This ex periment looked at the mortality rate and water weight loss in the Formosan subterranean termite, Coptotermes formosanus Shiraki, and the Eastern subterranean termite, Reticulitermes flavipes (Kollar), at various levels of wood moisture content (WMC). Ter mites were isolated in northern red oak (NRO) and southern yellow pine (SYP) wood blocks ranging from 5 to 35% WMC. Using destructive sampling, termite mortality and body water content were measured over a two week period. Separate bioassays were conduct ed to measure relative humidity in the air surrounding wood at 5 to 35% moisture content. Termite mortality decreased significantly with increase in WMC. Termites were unable to survive two weeks when placed in wood with <20% WMC. Termites in 30% WMC an d above is the point at which the wood is at or above full saturation, showed little to no mortality over the two week period in both NRO and SYP. Termite body water contents (BWC) decreased with a decrease in WMC. When BWC reached approximately 40%, rap id mortality occurred due to desiccatio n. Results showed that within air space

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9 resulted in significantly lower RH values. Termites stressed for moisture showed ability to conserve water by reabsorbing form their f eces, producing fecal pellets. Our results suggest that, while termites do require their ambient environment to be close to 100% relative humidity, they depend on free water in wood for long term survival when not in contact with other moisture sources as in aerial infestations.

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10 CHAPTER 1 INTRODUCTION Termites are structural pests that damage homes and buildings aesthetically and physically. Of the 2,300 species of termites that exist worldwide, about 50 are found in the United States, and about 18 are pests of structures (Su and Scheffrahn 1990). The Formosan subterranean termite, Coptotermes formosanus Shiraki is the most destructive and aggressive subterranean termite species found in the United States, whereas the Eastern subterranean termite, Ret iculitermes flavipes (Kollar), is one of the most widespread and important economic pests. All termites are social insects living in colonies that can consist of over a million individuals (Thorne 1998). In social colonies, each member completes certain tasks, dividing their work among specific members. Termite colonies have specific pheromones which are used to recognize nest mates of different castes and maintain separate existence from other colonies (Potter 2004). Castes are groups of individuals of distinct morphological characteristics that perform certain tasks within the colony. This division of labor results in an efficient, productive, and cooperative society that can continue for years (Thorne 1998). Termite castes consist mainly of reproduc tives, workers and soldiers. Colonies will also contain eggs and immatures. The subterranean life cycle begins as an egg which hatches into larvae These larvae later become workers which can then turn into separate castes, including nymphs, secondary r eproductives, or soldiers. Worker termites are considered the most important caste of the colony (Snyder 1948). They consist of both males and females and are blind. Workers consume wood and other cellulose based products, and are responsible to feeding and hydrating other caste members.

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11 They are responsible for tending the nursery, tunneling through soil, building mud tubes, and grooming the nest mates from mites and fungal infestations (Thorne 1998). These workers can Soldier termites are the defensi ve caste. They act aggressively toward invaders but sometimes can become passive and retreat from a confrontation (Thorne 1998). Their mandibles are very effective against certain predators such as ants, and certain species that have a chemical defense s ystem (Lee and Wood 1971). Colonies of C. formosanus consist of ~5 10% soldiers, whereas in the species R. flavipes soldiers are only ~1 2% of the colony (Haverty 1977, Haverty et al. 2000). Proportion of soldiers in a colony is dependent on food availabi lity, seasonal changes, worker nurturing capacity, and other factors (Su and La Fage 1987). Because soldier mandibles are developed for defense rather than for chewing food, soldiers are dependent on workers for nourishment. Although soldiers do not eat d irectly from the food source, they do forage and participate in food scouting (Potter 2004). Workers can also molt into apterous neotenics, which are wingless secondary reproductives. These are sexually mature and can reproduce if needed by the colony. Workers can also molt into nymphs. Nymphs are not sexually mature but have a small set of wing buds. These nymphs can lose their wing buds and molt back into a worker or can grow into an alate. Nymphs also have the ability to molt into brachypterous neo tenics, which are secondary reproductives with wing buds. Like the apterous neotenics, brachypterous neotenics are also sexually mature. The abdomen of these females becomes enlarged by physogastry, but they are less mature than the queen and king of the colony. Termites live underground in wood or in nests and will typically form tunnels in the soil in search of food. Underground, termites are able to acquire resources and are protected from predators, sunlight, air currents, and desiccation (Potter 20 04). In moist, loose sand, the

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12 subterranean termite will form tunnels by either pushing its head forward and then pressing the grains of soil from side to side with its head, body, and mandibles (Ebeling and Pence 1957) or by removing soil particles in co mpact soil. Su and Puche (2003) showed tunnel differences within different moisture gradients in R. flavipes and C. form osanus Termites tunnel more in sand with higher moisture. Termite species differ considerable in moisture requirements, some being ab le to live in wood above ground and in dry and hot environments while others are restricted to mesic or wetter regions (Collins 1969). Unlike other insects, which have a cuticle that forms an effective barrier to protect them from water evaporation, subte rranean termites are lightly sclerotized (Wigglesworth 1945). They are soft bodied insects and are prone to desiccation because of their inability to retain water very well; therefore, subterranean termites are more dependent on moisture for survival than other insects (Delaplane and La Fage 1989). Because subterranean termites lose water more readily from their integument rather than from respiration (Collins 1969), it is imperative that the moisture content of the ambient air is very high. If the sur rounding air is too dry, subterranean termites must leave that area in search of water; otherwise they will die (Collins 1969). If the relative humidity is near 100%, then the termites can survive and will continue to feed (Forschler 1999). In laboratory studies, Delaplane and LaFage (1989) found that subterranean termites showed a distinct preference for moister wood. To maintain this moisture, termites will bring moisture from their water source, such as soil, or deposit fecal material into the gallerie s (Collins 1969). Although subterranean termites typically obtain their moisture from the soil, they are capable of forming aerial nests, with no ground contact. If the moisture content of wood above ground level is high enough, subterranean termites can survive and multiply for an indefinite

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13 period with no soil contact (Potter 2004). For example, an intact colony of R. flavipes was found infesting a floating lake house and a houseboat in Kentucky, both completely suspended over water with no soil connec tion (Potter et al. 2000 and 2001). Aerial infestations of C. formosanus account for about 25% of structural infestations in Florida (Su and Scheffrahn 1986) and 50% of high rise buildings in Honolulu, Hawaii (Tamashiro et al. 1987). Because these two sp ecies are subterranean termites, they do require a source of moisture in order to survive. The moisture source can be from faulty plumbing, leaky roofs, condensation, or others. The relative humidity (RH) of the ambient air plays a large role in subterran ean termite survival. Other termite species, such as the drywood termite, Cryptotermes brevis (Walker) obtain the majority of its moisture from the ingested wood. The rectal pads in these species are more developed than those found in subterranean termi tes, allowing drywood termites to reabsorb most of the water from their feces (Collins 1969). When exposed to highly humid (Buxton 1932). Subterranean termites do not have rectal pads as specialized as drywood termites and depend on water from their food and environment. Other insects depend on the relative humidity of their environment, as well. The stored product beetles Sitophilus granary (Linnaeus) and S. ze amais (Motschulsky) both showed Tenebriodes mauritanicus (Linnaeus) however, had peak surviv al at RH between 65 and 80% (Kha n 1983). The cat flea, Ctenophelides felis (Bouche), is unable to live as a larva when RH is <45 or >95% (Bruce 1948). It has also been reported that in the desert fleas, Xenopsylla ramesis (Rothschild) and X. conformis (Wagner) humidity was the most important factor in determining survival. Low humiditi es cause a lower lifespan in these species, decreased larval activity, and soften

