Sexual selection is pervasive in nature and plays an important role in the evolution of biological diversity both within and among sexual species. However, while we have a good understanding of how competition for reproductive opportunities and mate choice can drive the evolution of exaggerated secondary sexual traits, much less is known about how sexual selection interacts with other forms of natural selection and the consequences such interactions may have for adaptation to novel environments, the purging of deleterious mutations, and population divergence/speciation. In my thesis, I carried out a series of experiments with the fruit fly Drosophila melanogaster to test hypotheses regarding the operation of sexual selection and to broaden our understanding of how sexual selection may influence adaptation and diversification. Theory suggests that natural and sexual selection may align to promote adaptation and the purging of deleterious mutations, although the harm imposed by sexual conflict may counter this. In two separate experiments, I find no evidence that sexual selection promotes adaptation to a novel environment and, rather than aligning with natural selection, I find that the effects of sexual conflict may cause sexual selection to hamper the purging of deleterious mutations. With respect to diversification, sexual conflict has been suggested to be an important, non-ecological driver of population divergence. However, the traits involved in sexual conflict may also affect nonsexual fitness and natural selection may therefore act to constrain diversification. Using an evolution experiment, I demonstrate ecologically-dependent parallel evolution of traits involved in sexual conflict, providing evidence for ecology’s importance in divergence via sexual conflict. Overall, my work has shed light on the interaction of natural and sexual selection and the consequences this may have beyond the evolution of exaggerated sexual displays and armaments.

Eavesdropping occurs when a receiver extracts information from an interaction without directly engaging with the signaler. Eavesdropping has been shown to be an effective way of evaluating the quality of potential mates and their abilities in male-male competition, without having to directly interact with them, thereby reducing energy costs and mating harassment. Girardinus metallicus is a livebearing poeciliid fish endemic to Cuba whose mating system is dominated by mating harassment in the form of sneak copulations, persistent displaying, and male-male aggression. G. metallicus has a male specific polymorphism in both melanin coloration and behavior. Males with melanin coloration are known as black morphs. Black morph males show persistent displaying and higher aggression, whereas plain morph males, the most common morph, do not have pronounced melanin patches and mate solely by sneak copulation. Plain morph males exhibit lower levels of aggression than black morph males. In Chapter 1, we created groups consisting of a female and two males differing in body size and exposed them to dichotomous choice tests of female preference before and after the females witnessed the males interacting with each other. We hypothesized that: 1) male morphological traits are sexually selected via female choice in G. metallicus because these traits indicate quality; 2) female G. metallicus eavesdrop on male aggression to make mate choice decisions because aggression may indicate the quality of the male and his propensity to harass females; and 3) male size classes differ in behavior and morphology (saturation, brightness, and gonopodium size), consistent with other poeciliid studies showing that body size influences phenotype and that these traits are intercorrelated. We predicted 1) females will associate more with more colorful males, males with shorter gonopodia, and the larger male, before eavesdropping on male-male interactions, after, or both; 2) females will spend more time associating with males that subsequently delivered more chases and bites to a competitor male; and 3) larger males would be more active, more persistent in mating attempts, be more aggressive, and have a larger gonopodium size, and greater saturation and brightness of their posterior, ventral, and dorsal body regions. We found that females prefer to associate with males whose body regions are highly saturated, before eavesdropping on the two males interacting, but females did not prefer saturation after eavesdropping. We also found that females had a preference for smaller gonopodia relative for a males’ body size after eavesdropping. We also found that as male size increases, gonopodium length is proportional to their standard length. This study is the first to show female preference for coloration traits within any morph of G. metallicus, suggesting that plain morph males are not as plain to females as their name suggests. Individual animals consistently vary in the average level of behavior exhibited across a range of contexts, which is also known as personality. Behavioral syndromes are correlations among personalities grouping them together. Personality traits have implications for mate choice, fitness, and predator avoidance. In Chapter 2, we addressed behavioral traits and personality in females and whether they influence how males respond to different degrees of boldness, activity, and aggression. We assessed female latency time to emerge from a refuge chamber, activity level, and aggressiveness to another female, on three successive days. We then quantified the degree of mating harassment each female experienced, when tested with a male. We hypothesized the following: 1) female G. metallicus exhibit personality across behavioral contexts (risk, activity, and resource competition), consistent with findings in other poeciliids including male G. metallicus; 2) the rank orders of boldness, activity, and aggression are positively correlated, consistent with other poeciliid studies that found evidence for behavioral syndromes; and 3) female personality traits mitigate male harassment because females that exhibit those personality traits are better at avoiding/retaliating against male harassment. We predicted that: 1) behavioral traits (latency time to emerge, boxes entered, and chases, bites, and fin flares delivered to a female competitor) measured within each context would be repeatable; 2) female rank orders of boldness, activity, and aggression personalities would be positively correlated with each other; 3) that larger females would experience less harassment; and 4) when females directly interact with a male, females that are bolder, more active, and more aggressive (bites and chases delivered to the male) are better able to mitigate male harassment. We found support for the hypothesis that some behavioral traits are repeatable in females; however, we found no evidence for behavioral syndromes. We found evidence to suggest that females that are less bold and less aggressive received less mating harassment from males, possibly because those females are of lower quality and not as attractive to males. Our most novel finding in this study was that activity and aggression were both consistent behavioral traits in females, and therefore constitute personalities; however, these personalities did not have a correlation grouping them together into a behavioral syndrome. Maybe there is a tradeoff: good females are bold and aggressive and get more food, but receive more mating harassment, whereas bad females are submissive and get less food, but avoid mating harassment.

