DIET and PREY PREFERENCE of GIANT GARTERSNAKES (THAMNOPHIS GIGAS) in the SACRAMENTO VALLEY of CALIFORNIA a University Thesis

DIET AND PREY PREFERENCE OF GIANT GARTERSNAKES (THAMNOPHIS GIGAS) IN THE SACRAMENTO VALLEY OF CALIFORNIA

A University Thesis Presented to the Faculty of California State University, East Bay

In Partial Fulfillment of the Requirements for the Degree Master of Science in Biological Sciences

By Julia Samiye Martins Ersan December, 2015

Abstract

The introduction of exotic species into an environment can introduce great change in the trophic dynamics of native species. This is of even greater concern if the native species are threatened or endangered. The giant gartersnake (Thamnophis gigas), an endemic predator of the Central Valley of California, is listed as threatened, its decline because of the conversion of its once vast wetland habitat to agriculture. Another anthropogenic factor contributing to this snake’s changing ecology and potentially its decline is the introduction of non-native prey species into their habitats. These introductions have resulted in a prey community that is almost completely composed of exotics. Exotic prey can be detrimental to predators, potentially causing injury, parasitic infection, and intraguild predation. A first step toward understanding the effect(s) of exotic prey on giant gartersnakes is to determine whether and the degree to which these snakes have an affinity toward and will actually consume these species. I examined dietary choices in the giant gartersnake using laboratory and field studies to determine whether these snakes are selecting their prey or if they are simply consuming what is abundant. The laboratory component of my investigation was comprised of two sets of behavioral trials in which naïve neonates of wild-caught adult females from three different areas served as the focal animals. I examined: 1) neonate prey preference in response to olfactory cues of prepared prey extracts and 2) what neonates actually consumed when provided with a simultaneous choice of different live prey items. The field studies involved an analysis of prey selection where I: 1) quantified the available prey species in several known giant gartersnake habitats and 2) examined stomach contents to reveal the composition of the

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snakes’ actual diet. Results from these studies revealed that giant gartersnakes both preferred and selected native Sierran treefrogs. These results will contribute to a further understanding of the giant gartersnake’s diet, its role as a predator and the degree of impact, if any, exotic species might be having on them. This, in turn, can direct management and conservation efforts.

Brian Halstead 12 / 2 / 15

Acknowledgments

I would like to thank Dr. Erica Wildy for serving as my major advisor and advocate in all matters related to the department. Thank you for your positivity, support and kindness.

I am most grateful and indebted to Dr. Brian Halstead. Thank you for your brilliant insight and guidance through developing the experimental design and the manuscript preparation. I am particularly appreciative for all of the direction and instruction in regard to the statistical analyses.

I thank Dr. Brian Perry for serving on my committee and for comments for revision on my thesis. I would like to thank my professors, Dr. Ellen Woodard and Dr.

Christopher Kitting, and colleagues Kristy Howe and Brianne Brussee for their opinions on specific sections or chapters.

I would like to thank Dr. Glenn Wylie and Mr. Mike Casazza for their advice with regard to presentations, general support of my project and my continued employment at

USGS.

A big thank you to all the landowners and land managers for allowing USGS to conduct research and capture snakes on their properties.

Another big thanks to the many employees and volunteers that contributed to the capture and care of the many snakes used in the study, including but not limited to: Justin

Demianew, Nicole Dotson, Allie Essert, Kristin Fouts, Anna Jordan, Dan Knapp, Brianna

Larsen, Desmond Mackell, Sam McNally, Kristin Ober, and Shannon Skalos.

To those in the Shaffer lab, particularly, Dr. Christopher Searcy for instilling in me a foundation to pursue herpetological study.

Thanks to my friends and family, above all my mom, a most excellent role model and cheerleader who provided me with opportunities that eventually allowed me to pursue a career in the field for which I am most passionate. Also, my dad and aunt for caring about my success and happiness. I thank my sisters Ari, Ayşe, Morgan, Juliana,

Zeka, Anna Joy, and Katie for emotional support, long phone conversations, and sleepovers.

I would like to thank my love, Andrew Sutherland. Thank you for challenging me, for the in-depth discussion and edits on my thesis, and for inspiring and sharing with me a love for nature and fauna.

I dedicate this thesis to the snakes.

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Table of Contents

Abstract ...... iii

Acknowledgments...... vi

List of Figures ...... x

List of Tables ...... xii

Chapter 1 Thesis introduction ...... 1

Statement of Thesis ...... 1

Biology of the Giant Gartersnake...... 2

Taxonomy and Description ...... 2

Natural History ...... 3

Distribution ...... 3

Threats and Conservation ...... 5

Factors Potentially Affecting Diet Selection in the Giant Gartersnake ...... 5

History of Methodology ...... 8

Significance of the Methodology ...... 9

Thesis Organization...... 10

Chapter 2 Prey Preference and Selection of Neonate Giant Gartersnakes (Thamnophis gigas) from the Sacramento Valley of California ...... 11

Introduction ...... 11

Materials and Methods ...... 13

Experiment 1 – Neonate Responses to Olfactory Chemical Cues of Potential Prey

(Day 15 to Day 20) ...... 14

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Experiment 2 – Neonate Prey Selection and Consumption (Multiple Aquatic Prey

Species; Day 21) ...... 17

Experiment 3 – Neonate Feeding Behavior (Single Terrestrial Prey Species; Day 21)

...... 21

Data Analysis ...... 22

Results ...... 24

Prey Preference Trials ...... 24

Prey Selection/ Consumption Trials ...... 28

Discussion ...... 33

Chapter 3 Giant gartersnake (Thamnophis gigas) prey selection ...... 42

Introduction ...... 42

Materials and Methods ...... 43

Field Methods ...... 43

Analytical Methods...... 50

Results ...... 51

Discussion ...... 54

Chapter 4 Thesis Discussion ...... 58

References ...... 63

Appendix ...... 76

List of Figures

Figure 1.1 Female giant gartersnake ...... 2

Figure 1.2 Giant gartersnake with three nematode cysts...... 7

Figure 1.3 Giant gartersnake with catfish spine protruding through body cavity. Photo

credit: S. Skalos ...... 8

Figure 2.1 Experimental setup for olfactory trials. See neonate in container and labeled

vials containing chemical cues...... 16

Figure 2.2. Neonate giant gartersnake tongue flick response to chemical cue...... 16

Figure 2.3 Experimental setup of consumption trials. Dishpan contains a bullfrog tadpole

(far lower left corner), a treefrog tadpole (visually above and near bullfrog

tadpole), a sunfish (closest to neonate), and a mosquitofish (farthest from

neonate)...... 19

Figure 2.4 Giant gartersnake neonate consuming a bullfrog tadpole ...... 21

Figure 2.5 Neonate giant gartersnakes release...... 22

Figure 2.6 Mean tongue-flick attack scores of neonate giant gartersnakes in response to

chemical cues of extracts in prey preference trials. Different letters represent

significant differences. Points represent posterior medians; error bars

represent 95% credible interval ...... 26

Figure 2.7 Consumption probabilities of neonate giant gartersnakes presented with prey

choices in consumption trials. 4 choice: n= 47, 3 choice: n=33, 2 choice: n= 7.

Different letters represent significant differences within groups. Points

represent posterior medians; error bars represent 95% credible intervals...... 31

Figure 2.8 Mean latency to attack and successful capture of prey by neonate giant

gartersnakes in consumption trials. Different letters represent significant

differences. Points represent posterior medians; error bars represent highest

posterior density intervals...... 32

Figure 3.1 Map of trapline locations where giant gartersnakes were captured. Each cluster

of traplines is grouped into Basin areas: Butte Basin (Butte County), Colusa

Basin (Colusa and northern Yolo counties), Sutter Basin (Sutter County),

American Basin (northeastern Sacramento and southern Sutter counties). Each

dot represents a trapline where snakes were captured and sampled...... 44

Figure 3.2 Traps used to catch giant gartersnakes ...... 48

Figure 3.3 Giant gartersnake regurgitation of a bullfrog metamorph...... 49

Figure 3.4 Giant gartersnake prey selection ratios (relative to bullfrog tadpole). Different

letters represent significant differences (95% credible interval of selection

ratio does not contain 1)...... 52

Figure 3.5 Giant gartersnake standardized prey selection ratio. Different letters represent

significant differences (95% credible interval of difference does not contain

0)...... 53

List of Tables

Table 2.1. Sums and proportions of prey types that neonate giant gartersnakes within

litters consumed. NA designates that the respective prey was not offered as a

choice for that litter...... 20

Table 2.2 Posterior quantiles of mean tongue flick attack scores of giant gartersnake

neonates in response to chemical cues in prey preference trials...... 27

Table 2.3 Posterior quantiles of individual variation of neonate giant gartersnakes in

response to chemical cues in prey preference trials...... 27

Table 2.4 Posterior quantiles of probability of selection of different prey types by neonate

giant gartersnakes that had four prey choices in consumption trials. AIW&A

designates that each litter within the group was analyzed independently,

weighted by size of litter and then averaged across group versus the pooled

method of analysis, where all individuals given the same number of prey

choices were pooled...... 29

Table 2.5 Posterior quantiles of probability of selection different prey types by neonate

giant gartersnakes that had three prey choices in consumption trials. AIW&A

designates that each litter within the group was analyzed independently,

weighted by size of litter and then averaged across group versus the pooled

method of analysis, where all individuals given the same number of choices

were pooled...... 29

Table 2.6 Posterior quantiles of probability of selection of neonate giant gartersnakes that

had two prey choices in consumption trials (fish only)...... 29

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Table 2.7 Odds ratio estimation of selection of different prey types by neonate giant

gartersnakes that had four prey choices in consumption trials...... 30

Table 2.8 Odds ratio estimation of selection of different prey types by neonate giant

gartersnakes that had three prey choices in consumption trials...... 30

Table 2.9 Posterior quantiles of mean latency to attack (in seconds) of neonate giant

gartersnakes to various prey types in consumption trials...... 33

Table 3.1 Trapline name, habitat type, snake and prey counts and trapping dates...... 45

Table 3.2 Raw counts of stomach contents from regurgitation of giant gartersnakes in

2014. Some individuals regurgitated multiple items...... 47

Table 3.3 Raw counts of stomach contents from regurgitation of giant gartersnakes in

2013. Some individuals regurgitated multiple items...... 47

Table 3.4 Prey selection model selection information ...... 51

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Chapter 1 Thesis introduction

Statement of Thesis

The introduction of exotic species into an environment can cause great change in the trophic dynamics of native species. This is because, in a given habitat, long-term interacting species coevolve to reach a balance in an ecosystem. Anthropogenic factors such as the introduction of exotic species, disease, pollution, or factors related to climate change can cause this balance to be disturbed, leading to the disruption of ecosystem functioning, alteration of community composition, and even extinction of populations.

Moreover, populations that have already been impacted by habitat loss or some other environmental factor(s) are particularly vulnerable to extirpation.

The giant gartersnake (Thamnophis gigas), an endemic predator of the Central

Valley of California, is listed as threatened at both the state and federal levels. This species has endured significant range reductions and fragmentation over the past several decades as a consequence of conversion of much of its wetland habitat to agriculture

(Halstead et al., 2010). An additional factor, possibly exacerbating habitat degradation, is the introduction of several nonnative fish and one frog species into the aquatic system of the Central Valley of California. Not surprisingly, this has sparked concern about the short-term and long-term effects that these introductions might have on giant gartersnake fitness. The research described in this thesis is designed to further investigate this issue.

In my study, I examine the dietary interest this native predator exhibits toward exotic versus native prey. Additionally, I describe the extent to which these exotic species comprise available prey for the giant gartersnake and measure actual prey selection.

Biology of the Giant Gartersnake

Taxonomy and Description

The giant gartersnake was originally described as a subspecies of Thamnophis ordinoides (Fitch, 1940). The species was later split in 1979 and the giant gartersnake became considered as a subspecies of Thamnophis couchii (Lawson & Dessauer, 1979).

It was then elevated to its own species, Thamnophis gigas, in 1987 (Rossman & Stewart,

1987).

The adult body length for this species ranges from 94 to 165 cm. Individuals usually exhibit a yellowish dorsal stripe and a lighter stripe on either side of a predominantly olive background (Stebbins & McGinnis, 2012). Pronounced checkering and orange instead of yellow stripes have been observed in certain populations (USGS, unpublished observation; Figure 1.1).

Figure 1.1 Female giant gartersnake

Natural History

The giant gartersnake is an endemic predator of wetlands in the Central Valley of

California. This species populates the slow moving waters of marshes and canals preying primarily on aquatic amphibians and fishes (Rossman et al., 1996). Their active season begins in early March and lasts until late September. Giant gartersnakes are sexually dimorphic for size with adult females larger than males (Wylie et al., 2010). These snakes are viviparous (Rossman et al., 1996) and females typically give birth between July and

September. Average litter sizes reported are 17 (Halstead et al., 2011) and 23 (Hansen &

Hansen, 1990). During their inactive season, giant gartersnakes brumate (enter a hibernation-like state) in rodent and crayfish burrows, often in levees or uplands near wetlands (Halstead et al., 2015).