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14 cocoons. Subterranean termites are also dependent on RH and so have developed ways to sustain the RH in aerial infestations. Subterranean termites are able to utilize a nest ing building material known as carton, consisting of soil and masticated wood cemented together with saliva and excrement, which allow them to sustain aerial nests. This nest carton, which helps retain and conserve water, can be found along walls, attics, or roofs of termite infested buildings. These nests provide termites with moisture, food, and protection away from the soil with no ground contact. Flat roofs are ideal locations for termites to start an aerial colony because of water pools in low areas giving termites the moisture they need to sust ain a colony (Su et al. 1990 ). Just as soil tubes are used as a guide to find active subterranean termite colonies, tubes can also be formed by termites living in an aerial nest. Because termites found in a erial infestations have no soil contact, these tubes will contain little to no soil and the majority of it will consist of saliva, feces, and masticated wood (Blasingame 1987). In urban southeastern Florida, 25% of all C. formosanus infestations are aeri al. In Honolulu, Hawaii, 50% of all C. formosanus infestations are found in high rise buildings in Honolulu, Hawaii (Su and Scheffrahn 1987). These infestations can be started by alates which find a suitable area above ground to start a colony. Another w ay aerial infestations can be formed is from a colony that is initially underground. Sometimes, worker termites will forage into an above ground area and find it more suitable than the original nest. In this situation, workers will transport the king and queen to the location above ground and the connection to the ground is severed. The third way aerial infestations can form is from secondary reproductives that get separated from the rest of the colony and are cut off from the ground. These secondary re productives are sexually mature males and females. When they are separated from their

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15 original colony, sometimes from pesticide applications, male and female secondary reproductives start new colonies by budding (Blasingame 1987; Su and Scheffrahn 1987). Wood moisture content plays a large role in colony survival (Delaplane and La Fage 1989). When wood is of lower moisture content, termites will rely on external sources of moisture. These external moisture sources allow subterranean termites to be able to survive in wood with moisture content less than 14.8% (Blasingame 1988). However, too much water can be lethal to termites resulting in reduction of foraging populations or cause termites to enter a state of quiescence for several hours (Potter 2004). If the wood containing termites is above 15% WMC, the interstitial air space occupied by the termites is believed to be near 100% relative humidity (Sponsler and Appel 1990). This relative humidity level should protect termites from cuticular desiccatio n. Depending on termite species, wood moisture content can be the factor determining whether or not termites can sustain activity. The subterranean termites Odontotermes sp. are known to survive best at 96% RH but can restore their moisture from the tim ber on which they are feeding (Milner et al. 1997). In a choice test of wood moisture preference, R. flavipes did not prefer moist wood over dry wood as long as they had a source of external moisture. However, C. formosanus preferred moist wood over drie r wood, even if there was an available external source of moisture (Delaplane and La Fage 1989). According to Forschler (1998), wood moisture readings above 15% could indicate conditions suitable enough to support subterranean termite activity with no e xternal water sources. Wood moisture content below 15% would be unsuitable for the termites and would result in death by cuticular water loss, if no external source of moisture was available. Because termites can be found infesting wood at 15% moisture c ontent, subterranean termites do not

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16 necessarily have to be in fully saturated wood in order to survive and continue to feed on the structure. Wood fiber saturation does not occur until the wood reaches 25 30% moisture content (Carll and Highly 1999). At WMC above 25 30% wood is to be above saturation point and free water is available for termites. Little research exists on wood moisture content required in order sustain aerial infestations without an external source of moisture (Potter 2004). Delaplane and La Fage (1989) showed that damp wood in buildings are more prone to sustain aerial termite infestations and that bait blocks should have higher moisture content in order to increase its effectiveness. McManamy et al. (2008) conducted a wood moisture e xperiment and concluded that wood moisture content needed to be at least 30% WMC in order for eastern subterranean termites to be able to survive more than six months. McManamy et al. stain an aerial infestation, but did not measure whether relative humidity had an effect. These authors also measured body weight and concluded that termite mortality drastically increased once termites dropped 30 40% of their body weight. Sponsler and A ppel (1990) obtained similar results and found that water loss at time of death was an average of 50.5% for C. formosanus and 53.5% for R. flavipes. This study was conducted to determine effects of wood moisture content on subterranean termites. Two ty pes of wood, northern red oak (NRO) and southern yellow pine (SYP), and two species of termites, C. fomosanus and R. flavipes, were used. My first objective was to determine the air relative humidity of a void inside the wood moistened at 5 35% and compar e our results between a hard wood, NRO, and a soft wood, SYP. My second objective was to determine the wood moisture content (WMC) needed for long term subterranean termite survival with no soil contact. For this experiment, two species of termites and th e two species of wood

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17 were compared and determine the wood moisture death kinetics for subterranean termites. Using these results, I attempted to correlate air relative humidity and subterranean termite mortality. Finally, I wanted to determine what perc ent body water loss that resulted in subterranean termite death, and to compare the results for the two termite species living on the two wood species.

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18 CHAPTER 2 M ATERIALS AND METHODS Insects Three colonies of Reticulitermes flavipes (Koller), separat ed by more than 1.5 km, were collected in Gainesville, FL as described by Tucker (2004). Briefly, termites were field collected in 6 L plastic buckets inserted in the ground and covered with a lid accessible just above the soil surface. Two rolls of corr ugated cardboard were inserted into the buckets and were checked every 7 10 days. At those times, cardboard rolls infested with termites were replaced with new rolls and infested rolls were taken back to the lab for termite removal. The termites were rem oved from cardboard rolls and placed on moist corrugated cardboard sheets and reared at room temperature (~23C) in plastic containers (27.0 by 19.0 by 9.5 cm) for < 1 wk before used in experiments. Coptotermes formosanus Shiraki were collected from the Au dubon Zoo in New Orleans, LA. Termites were collected from monitoring traps stocked with cylindrical pieces of wood. Infested wood was placed in 20 L plastic buckets and driven to the University of Florida in Gainesville, FL. Termites in infested wood w ere then kept in 30.5 by 14.0 by 20.3 cm Sterilite Show Offs TM (Sterilite Corporation, Townsend, MA) containers with 0.95 L builder sand at approximately 10% moisture. Termites collected from different collection sites in the New Orleans Zoo were kept in separate containers and assumed to represent different colonies. These containers were stored in a lit room at room temperature (~23C) for later use. Wood Blocks for Termite Bioassay Northern red oak (NRO) boards and southern yellow pine (SYP) stakes were cut into small blocks (7 by 4 by 2 cm). Three adjoining holes (1 cm deep) were drilled using a 1.9 cm Forstner bit on one side of each block to produce a void (5 by 1.9 by 1 cm). A separate hole, (0.5

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19 cm diam. by 1 cm deep) was drilled into the cor ner of each block 0.5 cm from one of the corners to allow moisture replacement. Blocks were oven dried at 60C for 48 hr to remove any water from the wood. Each block was then weighed and dry weights were recorded. Relative Humidity Experiment To measu re the relative humidity in the air space surrounding moist wood, a device was constructed to simulate the use of a sling psychrometer. A small fan, connected to either a 9V battery or electrical outlet, was placed in the center of a five sided 15.3 by 8. 8 by 7.0 cm wooden box. This fan moved the air and created the drying effect necessary to measure a differential of temperature between a dry and a wet thermocouple. The wooden box was placed in a 1.9 L Glad Gladware container (The Clorox Company, Oaklan d, CA). To create the wooden boxes, NRO boards or SYP stakes were cut into five 11.3 by 3.5 by 2.0 cm (long blocks) and four 8.8 by 3.5 by 2.0 cm (short blocks). Each block had a 0.8 cm deep and 2.5 cm wide drilled void to create blocks that were simila r to the ones described above which were used in moisture experiments with termites. The void was drilled so that it started 1.5 2 cm from one end of the block and continued to the opposite end. Wood blocks were placed in a 60C oven for 48 hr to dry and were then moistened to 5 35% WMC by weight in increments of 5%. Wood blocks were then individually placed in 0.5 L Ziploc bags which were placed into 3.8 L Ziploc bags to prevent moisture loss. Blocks were kept in plastic bags for 36 48 hr to allow wa ter to completely disperse within the wood block before blocks were used for experiments. One hour before the blocks were used for experiments, water was added to replace any evaporated water and reestablish the desired WMC. Nine wood blocks with the sa me WMC were used to create a five sided 15.3 by 8.8 by 7.0 cm wooden box. Blocks were positioned so the side with the void faced inward in the box. The length of the box consisted of two stacked long blocks and the width consisted for two