In this thesis, I explore four different ways in which relatedness affects sexual interactions in the red junglefowl Gallus gallus ssp., and the fruit fly Drosophila melanogaster. First, I show that in both species, inbreeding depression is sex-specific and modulated by parental age and gametic age. However, the sex that suffers higher inbreeding depression was trait- and species-dependent. Second, I examined patterns of inbreeding avoidance. I found no evidence of inbreeding avoidance in the fruit fly, but in the red junglefowl both males and females avoided mating with relatives, independently from sex-ratio of the social group. Third, I investigated whether relatedness amongst members of one sex affects mate choice in members of the opposite sex. Male fruit flies preferentially courted females unrelated to females with whom they had previously mated, while female flies displayed a weak preference for males related to their previous mates. In the red junglefowl, females exposed to male trios of two males related to each other and one unrelated male, displayed a marked preference for mating with the male unrelated to the other two males, and might also bias postcopulatory sperm utilization in favour of the unrelated male. Fourth, I explored the implications of male relatedness on the intensity of male-male competition. Male red junglefowl were less aggressive towards related competitors, but invested more sperm in females that had previously mated with a related male rather than with an unrelated male. In fruit flies, male relatedness had a strong impact on female life-history and offspring viability, although I found no evidence that these effects were modulated by changes in male-male competition. Collectively, the findings of these studies demonstrate the complex relationship between relatedness and other important biological phenomena as such senescence and sexual conflict.

Sexual conflict occurs when the fitness interests of males and females do not align with one another. The resolution of sexual conflict often depends on the level of control each sex has on the behavior in conflict. In Pogonomyrmex harvester ants with a genetically determined caste system, two separate lineages interbreed with one another during summer mating swarms. Diploid offspring sired by a single lineage develop into reproductive queens, while offspring sired by opposite-lineage parents develop into sterile workers. This results in sexual conflict, as males which mate with opposite lineage queens will produce only workers, resulting in no fitness benefit, while queens must mate with opposite-lineage males in order to obtain workers and survive. Despite these fitness differences, males do not discriminate between lineages prior to mating. One possible reason for the lack of male discrimination is that queens "mask" their identity cues, making discrimination difficult for males. In eusocial insects, identity cues are encoded by cuticular hydrocarbons (CHC’s) found on the exoskeleton of the insects. These cues contain information on the insect's reproductive status, sex, species, state, and nest membership. In addition to their communication functions, CHC’s also serve as desiccation-resistance molecules, preventing water from freely passing out of the cuticle. However, molecules that are best-suited for communication functions are poor desiccation resistance molecules, and molecules that are best-suited for waterproofing lack the diversity needed for communication; therefore, a tradeoff between these two functions is expected. In this dissertation, I explore sexual conflict in these ants and the chemical recognition cues that likely play a role in this conflict. To test for cryptic strategies in harvester ant mating swarms, I experimentally paired males and females from two interbreeding lineages of harvester ant with different fitness outcomes based on pairing, and measured the propensity to initiate copulation, pre-copulatory time, time in copula, and rate and amount of sperm transferred in each mating. Although females controlled copulation duration, males altered sperm transfer rates, resulting in no quantitative difference in total sperm transfer between lineages. To test for thermal constraints on the diversity and composition of cuticular hydrocarbon profiles, and changes in CHC profiles that occur in workers isolated from the queen, I surveyed the cuticular hydrocarbon profiles of a species complex of harvester ants. The CHC profiles of ants from more xeric environments showed evidence of constraints, while isolated workers differentiated from their queen-raised sisters, although not in queen-specific molecules. To test for queen identity masking and lack of discrimination ability in mating swarms, I tested for convergence in the CHC profiles of reproductives in two hybridizing lineages in response to the sexual conflict playing out in this species. Differences in CHC profiles were lost during the mating swarm, likely limiting male ability to discriminate between mates, limiting discrimination ability in mating swarms. To study the genetic regions that control CHC production, I created a physical linkage map of two of the interbreeding populations, and used that map to perform quantitative trait loci analysis on the cuticular hydrocarbon profile of recombinant males. One significant region associated with 13-methylnonacosane contained numerous odorant receptor genes, suggesting a link between that CHC production and the receptors that detect it, while a second region associated with n-pentacosane contained numerous genes that control expression levels. Overall, the genetic caste determination system in these ants leads to antagonistic coevolution between species. This coevolution is likely reinforced by the thermal constraints and exchange of recognition cues between species, lowering the ability of useful discrimination between lineages during mating swarms.