Distribution

California’s Central Valley was once rich with wetlands in the basins along the

Sacramento and San Joaquin Rivers. Since the late 1800s, the expansion of the United

States to include California and the associated rush to secure land and resources contributed to a dramatic increase in the human population in this region. This phenomenon, along with the fertile soil and temperate Mediterranean climate of the region, facilitated development of commercial agriculture; today California is the largest producer of many agricultural commodities. A long term consequence of this productivity is that an 91% of the original marsh and wetland areas in California have been converted and/or have had water diverted (Zedler, 1996) such that only isolated pockets of wetland habitat currently remain (Gilmer et al., 1982).

Different types of crops are grown in the northern and southern portions of the

Central Valley and this significantly affects the suitability of habitat for the giant gartersnake. Rice is the predominant crop in the Sacramento Valley (USDA-NASS,

2015), and because of its water-intensive requirements and associated canals, it provides suitable habitat for the giant gartersnake. Snakes emerge from brumation as early as

March and flooding of rice fields occurs in May. The presence of standing water and marsh-like conditions throughout the snake’s active season allows it to persist in areas where rice is the dominant crop. Additionally, rice harvest coincides with the end of the giant gartersnake’s active season, roughly matching the phenology of the snake.

In contrast, the San Joaquin Valley is dominated by cotton and other row crop production (USDA-NASS, 2015) that do not simulate the marsh-like environments that are produced by rice fields. Thus, while the historic range of the giant gartersnake in this region has been described as extending as far south as historic Kern and Buena Vista

Lakes in Kern County (Fitch, 1940), by 1980 habitat south of Fresno was deemed unsuitable (Hansen & Brode, 1980). This was the main cause of the drastic decline of the giant gartnersnake in this area and the primary reason the species was listed as threatened by the state in 1971 and federally in 1993 (U.S. Fish and Wildlife Service, 1993). One promising aspect of the circumstances that the giant gartersnake faces is that, although the southern portion of the population is almost extirpated because of the conversion of its habitat to non-rice agriculture and the associated water diversion, the northern population has been able to adapt and persist in zones of rice agriculture, which provides the aquatic habitat that this snake requires.

Threats and Conservation

Since the 1990s the United States Geological Survey (USGS) has conducted various studies that have explored factors affecting the ecology of the giant gartersnake, including habitat suitability, body condition and growth rate, temperature effects and thermoregulation, all while surveying for general demography and occupancy. These efforts have improved the understanding of the natural history of this elusive snake, which in turn contributes to improved management and agricultural practices. Despite these efforts, the diet of giant gartersnakes, a fundamental aspect to their conservation has not been thoroughly explored.

Factors Potentially Affecting Diet Selection in the Giant Gartersnake

For giant gartersnakes, the current selection of prey includes one native species, the Sierran treefrog (Pseudacris sierra), but is otherwise almost completely comprised of non-native species (Rossman et al., 1996). The list of introduced potential prey species includes, but is not limited to, many centrarchid fish species such as green sunfish

(Lepomis cyanellus), bluegill (Lepomis macrochirus), black and white crappie

(Pomoxis nigromaculatus and P. annularis), and largemouth bass (Micropterus salmoides); several species in the minnow family (Cyprinidae); and other fish species such as the Mississippi silverside (Menidia beryllina), western mosquitofish (Gambusia affinis), and black bullhead catfish (Ameiurus melas) (UC ANR, 2015). The most prolific invasive species in these wetland habitats is the American bullfrog (Lithobates catesbeianus). Although it is the most abundant prey source for giant gartersnakes, it also

appears to have significant negative impacts on the fitness of the giant gartersnake through increased rates of parasitic infection and intraguild predation (Wylie et al., 2003).

The bullfrog and the catfish, specifically, have poorly understood and potentially dangerous interactions with the giant gartersnake. Although the extent to which these exotic species affect the giant gartersnake is unknown, some preliminary studies and anecdotal observations elucidate the general nature of these impacts (Wylie et al., 2003;

Lichtenfels & Lavies, 1976). For example, based on USGS preliminary data, it appears that the giant gartersnake primarily eats bullfrog juveniles and tadpoles. Interestingly, it has been discovered that the predation is not one-sided. A study examining the stomach contents of 30 bullfrogs revealed that bullfrogs of varying sizes had consumed three giant gartersnake neonates (Wylie et al., 2003).

In addition to predation, it has been speculated that bullfrogs as well as introduced fish transmit nematodes (including Eustrongylides spp.) to giant gartersnakes (von Brand,

1944). Many giant gartersnakes exhibit cysts of larval nematodes that form subcutaneously (Figure 1.2). Though no studies have been conducted to examine the specific effect that parasites have on giant gartersnake fitness, nematode infections of a congener, the red-sided gartersnake (Thamnophis sirtalis infernalis), can cause contortion and mortality (Lichtenfels & Lavies, 1976). Furthermore, several captive giant gartersnakes from the American Basin died in association with the presence of this parasitic infection (U.S. Fish and Wildlife Service, 1999).

Figure 1.2 Giant gartersnake with three nematode cysts. The presence and activities of the bullfrog have other negative community-level effects leading to, among other things, the decline of native amphibians (Bury & Whelan,

1984; Adams & Pearl, 2007). This indirectly affects the giant gartersnake, whose diet includes native amphibian prey (Rossman et al., 1996).

Another introduced species that has a potential detrimental impact on giant gartersnakes and also serves as prey for them is the black bullhead catfish (Ameiurus melas). These fish are armed with dorsal and pectoral spines in their fins which can be erected as a defense mechanism. Occasionally, gartersnakes are found with these spines protruding through their skin (Figure 1.3). This can cause injury, blockages and potentially death (USGS, unpublished observation).

Figure 1.3 Giant gartersnake with catfish spine protruding through body cavity.

Photo credit: S. Skalos

The introductions of bullfrogs, catfish and other exotic species into the Central

Valley of California potentially have multifaceted, negative effects on giant gartersnakes that prey on them. Thus, it becomes critical that the scope and extent of the threat of these introduced species on giant gartersnakes are examined. My Master’s thesis investigates whether and how these and other exotic animals might be affecting diet selection and food consumption in giant gartersnakes.

History of Methodology

The first study examining the behavior of neonate snakes in response to chemical cues of prey was conducted by Gordon Burghardt in 1966 on the generalist species,

Thamnophis sirtalis. The development of quantifying attack and predator responses proceeded as did various methods of creating and testing extracts. Many procedures were tested and compared (Cooper & Burghardt, 1990; Cooper et al., 2000), and eventually, the influences of specific parameters (i.e. genetics, maternal diet, geographic variation, prior exposure) were able to be measured.

Responses to chemical cues of prey (and some predators) using extracts have been measured in many species within the genus Thamnophis, including but not limited to: T. elegans (Drummond & Burghardt, 1983), T. hammondii (Mullin et al., 2004; Hale, 2010),

T. sirtalis (Burghardt 1969, 1970, 1971, 1975; Arnold, 1978), and T. validus (de Queiroz et al., 2001), but have also been performed on other genera including, but not limited to:

Agkistrodon (Chiszar et al., 1979), Coluber (Cooper et al., 2000), Lampropeltis (Saviola et al., 2012; Williams & Brisbin, 1978), Nerodia (Burghardt, 1968; Gove & Burghardt,

1975; Czaplicki, 1975; Dunbar, 1979), Notechis (Aubret et al., 2006), and Pituophis

(Smith et al., 2015; Burger, 1991).

Significance of the Methodology

The laboratory and field studies are meant to be complementary and allow for a clearer understanding of giant gartersnake foraging ecology than either type of study could alone. While laboratory trials allow for a controlled setting to examine elements of particular interest, field investigations allow for a more comprehensive view of what actually occurs in nature with the influence of factors that cannot be controlled or are unknown to be significant. For the purposes of this thesis, laboratory trials allow for investigation of naïve olfactory prey preference and the opportunity for a unique visual

observation of neonate consumption unattainable in the field, while examination of field- collected data allow for measure of prey selection in nature, a situation that cannot be replicated in a laboratory.

Thesis Organization

To better understand aspects of giant gartersnake diet, I conducted an investigation involving two behavioral studies carried out in the laboratory and a two- staged field study. In Chapter 2, I describe the methodology and results for two sets of laboratory behavioral trials during which I collectively examined prey preference and consumption. The first set of trials was designed to determine prey preference and involved presentation of chemical cues to neonates via swabs dipped in prepared prey extracts. The second set of trials was designed to measure actual dietary choice and involved presenting neonates with different types of live prey items. In Chapter 3, I present the results from examination of stomach contents collected from adult snakes in the field. These data reveal the snakes’ actual selection among available prey in nature.

In Chapter 4, I discuss areas of potential improvement in the experimentation and suggest ideas for future research related to the current investigation.

Chapter 2 Prey Preference and Selection of Neonate Giant Gartersnakes

(Thamnophis gigas) from the Sacramento Valley of California

Introduction

To fully understand the effects that the introduction of non-native species has on an ecological system, an assessment of how species interactions as well as individual species are affected is critical (Sih et al., 2010). For example, foraging behavior (and the associated physiological processes) of predatory species can change in systems into which new species have been introduced (Britt et al., 2006). This not only directly affects the fitness of the individual consumers but can indirectly influence the long-term viability of the population to which those consumers belong (Manly et al., 2002). Thus, study and documentation of interactions between natives and exotics is crucial for understanding the short-term and long-term consequences of the introductions on native species. This is particularly true when the native species is of conservation concern.

The giant gartersnake (Thamnophis gigas), a state and federally threatened semi- aquatic snake precinctive to the wetlands of the Central Valley of California (Fitch,

1940), is a species that has experienced extensive human-mediated changes to its environment that affect its ecology. Giant gartersnake population decline has largely been attributed to a mass conversion of its wetland habitat to various types of agriculture

(U.S. Fish and Wildlife Service, 1993; Zedler, 1996). However, this snake has been able to persist in the northern part of its range, the Sacramento Valley, because of the presence of rice agriculture, which simulates the marsh-like conditions necessary for giant gartersnakes (Halstead et al., 2010).

The giant gartersnake is also faced with the introduction of many hardy, generalist species into its habitat, many of which can serve as potential prey items. The diet of the giant gartersnake has not been extensively investigated; thus, the question of whether and how these introductions might affect the snake and potentially contribute to the snakes’ population decline persists. Giant gartersnakes are generally thought to prey on anurans and fish (Stebbins & McGinnis, 2012; Rossman et al., 1996). Though historically these snakes preyed upon native fishes and frogs such as Sacramento blackfish (Orthodon microlepidotus), the now extinct thicktail chub (Gila crassicauda; NatureServe, 2013), and California red-legged frogs (Rana draytonii), the majority of prey now available to giant gartersnakes are introduced (Rossman et al., 1996). One native prey species that currently exists within the range of the giant gartersnake is the Sierran treefrog

(Pseudacris sierra) (Stebbins & McGinnis, 2012). Exotic fish species known to be consumed by these snakes include the Mississippi silverside (Menidia beryllina), western mosquitofish (Gambusia affinis), black bullhead catfish (Ameiurus melas) and several species in the minnow family (Cyprinidae) (USGS, unpublished observation). Many more introduced species, however, might comprise the diet of these snakes including many centrarchid fish species such as green sunfish (Lepomis cyanellus), bluegill

(Lepomis macrochirus), black and white crappie (Pomoxis species), and largemouth bass

(Micropterus salmoides) (UC ANR, 2015).

If giant gartersnakes are indeed consuming exotic species regularly, it is crucial that this activity is documented and the species identified so that any potential negative impact on giant gartersnake fitness can be examined. In an attempt to do the former, this

study is an examination of diet preference and consumption in giant gartersnake neonates, born from females originating from habitats into which non-native species, which might serve as potential prey along with existing native species, have been introduced. The use of captive born neonates allows for measure of a naïve response, as these snakes have never been exposed to any prey and therefore, have no learned experience. The objective of this study was to determine the relative strength of the predatory response of neonate giant gartersnakes to native and introduced prey. I achieved this using two sets of behavioral trials, one examining prey preferences of naïve neonate giant gartersnakes to chemical cues of different prey items, and a second quantifying the probability with which neonate giant gartersnakes consumed alternative prey in a controlled laboratory setting.

Materials and Methods

I collected three gravid female giant gartersnakes using modified minnow traps

(Halstead et al., 2013) from each of three sites in California’s Central Valley: Colusa

National Wildlife Refuge, Colusa County; Gilsizer Slough, Sutter County; and Natomas

Basin Conservancy Reserves, Sacramento and Sutter counties. I brought the snakes back to the USGS Dixon Field Station (Dixon, CA) where they were maintained in the laboratory until parturition. I housed the snakes in 38L glass aquaria lined with Astroturf that contained a metal water dish and an additional piece of rolled Astroturf that the snakes could use for shelter. Enclosures were cleaned with ≥ 3% quaternary ammonium

(Sparquat 256) after defecation. The laboratory had one window, which allowed for exposure to natural daylight; additional fluorescent lighting was also provided to

subsidize the natural lighting present from 0900 to 1700. The temperature was maintained between 23-28° C.