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20 stacked short b locks. The bottom of the box was a single long block, with the void facing upward. Size 16 rubber bands were used to secure the box. Formed and secured wooden boxes were placed open side up in the center of the 1.9 L Glad Gladware plastic containers, w hich were sealed with their plastic lids. Wooden boxes and containers remained in at room temperature for the remainder of the experiment. Two 3 mm holes, 1.0 cm apart, were created at the center of the lid of the plastic container to allow two thermocou ples to be inserted through the top of the lid. One of the two thermocouples was wrapped with a wet sponge (~0.4 cm in diam. and 1.5 cm long). Size 4 cotton thread was used to secure the sponge on the thermocouple with several knots. An additional hole was placed on the lid of the container to insert a disposable pipette. Tap water was placed in the pipette and the pipette was placed so its tip was touching the sponge wrapped thermocouple to allow water replenishment of the sponge throughout the experim ent. Temperature measurements from both dry and wet thermocouples were taken with an EasyView Dual Input Thermometer (Extech Instruments Corp., Waltham, MA) at 15 min increments until temperatures stabilized (3 4 hr). Using an online psychometric calcul ator (http://www.sugartech.co.za/psychro/index.php), wet bulb and dry bulb temperature values were used to compute the relative humidity within the wooden boxes constructed with wood at different WMC levels. Moisture Experiments Immediately after the wood blocks were oven dried, water was added to the large void of the wood blocks to achieve 5, 10, 15, 20, 25, 30, and 35% (wt/wt) WMC. Moistened blocks were placed into individual 0.5 L Ziploc bags (S.C. Johnson and Son, Inc, Racine, WI), which were k ept in larger 3.8 L Ziploc bags to further help prevent moisture absorption or loss for 36

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21 48 hours until added water was evenly distributed throughout the wood block. After this period, blocks were weighed and moist weight was recorded. Termite Mortal ity at Various Wood Moisture Contents Four separate experiments were conducted with R. flavipes or C. formosanus placed on NRO or SYP using the following procedure Termites were separated and counted to result in 101 R. flavipes or 110 C. formosanus ind ividuals in order to mimic natural field ratios of 1soldier for 100 workers for R. flavipes and 10 soldiers for 100 workers for C. formosanus After separation from the colony, termites were chilled over ice for ~1 min to slow the termites so they could b e placed into the large void in the wood blocks without escaping. Clear acetate paper (7.0 by 4.0 cm) was hot glued over the opening of the block void to prevent termite escape and allow for an enclosed air space. The total weight of the blocks, includin g termites, acetate paper and hot glue, was recorded immediately after setup, and blocks were sealed back into individual 0.5 L Ziploc bags. Bags with wood blocks were then placed in larger 3.8 L Ziploc bags and kept at room temperature (~23C) for the duration of the experiment. Water was added every 7 days throughout the experiments to the moisture reservoirs to maintain initial weight and WMC. Termite Body Water Content at Various Wood Moisture Contents Blocks were prepared for destructive sampling at 1, 2, 3, 5, 7, 10, and 14 days. At each sampling day, total block weights were recorded to allow an estimation of the amount of WMC that had been retained. Termite mortality was determined and all the surviving termites were weighed together, then kil led and dried at 60C for 30 minutes, and re weighed to a 0.01 mg precision. Percent body water content (BWC) was calculated by subtracting the termite dry mass from the initial mass of the living termites and dividing the result by the initial mass. Mass loss was assumed to be entirely from water loss.

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22 Data A nalysis Relative humidity was not measured directly from the wood blocks in which the termites were placed, but rather from a separate wooden box comprised of blocks of wood moistened at WMC levels 5 10, 15, 20, 25, 30, or 35% as described before. It was assumed that relative humidity in the air space of the wood blocks to which termites were exposed was the same as that within the wooden boxes used to measure relative humidity for each WMC. D eath an d survivorship curves were calculated for WMC 5 25% because it was at those levels that significant mortality in termites was observed, whereas no significant termite mortality occurred at 30 and 35% WMC. For the relative humidity experiment, a completel y random design was conducted, using two wood species and 7 moisture levels. Each experimental unit consisted of a wooden box built with nine blocks of wood placed in a plastic Gladware container with a psychrometer. There were three replicates totaling 42 experimental units. One way ANOVA was conducted to determine effects of wood species on ambient wood relative humidity. Using statistical analytical software (SAS Institute 2007), termite mortality and percent BWC were arcsine square root transformed. A randomized complete block design was used for the termite mortality experiment. Three replications, from different termite colonies, with 100 termite workers plus soldiers for each termite species, C. formosanus and R. flavipes were used. Using destru ctive sampling, termites were exposed to seven different WMC levels (5, 10, 15, 20, 25, 30, and 35%) and mortality was observed every 1, 2, 3, 5, 7, 10, and 14 th day. At each sampling date for each WMC level, BWC for remaining living termites was measur ed Signi ficant differences were determined by graphically comparing standard error bars surrounding the means. One way analysis of variance was used to view significance differences between treatments and significance of day and moisture within treatments Separation of means

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23 was conducted using student Neumann keuls. A t test was conducted to measure significant differences between termite species and wood species. The fundamental empirical model used to estimate thermal mortality kinetics (Johnson et al. 2009, Wang et al. 2002, Alderton and Snell 1970) was used to determine the effect of WMC on termite mortality. Time was measured in days in order to estimate kinetics of termite mortality when insects were exposed to different levels of WMC. Termite survival at each WMC was plotted against exposure days, and survival curves were obtained for each WMC by linear regression. A WMC death curve was obtained by plotting the observed minimum time in days [Log (time) plotted on y axis] needed to obtain 100% t ermite mortality at the different WMC (x axis). Plots were obtained with kinetic orders 0, 0.5, 1, 1.5, and 2 following the methods described by Wang et al. (2002). Coefficients of determination (r 2 ) were calculated for each curve. Based on these coeffici en ts, the kinetic order equations that best fitted the survival data were selected. Parameters for kinetic orders 0 and 0.5 order equations were used to determine the effect of WMC on termite mortality. These kinetic orders were chosen based on the best c ombination of coefficient of determinations, compa ring all kinetic orders Slope and intercept from the regression equations were used to estimate the number of days to 100% mortality at different WMC for each of four treatments ( C. formosanus on SYP, C. formosanus on NRO, R. flavipes on SYP, and R. flavipes on NRO). Estimated time to cause 100% termite mortality according to these c urves were plotted against WMC Confidence intervals (95%) of slope and intercept were used to compare significance in the regressions lines for each treatment. To determine the accuracy of these estimates, another WMC death curve was obtained plotting WMC against the number of days (LOG) where 100% mortality was actually observed

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24 in the wood blocks. Confidence intervals (95 %) of slope and intercept were used to compare regressions lines obtained for each termite species/wood type combination. Differences between treatments were determined to be significant when confidence intervals did not overlap. Termite BWC for each of the 7 sampling dates (1, 2, 3, 5, 7, 10, and 14 d) was grap hed for each WMC level (5 10, 35%) Significant differences between BWC within treatments were determined by comparing standard error values; means with overlapping standard errors were conside red not significantly different and means with standard errors that did not overlap were considered significantly different. The percent BWC relationship to survival of the termites was measured by comparing the amount of termites that survived at certain BWC levels. Termite BWC was grouped into increments of 5%. Number of sur viving termites measured at specific BWC was graphed