La compréhension des processus microévolutifs ayant lieu dans la nature nécessite la quantification des principales forces sélectives s'exerçant sur les populations sauvages. Ces 10 dernières années, les études à long terme et l'écologie de la résurrection (qui fait revivre des stades en dormance) ont été les principales approches pour étudier l'évolution des traits d'histoire de vie sur plusieurs générations dans les populations sauvages. Mon travail consiste à comprendre comment des facteurs écologies simples (p. ex. la température) et des interactions interspécifiques ou intraspécifiques plus complexes (p. ex. les interactions antagonistes hôte-parasite ou mâle-femelle) façonnent le processus évolutif des populations sauvages. Dans cette optique, j'ai utilisé l'Artémia comme un organisme modèle, en combinant des études sur le terrain et en laboratoire. Premièrement, j'ai étudié l'évolution de la niche thermique avec une approche d'écologie de la résurrection, en utilisant une série temporelle d'œufs de dormance d'une population d' introduite à partir de marais salants de régions tempérées dans des marais salants tropicaux dans les années 80. Cette étude montre que la survie aux températures élevées (caractéristiques du nouvel environnement) a augmenté linéairement au cours du temps à partir de l'introduction, suggérant un taux d'adaptation constant sur plus de 100 générations. Deuxièmement, j'ai utilisé une approche similaire pour étudier l'adaptation entre males et femelles dans une autre population d'Artémia. Cette étude suggère que les conflits sexuels provoquent une dynamique de coévolution fluctuante dans la nature sur une échelle d'environ 100 générations. Troisièmement, j'ai étudié les impacts respectifs de différents parasites (une espèce de cestode et deux espèces de microsporidie) sur la compétition entre une espèce d'hôte autochtone asexuée et une espèce d'hôte invasive sexuée. Chacun des trois parasites étaient soit spécialiste d'une espèce ou de certains génotypes d'hôte. De plus, l'espèce de cestode dont l'effet castrateur chez l'hôte est bien connu infectait uniquement l'espèce autochtone, suggérant que ce parasite joue un rôle majeur dans la compétition entre les espèces d'hôte autochtones et invasives. Par ailleurs, les trois espèces de parasite semblaient manipuler le comportement d'agrégation de leur hôte, très probablement pour augmenter leur transmission à de nouveaux hôtes. Enfin, j'ai réalisé des études de génétiques des populations d'espèces asexuées diploïdes et polyploïdes d'Artemia et d'espèces sexuées asiatiques proches. Les espèces asexuées diploïdes produisent des mâles rares et il semblerait que ceux-ci permettent une faible fréquence de reproduction sexuée. De plus, l'hybridation d'espèces d'Artémia éloignée phylogénétiquement a donné naissance à au moins trois lignées polyploïdes indépendantes. / Getting a comprehensive understanding of microevolution in natural populations requires proper quantification of the important selective forces exerted on these populations. Over the last decade, long-term studies and resurrection-ecology (revival of resting stages) have been the main approaches to study life history trait evolution over many generations in the wild. My work aims at understanding how simple ecological factors (e.g. temperature) and complex interactions between and within species (host-parasite and male-females antagonistic interactions) shape evolutionary processes in natural populations. To this end, I used the brine shrimp Artemia as a model system and combined laboratory and field studies. First, I investigated thermal niche evolution with a resurrection ecology approach, using dormant-egg time series from an Artemia population introduced from temperate to tropical salterns in the mid-80's. This experiment shows that survival at the high temperatures typical of the new environment increased linearly through time after the introduction, suggesting a sustained rate of adaptation over more than 100 generations. Second, I used the same approach to study adaptation between sexes in another Artemia population. I found that sexual conflicts result in fluctuating male-female coevolutionary dynamics in natura, over a time scale of ~100 generations. Third, I studied the relative role of one cestode and two microsporidian parasites in mediating the competition between a native asexual host and an invasive bisexual host. I found that all three parasites were either host- or genotype-specific and that the castrating cestode parasite specifically infected the native species, suggesting that this parasite actually played a major role in the competition between native and invasive hosts. Interestingly, all three parasites manipulated the swarming behavior of their host, most likely to increase their transmission. Fourth, I performed population genetic studies of diploid and polyploid Artemia parthenogenetica and their Asian bisexual close relatives. Diploid asexual Artemia produce rare males and I found indication that these males allow some rare sex in this otherwise parthenogenetic species. In addition, hybridization between divergent Artemia species has led to the origin of at least three independent polyploid lineages.