The adult snakes were offered field-caught live prey (adult or tadpole bullfrogs) from their respective site of capture once a day and fresh water ad libitum. The female snakes were maintained in captivity until they gave birth, after which they were released at their location of capture. Neonate snakes were retained and housed by clutch in the same manner described for the adults. I uniquely marked each neonate using a permanent marker as a means of identification and recorded their snout-vent length, tail length, and mass within two days of birth. Behavioral trials began 15 days after birth as has been convention in similar studies (Cooper et al., 2000; Dix, 1968). I conducted two different behavioral experiments, one involving a set of olfactory trials and a second involving a set of consumption trials.

Experiment 1 – Neonate Responses to Olfactory Chemical Cues of Potential Prey (Day

15 to Day 20)

In July and August of 2014, between the times of 1030 and 1700, 100 neonates, representing nine litters, were individually exposed to a series of eleven chemical cues in random order presented on individual cotton swabs soaked with extracts of a potential prey animal or control cue. The extracts were of the following: (1) adult and (2) tadpole

Sierran treefrog (Psuedacris sierra), (3) adult and (4) tadpole American bullfrog

(Lithobates catesbeianus), (5) mosquitofish (Gambusia affinis), (6) sunfish (Lepomis species), (7) black bullhead catfish (Ameiurus melas), (8) Louisiana red crayfish

(Procambarus clarkii), (9) water (visual control), (10) diluted mint extract (Mentha

species, olfactory control) and (11) California kingsnake (Lampropeltis californiae, predatory and snake control).

I created the extracts by soaking live prey items in deionized water at a ratio of

1:2 (grams of body mass:milliliters of water) for 30 minutes. These extracts were either used fresh or refrigerated at 8° C for up to six days. Extracts were allowed to reach room temperature in the morning before use to allow for maximum odor volatilization when presented to the neonates. Extracts were labeled by a third party so the observer was blind to which chemical cue was presented in each trial.

Before each trial, a neonate was placed in an arena consisting of a clean plastic container measuring 8 x 25 x 15 cm (Figure 2.1) with the container lid in place, and allowed to acclimate for five minutes. Following this habituation period, the lid was gradually moved enough to expose the neonate and a cotton swab was slowly introduced into the enclosure and brought within one cm of the neonate’s supralabials (Figure 2.2).

The time record started when the neonate flicked its tongue in the direction of the swab for the first time. During the trials, if a snake moved toward the swab, the swab was moved away from the snake to maintain the 1 cm buffer. Alternatively, if a snake turned away from the swab, the observer kept the swab still.

Figure 2.1 Experimental setup for olfactory trials. See neonate in container and labeled vials containing chemical cues.

Figure 2.2. Neonate giant gartersnake tongue flick response to chemical cue.

The degree of interest a neonate had in a particular extract was determined using the number of tongue flicks it exhibited in response to the swab. Tongue-flick scoring techniques are well established and broadly accepted as an indication of prey preference

in reptiles (Cooper & Burghardt, 1990). The method to gauge preference by measuring the response to chemical cues has been particularly well developed in the genus

Thamnophis. The methods described for this experiment most closely resemble those described in Cooper et al. 2000, through numerous studies performed by G.M. Burghardt

(1966, 1967, 1968, 1969, 1970, 1975).

Throughout each trial, tongue flicks exhibited by the neonate were counted until the snake turned away and tongue flicked more than 5 cm away from the swab or after a maximum of 45 seconds had passed, whichever occurred first. Gapes (opens mouth while remaining still) and lunges (opens mouth and sudden movement toward the cotton swab) were considered attacks and recorded. Each trial was replicated once a day, three times for each neonate between the ages of 15-20 days.

Experiment 2 – Neonate Prey Selection and Consumption (Multiple Aquatic Prey

Species; Day 21)

At the age of 21 days, the day after the neonates completed the olfactory trials, 99 individuals were subjected to a prey choice experiment. Before each trial began, neonates were placed in a clean plastic container (8 x 30 x 20 cm) containing a metal pan (20 x 20 x 5 cm) filled with 600 ml of water and a number of prey items (Figure 2.3). The prey items were weighed in a dry cup and then placed into the dish pan in random order before the snake was introduced. The metal pan contained a choice of either four prey items, three prey items, or two prey items, depending upon the availability of prey. The litters were grouped by the number of prey choices the individuals had: Group 1 (four litters, n=

47): four prey choices (a treefrog tadpole, a bullfrog tadpole, a sunfish and a

mosquitofish), Group 2 (three litters, n=33): three prey choices (a bullfrog tadpole, a sunfish and a mosquitofish), and Group 3 (two litters, n=7): two prey choices (a sunfish and a mosquitofish). Nine litters were represented in total (Table 2.1). Six individuals from litters CO4171 and BKS 6 that were offered a two-choice test did not select or consume a prey item after 45 minutes and were not included in the analyses. Neonates were placed in the container on the rim of the dishpan with their head oriented toward the water. Often, neonates would move off the rim and explore the container space around the dishpan for several minutes before returning to the rim to begin to hunt. The time record would only begin when the snake returned to the rim, oriented toward the prey items and tongue flicked (although sometimes a neonate would exhibit this behavior when first placed in the container in which case, the time record would start immediately). A clear sheet of thermoplastic was placed over the container and the trial was video recorded until the neonate caught one prey item or a maximum of 45 minutes passed, whichever occurred first (Figure 2.4). At this point, the neonate was removed.

The selected prey item was replaced with a fresh individual of that prey type, but the remaining prey items and the water in the dishpan remained and were reused in consecutive trials. For this experiment, each neonate was tested just once. The video was watched in real time outside the room to monitor activities and then rewatched (by the same observer) to record latency to attacks and the identity of the prey item involved for all attacks, both successful and unsuccessful.

Figure 2.3 Experimental setup of consumption trials. Dishpan contains a bullfrog tadpole (far lower left corner), a treefrog tadpole (visually above and near bullfrog tadpole), a sunfish (closest to neonate), and a mosquitofish (farthest from neonate).

Table 2.1. Sums and proportions of prey types that neonate giant gartersnakes within litters consumed. NA designates that the respective prey was not offered as a choice for that litter.

Four choice group Treefrog Bullfrog Litter (n=47) tadpole tadpole Sunfish Mosquitofish Size LN2224 4 1 0 0 5 Butler 4 15 1 0 20 LN2203 9 7 0 1 17 BKS 6 0 5 0 0 5 Three choice group (no treefrog) (n=33) CO4177 NA 11 3 0 14 GI NA 7 0 0 7 CO4176 NA 12 0 0 12 Two choice group (no frogs) (n=7) Matt NA NA 6 1 7 CO4171 NA NA 0 0 4 BKS 6 NA NA 0 0 2 Sum 17 58 10 2 Proportion 36.17% 72.5% 11.49% 2.3% N = 87

Figure 2.4 Giant gartersnake neonate consuming a bullfrog tadpole

Experiment 3 – Neonate Feeding Behavior (Single Terrestrial Prey Species; Day 21)

In a separate set of trials, I offered Sierran treefrog metamorphs to neonate giant gartersnakes to test the feeding response of the snakes to the terrestrial life stage of this prey species and to determine whether there is a difference in the ease with which neonates captured this stage of these prey animals. I conducted these trials separately from the trials with aquatic prey so that I could examine this issue without the additional complication of having multiple habitat regions within the testing arena.

To start, I placed one treefrog metamorph under a 125 mL cup (diameter 65 cm) in a clean plastic container (8 x 30 x 20 cm) with no metal pan and containing no water.

Then, the neonate was placed in the container and the cup removed to reveal the metamorph. Latency to a successful attack was recorded. Nine neonates in total were tested. Prior to the consumption trial and one day prior to release (20 days old), the snout- vent length, tail-vent length, and mass were again recorded for all neonates. The day after feeding, the animals were released at the location their mother was found (Figure 2.5).

Figure 2.5 Neonate giant gartersnakes release.

Data Analysis

The olfactory data were analyzed using Bayesian hierarchical models with a

Poisson distribution and a log link function in program JAGS 3.4.0 (Plummer, 2013 http://mcmc-jags.sourceforge.net) called through R 3.1.0 (R Core Team, 2014) using packages ‘rjags’ (Plummer, 2014), ‘snow’ (Tierney et al., 2013) and ‘emdbook ’(Bolker,

2008, 2015). The tongue-flick attack score (TFAS) was calculated by subtracting the latency to attack (if any) from the length of the trial and adding the maximum number of tongue flicks for that individual from any trial (Cooper & Burghardt, 1990). Variation in

TFAS was estimated across litters, individuals, and trials within individuals as log- normal random effects. Repeated measures and overdispersion were accounted for with a log-normal observation-level random effect. The effect of age on TFAS was also

included in the model. Goodness-of-fit was tested with a Bayesian p-value using Pearson residuals and data simulated from the model. Uninformative priors included a uniform (0,

10) (upper, lower) distribution for standard deviations of random effects and a normal (0,

10) (mean, standard deviation) distribution for the intercept and coefficients. The model ran using 5 independent chains, a 99,000 iteration burn-in, plus 100,000 iterations after the burn-in. The output was thinned by a factor of 50. I examined history plots and the

Gelman-Rubin diagnostic (Gelman & Rubin, 1992) to assess convergence, and found no evidence for lack of convergence (Gelman-Rubin diagnostic < 1.1 for all monitored parameters).

The consumption data were analyzed using Bayesian hierarchical models with a multinomial (three or more prey types offered) or binomial (two prey types offered) distribution for each set of consumption trials in program OpenBUGS 3.2.3 (Thomas,

2004; Thomas et al., 2006) called through R 3.1.0 using package ‘R2OpenBUGS’ (Sturtz et al., 2005) and ‘coda’ (Plummer et al., 2006). The litters were analyzed independently

(with weighted averages based on litter size) and then analyzed as a pooled group for comparison. Odds ratios were calculated for pooled groups. Litters in which all individuals consumed the same prey were interpreted as 100% likely to consume that particular prey item (no analyses were performed). As an uninformative prior, the probabilities for selection of each type were equally likely and between 0 and 1 (e.g. for two prey types, each prey type had a prior probability of 0.5; for three prey types, each prey type had a prior probability of 0.33, etc.). A binomial distribution was used for the group with two prey choices and a multinomial distribution was used for groups with

three or four prey choices. The model ran using five independent chains, a 20,000 iteration burn-in, plus 20,000 iterations after the burn-in. The output was thinned by a factor of 10. History plots depicted no evidence for lack of convergence.

Latency to successful capture data were analyzed using a fixed effects ANOVA in program JAGS 3.4.0 called through R 3.1.0 using package ‘runjags’ (Denwood, in review) and ‘modeest’ (Poncet, 2012). Latencies were log-transformed to meet normality assumptions. Uninformative priors included a normal (0, 100) distribution for mean latency to attack for each prey type, and a uniform (0, 100) distribution for the standard deviation of the residual error. Goodness-of-fit was tested with a Bayesian p-value using

Pearson residuals and data simulated from the model. The model ran using 5 independent chains, a 9,000 iteration burn-in, plus 20,000 iterations after the burn-in. History plots and the Gelman-Rubin diagnostics (Gelman & Rubin, 1992) were assessed for convergence, and no evidence for lack of convergence was found (Gelman-Rubin diagnostic < 1.1 for all monitored parameters).

Results

Prey Preference Trials

The three native species elicited the strongest responses from the neonates. The strongest tongue flick responses were exhibited toward the native prey species, P. sierra treefrog metamorphs and tadpoles, followed by the native predator, the California kingsnake (Figure 2.6, Table 2.2). The weakest responses were exhibited toward the visual and olfactory controls (Figure 2.6, Table 2.2). All other responses were to introduced species and intermediate in strength with respect to controls and natives

(Figure 2.6, Table 2.2). Responses were more consistent among litters (0.202 [0.099 -

0.435]) and individuals (0.281 [0.234 - 0.338]) than within individuals (0.646 [0.623 -

0.669]). Variation among individuals in their response to cues was least for the native treefrog metamorph extract and greatest for the introduced catfish extract and controls

(Table 2.3). As neonates increased in age by one day, TFAS decreased 1.107 [1.091 -

1.13] times. Attacks were elicited in 25 of 100 neonates and occurred in 45 of 3086

(0.015 %) trials.

errorcrediblerepresent 95% interval bars prey in Diff preference trials. Figure

Mean tongue Mean

flick attack scores of neonate giantgartersnakes flick attack response scores neonate of chemical to extracts of cues in

erent letters represent significant represent erentletters Points represent medians; differences. posterior

Table 2.2 Posterior quantiles of mean tongue flick attack scores of giant gartersnake neonates in response to chemical cues in prey preference trials.