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25 CHAPTER 3 RESULTS Relative Humidity Relative humidity (RH) in the air surrounding the wood box arenas showed a significant inc rease with an increase in WMC, reaching at plateau at 25% WMC for both NRO and SYP ( Fig. 3 1 ). There was no significant difference in RH between the two types of wood ( df =1; F= 0.10; P= above 98% were measured using the methods in these studies. Relative humidities for 25, 30 and 35% WMC were not significantly different on NRO or SYP. Termite Mortality at Different Wood Moisture Content Levels An increase in WMC yielded a decrease in ter mite mortality and increase in longevity for both R. flavipes and C. formosanus on NRO and SYP (Table 3 1 ). Termite mortality was significantly lower at higher WMC for all treatments (Table 3 2 ). Analysis of variance showed a significant relationship bet ween WMC and te rmite mortality, representing a significant decrease in mortality with increase in WMC. Neither species of termites were able to sustain activity Mortality of R flavipes o n NRO (Fig. 3 2 A) decreased with an increase in WMC yet a min imum of 25% WMC was required to yield some survival for the 14 day duration of the experiment. Rapid mortality occurred at 5 20% WMC within the first 7 days. By 7 d, all termites had died at up to 15% WMC. At 20% WMC, termite mortality was <10% within the first 5 d but increased to 72% by 7 d There was a slight increase in mortality at 25% WMC on 10 d but no significant differences, with less than 10% mortality, occurred between 25, 30, and 35% WMC a t 14 d for R. flavipes on NRO ( df =2 ; F= 2.00 ; P= 0.2160).

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26 Similarly, R. flavipes on SYP at 5 15% WMC (Fig. 3 2 B) were not able to survive longer than 7 d. At 20% WMC, rapid mortality occurred between 3 and 10 d with an average of 89% population mortality. R. flavipes mortality at 25% was not observed until after 7 d, averages for mortality were 0.3% at 7 d, 19.8 % at 10 d, and 43.2% at 14d. There were no significant differences in mortality between sampling days at 30% ( df =6 ; F= 0.35 ; P= 0.8953) or 35% (df=6; F=0.80; P=0.5890) WMC both with <3 mortality throughout the experiment. Coptotermes formosanus on NRO (Fig. 3 2 C) were not able to survive 5 15% WMC longer than 7 d. At 20% WMC, termites had <20% mortality for the first seven days but at 10 d, mortality was 100%. At 25% WMC, mortality slowly increased from 8. 5% at 7 d, to 11.2 % at 10d and rapidly increased to 41.2 % at 14 d. At 30 and 35% WMC, mortality maintained below 10% and showed no significant differences between sampling days (30 % WMC : df =6; F= 0. 43; P =0.8447; 35 % WMC : df =6; F= 0.77; P =0.6029) Coptotermes formosanus on SYP at 5 15% WMC (Fig. 3 2 D) were also unable to survive past 7 d. When termites were placed in SYP at 20% WMC, C. formosanus mortality began to increase rapidly from 5 to 10 d to an C. formosanus SYP were unable to survive 14 d. A t 25% WMC, the majority of mortality for C. formosanus was observed between 7 and 14 d with an average of 53% mortality. C. formosanus mortality on SYP at 25% WMC slowly i ncreased from 5.45% at 7 d to 10.9 1% at 10 d and increased to 61.82% at 14 d. There was significant mortality at 30% WMC and at 35% WMC at 14 d. However, there was no significant difference between 30 and 35% WMC mortalities at 14 d. One way analysis of variance showed significant differences in mortality between treatments ( df =3; F =4 .64; P =0.0033). Separation of means showed no significant differences between C. formosanus on SYP, R. flavipes on NRO, and R. flavipes on SYP. However, C.

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27 formosanus on N RO did have significantly lower mortality. There were no significant differences in mortality between the two termite species ( df =559; T = 1.47; P =0.1416) However mortality was significantly greater on SYP than NRO (df =559; T= 2.73; P=0.0064) For R. fl avipes at 5% WMC on NRO, the air RH was 54% which lead to 11% termite mortality by 1 d, 96% by 2 d, and 100% mortality by 3 d (Table 3 1) A 5% increase in WMC increased air RH to 75% at 10% WMC, which caused 98% termite mortality by 3 d and 100% by 5 d. At 15% WMC, air RH in wooden boxes had increased to 85% though it was not enough to allow termites to survive beyond 7 d. Mortality at 10 d for R. flavipes on NRO was 96% at 20% WMC and 1.3% at 25% WMC, though air RH for both test arenas were estimated t o be 95% (Table 3 1 ). Wood en NRO boxes with 30 and 35% WMC were measured to have 97% air RH and resulted in <2% termite mortality throughout the 2 week period (Table 3 1 ). 15% WMC were not significantly diff (Table 3 1) yet R. flavipes mortalities were significantly higher on SYP than on NRO. On SYP at 5% WMC, air RH was 52% and resulted in 78% mortality at 1 d and 100% mortality by 2 d. On SYP at 10% WMC, a ir RH in wood boxes was 73% which resulted in 25% R. flavipes mortality on 1 d, 64% at 2 d, 99% at 3 d, and 100% by 5 d. At 25% WMC, RH was 95% and <2% termite mortality was observed until 14 d when there was 43% mortality (Table 3 1 ). Coptotermes form osanus on NRO were not able to survive >10 d when the RH in the air surrounding the wood was assumed to be from the RH data at <20% WMC (Table 3 1) When the RH was 95% (20% WMC), C. formosanus mortality reached 100% at 10 d. Relative humidity was n ot significantly different between 20 and 25% WMC on NRO however, termite

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28 mortality was significantly lower at 25% WMC than 20% WMC At 98% RH (35% WMC on NRO), C. formosanus week period. For C. formosanus on SYP at 25% WMC, though the air RH in the arenas was measured in wooden boxes to be 95%, as was seen in NRO, 64% mortality occurred at 14 d (Table 3 1). At 20 and 25% WMC ( 97% RH ) mortality slowly increased to 13% by 14 d at 30% WMC, and to 18% at 14 d at 35% WM C. Kinetics of Wood Moisture Effects on Termite Survival Daily mortality rates for both termite species on both wood species at 30 and 35% WMC were not estimated because the estimated slopes for the mortality lines were not different from 0. Coefficient s of determinations (r 2 ) were determined to choose the best kinetic model for death curve estimates (Table 3 3). T he 0.5 order kine tic model (Wang et al. 2002) were used for R. flavipes on NRO (Fig. 3 3 ). At 5% WMC, mortality was estimated to begin to occ ur before 1 d and to reach 100% at 3 d, increasing at a rate of 45.4% per day. For 10% WMC, mortality was also estimated to begin before 1 d and to reach 100% at 4.6 d. A t 10% WMC live populations were estimated to decrease by 24.5% per day. When NRO ha d 15% WMC, mortality was estimated to begin at 1.1 d and to reach 100% at 9.5 d, with populations decreasing by 11.9% a day. At 20% WMC, mortality was estimated to begin at 4.4 d, and to reach 100% mortality by 10 d, with the population decreasing 16.4% p er day. For wood at 25% WMC, mortality was estimated to begin at 4.8 d. Populations were not estimated to reach 100% mortality until 52.3 d and were estimated to reach 19% mortality at 14 d increasing at a rate of at 2.1% per day. The 0 o rder kinetic model was used for R. flavipes on SYP and C. formosanus on NRO and SYP (Fig. 3 4, 3 5 and 3 6 ). Survivorship curves for all three treatments showed a decrease in their rate of mortality with an increase in WMC. For R. flavipes on SYP (Fig. 3 4 ), at 5% WM C, mortality was estimated to begin within the first day and reach 100% by 2 d. A t 15%