Sexual conflict, the different interests of males and females over reproduction, is a potent evolutionary force. Here I investigate sexual conflict in the context of parental care by focussing on two questions: (i) which behavioural, morphological and environmental traits influence the parents’ decision to care for the brood or desert? (ii) How does sexual conflict influence the evolution of behaviour and morphology? I investigate both questions using a small, polygamous passerine bird, the Eurasian penduline tit Remiz pendulinus, which exhibits intense sexual conflict over parental care such that either the male, the female or both parents desert the clutch. Using detailed behavioural observations during the crucial few days preceding desertion, I argue that it may be in the parents’ best interest to conceal their intention to care for (or desert) their brood. The rapid resulting process that leads to variable parental care resembles a coordination game in which either parent may desert first. I developed a game-theoretical model that suggests that a key to resolving the conflict between parents is the sex difference in reproductive payoffs for given parental care strategies, rather than differences in parental quality per se. Environmental variables (e.g. food availability and mating opportunities) seem only subsidiary in the decision-making process of parents. My final chapter explores ramifications of sexual conflict at an evolutionary timescale. By comparing the highly polygamous Eurasian penduline tit with the monogamous Cape penduline tit, I show that morphological and behavioural differences between these two species are consistent with predictions of sexual conflict theory. During my PhD I also identified that there is considerable variation in breeding systems within different species of penduline tits. I argue that by studying these systems new insights will emerge into (i) the drivers of breeding systems, and (ii) neural and genomic traits that underlie breeding system evolution.

Sexual conflict has been proposed to be important for evolution, and is often implicated in population divergence and speciation through sexually antagonistic co-evolution (SAC). However, empirical tests of these ideas on field populations are few. How sexual conflict, and SAC, operates in the wild, remains an important unanswered question if we are to fully understand the role of sexual conflict in evolution in nature. Here, I studied sexual conflict over mating in the seed feeding bugs Lygaeus equestris and Lygaeus simulans. Firstly, I show that laboratory adapted populations of L. equestris that differ in the magnitude of sexual conflict also differ in aspects of their reproductive development and mating propensity, with the population displaying greater conflict load also mating more readily. Study of female receptivity to mating as an evolvable trait, that could be involved in conflict over mating, revealed moderate to low heritability at two age groups. To better understand variation in the expression of sexual conflict in the wild, field caught populations of L. equestris, ranging across its distribution, and also of its sister species, L. simulans, were assayed for the magnitude of sexual conflict over mating in common garden laboratory experiments. High female mating costs were apparent across the populations, but the magnitude of these costs did not vary. No consistent patterns of mating costs and life history variation were found however, arguing against close links between mating costs and life-history. Finally, I investigated whether populations displaying sexual conflict over mating have begun to diverge, and evolve reproductive isolation. I found no evidence of reproductive isolation, or variation in mating propensity, between populations of L. equestris when crossed in reciprocal no-choice mating trials. However, L. equestris and L. simulans did show pre-zygotic reproductive isolation albeit with asymmetries between the reciprocal crosses (L. simulans males were able to mate L. equestris females, but male L. equestris were largely unable to mate L. simulans females). As expected for close taxa that perhaps have not been diverged for long, pre-zygotic isolation was perhaps stronger than post-zygotic isolation, as F2 offspring were generated by some of the inter-specific crosses: gene flow can therefore occur between these species contrary to previous studies. My data suggest that sexual conflict over mating may reduce the likelihood of speciation through the evolution of male persistence, as well as promote it through population divergence.