Prey Type 2.50% 50% 97.50% Mint 5.374801 6.536248 7.908682 Water 5.793157 7.033123 8.400995 Bullfrog adult 6.974774 8.416102 10.06694 Crayfish 7.46051 8.95984 10.74252 Sunfish 8.569868 10.37063 12.46474 Bullfrog tadpole 9.640639 11.5518 13.76571 Mosquitofish 9.718101 11.70947 14.01612 Catfish 10.12227 12.2811 14.8264 Kingsnake 10.96176 13.23642 15.86882 Treefrog tadpole 12.72068 15.26972 18.19816 Treefrog metamorph 14.54217 17.39044 20.7477

Table 2.3 Posterior quantiles of individual variation of neonate giant gartersnakes in response to chemical cues in prey preference trials.

Cue Standard Deviation (Log Scale) 2.50% 50% 97.50% Water 0.234 0.281 0.338 Bullfrog adult 0.005 0.121 0.287 Crayfish 0.005 0.08 0.234 Sunfish 0.01 0.163 0.327 Bullfrog tadpole 0.004 0.082 0.233 Mosquitofish 0.012 0.148 0.308 Catfish 0.104 0.287 0.423 Kingsnake 0.006 0.157 0.314 Treefrog tadpole 0.004 0.071 0.216 Treefrog metamorph 0.002 0.059 0.182

Prey Selection/ Consumption Trials

Little difference in estimation existed between the pooling method versus calculating probabilities independently for each litter, then calculating the weighted mean

(weighting litters by size; Tables 2.4, 2.5, 2.6). In the group that had four prey choices, neonates did not consume either tadpole species more than the other (odds of consuming bullfrog over treefrog: 0.363 [0.116 - 1.153]) or either fish type more than the other (odds of consuming sunfish over mosquitofish: 1.076 [0.023 - 40.981]; Figure 2.7, Table 2.7).

There was, however, significant selection of both tadpoles species over both fish species in general (odds of consuming either tadpole over either fish 746.195 [57.038 -

32610.581]). In the group with three prey choices (treefrog tadpole not offered), the bullfrog tadpole was heavily selected over both fish types (odds of consuming bullfrog over sunfish: 15.900 [1.423 - 330.754], odds of consuming bullfrog over mosquitofish:

5.12E+31 [1153.38 - ∞], odds of consuming bullfrog over either fish: 165.578 [16.243-

1891.720]; Figure 2.7, Table 2.8) and there also was a significantly higher probability of consuming sunfish over mosquitofish (odds of consuming sunfish over mosquitofish:

2.39E+30 [81 - ∞]; Table 2.8). However, in the group with only sunfish and mosquitofish offered, there was no evidence for selection of either fish species over the other (odds of consuming sunfish over mosquitofish: 3.993 [0.891 - 31.160]). In this group, 12 individuals in total, and all individuals in one litter, were not able to catch a prey item after 45 minutes and were not included in analyses.

Table 2.4 Posterior quantiles of probability of selection of different prey types by neonate giant gartersnakes that had four prey choices in consumption trials. AIW&A designates that each litter within the group was analyzed independently, weighted by size of litter and then averaged across group versus the pooled method of analysis, where all individuals given the same number of prey choices were pooled.

Four choice group AIW&A Pooled AIW&A Pooled AIW&A Pooled Percentile 2.5% 2.5% 50.0% 50.0% 97.5% 97.5% Treefrog 0.132 0.234 0.301 0.354 0.485 0.502 Bullfrog 0.449 0.453 0.638 0.606 0.828 0.729 Sunfish 0.001 0.001 0.030 0.017 0.129 0.08 Mosquitofish 0.001 0.001 0.015 0.015 0.075 0.081

Table 2.5 Posterior quantiles of probability of selection different prey types by neonate giant gartersnakes that had three prey choices in consumption trials.

AIW&A designates that each litter within the group was analyzed independently, weighted by size of litter and then averaged across group versus the pooled method of analysis, where all individuals given the same number of choices were pooled.

Three choice group AIW&A Pooled AIW&A Pooled AIW&A Pooled Percentile 2.5% 2.5% 50.0% 50.0% 97.5% 97.5% Bullfrog 0.807 0.787 0.915 0.931 0.978 0.980 Sunfish 0.022 0.020 0.085 0.069 0.192 0.213 Mosquitofish 0.000 0.000 0.000 0.000 0.001 0.003

Table 2.6 Posterior quantiles of probability of selection of neonate giant gartersnakes that had two prey choices in consumption trials (fish only).

Two choice group 2.50% 50% 97.50% Sunfish 0.471 0.8 0.969 Mosquitofish 0.031 0.2 0.529

Table 2.7 Odds ratio estimation of selection of different prey types by neonate giant gartersnakes that had four prey choices in consumption trials.

Four choice group Likelihood to Select: 2.50% 50% 97.50% Bullfrog over Treefrog 0.116 0.363 1.153 Bullfrog over Sunfish 13.286 91.858 2778.374 Bullfrog over Mosquitofish 14.535 102.105 2671.388 Treefrog over Sunfish 5.637 33.598 1162.986 Treefrog over Mosquitofish 5.240 35.590 995.802 Sunfish over Mosquitofish 0.023 1.076 40.981

Table 2.8 Odds ratio estimation of selection of different prey types by neonate giant gartersnakes that had three prey choices in consumption trials.

Three choice group Likelihood to Select: 2.50% 50% 97.50% Bullfrog over Sunfish 1.423 15.920 330.754 Bullfrog over Mosquitofish 1552.38 5.12E+31 ∞ Sunfish over Mosquitofish 81.00 2.39E+30 ∞

Figure 2.7 Consumption probabilities of neonate giant gartersnakes presented with prey choices in consumption trials. 4 choice: n= 47, 3 choice: n=33, 2 choice: n= 7.

Different letters represent significant differences within groups. Points represent posterior medians; error bars represent 95% credible intervals.

The neonates’ average latency to successful attack differed significantly among the prey types (Figure 2.8, Table 2.9). The mean latency to attack response for treefrog metamorphs (29.288 s [12.020 – 70.536 s]) was significantly shorter than all other prey types (Figure 2.8, Table 2.9). The mean latency to successful attack did not differ among

treefrog tadpoles, bullfrog tadpoles, and mosquitofish, but latency to successful attack was shorter for treefrog tadpoles (104.366 s [51.686 – 212.358 s]) than sunfish (426.026 s

[191.955 – 948.846 s]; Figure 2.8, Table 2.9).

Figure 2.8 Mean latency to attack and successful capture of prey by neonate giant gartersnakes in consumption trials. Different letters represent significant differences. Points represent posterior medians; error bars represent highest posterior density intervals.

Table 2.9 Posterior quantiles of mean latency to attack (in seconds) of neonate giant gartersnakes to various prey types in consumption trials.

Prey Type 2.5% 50% 97% Treefrog metamorph 12.01993 29.28865 70.53589 Treefrog tadpole 51.68576 104.3656 212.3581 Bullfrog tadpole 146.9737 212.2362 307.3595 Mosquitofish 39.25757 257.6752 1661.852 Sunfish 191.9554 426.0258 948.8161

Discussion

The results from the olfactory trials in this study indicate that giant gartersnakes prefer native prey and can distinguish a native predator. The focal result suggests that giant gartersnakes innately prefer native treefrogs and their larvae. The preference for native prey species also exists in other snakes, including T. hammondii (Hale, 2010), T. elegans (Arnold, 1977), Coluber constrictor (Cooper et al., 2000), and Natrix [Nerodia] sipedon (Gove & Burghardt, 1975), and the lizard Phrynosoma coronatum (Suarez et al.,

2000). It is not particularly surprising that a predator would innately prefer native prey, however preference in this context might simply mean that neonates innately recognize treefrogs as prey and might not recognize the other species as prey. Although the mechanisms leading to preference of native prey are unknown and potentially complex, neonate giant gartersnakes clearly exhibit the strongest response to cues from native prey

Neonates exhibited the third highest response towards the native predator, the

California kingsnake. A response of this nature is not unprecedented. High TFASs toward ophiophagous snakes were also observed in studies on other snakes in the genus

Thamnophis (Weldon, 1982) and Coluber constrictor (Cooper et al., 2000) and were

interpreted as recognition of a predator. The addition of a predator extract in the olfactory trials was for comparison to bullfrog adults, which also prey on giant gartersnake neonates (Wylie et al., 2003). Because the response of neonates to the bullfrog adult was substantially less than that exhibited toward the California kingsnake, the data do not support innate recognition of adult bullfrogs as predators. The occurrence of intraguild predation between giant gartersnakes and bullfrogs might cause selection pressures to counter each other and thus result, in a neutral-type response. Alternatively, interpretation of a high TFAS towards the California kingsnake could be attributed to the condition of it being native and therefore innately recognized in general.

In the consumption trials, treefrog tadpoles were not found to be selected significantly more often than bullfrog tadpoles. This seemingly contradicts the results of the olfactory trials where neonates exhibited a preference for native treefrog tadpoles and metamorphs over bullfrog tadpoles. Alternatively, these data suggest that although bullfrog tadpoles are less preferred, they might be easier to catch. No locomotory measurements were made in this study, but different tadpole species differ in swimming kinematics (Wassersug, 1989). Furthermore, though attempts were made to reduce size variation among prey items, bullfrog tadpoles tended to be larger (as much as 0.5 grams) than other prey items. This could have influenced both locomotory performance among tadpole individuals as well as selection by neonates for larger prey items (Arnold, 1993).

Additionally, naïve larvae of three frog species have been shown to vary in their refuge- seeking response when exposed to chemical cues of native and introduced predators

(Pearl et al., 2003), thus supporting the hypothesis of additional behavioral variability

between tadpole species. Perhaps treefrogs, which have evolved with giant gartersnakes, have more developed antipredatory responses to giant gartersnakes than do bullfrogs, which did not evolve with giant gartersnakes (but did evolve with other natricines).

Results from consumption trials indicated that although neonates will consume fish, they are consumed with lower probability than anuran prey. Alfaro (2002) examined aquatic hunting modes in three Thamnophis species. He concluded that piscivorous specialists (T. couchii and T. rufipunctatus) have highly evolved hunting strategies that differ from that of the generalist T. sirtalis. Although more aquatic than T. sirtalis, giant gartersnakes have also not evolved as exclusively piscivorous; thus, they might be less efficient at catching fish than larval anurans.

An additional variable that could have contributed to the modest, albeit present selection of fish by neonates, could be the design of the experimental setup itself. In another highly aquatic snake species, Nerodia clarkii, individuals varied in fish capture success with differences in mangrove density (Mullin & Mushinsky, 1995). It is possible that the limited size and structural simplicity of the dishpan made it harder (or easier) for giant gartersnake neonates to capture fish they were offered relative to what might be observed in their natural habitats, which are more structurally complex. It is also important to note that no native fish were used in trials because few native fish occurred in sampled areas. Additionally, non-native fish that evolved in the presence of exclusively piscivorous snake(s) could arguably have better anti-predatory defenses than native fish, such as those inhabiting the Central Valley, and did not evolve in such circumstances.

Giant gartersnake foraging ecology has not been extensively investigated, so their hunting strategies are unknown. In a study comparing four species in the genus

Thamnophis, two aquatic specialists (T. couchii and T. melanogaster) and two terrestrial- aquatic generalists (T. sirtalis and T. elegans), the aquatic specialists had superior aquatic hunting strategies and caught more fish when compared to the terrestrial-aquatic generalists (Drummond, 1983). Qualitative comparisons of generalist versus specialist predators are really most useful when contrasted with another species (Futuyma &

Moreno, 1988). That type of characterization cannot be made based on this study alone, but these examples demonstrate that differences exist in the ability of predators to catch different types of prey and why those differences might exist. The lower mean latency to successful attack for treefrog metamorphs suggests that they might be easier for neonate giant gartersnakes to catch. Perhaps this is because they are more often encountered on land than water as compared to chiefly aquatic prey, such as tadpoles and fish, although they might be easier to catch in water as well. Additionally, because of differences in aquatic and terrestrial conditions, transmission of olfactory and visual cues might contribute to prey recognition and attack precision, thus reducing latency to successful attack in terrestrial conditions.

Another factor affecting the interaction of giant gartersnakes with treefrogs is the overlap of the timing in the commencement of their life stages (i.e. giant gartersnake neonates and treefrog metamorphs). Treefrogs generally breed January through July, and can lay up to three clutches per season; however, their reproduction is highly adapted to a given locality and its respective precipitation and temperatures (Stebbins & McGinnis,

2012). Thus, with warmer temperatures such as those in the Sacramento Valley, the breeding season can be long and larval development can be fast. Treefrog tadpoles were observed at sampled sites in the Sacramento Valley as early as April 18th and as late as

September 25th in 2014 (USGS, unpublished observation). Female giant gartersnakes give birth approximately mid-July to early October (Halstead et al., 2011), likely when metamorphs are at the peak of their abundance. The shorter latency to attack and capture provides evidence that these metamorphs might be an important prey source, particularly for neonate snakes. Another aquatic snake, Nerodia fasciata, shifts from selecting aquatic salamanders to seasonally available treefrogs and cricketfrogs (Plummer & Goy, 1984) which supports the hypothesis that seasonally available amphibian metamorphs can be important prey for natricine snakes.