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29 WMC, mortality was estimated to begin before 1 d and to reach 100% by 7 d, with populations decreasing at 16.8% per day. With 20% WMC, mortality was estimated to begi n at 1.3 d and to reach 100% mortality by 12 d, decreasing 8.9% a day. At 25%, mortality was estimated to begin at 6.9 d and to reach 43% at 14 d. Using this death curve, it was estimated that R. flavipes at 25% WMC in S YP would reach 100% mortality by 2 3.3 d, with a mortality rate of 6.1 % per day. The wood moisture death kinetics for C. formosanus on NRO (Fig. 3 5 ) estimated that termites started dying before 1 d at 5% WMC and that mortality reached 100% mortality at 2 d, with populations decreasing 6 8.7% per day. At 10% WMC, mortality was estimated to begin before 1 d and reach 100% at 4.5 d. Populations of C. formosanus are estimated to decrease by 22.7% per day when placed on NRO at 10% WMC. At 15% WMC, C. formosanus were estimated to start dying at 1.9 d and to reach 100% mortality by 6.9 d, with populations decreasing by 19.7% per day. When C. formosanus termites were placed in NRO at 20% WMC, mortality was estimated to begin at 4.7 d and reach 100% mortality 10.6 d, with populations decreasing by 17.1% per day. At 25% WMC, mortality was estimated to begin at 6.1 d and reach 38% mortality at 14 d. Using this curve, 100% mortality was estimated to occur at 26.8 d. When C. formosanus was placed in SYP (Fig. 3 6 ) at 5% WMC, mortality was estima ted to begin before 1 d and 100% mortality was estimated to occur by 2.7 d. Populations were estimated to decrease at a rate of 29.8% per day when placed in 5% WMC. At 10% WMC, mortality was estimated to start before 1 d and reach 100% by 4.2 d, with pop ulations decreasing by 18.8% per day. At 15% WMC, mortality was estimated to start before 1 d and reach 100% at 6.8 d, with populations decreasing by 14.4% per day. For 20% WMC, the wood moisture death curve estimated mortality to begin at 1.2 d and reac h 100% by 11.0 d, with mortality rates of

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30 10.2% per day. At 25% WMC, mortality was estimated to begin at 4.8 d and reach 62% at 14 d. When the SYP was 25% WMC, mortality for C. formosanus was not estimated to reach 100% at 19.7 d. The WMC death curves pl otted using the slopes and intercepts from the model estimated survival curve s (Fig. 3 7 ) showed that there were no significant difference s in kinetics of WMC effects between R. flavipes and C. formosanus on NRO On SYP R. flavipes was estima ted to surv ive longer t han C. formosanus for each respective WMC The 95% CI for the wood moisture death curves generated with the observed data on time to reach 100% mortality (Fig. 3 8 ) showed that survival of both termite species on SYP and NRO were not significa ntly different. A comparative graph was made combining death curves from the model estimated survival curves and death curves generated with observed data (Fig. 3 9). No differences were observed between mortality estimates based on model estimated kinet ics and actual mortality data. Body Water C ontent Body water content (BWC) in termites started to decrease within the first day for all trea tments at all WMC levels (Fig. 3 10 ). Termites confined to blocks with higher WMC lost BWC at a slower rate. The average initial BWC for R. flavipes was 77%. On NRO, termite mortality for WMC 5 20% occurred rapidly after termite BWC decrea sed to an average of 40% (Fig. 9 A). In SYP at 5% WMC (Fig. 3 10 B), R. flavipes lowest BWC measurement was 39% at 1 d. On SYP, termites decreased to a BWC average of 38% when place d on wood at 5 20% WMC before reaching 10 0% mortality. There were only two remaining live termites after 3 d from one rep at 10% WMC which were measured to have reduced their BWC to 22% before reaching 100% mortality. At 30 and 35% W MC, termite BWC reached levels ~ 70% before recovering previously lost BWC.

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31 Coptotermes formosanus initial average BWC was 71%. The lowest BWC measurement of live term ites on NRO between 5, 10, and 15 % WMC was between 40 and 50% (Fig. 3 10 C) before all ter mites in the treatment died Termites reached an average of 39 % BWC before reaching 100% mortality at 5 20% WMC. Live t ermites on wood at 25 35% WMC did not replenish all the BWC lost and were measured to have an average BW C of 54% at 14 d. On SYP, C. formosanus BWC decreas ed on the first day for all WMC levels (Fig. 3 10 D). The lowest BWC recorded for termites at 5, 10, and 20% WMC was ~45%. Four remaining surviving termites subjected to 15% at 5 d were measured to have 31.63% C. formosanus BWC had decreased to an average of 41% for insects on wood at 5 20% WMC before reaching 100% mortality. After 14 d, termites in 25 35% WMC had an average of 52% BWC. Standard error differentiation showed these values to be significa ntly lower than initial BWC. There was a positive correlation between termite BWC and survival for all treatments (Fig. 11 ). Survival of R. flavipes on NRO (Fig. 3 11 A) was highest when BWC was 51 55%. For this treatment, there was a n increase of 49% i n survival between 36 40% and 41 45% BWC. For R. flavipes on SYP, (Fig. 3 11 B), termite survival increased more gradually with increase in BWC, however, <50% of termites were able to survive at BWC lower than 41 45%. Similarly, with C. formosanus on NRO ( Fig. 3 11 C), an average of 44% of termites were able to survive when BWC was 41 45% or less. C. formosanus survival on NRO was highest when BWC was 51 55% and greater. For C. formosanus on SY P (Fig. 3 11 D), survival also showed a gradual increase with an increase in termite BWC. C. fomosanus survival on SYP was >50% when BWC was 36 40% and 68% survival was observed in termites with 41 45% BWC. When BWC was above 50%, termite mortality became minimal (89% survival or higher).

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32 Figure 3 1 Percent relative humidity (RH) of space enclosed by Southern Yellow Pine (SYP) and Northern Red Oak (NRO) at 5, 10, 15, 20, 25, 30, and 35% wood moisture content (WMC).

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33 A B C D Figure 3 2 Effect of wood moisture on mortality of Reticul itermes flavipes and Coptotermes formosanus placed in southern yellow pine (SYP) and northern red oak (NRO) wood blocks with 5, 10, 15, 20, 25, 30 and 35% moisture contents: A) R. flavipes on NRO; B) R. flavipes on SYP; C) C. formosanus on NRO; D) C. formo sanus on SYP. Percent Mortality Day Day Percent Mortality

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34 Figure 3 3 Survival curves of Reticulitermes flavipes placed in northern red oak moistened at 5, 10, 15, 20, and 25% moisture contents.

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35 Figure 3 4 Survival curves of Reticulitermes flavipes placed in southern yellow pine mois tened at 5, 10, 15, 20, and 25% moisture contents.

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36 Figure 3 5 Survival curves of Coptotermes formosanus placed in northern red oak moistened at 5, 10, 15, 20, and 25% moisture contents.

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37 Figure 3 6 Survival curves of Coptotermes formosanus placed in southern yellow pine moistened at 5, 10, 15, 20, and 25% moisture contents.

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38 A B C D Figure 3 7 D eath curve using the kinetic order model (n) representing LOG of days to reach 100% mortality of Reticulitermes flavipes and Copto termes formosanus placed in southern yellow pine (SYP) and northern red oak (NRO) blocks with 5, 10, 15, 20, and 25% wood moisture contents (WMC), dotted lines represent 95% confidenc e interval of regression line: A ) R. flavipes on NRO, B ) R. flavipes on S YP, C ) C. formosanus on NRO, D ) C. formosanus on SYP. Treatment time (Log[days]) Percent Wood Moisture Content

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39 A B C D Figure 3 8 D eath curves representing observed time (Log[days]) and wood moisture content (WMC) combinations in Reticulitermes flavipes and Coptotermes formosanus yielding 100% mortality, dotted lines represent 95% confidence interval of regression line: A) R. flavipes on NRO; B) R. flavipes on SYP; C) C. formosanus on NRO; D) C. formosanus on SYP. Treatment time (Log[days]) Percent Wood Moisture Content

