A wide range of environmental and physiological factors influence the type and extent of sexual dimorphism found in animals. Influential factors include variable climate, competition for resources and mates, mate choice, and parental investment. In addition, some investigators attribute differences in body size to physiological factors influenced by genetic and environmental variation. Since Darwin's time researchers studying the evolution of animal life-history have discussed the interactions that exist between this wide array of influences on sexual dimorphism in the context of natural selection. Much attention has been paid to the interrelationships of parental size, offspring size and number of offspring per reproductive bout. Spatial and temporal variation among these parameters have been investigated. However, these relationships have not been adequately examined while accounting for differences in offspring gender. In order to characterize the nature and extent of sexual size dimorphism found in adult red-spotted garter snakes, 108 adult females producing 782 female offspring and 790 male offspring were utilized to examine the relationships of the number and size of offspring, clutch mass and maternal size. The second facet of our investigation involved the hormonal manipulation of growth over the first year of post-natal life. To gain an understanding of which endogenous hormones may influence growth in this species we constructed a hormone profile of estradiol and testosterone from birth to 15 weeks of age. We experimentally tested the effects of exogenous hormone as well as the removal of endogenous hormone on growth with the use of estrogen and androgen antagonists. We have found that the extent of sexual size dimorphism observed in adult red-spotted garter snakes is not present at birth. The relative number of males or females born varies differentially with maternal size and age such that larger, older females produce predominately more male offspring. There are no differences in endogenous levels of testosterone and estradiol from birth though 15 weeks and at 36 weeks of age between the sexes. However, these sex steroids do differentially affect the growth of males and females. Finally, we suggest that while all of these factors contribute to sexual size dimorphism in this species, embryonic and environmental influences need to be explored. / Graduation date: 1999

Graduation date: 1994

Pheromones are chemical cues produced by organisms that affect the behavior and/or physiology of conspecifics. The orchestration of reproductive behaviors in many animals depends on the expression of sex pheromones. In insects, intraspecific variation in sex pheromone expression is commonly observed and often influences social interactions between individuals. To what extent similar variation is present in vertebrate sex pheromone systems is not well understood. This thesis investigated the occurrence and significance of phenotypic variation in the sexual attractiveness pheromone of the red-sided garter snake, Thamnophis sirtalis parietalis. Composed of a homologous series of saturated and unsaturated methyl ketones sequestered in the skin lipids of females, this pheromone elicits male courtship behaviors during the breeding season. Variation in the expression of the sexual attractiveness pheromone was examined at three levels: seasonal, individual and populational. Seasonal variation in pheromone expression was examined with respect to trailing behavior. In field experiments, males followed female trails during the breeding season but not during the non-breeding season. Skin lipid analysis revealed marked differences in pheromone composition between the two seasons, suggesting a role for this pheromone in regulating male trailing behavior. Individual variation in pheromone expression was examined with respect to mate choice. In arena trials, males displayed a courtship preference for larger females and continued to demonstrate this preference when visual, tactile, and behavioral cues from the females were removed through the use of skin lipid extracts. Analysis of pheromone profiles from individual females showed size-specific variation in composition, demonstrating that this pheromone can function as a reliable indicator of female size. Finally, populational variation in pheromone expression was examined with respect to sexual isolation among hibernacula. In arena and trailing experiments, males preferred to court and trail resident (same den) females over non-resident (distant den) females. Chemical analysis showed significant inter-den variation in pheromone composition, suggesting that this pheromone represents the cue that males use to identify females from their own population. In summary, these studies demonstrate that significant variation exists in the female sexual attractiveness pheromone of the red-sided garter snake and suggest that this variation has functional significance for this species. / Graduation date: 2002

Natural history is a cornerstone of both theoretical and applied ecology, and provides the context for comparative and experimental studies. I studied the natural history of the Plains Garter Snake (Thamnophis radix) near the northern limit of its range in Miquelon Lake Provincial Park. Alberta, Canada. As is typical of garter snakes, T. radix at Miquelon were sexually dimorphic as adults - females grew faster, reached larger sizes and had relatively shorter tails than males. Mating occurred in spring and females gave birth in August; litter size ranged from 8-50 and was correlated with size of female. These snakes were generalist predators, but fed mainly on anurans when small, adding mammals and other prey as they grew. Despite sexual size dimorphism, I found no difference in diet between adult males and females. Feeding frequency was low overall (22%). especially in gravid females. In summer, snakes were found mainly near cover on south-facing field edges or in marshes, but not in forests, and moved relatively little. However, movement distances increased in fall as snakes sought hibernating sites. Snakes hibernated for up to eight months, either individually or in pairs, in rodent burrows. This study also revealed the importance of radiotelemetry when studying movements of small, secretive animals, as there were significant differences between the results obtained through opportunistic captures versus telemetry: habitat descriptions from opportunistic captures therefore were biased, although they still captured some elements of microhabitat use by T. radix. The data from this study will be important for comparative studies of the ecology of garter snakes and for the management of this species on the human-altered landscape surrounding Miquelon Lake.