Additional evidence for the predilection of native prey was found in the consumption trials where neonates were offered treefrog metamorphs. The data collected during the different series of consumption trials cannot be directly compared since they were independently run and because of the differences in aquatic and non-aquatic conditions of the trial designs. That notwithstanding, latencies to attack of metamorphs were substantially shorter than in trials where neonates were offered the aquatic prey in dish pans. Moreover, with aquatic prey, several failed attempts typically occurred before capture (if capture even occurred) whereas, in the treefrog metamorph prey trials, every neonate was able to capture the prey item during their first attempt.

Preference for native prey might not be constant throughout an individual’s life.

For example, innate preference for native prey by T. hammondii was shown to be

modified such that they learned to prefer nonnative prey (Hale, 2010). This confirmed the finding that wild-caught T. hammondii preferred nonnatives including centrarchids and an anuran, Xenopus laevis (Mullin et al., 2004). Bullfrogs were introduced to California in 1896 (Heard, 1904), which might have provided giant gartersnakes enough time to evolve to innately recognize them as alternative prey, especially because giant gartersnakes might have historically consumed a related species, the California red- legged frog (Rana draytonii) (Rossman et al., 1996).

Developing a preference for non-native prey might have a neutral or even positive effect on native predators. For example, bullfrogs can serve as prey (Rossman et al.,

1996) or predators (Wylie et al., 2003), which complicates their role as a positive or negative influence in giant gartersnake ecology. In one respect, their presence as prey might positively affect giant gartersnakes. This was the case for a threatened watersnake

(Nerodia sipedon insularum), which experienced an increase in body size and fecundity as a consequence of a new invasive prey source, the round gobie (Neogobius melanostomus) (King et al. 2006, 2008). As the most prolific invasive species living sympatrically with giant gartersnakes in Central Valley wetland habitats, bullfrogs can also negatively affect the giant gartersnake. Adult bullfrogs can prey on giant gartersnake neonates (Wylie et al., 2003), but how significant of a threat these animals are to giant gartersnake populations is not known. The presence and activities of bullfrogs also have negative community-level effects which have ultimately led to, among other things, the decline of native amphibians (Bury & Whelan, 1984; Adams & Pearl, 2007).

Olfactory trials showcased a moderate mean TFAS toward fish (which were all non-native), suggesting that neonates might view fish as suitable prey. However, the highest response among fish and crayfish was toward black bullhead. Bullheads are another non-native prey species that serve as a potential threat if consumed by the giant gartersnake. These fish are armed with dorsal and pectoral spines in their fins which can be erected as a defense mechanism (Forbes, 1989). Occasionally, gartersnakes are found with these spines protruding through their skin (Figure 1.3). Though some snake species, such as those within the genus Nerodia, can safely consume catfish (Plummer & Goy,

1984), giant gartersnakes have not coevolved with the catfish; thus their attempt to consume catfish might cause them injury, blockages and potentially death (USGS, unpublished observation).

In addition to directly affecting giant gartersnakes, non-native fishes and bullfrogs may potentially harm giant gartersnake populations through indirect means. For example, there is some evidence that bullfrog tadpoles negatively affect treefrog survival and size at metamorphosis (Kupferberg, 1997). Since the giant gartersnake consumes native treefrog adults and tadpoles, and treefrogs can be found co-occurring with bullfrog adults and tadpoles in both terrestrial and aquatic habitats, it stands to reason that bullfrogs might cause declines in treefrog populations, and perhaps reduce prey abundance for giant gartersnakes. On a related note, numerous studies have demonstrated overall adverse effects of nonnative fish on treefrog tadpoles (Adams 2000, Pearl et al. 2005,

Preston et al. 2012, Gilliland 2010). Additionally, nonnative fish have also been shown to facilitate bullfrog abundance (Adams et al. 2003). Due to the extensive stocking of

mosquitofish as mosquito control in sampling locations and the overwhelming abundance of nonnative fish and bullfrogs, these species likely have adverse and perhaps synergistic effects on treefrog tadpole success in the community of the giant gartersnake as well.

Although treefrog decline is not yet of regulatory concern in the Central Valley (Fisher &

Shaffer, 1996), treefrogs are sensitive to nitrates in agricultural runoff (Bishop et al.,

2010; De Jong Westman et al., 2010). Based on the findings in the current study, management to increase the abundance of treefrogs would likely benefit giant gartersnake populations.

Non-natives can also serve as vectors of parasites and/or disease. For example, bullfrogs, Gambusia spp. (Coyner et al., 2003), and centrarchid species such as Lepomis gibbous (Cone & Anderson, 1977) can serve as intermediate hosts of nematodes (i.e.

Eustrongylides spp.), and can transmit these parasites to their ophidian predators

(Jiménez-Ruiz et al., 2002). Many giant gartersnakes exhibit cysts of larval nematodes that form subcutaneously (Figure 1.2). Although effects of nematodes on giant gartersnake fitness are unknown, nematode infestations have been shown to cause contortion and mortality in a congener, Thamnophis sirtalis (Lichtenfels & Lavies, 1976).

In addition to observations in related species, several captive giant gartersnakes died in association with the presence of this parasitic infection (U.S. Fish and Wildlife Service,

1999).

The introductions of bullfrogs and non-native fish into the Central Valley of

California have multifaceted community effects. Thus, it is important to develop a full understanding of the relationship between these new species and the existing native species, especially those that are of conservation concern. Although in nature, the giant gartersnake has exhibited both habitat and dietary plasticity by persisting in rice agriculture and consuming these available non-native prey types, it remains important to understand the relationship between a threatened native predator and its prey in this complex modified system. The knowledge of giant gartersnake innate prey preference for a native amphibian contributes to the understanding of this predator-prey interaction (Sih

& Moore, 1990) and provides support for better conservation of both species.

Chapter 3 Giant gartersnake (Thamnophis gigas) prey selection

Introduction

Animals typically use resources in different proportions than their availability in the environment, and the consideration of availability is necessary to measure selection.

Prey selection, in particular has important consequences for understanding the relationships of animals with their environment and each other (Manly et al., 2002).

Predator-prey interactions have many factors that ultimately influence prey selection (Sih

& Moore, 1990). The introduction of non-native prey species can alter prey communities, prey availability, and therefore trophic dynamics. Documenting the selection or avoidance of specific prey, including non-natives, can help inform management tactics and therefore, conservation for rare species.

The giant gartersnake is currently listed as a threatened species at both the state and federal levels, having endured significant range reductions and fragmentation over the past several decades (Halstead et al., 2010). The introduction of several non-native prey species is another potential pressure that could alter the foraging ecology and ultimately the fitness of this snake. Despite its conservation status, no studies have examined the diet or prey selection of giant gartersnakes even though a changing prey base might contribute to the decline of this species.

The giant gartersnake diet generally consists of fish and anurans (Rossman et al.,

1996). Along with one native anuran, the Sierran treefrog (Pseudacris sierra), current prey available to giant gartersnakes include a non-native frog, the American bullfrog

(Lithobates catesbeianus), several non-native fish species including, but not limited to,

many species in the sunfish family (Centrachidae), such as green sunfish (Lepomis cyanellus), bluegill (Lepomis macrochirus), and largemouth bass (Micropterus salmoides); other non-native fish species, such as the Mississippi silverside (Menidia beryllina), western mosquitofish (Gambusia affinis), and black bullhead catfish

(Ameiurus melas); and several native and non-native species in the minnow family

(Cyprinidae) (UC ANR, 2015; McGinnis, 2006).

The objective of my study was to estimate the diet selection of giant gartersnakes.

In particular, I was interested in the extent to which giant gartersnakes select native prey, despite the numerical abundance of non-natives. Understanding the degree to which giant gartersnakes are selective of native and exotic prey will enable resource managers to better conserve this rare snake.

Materials and Methods

Field Methods

Sampling locations (Figure 3.1) for this study were divided into four areas in the

Sacramento Valley: the American, Butte, Colusa, and Sutter Basins. In the American

Basin, all sites were located within the Natomas Basin Preserve (a mitigation site with past and future wetland restoration) located in Sacramento and Sutter counties. In the

Butte Basin, trapping occurred on private property. In the Sutter Basin, trapping was conducted on privately owned rice farms in addition to Gilsizer Slough Giant Gartersnake

Conservation Complex (a mitigation site with preserved and restored wetlands) in Sutter

County. In the Colusa Basin, trapping was conducted on privately owned rice farms in addition to the Colusa National Wildlife Refuge, both in Colusa County. Detailed trapline

names, locations, trapping dates, and respective number of snakes captured are provided in Tables 3.1 and 3.2.

Figure 3.1 Map of trapline locations where giant gartersnakes were captured. Each cluster of traplines is grouped into Basin areas: Butte Basin (Butte County), Colusa

Basin (Colusa and northern Yolo counties), Sutter Basin (Sutter County), American

Basin (northeastern Sacramento and southern Sutter counties). Each dot represents a trapline where snakes were captured and sampled.

Table 3.1 Trapline name, habitat type, snake and prey counts and trapping dates.

Number Number Trapping Trapping of of Prey Start End Year Basin Trapline Habitat Snakes Items Date Date Bennett S American Wetland 1 1 Aug-13 Aug-13 2013 Wetland Bennett S American Wetland 1 1 Aug-14 Sep-14 2014 Wetland 2013 American BKS 3 Canal 3 4 May-13 Jun-13 2014 American BKS 3 Canal 4 6 Apr-14 Jun-14 2013 American BKS 4 Wetland 1 8 May-13 Jun-13 2013 American BKS 5 Wetland 2 1 May-13 Jun-13 2014 American BKS 5 Wetland 13 22 Apr-14 Jun-14 2014 American BKS 6 Wetland 5 7 Apr-14 Jun-14 2013 American BKS NE Wetland 2 7 Aug-13 Aug-13 2014 American BKS NE Wetland 1 1 Aug-14 Sep-14 BKS Pond American Wetland 1 1 Jul-14 Aug-14 2014 Q 2014 American Elsie Canal 2 5 Sep-14 Sep-14 Huffman American Canal 1 2 Jul-13 Aug-13 2013 West 2013 American LN 2 Canal 1 2 Apr-13 Jun-13 2014 American LN 2 Canal 3 3 Apr-14 Jul-14 2013 American LN 3 Wetland 4 4 Apr-13 Jun-13 2014 American LN 3 Wetland 1 1 Apr-14 Jul-14 LN American Wetland Wetland 1 1 Aug-14 Sep-14 2014 NE 2013 American LS 3 Wetland 1 1 Apr-13 Jul-13 2013 American NF 1 Wetland 2 2 Apr-13 Jul-13 2014 American NF 3 Wetland 1 1 Apr-14 Jun-14 2014 American Sills 2 Canal 1 1 May-14 Jul-14 2013 American Sills 3 Canal 1 1 Jun-13 Jul-13 2014 American Sills 3 Canal 1 1 Jun-14 Jul-14 2013 Butte Pritchard 3 Canal 1 1 Jul-13 Aug-13 2013 Butte Pritchard 4 Canal 1 1 Jul-13 Aug-13 2013 Butte Pritchard 5 Canal 1 1 Aug-13 Aug-13

Number Number Trapping Trapping of of Prey Start End Year Basin Trapline Habitat Snakes Items Date Date 2013 Colusa 24-11 Wetland 4 5 Jul-13 Aug-13 2013 Colusa 24-2 Wetland 2 2 Jul-13 Aug-13 2014 Colusa 24-7 Canal 1 1 Jul-14 Sep-14 Bransford Colusa Canal 4 10 May-14 Jun-14 2014 1 Bransford Colusa Canal 1 2 Jul-13 Jul-13 2013 2 Bransford Colusa Canal 3 4 May-14 Jun-14 2014 2 2014 Colusa Brennan 1 Canal 2 2 May-14 Jun-14 2014 Colusa Brennan 4 Canal 2 3 Jun-14 Jul-14 2014 Colusa Brennan 5 Canal 1 1 Jun-14 Aug-14 2014 Colusa Brennan 7 Canal 1 1 Aug-14 Sep-14 2013 Colusa East Pond Wetland 3 3 Jul-13 Aug-13 2014 Colusa East Pond Wetland 13 13 Jul-14 Sep-14 2013 Colusa GCID Canal 4 5 Jul-13 Aug-13 2014 Colusa Griffith 1 Canal 1 1 Jun-14 Jul-14 2013 Colusa J-Drain Canal 1 1 Jul-13 Aug-13 2013 Sutter BHTD Canal 8 10 May-13 Aug-13 2014 Sutter BHTD Canal 1 1 Jun-14 Aug-14 2014 Sutter Butler 1 Canal 4 5 May-14 Jun-14 2014 Sutter Butler 2 Canal 5 23 May-14 Jul-14 2014 Sutter Butler 3 Canal 1 1 Jun-14 Jul-14 2014 Sutter Cole 1 Canal 3 3 May-14 Jul-14 NCookies Sutter Wetland 1 1 May-13 Aug-13 2013 Wetland 2013 Sutter NTD Canal 2 1 May-13 Aug-13 2014 Sutter NTD Canal 1 1 Jun-14 Aug-14 2014 Sutter RGF 1 Canal 2 2 May-14 Jun-14 2014 Sutter RGF 2 Canal 4 5 May-14 Jun-14 2013 Sutter Wetland 2 Wetland 4 4 May-13 Aug-13 2014 Sutter Wetland 2 Wetland 1 1 Jun-14 Jul-14

Table 3.2 Raw counts of stomach contents from regurgitation of giant gartersnakes in 2014. Some individuals regurgitated multiple items.