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40 A B C D Figure 3 9 Comparison of estimated (Fig. 3 7) and actual (Fig. 3 8) death curves representing time (Log[days]) and wood moisture content (WMC) combinations in Reticulitermes flavipes and Coptotermes formosanus yielding 100% mortality, dotted lines represent 95% confidence interval of regression line A) R. flavipes on NRO; B) R. flavipes on SYP; C) C. formosanus on NRO; D) C. formosanus on SYP Treatment time (Log[d ays]) Percent Wood Moisture Content

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41 A B C D Figure 3 10 Effect of wood moisture on body water content of Reticulitermes flavipes and Coptotermes formosanus pla ced in southern yellow pine (SYP) and northern red oak (NRO) wood blocks with 5, 10, 15, 20, 25, 30 and 35% moisture contents: A ) R. flavipes on NRO, B ) R. flavipes on SYP, C ) C. formosanus on NRO, D ) C. formosanus on SYP. Percent Body Water Content Day

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42 A B C D Figur e 3 11 Relationship between classes of percent body water content (BWC) and survivorship of Reticulitermes flavipes and Coptotermes formosanus confined to wood at different WMC. A) R. flavipes on NRO; B) R. flavipes on SYP; C) C. formosanus on NRO; D) C formosanus on SYP. Percent Body Water Content Proportion of Survivorship

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43 Table 3 1 Effect of WMC, and estimated relative humidity, on the mortality of Reticulitermes flavipes and Coptotermes formosanus over 14 d in northern red oak (NRO) and southern yellow pine (SYP) Reticulitermes flavipes Copt otermes formosanus Days Days wood species % WMC Estimated % Relative Humidity 1 2 3 5 7 10 14 1 2 3 5 7 10 14 NRO 5 54 11 96 100 31 100 10 75 2 45 98 100 08 44 85 100 15 86 0 23 23 64 98 100 05 09 11 68 100 20 95 0 06 02 03 72 96 100 02 04 1 19 14 100 25 96 0 03 04 02 08 19 09 03 02 03 08 08 11 41 30 95 0 0 02 0 01 01 04 01 04 02 03 05 07 04 35 98 0 0 02 0 01 02 0 01 01 0 03 02 02 05 SYP 5 52 78 100 4 99 100 10 73 25 64 99 100 15 78 96 100 15 86 02 08 58 77 99 100 01 16 63 72 99 100 20 90 01 03 06 3 6 95 100 0 05 34 27 49 98 100 25 95 0 01 02 01 0 2 43 0 04 07 05 11 31 64 30 97 01 01 01 0 0 0 0 0 06 06 08 08 13 13 35 97 0 01 01 01 0 02 01 0 03 06 07 05 11 18

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44 Table 3 2. Analysis of Variance results for termite mortality in 5 35% wood moisture content Treatment DF MS F value P value R. flavipes on NRO 6 6.51 33.45 <0.0001 R. flavipes on SYP 6 7.77 50.3 9 <0.0001 C. formosanus on NRO 6 1.79 12.96 <0.0001 C. formosanus on SYP 6 6.38 32.82 <0.0001

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45 Table 3 3 Coefficients of determination (r ) used to determine N (kinetic orders) used for survivorship curves to estimate termites mortality at any given wood moisture content. Highlighted values represent those used to develop death curves Wood Moisture Content Treatment N 5 10 15 20 25 30 35 R. flavipes NRO 0 1 0.8743 0.9656 0.8011 0.8718 0.2763 1 0.5 1 0.979 0.9505 0.8204 0.8306 0.3027 1 1 1 0.933 0.8067 0.8357 0.8521 0.3312 1 1.5 1 0.8066 0.7035 0.8413 1 0.3621 1 2 1 0.7755 0.6854 0.842 0.8379 0.3951 1 R. flavipes SYP 0 0.7667 0.6665 0.8509 0.8828 0.9791 0.843 0.9992 0.5 0.8089 0.7984 0.9388 0.8193 0.9808 0.8398 0.9998 1 0. 8909 0.9201 0.9646 0.7151 0.7158 0.8366 1 1.5 0.946 0.9469 0.8972 0.6542 0.1016 0.8335 0.9998 2 0.9608 0.9493 0.8571 0.6432 0.0342 0.8303 0.999 C. formosanus NRO 0 0.7829 0.7753 0.9122 0.8678 0.9944 0.9301 0.1263 0.5 0.8488 0.8167 0.9344 0 .9152 0.9872 0.9265 0.1241 1 0.9684 0.8701 0.9010 0.9665 0.9778 0.9229 0.1219 1.5 0.9970 0.8872 0.8549 0.9846 0.9666 0.9192 0.1197 2 0.9777 0.8893 0.8444 0.9867 0.9543 0.9155 0.1175 C. formosanus SYP 0 1.0000 0.9987 0.9096 0.9376 0.9992 1. 0000 0.2500 0.5 1.0000 0.9728 0.9629 0.9520 1.0000 1.0000 0.2421 1 0.7500 0.8426 0.8755 0.8980 0.9995 1.0000 0.2344 1.5 0.7500 0.8706 0.6943 0.8371 0.9975 0.8176 0.2267 2 0.7500 0.8842 0.6346 0.9268 0.7784 0.8176 0.2192

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46 C HAPTER 4 DISCUSSION Tho ugh studies have been conducted demonstrating that wood moisture content needs to be high in order for termites to be able to sustain activity, the exact wood moisture content needed, with out external moisture sources was unknown. My study aimed at determ ining the wood moisture content effects on termite survival and body water content. Previous studies have shown that termites prefer moist wood over dry wood (Delaplane and La Fage 1989), even if given an additional source of water. However, these author s did not examine the effects the lower moisture wood had on subterranean termites. WMC affects the relative humidity in the ambient environment around the block of wood. Though it has been noted that termites are able to survive in wood as long as the moisture content is >16% (Forschler 1999), my data suggests otherwise. It has been suggested that when wood has >16% WMC, the relative humidity in the interstitial spaces is at or near 100% and, therefore, termites should not die from water loss thr ough the cuticle However, my results show that even in cases where the wood is >16% WMC, and even at 25% WMC termites still lose body water. I estimated the relative humidity of the air within 15% WMC blocks to be <90%, which explains why termi tes were una ble to survive for a week at 15% WMC or below. At 20% WMC, neither R. flavipes nor C. formosanus were able to survive longer than 10 days, where relative humidity was lower than that found at 25% WMC. This suggests that relative humidity needs to be >95% in order for termites to sustain activity as shown before. McManamy et al. (2008) found that WMC had to be >24% in order to sustain subterranean termite aerial infestations with no soil contact. However, the authors assumed that the relative humidity wi thin the wood was at or near

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47 than previously expected when the wood has >24% WMC, therefore, termites can lose cuticular body water. Relative humidity result s peaked to 98% relative humidity at 25% WMC and there was no significant difference between the relative humidity in the area surrounding wood boxes moistened to 30 and 35% WMC. I was unable to obtain a relative humidity value of 100% when wood was moist ened at a point higher than saturation. This may have occurred because tap water was used to wet the wood blocks. Wood satur ation with distilled water may have given a close relative humidity of 100% (Almeida and Hernandez 2005). Tap water may have cont ained dis solved salts which prevented measurements of ambient environment from reaching 100%. Studies have shown that tap water contains higher fluoride content than distilled water (Lalumandier et al. 2000). Sponsler and Appel (1990) exposed termites to conditions of very low relative humidity (0 2%) Our results show that termites were able to lose more BWC throughout time than previous studies before death occurred. The longer survival observed in my studies may have been a result of moisture from wood maintaining a higher ambient relative humidity for a longer period of time. A higher relative humidity would have allowed termites to retain their BWC for a longer time, maintain ing BWC above a critical level at which death occurs. Relative humidit y and the temperature to which termites are exposed cause changes in termite BWC which determine survivorship of the insect. Previous studies have also shown that R. flavipes will live 5.1 hr at 30C and 0 2% RH, with 53.5 % BWC at time of death, and C. fo rmosanus will live 7.1 hr, with 50.5 % BWC at time of death (Sponsler and Appel 1990). Collins (1969) showed that R. flavipes workers were able to l ive 3.0 6.2 hr and drop to 56.5 % BWC before 100% mortality in 34 35C and 0 4% relative humidity conditions.