Sexual conflict over mating duration drives the evolution of male and female adaptations that facilitate the manipulation of mating interactions in their favor. This conflict drives the evolution of traits that improve the fitness of the focal sex despite inflicting costs on mates. However, males can express multiple traits that increase and decrease female fitness simultaneously. When the effects of male traits on female fitness increase or decrease with duration of exposure, females traits that influence mating duration are selected upon. Females of Callosobruchus maculatus, a bruchid bean beetle, kick mates to forcefully end copulation. Although both negative effects of male genital spines and positive of effects ejaculatory materials on female fitness have been documented, it is not yet clear how these male traits interact to influence the timing of female kicking. In this study, we observed the effect of male genital spine size, ejaculate size and mating history, and manipulated mating duration to disentangle the effects of male traits on the timing of female kicking behavior. We found that male mating history and mate body size dimorphism predicted the timing and duration of female kicking, but that male ejaculate size and spine length did not predict female kicking timing.

Phenotypic differences between the sexes evolve largely because selection favours a different complement of traits in either sex. Theory suggests that, despite its frequency, sexual dimorphism should be generally constrained from evolving because the sexes share much of their genome. While selection can lead to adaptation in one sex, correlated responses to selection can be maladaptive in the other. In this thesis I use Drosophila to examine the extent to which the shared genome constrains the evolution of sexual dimorphism and whether the sex chromosomes might play a special role in resolving intralocus sexual conflict. Gene expression data shows that intersexual genetic correlations are generally high, suggesting that genes often affect both sexes. The intersexual genetic correlation is negatively associated with sex-bias in expression in D. melanogaster, and the rate of change in sex-bias between D. melanogaster and six closely related species, showing that a sex-specific genetic architecture is a prerequisite for the evolution of sex difference. In further studies I find that genetic variance affecting lifespan is found in the male-limited Y chromosome within a population, which could offer a route to the evolution of further sexual dimorphism in lifespan, though the amount of variance was small suggesting adaptive potential from standing genetic variance is limited. Genetic variance on the X chromosome is also expected to be depleted once the sex chromosomes evolve, but here I find no evidence of depletion in either sex. Dosage compensation does not appear to double the male X-linked genetic variance, but this effect may be complex to detect. Finally, the X chromosome appears to be enriched for sex-specific genetic variance, and the consequences of this are explored using a variety of analytical methods to test biologically meaningful aspects of G-matrix structure. In summary, this thesis suggests that the evolution of sexual dimorphism is generally constrained by the shared genome, but intralocus sexual conflict could be resolved by novel mutations on the Y chromosomes, and by standing sex-specific genetic variance on the X chromosome. It highlights a special role for the X chromosome in the evolution of sexual dimorphism.

Reproductive traits evolve rapidly at the morphological, physiological and molecular levels, a taxonomically robust pattern that is thought to arise from sexual selection. In internally fertilizing organisms, female promiscuity results in competition between multiple male ejaculates for fertilizations in the same female reproductive tract, extending sexual selection past courtship and copulation. In this post-copulatory arena, biochemical interaction between male ejaculates and female reproductive tracts form a dynamic molecular interface that modulates female post-mating responses essential to reproductive fitness. Consistent with the hypothesis that these interactions are subject to sexual selection, sperm and seminal proteins are known to evolve rapidly in a broad range of taxa. The female role in this process, however, in terms of both molecular mechanisms and evolutionary dynamics, remains unclear.The presented dissertation research examines the biochemical nature and evolutionary consequences of post-copulatory sexual selection in two sister-species of cactophilic Drosophila, D. mojavensis and D. arizonae. I first present data that female post-mating response in crosses between these to species is perturbed, severely reducing the reproductive output of heterospecific crosses. A breakdown of reproductive tract interactions in matings between divergent lineages suggests that male and female contributions to reproductive outcomes are coadapted. Next, I use a combination of bioinformatic analyses, comparative sequence analyses, and biochemical assays to elucidate candidate female reproductive tract proteins that may be involved in ejaculate-female dynamics. 241 candidate female reproductive proteins are identified, the most intriguing of which are recently-duplicated secreted proteases. Finally, I explore the evolutionary history of two families of secreted proteases within geographically isolated populations of D. mojavensis. I show that both families evolve rapidly through a complex process involving gene duplication, gene conversion, pseudogenation and positive selection, a unique pattern never before documented in reproductive proteins.Collectively, my dissertation research suggests that females are active participants in the evolution of reproductive tract interactions. Further exploration of how sexual reproduction coevolves between males and females, both in terms of interacting biomolecules, and dynamic evolutionary histories, remains an important challenge for future research.