2014 Basin Sums of Items by Prey Type American Sutter Colusa Type American bullfrog (metamorph) 7 6 13 26 American bullfrog (tadpole) 13 3 2 18 Sierran Treefrog 7 1 0 8 Mosquitofish 6 9 4 19 Centrarchids 6 1 2 9 Cyprinids 0 14 12 26 Ictalurids 0 1 1 2 Silverside 3 0 0 3 Sum of Items by Site 41 35 34 108 Sample Size (Number of Snakes) 36 20 26 82

Table 3.3 Raw counts of stomach contents from regurgitation of giant gartersnakes in 2013. Some individuals regurgitated multiple items.

2013 Basin Sums of Items by Prey Type American Sutter Colusa Butte Type American bullfrog (metamorph) 8 13 13 0 34 American bullfrog (tadpole) 8 2 0 2 12 Sierran Treefrog 2 0 0 1 3 Mosquitofish 3 2 4 0 9 Centrarchids 1 0 0 0 1 Cyprinids 0 0 0 0 0 Ictalurids 0 0 0 0 0 Silverside 1 0 0 0 1 Sum of Items by Site 23 17 17 3 60 Sample Size (Number of Snakes) 17 14 15 3 49

As part of routine protocols for occupancy and abundance surveys, snakes were captured in modified minnow traps (Figure 3.2), processed and then released. California

SCP # 10779, USFWS Recovery Permit # TEI57216-2, CSUEB Institute for Animal

Care and Use Protocol, and U.S. Geological Survey ACUC Protocol WERC-2014-01 were approved in advance. Trap modifications involved attaching Styrofoam so that the funnel of the trap was half above and half below the water surface, attaching hardware cloth extensions to increase funnel area and therefore trap efficacy, and attaching zip ties to the funnel hole to increase snake retention (Figure. 3.2; Halstead et al., 2013).

Figure 3.2 Traps used to catch giant gartersnakes

Giant garter snakes collected from traps were marked with unique microbrands and passive integrated transponder tags, and morphometric data were collected. In addition to these procedures, non-gravid adult snakes in healthy condition were gently palpated to encourage regurgitation for identification of stomach contents (Figure 3.3).

Palpation to encourage regurgitation is a well-established and commonly practiced technique that has no long term negative effect on the snake (Fauvel et al., 2012).

Figure 3.3 Giant gartersnake regurgitation of a bullfrog metamorph.

I measured prey availability by considering prey species that were captured as bycatch in the traps used to catch snakes as available prey. Specifically, by-catch was identified to species, genus, or family, counted, and released daily in every fifth trap of traplines typically containing 50 traps.

Analytical Methods

Stomach contents data from 131 individual snakes (Table 3.2 and 3.3) were analyzed using Bayesian hierarchical models with a Poisson distribution and a log link function in program JAGS 3.4.0 called through R 3.1.0 using package ‘runjags’.

Uninformative priors included a Uniform distribution (min = 0, max = 10) for the standard deviation describing variation among individuals and a Normal distribution

(mean = 0, standard deviation = 10) for the mean intercepts and coefficients for each variable. Several models were fit to see if specific effects influenced selection including a null model (no selection), constant selection, snake length, sex, habitat (wetland or canal), date (within year), and year (2013 and 2014). All models included random effects to account for differences among individuals in the number of available prey, the number of prey consumed, and in the case of models involving selection, individual variation in the selection of prey. Goodness-of-fit was tested with a Bayesian p-value using Pearson residuals and simulated data. The deviance information criterion (DIC) was compared to see which model best fit the data. The null model ran using 3 independent chains, a 9,000 iteration burn-in, plus 100,000 iterations after the burn-in.. The other models ran using 3 independent chains, a 9,000 iteration burn-in, plus 100,000 iterations after the burn-in.

The output was thinned by a factor of 10. Additional iterations were added 100,000 at a

time until the generated effective samples size for each measured parameter surpassed

10,000. We found no evidence for lack of convergence (Gelman and Rubin 1992); the

Gelman-Rubin diagnostic was less than 1.1 for all monitored parameters and history plots

were well-mixed.

Table 3.4 Prey selection model selection information

Model Mean Penalty Deviance Information Difference from Variant Deviance Criterion (DIC) Top Model Constant 4131.51 848.23 4980.33 Top Model Null 4563.76 750.96 5314.72 334.39 Sex 6960.01 878.07 7837.31 2856.98 Habitat 7011.01 847.97 7859.03 2878.70 Year 7020.00 867.12 7887.12 2906.79 Date 10099.44 836.95 10936.39 5956.06 Length 10082.15 861.57 10941.45 5961.12

Results

The constant selection model had the lowest DIC, meaning that the snakes are

selecting their diet independent of the variables tested (sex, habitat, year, date, and

length; Table 3.4). Although the giant gartersnake selection ratio was largest for treefrog

metamorphs (12.502 [0.00004 – 26.977] times more likely to choose a treefrog

metamorph over a bullfrog tadpole; Figure 3.4), the strength of selection for treefrog

metamorphs was very uncertain. Bullfrog adults were more likely to be selected than all

other prey except treefrog metamorphs (Fig. 3.4). Bullfrog tadpoles and cyprinids were

less likely to be selected than bullfrog adults, but more likely to be selected than

mosquitofish and centrarchids (Fig. 3.4).

treefrog metamorph

Figure 3.4 Giant gartersnake prey selection ratios (relative to bullfrog tadpole).

Different letters represent significant differences (95% credible interval of selection ratio does not contain 1).

Standardized selection ratios, which indicate the probability that the next prey item chosen would be of a particular type given equal availability, varied among prey items (Figure 3.5). Treefrog metamorphs (standardized selection ratio = 0.596 [0.0254 -

0.855]) and bullfrog adults (0.291 [0.008 - 0.448]) were significantly more likely to be selected than all other prey. Bullfrog tadpoles (0.061 [0.002 - 0.118]) were more likely to

be selected than mosquitofish (0.004 [0 - 0.010]), bullheads (0.024 [0 - 0.109]), and centrarchids (0.002 [0 - 0.005]), and cyprinids (0.046 [0 - 0.101]), also were more likely to be selected than mosquitofish and centrarchids. Silversides (0.026 [0 - 0.073]) only differed from treefrog metamorphs and bullfrog adults in their standardized selection ratio.

treefrog metamorph

Figure 3.5 Giant gartersnake standardized prey selection ratio. Different letters represent significant differences (95% credible interval of difference does not contain 0).

Discussion

Giant gartersnakes select anuran prey more than fish. Giant gartersnakes particularly selected juvenile and adult anurans. Given the description of giant gartersnakes as aquatic and their high affinity for water (refs.), this result is somewhat surprising. Large adults of some other aquatic snake species also predominantly consume frogs over fish (Santos et al., 2000; Mushinsky et al., 1982). More specifically, ontogenic diet shifts have been observed. Nerodia erythrogaster prey selection shifts from fish to frogs as snakes increase in size (Mushinsky et al., 1982). In a study examining the influence of a restricted diet on preference over time, Nerodia fasciata was found to have a strong preference for fish until the age of 6 months despite diet, after which diet was found to influence preference (Mushinsky & Lotz, 1980). The current study produced little evidence for the hypothesis of ontogenic dietary shift from fish to frogs in giant gartersnakes. However, due to the threatened status of this species, gravid females and particularly small snakes (< 12 grams) were not encouraged to regurgitate, thus restricting the sample size in those respective age class categories.

The fish identified in this study were assigned into functional groups (by family) because of difficult definitive species identification and similar life histories and thus, they cannot be absolutely discussed based on their indigenousness. Cyprinids were more likely to be selected than other fish groups. The selection of cyprinids relative to other fish groups might be caused, in part, by the historic presence of several native cyprinids in the Sacramento Valley. In addition to the verified (USGS observation) introduced cyprinid species captured in traps including, common carp (Cyprinus carpio), fathead

minnow (Pimephales promelas), red shiner (Cyprinella lutrensis), and golden shiner

(Notemigonus crysoleucas), the possible current native Cyprinid species occurring within giant gartersnake range include, but are not limited to, hitch (Lavinia exilicauda),

California roach (Hesperoleucus symmetricus), speckled dace (Rhinichthys osculus), hardhead (Mylopharodon conocephalus), Sacramento splittail (Pogonichthys macrolepidotus), and sacramento blackfish (Orthodon microlepidotus) (McGinnis, 2006;

UC ANR, 2015). Further investigation to identify native and non-native cyprinids in the giant gartersnake’s range would be valuable since cyprinids seems to hold some significance in the giant gartersnake diet composition.

Centrarchids and mosquitofish were least likely to be selected. There currently exists one native centrarchid, the Sacramento Perch (Archoglites interruptus; UC ANR,

2015) that once ranged throughout the Sacramento Valley waterways, but is now restricted to Clear Lake and the Sacramento-Dan Joaquin River Delta (McGinnis, 2006).

It has never been identified in USGS traps. Another native, the Tule Perch

(Hysterocarpus traski) has been confirmed in some locations sampled in this study, however it was included in the centrarchid family as it has a similar body style and life history and is often associated with centrarchids (UC ANR, 2015). In the consumption trials in the previous chapter, neonates were shown to have selected mosquitofish and centrarchids significantly less often when given the choice of both fish and anuran prey.

Additionally, when given a fish-only prey choice, although neonates significantly selected for centrarchids over mosquitofish, several neonates did not consume any prey at all within the 45 minute trial. Both of these studies provide evidence that centrarchids and

mosquitofish are likely not as important as anurans or Cyprinids within the giant gartersnake diet. This is somewhat enlightening as current management practices for giant gartersnake diet focus on fish availability (J. Roberts, The Natomas Basin

Conservancy, Personal Communication). Further, this practice is concerning since it has been shown that introduced fish can degrade amphibian communities (Adams, 2000;

Adams et al., 2003). This finding is important for improved management of giant gartersnake habitats.

The top model suggests that giant gartersnake prey selection was not influenced by any of the covariates tested. More specifically, I found that neither size, sex, habitat, date or year were influencing selection. However, similar studies on other natricine snakes found that both sex and size were factors that influence diet selection. Santos et al.

(2000) found prey partitioning where male snakes (Natrix maura) selected carp, females selected adult frogs, and juveniles selected tadpoles. These covariates might not have been revealed as influencing factors in the current study because of some potential limitations in the sampling techniques.

In addition to the aforementioned inability to sample all captured snakes, there are some ramifications within the trapping methods of prey. Anuran sampling might be biased toward more aquatic species (i.e. adult treefrogs are not as aquatic as adult bullfrogs). Snakes captured in traps might have eaten prey items that were captured as bycatch in the trap; thus diet information from snakes in traps might not be fully representative of giant gartersnake diet in general. More specifically, it might be that the giant gartersnakes sampled for this study had a higher prevalence of fish as stomach

contents because of a potential increase in ease of capture inside a trap and because so many fish were caught in the traps. Evidence of this discussed in the previous chapter where some neonates that were presented with a choice of only two fish species experienced such extreme difficulty catching a fish to the point where they did not consume either species.

In another highly aquatic snake species, Nerodia clarkia, the snakes have varying success in fish capture because of differences in mangrove density (Mullin & Mushinsky,

1995). Regurgitation of hand caught snakes would have been a useful comparison though this species is very wary (Wright & Wright, 1957; Fitch, 1940); therefore obtaining sufficient sample sizes by means of hand capture would have been very difficult. Despite these potential limitations, likelihood of selection for frogs indicated in the current study is consistent with the neonate consumption findings revealed in the previous chapter.

To be able to address the conservation needs of wildlife, the natural history of species of concern must be understood. This includes developing a comprehensive knowledge of the ecological roles these species occupy in their environment. Current management practices for giant gartersnakes focus on addressing elements of their habitat, specifically on the availability of water and vegetation. To date, little consideration has been given to prey communities, despite evidence that some prey species can be detrimental to giant gartersnakes. Management that increases the abundance of anurans, especially native anurans, will likely benefit giant gartersnakes.