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48 Although a surviving R. flavipes termite on SYP at 10% WMC was measured as having 21.7% BWC, termites began to die rapidly at approximately 35 49% body weight loss in my experiment, which is lower than the BWC observed at time of death by Collins (1969) and Sponsler and Appel (1990). McManamy et al. (2008) found similar results where 20% WMC caused termites to drop approximately 40% body weight which led to rapid mortality. In addition to ambient relative humidity, temperature also has an effect on te rmite mortality. Our experiment were maintained at room temperature (23C), contrary to those in previous mentioned experiments (Collins 1969 ; Sponsler and Appel 1990) where termites were held in termite mortality, allowing termites to survive longer. Unlike Collins (1969) who exposed termites to hig h temperatures (30C) and low relative humidities (0 2%), my experiment maintained termites at room temperature and relative humidities their BWC over a longer t ime before reaching 100% mortality. My experiment showed that even though BWC decreased, termites were able to survive longer than 24 hr with ambient relative humidity <60%. As shown by Mellanby (1939), termites were able to lower their BWC and still be able to sustain activity. My results showed that even though there was no significant difference between termite mortality at 30, and 35% WMC, mortality was significantly greater at 25% WMC for termites in SYP McManamy et al. (2008) reported that wood moisture content needed to be at least 30% in order for subterranean termites to sustain an aerial infestation. Wood fiber saturation does not occur until the wood reaches 25 30% moisture content (Carll and Highly 1999); any moisture content above that is free water in the wood. This suggests that termite mortality is not only dependent on

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49 ambient relat ive humidity but also require access to free water. Both R. flavipes and C. formosanus on NRO showed no significant difference in mortalities between 25 a nd 30% WMC at the end of the experimental period. Insects are able to draw upon their hemolymph as a liquid reserve without impairing their locomotion or respiration (Mellanby 1939). Termites are able to do this and have been seen walking after substant ial hemolymphic fluid loss following flattening of the abdomen (Collins 1969). By flattening their abdomens, termites were able to sustain more body water weight loss while being exposed to lower relative humidities. This explains why termites in my expe riments were able to live for days at lower relative humidities Another source of moisture when stressed for water may be the feces of the subterranean termite. Subterranean termite feces is usually a muddy texture, with no solid shape, unlike drywood t ermite feces. Nakayama et al (2004) suggested that when subterranean termites become stressed for moisture, they will excrete liquid feces to increase ambient relative humidity. Contrary to these reports, solid pellets of feces were found in blocks with WMC <25% in my experiments (Fig. 4 1) Similar observations were made where species of Kalotermitidae showed a plug of hard material protruding from the anus following experimental drying (Collins 1969). This suggests that, when stressed for moisture, su bterranean termites may be able to absorb extra moisture from their feces. McManamy et al (2008) suspected termites to be losing water through their feces, which would be minimized if termites were excreting solid feces, as I observed. Dead termites expo sed to low relative humidities died from desiccation A closer observation to those termites showed pellets of feces protruding from their anus (Fig. 4 2). This proves that when stressed for moisture, subterranean termites will try to retain as much mois ture as possible from their feces.

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50 Additionally, termites may have better access to moisture depending on the type of wood they are feeding on. C. formosanus prefers hard wood over southern yellow SYP (Morales Ramos and Rojas 2001), however R. flavipes ha s not shown to have any preference. Feeding preference in previous studies may have been affected by the way water is retained in wood. Though hardwoods have tighter wood fibers than soft woods, hard woods contain vesicles which are able to retain more i solated amounts of water, allowing termite access to more water (Manchester 1996) This fact would further explain why both termite species were able to survive in NRO at lower wood moisture contents than SYP. Using the thermal kinetic method was an ef fective way to estimate termite mortality. Slopes for all estimated survival curves using the kinetic equations decreased with an increase in wood moisture content for all treatments. This shows that, with an increase in wood moisture content, termites ar e able to survive longer. This occurs because, with an increase in wood moisture content, there is an increase in water uptake as termites consume the wood. The more water they are able to consume, the longer they are able to survive, decreasing the slop e of the survival curve. At 30 35 WMC, termite life span is not limited by lack of moisture and termite colonies should be able to sustain an aerial infestation without other water sources. Another factor that affects termite survival is group size. Stu dies have shown that termites exposed to dry conditions will live longer if tested in groups rather than individually (Collins 1969) Different amounts of total insects were used per wood block per species; 101 R. flavipes and 110 C. formosanus This cou ld explain why C. formosanus on NRO showed lower mortality rates than all other treatments In our experiments, because 101 110 termites were used in each individual block, mortality in block with WMC at 30 and 35% may have b een caused by cannibalism, mol d ( which was formed in the wood ) or other natural causes. In field settings, termites have the ability to move

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51 from an area heavily infested with mold. However in my experiments, termites were confined to an area where mold may have affected the longevi ty of the termites. Also, because both termite species were field collected and age was not determined, that may have had an effect on termite mortality. Wood moisture content is a very important factor in determining whether or not a termite colony is able to survive in aerial infestations without soil contact or other water sources. Because these infestations have no soil contact, it is imperative that that moisture content of the nest and relative humidity within the nest are high enough to sustain t he colonies. My research shows that relative humidity needs to be <95% in order to sustain long term termite survival in wood with no external source of moisture. Furthermore, termites seem to need free water in the wood to be able to survive requiring WMC to be >25%. Termites in wood with low WMC were probably not able to obtain enough water to replace water they were losing through the cutic le due to low ambient relative humidities. My experiments showed that when subterranean termites are stressed f or moisture, they are capable of conserving water by reabsorbing it from their feces, forming pellets. However, once their BWC lowers to ~35 45%, subterranean termi tes cannot survive and rapid mortality occurs For this reason, termites rely greatly on e xternal sources of moisture such as leaky roofs, faulty plumbing, or excess rain water to live long term in aerial infestations

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52 Figure 4 1. Dry fecal pellets obtained from Reticulitermes flavipes.

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53 Fig ure 4 2. Desiccated Reticulitermes flavipes w orker with dried fecal pellet protruding from anus

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54 LIST OF REFERENCES Alderton, G., and N. Snell. 1970. Chemical states of bacterial spores: heat resistance and its kinetics at intermediate water activity. Appl. Microbiol. 19: 565 572. Almeida, G., and R. E. Hernandez. 2006. Changes in physical properties of tropical and temperate hardwoods below and above the fiber saturation point. Wood. Sci. Technol. 40: 599 613. Blasingame, W. 1988. Detecting above ground infestations of subterranean termites. Pest Manage. 7: 19 21. Bruce W. N. 1948. Studies on the biological requirements of the cat flea. Ann. Entomol. Soc. Am. 41: 346 352. Buxton, P. A. 1932. The relation of the adult Rhodnius prolixus (Reduviidae, Rhynchota) to atmospheric humidity. Parasitol. 24: 429 439. Carll, C. G., and T. L. Highley. 1999. Decay of wood and wood based products above ground buildings. J. Testing Evaluat. 27: 150 158. Chambers, D. M., P. A. Zungoli, and H.S. Hill. 1988. Distribution and habits of the Formosan subterranean termite in South Carolina. J. Econ. Entomol. 81:1611 1619. Collins, M. S. 1969. Water relations in termites. In: Krishna, K, and F.M. Weesner [eds.] Biology of termites. Academic Press, New York, NY. 433 435 Delaplane, K. S., and J. P. La Fage 1989. Pre ference of moist wood by the Formosan subterranean termite (Isoptera: Rhinotermitidae). Ann. Entomol. Soc. Amer. 62: 1274 1284. Ebeling, W. and R.J. Pence. 1957. Relation of particle size to the penetration of subterranean termites through barriers of san d or cinders. J. Econ. Entomol. 50: 690 692. Forbes, C. F., and W. Ebeling. 1987. Update: use of heat for elimination of structural pests. The IPM Practitioner. 9: 1 6. Forschler, B. T. 1999. Subterranean Termite Biology in Relation to Prevention and Re moval of Structural Infestations. National Pest Control Association research report on subterranean termites. Dunn Loring, VA: National Pest Control Association Publications. 31 50. Haverty, M. I. 1977. The proportion of soldiers in termite colonies: a li st and a bibliography (Isoptera). Sociobio. 2: 199 216.