Canada is the second highest producer of hydroelectric energy in the world. Nearly 50 of the hydroelectric reservoirs in the country have a capacity larger than 1 billion m3. Despite the great number and extent of hydropower developments in Canada and around the world, relatively little is known about how dams and their operations influence terrestrial and semi-aquatic wildlife. Reservoirs at northern latitudes are characterized by large fluctuations in water level, which create modified shorelines called drawdown zones. To evaluate the impact of these disturbances on amphibians and reptiles, I conducted visual encounter surveys at two sites in the drawdown zone of Kinbasket Reservoir, near Valemount, B.C. From April to August of 2010 and 2011, I documented the habitat use, reproductive phenology, and body condition of two amphibian species (Anaxyrus boreas and Rana luteiventris) as well as the growth, movements, diet, and distribution of one species of garter snake (Thamnophis sirtalis). At two sites in the drawdown zone, A. boreas and R. luteiventris were present for the duration of the summer and utilized several ponds for reproduction. The presence and abundance of Rana luteiventris eggs were generally associated with ponds that had higher mean temperatures, higher mean pH, and the presence of fish. In 2010, there was sufficient time for amphibian breeding and metamorphosis to occur before the reservoir inundated the drawdown zone, but low precipitation levels in that year led to desiccation of many breeding ponds. In 2011, high rainfall and snowmelt led to early inundation of breeding ponds, and thousands of tadpoles were presumably swept into the reservoir. Gravid Thamnophis sirtalis were found at just one of two sites in the drawdown zone, but both sites were frequented by foraging individuals of this species. Anaxyrus boreas appears to be the primary prey of T. sirtalis in the drawdown zone. An improved understanding of how the amphibians and reptiles at Kinbasket Reservoir have persisted in this highly disturbed environment may be vital to their conservation — the activation of a new generating unit at Mica Dam in 2014 will alter the pattern and timing of reservoir inundation for the first time since it was constructed 40 years previously. / Graduate

Understanding the nature of adaptive evolution has been the recent focus of research detailing the genetic basis of adaptation and theoretical work describing the mechanics of adaptive evolution. Nevertheless, key questions regarding the process of adaptive evolution remain. Ultimately, a detailed description of the ecological context, evolutionary history, and genetic basis of adaptations is required to advance our understanding of adaptive evolution. To address some of the contemporary issues surrounding adaptive evolution, I examine phenotypic and genotypic changes in a snake feeding adaptation. Adaptations can arise through fixation of novel mutations or recruitment of existing variation. Some populations of the garter snakes Thamnophis sirtalis, T. couchii, and T. atratus possess elevated resistance to tetrodotoxin (TTX), the lethal toxin of their newt prey. I show that TTX resistance has evolved independently through amino acid changes at critical sites in a voltage-gated sodium channel protein (Nav1.4) targeted by TTX. Thus, adaptive evolution has occurred multiple times in garter snakes via de novo acquisition of beneficial mutations. Detailing the genetic basis of adaptive variation in natural populations is the first step towards understanding the tempo and mode of adaptive evolution. I evaluate the contribution of Nav1.4 alleles to TTX resistance in two garter snake species from central coastal California. Allelic variation in Nav1.4 explains 29% and 98% of the variation in TTX resistance in T. atratus and T. sirtalis, respectively, demonstrating that Nav1.4 is a major effect locus. The simple genetic architecture of TTX resistance in garter snakes may significantly impact the dynamics of trait change and coevolution. Patterns of convergent evolution are cited as some of the most compelling examples of the strength of natural selection in shaping organismal diversity. Yet repeated patterns may tell us as much about the constraints that restrict evolution as about the importance of natural selection. I present data on convergent molecular adaptations in parallel arms races between diverse snakes and amphibians from across the globe. Six snake species that prey on TTX bearing amphibians have independently acquired amino acid changes in Nav1.4. The derived mutations are clustered in two portions of the gene, often involving the same sites and substitutions. While a number of amino acid changes can make Nav1.4 insensitive to TTX, most of these negatively impact or abolish the ion-conducting function of the protein. Thus, intramolecular pleiotropy likely prevents most replacements from becoming fixed and imposes limits on protein evolution.