Chapter 4 Thesis Discussion

This thesis is meant to fill a major gap in what is known about the basic natural history of the giant gartersnake, a threatened predator native to the Central Valley of

California. Past studies on giant gartersnakes have focused on several aspects on their ecology including habitat suitability (Halstead et al., 2010) and use (Halstead et al.,

2015), demography (Wylie et al., 2010) and population structure (Paquin et al., 2006), reproductive ecology (Halstead et al., 2011), body condition (Coates et al., 2009), and even thermoregulatory behavior (Wylie et al., 2009). Despite the information generated by these previous studies, it is clear that additional studies further clarifying the basic natural history and behavior of these animals is critical to fully understand the various factors with which they contend in nature and how these snakes may be affected by them.

This is especially true given that these snakes typically exist in modified habitats and with introduced species. The research presented in this thesis, an investigation into the dietary preference and selection of giant gartersnakes, is an examination of a poorly understood aspect of this species’ habits.

Overall results of the laboratory and field studies in this thesis provide evidence that giant gartersnakes have a preference for the Sierran treefrog, a native prey species, but will readily consume non-native bullfrogs and, to some extent, non-native fish.

Based on this study alone, it is unclear whether treefrogs are as important a prey source for adult giant gartersnakes as it seems they are for neonates. However, it should be considered that perhaps if treefrogs were more abundant, survival and recruitment of neonate giant gartersnakes into adulthood would be bolstered. Furthermore, although

bullfrog tadpoles are abundant within the Sacramento Valley, tadpoles that are small enough for neonates to consume might not be plentiful (USGS, unpublished observation) because of the fast growth rate of bullfrog tadpoles and their life history. Bullfrog tadpoles overwinter in their first year (i.e. they do not metamorphose until their second year); thus depending on the survival from a given year to the next, the majority of the composition of this stage might not be of suitable size to be considered feasible prey for giant gartersnake neonates.

On a related note, the study of locomotory performance among prey items would contribute to an even better understanding of giant gartersnake prey selection. It is presumed that tadpoles are easier to capture than fish. The trial testing neonates’ latency to attack and capture of treefrog metamorphs in a non-aquatic environment suggests that capture of metamorphs might be easier than exclusively aquatic prey. It stands to reason then, that treefrog metamorphs might also be easier to capture in water than tadpoles and fish, though this circumstance was not tested. Selection of bullfrog tadpoles in consumption trials, despite lower preference in preference trials, suggests that some other variable is contributing to selection. Investigation of prey locomotory performance might elucidate if ease of capture does indeed contribute to prey selection in this system.

Consideration to the design of the current studies can also be of value. The specific methodology of the prey preference trials was better described in this thesis than in the current literature. Cooper et al. (2000) described a procedure in which the observer would maintain a one centimeter distance between the swab and the snake’s supralabials and count tongue flicks directed at the swab until the snake “moved away.” In pilot studies, I

began to try to distinguish between direct versus indirect tongue flicks toward the cue, but this felt somewhat subjective. I decided to establish a five centimeter radius around the swab tip to objectively categorize if the neonate’s tongue flick was toward the swab and thus to the chemical cue. However, the decision to end the trials if the neonate tongue flicked at a distance of five centimeters or greater was somewhat arbitrary and that establishment could have been made with more consideration to qualities of volatility of the chemical cues in the experimental setup. Additionally, it is possible that for some of the chemical cues (e.g. potential predators: California kingsnake, bullfrog adult, and crayfish), moving away from the swab could be interpreted as evasive behavior and not just disinterest.

For consumption trials, the availability of certain prey items (e.g. treefrog and small bullfrog tadpoles) was limited because of acquisition and retention difficulties.

Therefore, individual prey items were reused in consecutive trials. Ideally, if available, prey items used in one trial should be replaced with new individuals. This would better control for individual prey becoming tired and stressed from the pursuit from neonates and potential limited availability of oxygen from remaining in a small dishpan with limited water for several hours. The addition of a cyprinid, or better, both a native and non-native cyprinid would have been insightful in this trial.

Some improvements in the field studies could include the collection of more specific information about bullfrogs. The size of bullfrog adults and tadpoles could have provided a more detailed understanding of how this species serves as prey (and predator) for giant gartersnakes. Additionally, identification to species within the cyprinid family

(native versus non-native) and of tule perch would have provided a more detailed understanding of the abundance and distribution of certain species and how they are selected for consumption by snakes.

The findings yielded by this study invite additional questions on which new studies might be built. For example, it would be of value to explore the response of neonates to the chemical cues of additional prey and predator species in olfactory trials and the addition of other prey in the consumption trials. In the prey preference trials, different prey species that were not included in the current studies but would be worth examining in future studies include tule perch, Sacramento blackfish, and other natives

(specifically cyprinids) that coexist with giant gartersnakes. However, when increasing the number of prey species tested, one also must increase the sample size of neonates or restrict the number of chemical cues for such a project. A smaller trial could conceivably be performed to see if a significant differential response among species within Cyprinidae and Centrarchidae (separate from their status as native or non-native) even exists.

Moreover, prey preference trials testing wild-caught and/or laboratory raised juveniles and adults would address the existence of a potential ontogenic diet shift and provide insight to the influence of experience into selection as was shown in Thamnophis hammondii (Mullin et al., 2004; Hale, 2010).

Lastly, parasitology in giant gartersnakes has not been extensively investigated. A recent collaboration with a veterinarian, Dr. Ray Wack from the Sacramento Zoo, involved health evaluations of live snakes and necropsies of dead snakes involved in

USGS research. They revealed a high load of several species of parasites (yet to be

identified) which likely contributed to the death of some snakes by means of decreased immunity. Further exploration of parasitology in giant gartersnakes will contribute to an understanding of survival in this snake and potentially how and if non-native prey species are playing a role in this situation.

Many factors contribute to prey selection in the giant gartersnake. The results of my studies provide important baseline information on giant gartersnake foraging ecology.

Because of the level of complexity of the sampled habitats caused by the introduction of non-native species, more refined investigations could provide answers to more detailed questions and inform best management practices for this threatened species.

References

Adams, M.J. (2000). Pond permanence and the effects of exotic vertebrates on anurans.

Ecological Applications, 10(2): 559-568.

Adams, M.J., Pearl, C.A., & Bury R.B. (2003). Indirect facilitation of an anuran invasion

by non-native fishes. Ecology Letters, 6(4): 343-351.

Adams, M.J. & Pearl, C.A. (2007). Problems and opportunities managing invasive

Bullfrogs: is there any hope? In Biological invaders in inland waters: Profiles,

distribution, and threats (pp. 679-693). Springer Netherlands.

Alfaro, M.E. (2002). Forward attack modes of aquatic feeding garter snakes. Functional

Ecology, 16(2): 204-215.

Arnold, S.J. (1977). Polymorphism and geographic variation in the feeding behavior of

the garter snake Thamnophis elegans. Science, 197(4304), 676-678.

Arnold, S.J. (1978). Some effects of early experience of feeding responses in the

common garter snake, Thamnophis sirtalis, Animal Behaviour, 26: 455-462.

Arnold. S.J. (1993). Foraging theory and prey size - predator size relation in snakes.

Snakes: Ecology and Behavior, 87-115.

Aubret, F., Burghardt, G.M., Maumelat, S., Bonnet, X., & Bradshaw, D. (2006). Feeding

preferences in 2 disjunct populations of tiger snakes, Notechis scutatus

(Elapidae). Behavioral Ecology, 17(5), 716-725.

Bishop, C.A., Ashpole, S.L., Edwards, A.M., Van Aggelen, G., & Elliott, J.E. (2010).

Hatching success and pesticide exposures in amphibians living in agricultural

habitats of the South Okanagan Valley, British Columbia, Canada (2004–

2006). Environmental toxicology and Chemistry, 29(7): 1593-1603.

Britt, E.J., Hicks, J.W., & Bennet, A.F. (2006). The energetic consequences of dietary

specialization in populations of the garter snake, Thamnophis elegans. The

Journal of Experimental Biology, 209: 3164-3169.

Bolker, B. (2008). Ecological Models and Data in R. Princeton University Press.

Bolker, B. (2015). emdbook: Ecological Models and Data in R; R package version 1.3.8.

Burger, J. (1991). Response to prey chemical cues by hatchling pine snakes (Pituophis

melanoleucus): effects of incubation temperature and experience. Journal of

Chemical Ecology, 17(6): 1069-1078.

Burghardt, G.M. (1966). Stimulus control of the prey attack response in naive garter

snakes. Psychonomic Science, 4(1): 37-38.

Burghardt, G.M. (1967). Chemical-cue preferences of inexperienced snakes: comparative

aspects. Science, 157(3789): 718-721.

Burghardt, G.M. (1968). Chemical preference studies on newborn snakes of three

sympatric species of Natrix, Copeia, 1968(4): 732-737.

Burghardt, G.M. (1969). Prey-Attack Studies in Newborn Snakes of the Genus

Thamnophis. Behaviour, 33(½): 77-114.

Burghardt, G.M. (1970). Intraspecific geographical variation in chemical food cue

preferences of newborn garter snakes (Thamnophis sirtalis). Behaviour, 36(3):

246-257.

Burghardt, G.M. (1971). Chemical-cue preferences of newborn snakes: influence of

prenatal maternal experience. Science: New Series, 171(3972): 921-923.

Burghardt, G.M. (1975). Chemical prey preference polymorphism in newborn garter

snakes Thamnophis sirtalis. Behaviour, 52 (¾): 202-225.

Bury, R.B., & Whelan, J.A. (1984). Ecology and management of the bullfrog. Smith VII.

Chiszar, D., Simonsen, L., Radcliffe, C., & Smith, H. M. (1979). Rate of tongue flicking

by cottonmouths (Agkistrodon piscivorus) during prolonged exposure to various

food odors, and strike-induced chemosensory searching by the cantil (Agkistrodon

bilineatus). Transactions of the Kansas Academy of Science, 82(1): 49-54.

Coates, P.S., Wylie, G.D., Halstead, B.J., & Casazza, M.L. (2009). Using time‐dependent

models to investigate body condition and growth rate of the giant

gartersnake. Journal of Zoology, 279(3): 285-293.

Cone D.K. & Anderson R.C. (1977). Parasites of pumpkinseed (Lepomis gibbosus L.)

from Ryan Lake, Algonquin, Ontario. Canadian Journal of Zoology, 1410-1423.

Cooper W.E. & Burghardt G.M. (1990). A comparative analysis of scoring methods for

chemical discrimination of prey by Squamate reptiles. Journal of Chemical

Ecology, 16(1): 45-65.

Cooper W.E., Burghardt G.M., & Brown W.S. (2000). Behavioural responses by

hatchling racers (Coluber constrictor) from two geographically distinct

populations to chemical stimuli from potential prey and predators. Amphibia-

Reptilia, 21(1): 103-115.

Coyner, D.F., Spalding, M.G., & Forrester, D.J. (2003). Epizootiology of Eustrongylides

idnotus in Florida: transmission and development in intermediate hosts. Journal

of Parasitology, 89(2): 290-298.

Czaplicki, J. (1975). Habituation of the chemically elicited prey-attack response in the

diamond-backed water snake, Natrix rhombifera rhombifera. Herpetologica,

31(4): 403-409.

Denwood, M. J. (In Review). runjags: An R package providing interfaceutilities, parallel

computing methods and additional distributions for MCMC models in JAGS.

Journal of Statistical Software. URL:http://runjags.sourceforge.net

De Jong Westman, A., Elliott, J., Cheng, K., van Aggelen, G., & Bishop, C. A. (2010).

Effects of environmentally relevant concentrations of endosulfan,

azinphosmethyl, and diazinon on Great Basin spadefoot (Spea intermontana) and

Pacific treefrog (Pseudacris regilla). Environmental Toxicology and

Chemistry, 29(7): 1604-1612. de Queiroz, A., Henke, C., & Smith, H. M. (2001). Geographic variation and ontogenetic

change in the diet of the Mexican Pacific lowlands garter snake, Thamnophis

validus. Copeia, 2001(4): 1034-1042.

Dix, M. W. (1968). Snake food preference: innate intraspecific geographic variation.

Science, 159(3822): 1478-1479.

Drummond, H. (1983). Aquatic foraging in garter snakes: a comparison of specialists and

generalists. Behaviour, 86(1): 1-30.

Drummond, H., & Burghardt, G.M. (1983). Geographic variation in the foraging

behavior of the garter snake, Thamnophis elegans. Behavioral Ecology and

Sociobiology, 12(1): 43-48.

Dunbar, G. L. (1979). Effects of early feeding experience on chemical preference of the

northern water snake, Natrix s. sipedon (Reptilia, Serpentes, Colubridae). Journal

of Herpetology, 13(2): 165-169.

Fauvel, T., Brischoux, F., Briand, M. J., & Bonnet, X. (2012). Do researchers impact

their study populations? Assessing the effect of field procedures in a long term

population monitoring of sea kraits. Amphibia-Reptilia, 33(3-4): 365-372.