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55 Haverty, M. I., G. M. Getty, K. A. Copren, and V. R. Lewis. 2000. Size and dispersion of colonies of Reticulitermes spp. (Isoptera: Rhinotermitidae) in a wild land and a residential location in north ern California. Environ. Entomol., 29: 241 251. Johnson, J. A., S. Wang, and J. Tang. 2003. Thermal death kinetics of fifth instar Plodia interpuctella (Lepidoptera: Pyralidae). J. Econ. Entomol. 96: 519 524. Kettering, J. D., J. A. Stephens, C. A. Munoz Viveros, and W. P. Naylor. 2002. Reducing bacterial counts in dental unit waterlines: tap water vs. distilled water. J. Cont. Dent. Prac. 3: 1 11. Khan, M. A. 1983 Effect of relative humidity on adults of 10 different species of stored product beetles. Z. Angew. Entomol. 95: 217 227. Krasnov, B. R., I. S. Khokhlova, L. J. Fielden, and N. V. Burdelova. DATE. Effect of air temperature and humidity on the survival of pre imaginal stages of two flea species (Siphonaptera: Pulicidae). J. Med. Entomol. 38: 6 29 637. Lalumandier J. A., and L. W. Ayers. 2000. Fluoride and bacterial content of bottled water vs. tap water. Amer. Med. Assoc. 9: 246 250. Lee, K. E., and T. G. Wood. 1971. Termites and soils. Academic Press, New York, NY. Manchester, S. R. 1996 Collecting fossil plants in Florida. Pap Fla Palentol. 8: 1 8. McManamy, K., P. G. Koehler, D. D. Branscome, and R. M. Pereira. 2008. Wood moisture content affects the survival of eastern subterranean termites (Isoptera: Rhinotermitidae), under saturate d relative h umidity conditions. Sociobio 52: 1 12. Mellanby, K. 1939. The functions of insect blood. Bio. Rev. 14: 243 260. Milner, R. J., J. A. Staples, and G. G. Lutton. 1997. The effect of humidity on germination and infection of termites by the hy phomycete, Metarhizium anisopliae J. Invert. Pathol. 69: 64 69. Minnick, D. R., S. H. Kerr, and R. C. Wilkinson. 1973. Humidity behavior of the drywood termite Crytotermes brevis Environ. Entomol. 2: 597 601. Morales, J. A., and M. G. Rojas. 2001. Nu tritional ecology of the Formosan subterranean termite (Isoptera: Rhinotermitidae): Feeding response to commercial wood species. J. Econ. Entomol. 94: 516 523. Nakayama, T., T. Yoshimura, and Y. Imamura. 2004. The optimum temperature humidity combination for the feeding activities of Japanese subterranean termites. J. Wood Sci 50: 530 534.

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56 Pereira, R. M., P. G. Koehler, M. Pfiester, and W. Walker. Lethal effects of heat and use of localized heat treatment for control of bed bug infestations. J. Econ. Ent omol. 102: 1182 1188. Potter, M. F., and R. T. Bessin 2000. Termites and public attitudes. Pest Control Technol. 29(2): 39 43, 43, 48, 50, 52, 56. Potter, M. F., C Asberry, and A. J. Grieco 2001. Houseboat horror. Pest Control Technol. 29(8): 68, 70, 72. Potter.M. F. 2004. Termites. In: S.A. Hedges [ed.], Handbook of pest control: the behavior, life history and control of household pests. Gie Media Inc., Richfield, OH. 217 361 Snyder, T. E. 1948. Our enemy the termite. [rev. ed.] Comstock Publ. Co ., Inc., Ithaca, NY. Sponsler, R. C. and A. G. Appel 1990. Aspects of the water relations of the Formosan and eastern subterranean termites (Isoptera: Rhinotermitidae). Environm. Entomol. 19: 15 20. Su, N. Y., and J. P. La Fage. 1987. Effects of soldier proportion on the wood consumption rate of the Formosan subterranean termite (Isoptera: Rhinotermitidae). Sociobiology 13: 145 151 Su, N. Y., and H. Puche. 2003. Tunneling activity of subterranean termites ( Isoptera: Rhinotermitidae) in sand and moisture gradients. J. Econ. Entomol. 96: 89 93. Su, N. Y., and R. H. Scheffrahn. 1986. The Formosan subterranean termite, Coptotermes formosanus (Isoptera: Rhinotermitidae), in the United States 1907 1985 pp 31 38 In : Proceedings of the National Conference on Urban Entomolog y, College Park, MD. Su, N. Y., and R. H. Scheffrahn. 1990. Economically important termites in the United States and their control. Sociobiology. 17: 77 94. Su, N. Y., R. H. Scheffrahn, and P. M. Ban. 1990. High rise termites. Pest Managem. 9: 22 24. Tamashiro, M., J. R. Yates, and R.H. Ebesu 1987. The Formosan subterranean termite in Hawaii: problems and control pp. 15 22 In : M. Tamashiro and N.Y. Su, Eds., Biology and control of the Formos an Subterrnean termite. College of Tropical Agriculture and Human Resources, University of Hawaii, Honolulu, HI Thorne, B. 1998. Biology of the subterranean termites of the genus Reticulitermes National Pest Control Association research report on subter ranean termites. Dunn Loring, VA: National Pest Control Association Publications. 1 29

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57 Tucker, C. L., P. G. Koehler, and F. M. Oi. 2004. Influence of soil compaction on tunnel network construction by the eastern subterranean termite (Isoptera: Rhinotermit idae). J. Econ. Entomol. 97: 89 94. Wang, S., J. Tang, J. A. Johnson, and J. D. Hansen. Thermal death kinetics of fifth instar Amyelois transitella (Walker) (Lepidoptera: Pyralidae). J. Stored Prod. Res. 38: 427 440. Wigglesworth, V. B. 1945. Transpirat ion through the cuticle of insects. J. Exptl. Biol. 21: 97 114.

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58 BIOGRAPHICAL SKETCH Wai Han Chan grew up in Miami, FL. She is the second of four daughters to Man Tat Chan and Leyon Chan. She attended Gulliver Preparatory for high school. Upon graduation in 2003, she attended Purdue Uni versity where she obtained her B achelor of Science in e ntomology in 2007. While studying at Purdue University, she was chosen for the Pat Baker/ pow Pest Control, Inc. Memorial Scholarship in 2005 a nd the Oser Family Scholarship for e ntomology in 2007. She then earned her Master of Science in a gricultural and l ife s ciences in u rban e ntomology Summer of 2010 from the University of Florida under the mentorship of Dr. Philip Gene Koehler and Dr. Robe rto Manoel Pereira. While obtaining her research degree studying the wood moisture effects on subterranean termites, she had the opportunity to give six professional presentations at annual meetings including the Entomological Society of America, Florida Pest Management Asso ciation, Southeastern pest management conference and the Florida Entomological Society where she won 2 nd place in a student paper competition In 2008, she was elected president of Urban Entomological Society while being a member of t he Entomological Society of America and Florida Entomological Society. Upon graduating from the University of Florida, she has completed six publications in Florida Pest Pro magazine, University of Florida Featured Creatures, and Pests in and Around the h ome. Wai Han Chan currently lives in Melbourne, FL. She is currently working at TyraTech Laboratories as a Product D evelopment Staff Scientist. She plans and executes scientific studies and interpr e ts data for the development of insecticidal products made from plant essential oils in applied entomology testing on urban and agricultural test species.