Projet de recherche réalisé avec Bernard Angers comme directeur de maîtrise, Denis Réale en tant que co-directeur et grâce à la collaboration active d'Emmanuel Milot. / La région métropolitaine de Montréal est formée de nombreuses îles à la jonction du fleuve Saint-Laurent et de la rivière des Outaouais, isolant ainsi les populations insulaires en fonction de distances respectives ainsi que des courants. Ce système offre un contexte idéal pour évaluer l’effet de la perte d’habitat liée à la pression d'urbanisation dans un paysage métropolitain insulaire ou en situation d’archipel. La présente étude a pour objectif de comparer l’effet de la perte d’habitat sur la diversité génétique de deux serpents très distincts, Storeria dekayi et Thamnophis sirtalis. Des analyses réalisées à l’aide de marqueurs microsatellites révèlent une plus importante structure génétique entre les populations de S. dekayi (FST=0,19) qu’entre celles de T. sirtalis (FST=0,07) dans la région montréalaise. Chez les deux espèces étudiées, la majorité des populations des habitats réduits présente une richesse allélique moyenne comparable à celle observée dans les habitats plus vastes. Néanmoins, certaines populations présentent des réponses différentes, dont des traces de goulots d’étranglement, une perte de richesse allélique ou encore une importante modification des fréquences alléliques. Au niveau régional, les résultats présentent une importante perte de diversité génétique chez les couleuvres se trouvant sur le continent alors que les populations insulaires de la région montréalaise constituent désormais un réservoir de diversité génétique. Les résultats observés auprès des populations insulaires démontrent que les effets de la perte d’habitat peuvent s’avérer très spécifiques à chaque situation et que la détection de traces génétiques d’un tel phénomène peut nécessiter un contexte logistique très particulier. Un nombre croissant de publications reportent une absence de signature génétique suite à la perte d’habitat chez des oiseaux et des mammifères. Il s’agit de la première étude témoignant de ce phénomène chez les reptiles. Une note est fournie en annexe à l’intention des gestionnaires au sujet de la conservation de la couleuvre brune, S. dekayi. / The Montreal metropolitan community includes numerous islands located at the confluence of the Saint-Lawrence and Ottawa Rivers. In such a fragmented landscape, dispersal of animals is limited by the distance between islands as well as the currents. This system offers an ideal context for the study of the effects of habitat loss on the genetic diversity of animal populations located on islands or archipelagos. This study seeks to assess the effects of habitat area by comparing the organization of genetic diversity of two highly distinct snake species, Storeria dekayi and Thamnophis sirtalis. Analysis realized with microsatellite markers reveals a much stronger genetic organisation in S. dekayi (FST=0.19) than in T. sirtalis (FST=0.07) in the Montreal area. For both studied species, most populations found in reduced habitats showed similar genetic diversity to what was observed in larger habitats. Nevertheless, some populations showed different responses to the loss of habitat, including traces of genetic bottlenecks, a loss in mean allelic richness or an important alteration of their allelic frequencies. This study also reveals an important loss of genetic diversity in the continental snake populations. At the regional scale, the results reveal an important loss of genetic diversity in the continental snake populations and that the insular populations of the Montreal area now constitute a reservoir of the remnant genetic diversity. Moreover, this study not only demonstrates that the genetic response to habitat loss can be very case-specific, but also that to detect traces of such a phenomenon can require a very particular framework. A growing number of publications based on birds and mammals have reported the absence of a genetic signature following a habitat loss. This is the first study to report this phenomenon in reptiles. A note intended for managers is provided about the conservation of the Dekay’s brown snake, S. dekayi.