Fisher, R.N., & Shaffer, H.B. (1996). The decline of amphibians in California’s Great

Central Valley. Conservation Biology, 10(5), 1387-1397.

Fitch, H.S. (1940). The biogeographical study of the ordinoides Artenkreis of garter

snakes (Genus Thamnophis). University of California Publications in Zoology,

44: 69-73.

Forbes, L.S. (1989). Prey defenses and predator handling behaviour: the dangerous prey

hypothesis. Oikos, 55(2): 155-158.

Futuyma, D.J. & Moreno, G., (1988). The evolution of ecological specialization. Annual

Review of Ecology and Systematics, 19: 207-233.

Gelman, A. & Rubin D. B. (1992). Inference from iterative simulation using multiple

sequences. Statistical Science, 7: 457-472.

Gilliland, K.E. (2010). The presence of Micropterus salmoides (Largemouth Bass)

influences the populations of Rana draytonii (California Red-legged frog) and

Pseudacris regilla (Pacific treefrog) in two ponds in Santa Barbara County,

California. Master's Theses and Project Reports, 244.

Gilmer, D.S., Miller, M.R., Bauer, R.D., & LeDonne, J.R. (1982). California's Central

Valley wintering waterfowl: concerns and challenges. US Fish & Wildlife

Publications, 41.

Gove, D., & Burghardt, G.M. (1975). Responses of ecologically dissimilar populations of

the water snake Natrix s. sipedon to chemical cues from prey. Journal of

Chemical Ecology, 1(1): 25-40.

Hale, K.B. (2010) Prey preferences as a function of feeding experience and prey type in

neonate gartersnakes (Colubridae: Thamnophis). Masters Theses & Publications.

Paper 61.

Halstead, B.J., Skalos, S.M., Wylie, G.D., & Casazza M.L. (2015). Terrestrial ecology of

semi-aquatic giant gartersnakes (Thamnophis gigas). Herpetological

Conservation and Biology. In Press.

Halstead, B.J., Wylie, G.D., & Casazza, M.L. (2010). Habitat suitability and conservation

of the giant gartersnake (Thamnophis gigas) in the Sacramento Valley of

California. Copeia, 2010(4): 591-599.

Halstead, B.J., Wylie, G.D., & Casazza, M.L. (2013). Efficacy of trap modifications for

increasing capture rates of aquatic snakes in floating aquatic funnel traps.

Herpetological Conservation and Biology, 8(1):65-74.

Halstead, B. J., Wylie, G.D., Casazza, M.L., & Coates, P.S. (2011). Temporal and

maternal effects on reproductive ecology of the Giant Gartersnake (Thamnophis

gigas). The Southwestern Naturalist, 56(1): 29-34.

Hansen, G.E. & Brode, J.M. (1980). Status of the Giant Garter Snake Thamnophis couchi

gigas (Fitch) State of California, The Resources Agency, Department of Fish and

Game.

Hansen, R.W. & Hansen, G.E. (1990). Thamnophis gigas (giant garter snake)

reproduction. Herpetological Review, 21(4): 93-94.

Heard, M. (1904). A California frog ranch. Out West, 21:.20-27.

Jiménez-Ruiz, F.A., García-Prieto, L., & Pérez-Ponce de León, G. (2002). Helminth

infracommunity structure of the sympatric garter snakes Thamnophis eques and

Thamnophis melanogaster from the mesa central of Mexico. Journal of

Parasitology, 88(3): 454-460.

King, R.B., Ray, J.M., & Stanford, K.M. (2006). Gorging on gobies: beneficial effects of

alien prey on a threatened vertebrate. Canadian Journal of Zoology, 84(1), 108-

115.

King, R.B., Stanford, K.M., Ray, J.M. (2008) Reproductive consequences of a changing

prey base in island watersnakes (Reptilia: Colubridae). South American Journal of

Herpetology, 3(2): 155-161.

Kupferberg, S. J. (1997). Bullfrog (Rana catesbeiana) invasion of a California river: role

of larval competition. Ecology, 78: 1736-1751.

Lawson, R. & Dessauer, H.C. (1979). Biochemical genetics and systematics of garter

snakes of the Thamnophis elegans-couchii-ordinoides complex. Louisiana State

University.

Lichtenfels, J.R. & Lavies, B. (1976). Mortality in red-sided garter snakes, Thamnophis

sirtalis parietalis, due to larval nematode, Eustrongylides sp.. Laboratory Animal

Science, 26(3): 465-467.

Manly, B.F.J., McDonald, L.L., Thomas, D.L., McDonald, T.L., & Erickson, W.P.

(2002). Resource selection by animals: statistical analysis and design for field

studies. Nordrecht, The Netherlands: Kluwer.

McGinnis, S.M. (2006). Field guide to freshwater fishes California (No.77). University

of California Press.

Mullin, S.J., Imbert, H., Fish, J.M., Ervin, E.L., & Fisher, R.N. (2004). Snake

(Colubridae: Thamnophis) predatory responses to chemical cues from native and

introduced prey species. The Southwestern Naturalist, 49(4): 449-456.

Mullin, S.J., & Mushinsky, H.R. (1995). Foraging ecology of the mangrove salt marsh

snake, Nerodia clarkii compressicauda: effects of vegetational density. Amphibia-

Reptilia, 16(2), 167-175.

Mushinsky, H.R., Hebrard, J.J., & Vodopich, D.S. (1982). Ontogeny of water snake

foraging ecology. Ecology, 1624-1629.

Mushinsky, H. R., & Lotz, K. H. (1980). Chemoreceptive responses of two sympatric

water snakes to extracts of commonly ingested prey species. Journal of Chemical

Ecology, 6(3): 523-535.

NatureServe (2013). Gila crassicauda. The IUCN Red List of Threatened Species.

Version 2015.2. . Downloaded on 23 July 2015.

Paquin, M. M., Wylie, G. D., & Routman, E. J. (2006). Population structure of the giant

garter snake, Thamnophis gigas. Conservation Genetics, 7(1): 25-36

Pearl C.A., Adams, M.J., Leuthold, N., & Bury, R.B. (2005) Amphibian occurrence and

aquatic invaders in a changing landscape: implications for wetland mitigation in

the Willamette Valley, Oregon, USA. Wetlands, 25(1):76-88.

Pearl C.A., Adams, M.J., Schuytema, G.S., & Nebeker, A.V. (2003) Behavioral

responses of anuran larvae to chemical cues of native and introduced predators in

the Pacific Northwestern United States, Journal of Herpetology, 37(3): 572-576.

Plummer, M. (2013). JAGS: A program for analysis of Bayesian graphical models using

Gibbs sampling, Proceedings of the 3rd International Workshop on Distributed

Statistical Computing (DSC 2003), March 20-22, Vienna, Austria. ISSN 1609-

395X. http://mcmc-jags.sourceforge.net

Plummer, M. (2014). rjags: Bayesian graphical models using MCMC. R package version

3-14. http://CRAN.R-project.org/package=rjags

Plummer, M., Best, N., Cowles, K., & Vines, K. (2006). CODA: Convergence Diagnosis

and Output Analysis for MCMC, R News, vol 6: 7-11.

Plummer, M.V., & Goy, J.M. (1984). Ontogenetic dietary shift of water snakes (Nerodia

rhombifera) in a fish hatchery. Copeia, 1984(2): 550-552.

Poncet, P. (2012). modeest: Mode Estimation. R package version 2.1. http://CRAN.R-

project.org/package=modeest

Preston, D.L., Henderson, J.S., & Johnson, P.T. (2012). Community ecology of

invasions: direct and indirect effects of multiple invasive species on aquatic

communities. Ecology, 93(6): 1254-1261.

R Core Team (2013). R: A language and environment for statistical computing. R

Foundation for Statistical Computing, Vienna, Austria. URL http://www.R-

project.org.

Rossman, D.A., Ford, N.B., & Seigel, R.A. (1996). The garter snakes: evolution and

ecology. University of Oklahoma Press.

Rossman, D.A. & Stewart, G.R. (1987). Taxonomic reevaluation of Thamnophis couchii

(Serpentes: Colubridae). Occ. Pap. Mus. Zool. Louisiana State Univ. 63: 1-25.

Santos, X., González-Solís, J., & Llorente, G.A. (2000). Variation in the diet of the

viperine snake Natrix maura in relation to prey availability. Ecography, 185-192.

Saviola, A.J., McKenzie, V.J., & Chiszar, D. (2012). Chemosensory responses to

chemical and visual stimuli in five species of colubrid snakes. Acta

Herpetologica, 7(1): 91-103.

Sih, A., & Moore, R.D. (1990). Interacting effects of predator and prey behavior in

determining diets. In Behavioural mechanisms of food selection (pp. 771-796).

Springer Berlin Heidelberg.

Sih, A., Stamps, J., Yang, L.H., McElreath, R., & Ramenofsky, M. (2010). Behavior as a

key component of integrative biology in a human-altered world. Integrative and

Comparative Biology, 50(6): 934-944.

Smith, K.P., Parker, M.R., & Bien, W.F. (2015). Behavioral variation in prey odor

responses in northern pine snake neonates and adults. Chemoecology, 5027(650):

1-10.

Stebbins, R.C. & McGinnis, S.M. (2012). Field guide to amphibians and reptiles of

California (Vol. 103). University of California Press.

Sturtz, S., Ligges, U., & Gelman, A. (2005). R2WinBUGS: A Package for Running

WinBUGS from R. Journal of Statistical Software, 12(3), 1-16.

Suarez, A.V., Richmond, J.Q., & Case, T.J. (2000). Prey selection in horned lizards

following the invasion of Argentine ants in southern California. Ecological

Applications, 10: 711-725.

Thomas, A. (2004). OpenBUGS. URL http://www.openbugs.net/.

Thomas, A., O Hara, B., LIgges, U., & Sturtz S. (2006). Making BUGS Open. R News 6:

12-17.

Tierney, L. Rossini, A. J., Li, N., & Sevcikova, H. (2013). snow: Simple Network of

Workstations. R package version 0.3-13. http://CRAN.R-

project.org/package=snow.

University of California, Division of Agriculture and Natural Resources. (2015).

California Fish Website. http://calfish.ucdavis.edu/species

U.S. Department of Agriculture, National Agriculture Statistics Service.

(2015). County Maps.

http://www.nass.usda.gov/Charts_and_Maps/Crops_County/index.php

U.S. Fish and Wildlife Service (1993). Endangered and threatened wildlife and plants:

determination of threatened status for the Giant Garter Snake. Federal Register

58:54053-54066.

U.S. Fish and Wildlife Service (1999). Draft Recovery Plan for the Giant Garter Snake

(Thamnophis gigas) U.S. Fish and Wildlife Service, Portland, Oregon. ix+ 192

pp. von Brand, T. (1944). Physiological observations upon a larval Eustrongylides: VI.

Transmission to various coldblooded intermediate hosts. Catholic University of

America.

Wassersug, R.J. (1989). Locomotion in amphibian larvae (or "Why aren't tadpoles built

like fishes?"). Integrative and Comparative Biology, 29(1): 65-84.

Weldon, P.J. (1982). Responses to ophiophagous snakes by snakes of the genus

Thamnophis. Copeia, 788-794.

Williams Jr, P.R., & Brisbin Jr, I.L. (1978). Responses of captive-reared Eastern

Kingsnakes (Lampropeltis getulus) to several prey odor stimuli. Herpetologica,

34(1): 79-83.

Wright, A.H., & Wright, A.A. (1957). Handbook of snakes of the United States and

Canada.

Wylie, G.D., Casazza, M.L., Gregory, C.J., & Halstead, B.J. (2010). Abundance and

sexual size dimorphism of the Giant Gartersnake (Thamnophis gigas) in the

Sacramento valley of California. Journal of Herpetology, 44(1): 94-103.

Wylie, G.D., Casazza, M.L., Halstead, B.J., & Gregory, C.J. (2009). Sex, season, and

time of day interact to affect body temperatures of the Giant Gartersnake. Journal

of Thermal Biology, 34(4): 183-189.

Wylie G.D., Casazza, M.L., & Carpenter, M. (2003). Diet of bullfrogs in relation to

predation of giant garter snakes at Colusa National Wildlife Refuge. California

Fish and Game, 89: 139-145.

Zedler, J.B. (1996). Ecological Issues in Wetland Mitigation: An Introduction to the

Forum. Ecological Applications, 6: 33-37.

American Sutter Colusa American Sutter Sutter American Ame Colusa Colusa Sutter American American Basin

Table of a Random Selection of TableSnakes’ of Prey Stomach and of a Content Sums Random Selection Available. ThatWere

rican

Date

5/14/2013 7/14/2013 8/22/2013 6/26/2013 5/15/2013 4/29/2014 5/25/2014 8/22/2014 7/11/2014 4/20/2014

6/7/2014 6/7/2014 6/7/2014

7/7/2014

2228 223 4108 2136 466 436 2160 2228 3123 3246 489 2148 2247 ID

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Regurgitated

Appendix

tadpole

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frog

13 98

1 3 2 3 0 1 1 1 0 0 0

morph