Movements of animals presumably reflect their changing needs and the changing availability of necessary resources. In cold climates, snakes often make long seasonal migrations between hibernacula and summer habitats, Finding suitably deep hibernacula with minimal delay could be critical. I hypothesized that such animals should have highly developed navigational ability. By contrast, snakes living in mild climates can hibernate in shallow sites and probably do not migrate; if so, they might be expected to show poorly developed orientation mechanisms!. The objectives of this study were to determine movement patterns and navigational ability of garter snakes (Thamnophis) living in a mild climate and compare them with a congeneric population known to be migratory. From 1986 - 1988 I examined, using mark-recapture, movement behaviour of two populations of garter snakes at Spectacle Lake Provincial Park (SLPP) on Vancouver Island, British Columbia, focusing on several components typically associated with migratory behaviour - distances travelled, population directionality, fidelity to seasonal sites, distinctness of seasonal habitats, and route directness. Thamnophis sirtalis, the common garter snake, is the most widely distributed North American snake species and high latitude populations are migratory. Thamnophis ordinoides, the northwestern garter snake, is restricted to the Pacific northwest and migratory behaviour has never been reported. Both species displayed combinations of traits clearly suggesting nonmigratory behaviour. These included short-distance (< 500 m), random movements, a lack of den fidelity, and variation in the maintenance of specific home ranges between successive years. Home ranges overlapped between individuals, averaged less than 0.3 ha measured over a single active season, and were not clearly distinct from denning areas. Although some directionality of movement was evident, it was likely related to foraging strategy and unlike the typical unidirectional movements undertaken by migrating snakes, Sexual and reproductive differences in any of these traits generally were not significant. The navigational abilities of a migratory population of T. sirtalis from Wood Buffalo National Park (WBNP) in northern Alberta were examined as were those of the nonmigratory populations of snakes from SLPP. Displacement studies were carried out during the active seasons of 1986 - 1988 to determine the level of orientational abilities present in each population and to examine potential orientation cues. Snakes were displaced from their home range and tested in an arena under a variety of conditions, The results demonstrated that T. sirtalis from both SLPP and WBNP possessed advanced navigational abilities. Advanced skills may be absent in T. ordinoides. Thamnophis sirtalis at both study sites demonstrated time-compensated solar orientation as determined by 6 hr phase-delayed tests. Pheromone trails produced by recently copulated females (but not unmated females) also provided an orientation guide for displaced WBNP males, but results from SLPP were less conclusive. Thamnophis ordinoides did not respond in a discernible way to either cue. Navigational skills thus vary relatively little between migrating and nonmigrating populations of the same species but may be poorly developed in completely nonmigratory species. / Graduate

Postcopulatory sexual selection—sperm competition and cryptic female choice—has become a major area of research over the past 40 years. Within this field there are many outstanding questions at every level of analysis, from proximate to ultimate. The fitness consequences for both sexes in the period after copulation and before fertilization are considerable, but are obscured within the female reproductive tract. Our understanding of postcopulatory mechanisms is especially sparse in taxa other than birds and insects. Nearly nothing is known in reptiles except that multiple paternity is common and widespread, and often results from long-term sperm storage across breeding seasons. We present some of the very first data on the determinants of fertilization success in the context of sperm competition in reptiles, a group that accounts for 30% of terrestrial vertebrates. In the first chapter, "Asymmetric gametic isolation between two populations of red-sided garter snakes", we discuss the use of between-population crosses to reveal gametic isolation. The effect of population density and operational sex ratios on mating systems and the speciation process has fueled theoretical debate. We attempted to address these issues using two populations of red-sided garter snakes (Thamnophis sirtalis parietalis) from Manitoba, Canada. Our study populations differ markedly in their density mating aggregations, with a 10-fold difference between them. Using microsatellite markers for paternity analysis of litters produced from within and between population crosses. We found that the population with highest aggregation density, and presumably with the highest level of sexual conflict (i.e., when the evolutionary interests of the sexes differ) over mating, was also the population that exhibited homotypic sperm precedence. The less dense population showed a distinct postcopulatory male-size advantage. We also demonstrated that sperm stored within the female over hibernation can father 20-30% of offspring in a litter. In the second chapter, "Sperm competition and mate-order effects in red-sided garter snakes", we test whether females use mate-order effects to ensure that a larger (fitter) male will sire her offspring. Does that second male should have precedence in sperm competition? We tested for second-male precedence using singly-mated females that mated with a second male. Average proportion of paternity was shared equally among the first (P₁, i.e., proportion of offspring from a litter fathered by the first male to mate) and second males (P₂) to mate, and stored sperm (P[subscript ss]). This may be a case where last male precedence breaks down with more than two males. All females were spring virgins (they had not mated that spring, but may have stored sperm from fall matings); thus sperm stored presumably from fall matings is important in this system. As the interval between matings increased P₁ increased at the expense of P[subscript ss]. As the second male to mate's copulation duration increased, P₁ also increased at the expense of P₂. This last result may indicate female influence over sperm transfer during coerced matings. Copulatory plugs (CPs) are found in many taxa, but the functional significance is debated. Male garter snakes produce a gelatinous copulatory plug during mating that occludes the opening of the female reproductive tract for approximately two days. In chapter three, "Not just a chastity belt: the role of mating plugs in red-sided garter snakes revisited", we experimentally tested the role of the CPs. In snakes, sperm are produced in the testes and delivered through the ductus deferens, and the copulatory plug is thought to be produced by the sexual segment of the kidney and conveyed through the ureter. We manipulated the delivery of the two fluids separately by ligating the ducts. We confirmed that the CP is not formed in ureter-ligated males and that sperm leaks out immediately after copulation. The CP is analogous to a spermatophore. The protein matrix contains most of the sperm which are liberated as the plug dissolves within the female's vaginal pouch. One of the fundamental principles in sperm competition is that increased sperm numbers increase the odds of winning in competitions for fertilization success and males will adjust their ejaculate relative to competition and the quality of his mate. In chapter four, "Sperm depleted males and the unfortunate females who mate with them", we detect significant among-male variation in the number of sperm ejaculated, and that male mate-order reduces sperm numbers. Male sperm numbers drop significantly from one mating to the next, and this has implications for sperm competiveness, as Thamnophis sirtalis exhibits a disassociated reproductive tactic, in that sperm stores are produced outside the breeding season, and thus cannot be replenished after mating. Interestingly, however, the on average the mobility of the sperm increased for a male's second mating. Therefore, increased sperm quality may compensate for reduced numbers in a competitive context. Further, females increase their remating rate when mating with males that are unable to deliver sperm. In chapter five, "Sexual conflict during mating in red-sided garter snakes as evidenced by genital manipulation", we revisited the CP in the context of sexual conflict. Sex-differences in optimal copulation duration can be a source of conflict, as increased copulation duration may be advantageous for males as it delays female remating. Males of many species actively guard females to prevent them from remating, and in some cases males produce copulatory plugs to prevent remating. If precopulatory choice is limited at the time of her first mating, conflict may be especially onerous to a female. The size of the plug is influenced by the copulation duration. We experimentally tested the contribution of male and female control over copulation duration. We ablated the largest basal spine on the male's hemipene and found a reduction in copulation duration and an increase in the variation of plug mass. Further, we anesthetized the female's cloaca and found copulation duration increased, which suggests that males benefit from increased copulation duration while females actively try to reduce copulation duration. Therefore, sexual conflict is manifest in divergent copulation duration optima for males and females. / Graduation date: 2013