Stuart Wigby

Seminal fluid proteins (SFPs) play crucial roles in reproduction by shaping female post‐mating physiology and behaviour, male sperm competition and sexual conflict between the sexes. However, we know little about how abiotic ecological factors regulate SFP expression and transfer. Here, we use quantitative proteomics in adult flies from a wild Drosophila melanogaster population to test how exposure to natural temperature variation (20°C, 24°C and 28°C) across two time scales (48 h and 13d), and under either no‐ or high‐sperm competition risk, interact to shape SFP production and transfer. Short‐term exposure to different temperatures resulted in lower SFP production and transfer at 24°C under no sperm competition risk, but no differences under high sperm competition risk. In contrast, both the production and transfer of SFPs were consistently reduced across sperm competition treatments after long‐term (13d) exposure to low (20°C) and high (28°C) temperatures, including for key proteins in the sex peptide and ovulin networks that mediate female post‐copulatory responses, sperm competition and sexual conflict. Our results show that natural temperature fluctuations can have a strong effect on SFP production and transfer, and thus on post‐copulatory sexual selection dynamics.

Whether sexual selection facilitates or hinders adaptation to novel environments is a question of fundamental importance due to current rapid environmental change. Using phylogenetic comparative analysis in 42 Drosophila species, we found that sperm length – a sexually selected ornament – positively associates with the ability of males to survive high temperatures. However, the ability to remain fertile does not increase with sperm length to the same extent, meaning that longer sperm species experience a larger gap between temperatures that kill and temperatures that sterilize males. The findings are consistent with the idea that strong sexual selection on condition-dependent exaggerated traits may facilitate somatic resilience to environmental stressors, but that susceptibility to heat-induced infertility may be a cost of extreme sperm ornamentation. Overall, the data suggests multifaceted links between postcopulatory sexual selection and climate change.

When females mate with multiple males, ejaculates from different partners compete within the female reproductive tract. Males gain a competitive advantage by transferring seminal fluid proteins (Sfps) that manipulate female post-mating responses, such as reducing female propensity to remate. But in monandrous species, where females typically mate only once, post-mating sexual selection is absent, raising questions about the complexity and identity of Sfps in monandrous seminal proteomes. To address this, we characterized the seminal proteome of the monandrous fly Drosophila subobscura. Using a label-free quantitative proteomics approach, we identified 172 Sfps, comparable in number to the 153 Sfps previously reported in polyandrous Drosophila melanogaster using a similar approach. The D. subobscura seminal proteome contains all the D. melanogaster sex peptide network—proteins that control the female post-mating response—and many proteins known to be involved in post-copulatory sexual selection. We also found a surprising over-abundance of proteins previously shown to be downregulated under monogamy selection in D. melanogaster. Collectively, the patterns suggest that the D. subobscura seminal proteome is characterized more by conflict than cooperation and, despite differences in mating systems, is compositionally similar to that of D. melanogaster, with a complex proteome rich in receptivity-inhibiting Sfps.

Trade-offs between lifespan and reproduction are a central theme in evolution and ecology, often attributed to physiological constraints on the allocation of limiting resources. However, recent work suggests such trade-offs may depend on macro- and micronutritional context. We examined how macronutrient balance and cholesterol availability shape lifespan and reproductive performance in male Drosophila melanogaster , explicitly testing both unmated males and those permitted to mate freely throughout life—a design rarely applied. We found that cholesterol deprivation substantially reduced lifespan, but only in frequently mating males, indicating that sterols play a key role in mediating the lifespan costs of mating, consistent with their known role in sperm and seminal fluid. Conversely, the inclusion of dietary cholesterol reduced lifespan in unmated males maintained on low protein-to-carbohydrate diets, suggesting that male reproductive investment is sufficiently high to generate sterol-dependent costs to the soma. Lifespan was maximized at higher protein-to-carbohydrate ratios than previously estimated, while reproductive performance in aged males was favored by carbohydrate-biased diets, demonstrating a nutritional trade-off. Males preferentially consumed macronutrient ratios that promoted reproductive success rather than longevity and discriminated the presence of cholesterol in food. Together, these findings reveal sterols as a key mediator of male longevity, with effects that depend critically on both diet and reproductive context. More broadly, by integrating sterols, macronutrient balance, and sustained reproductive investment, our study adds to the growing evidence that male reproductive costs can be substantial and refines our understanding of the evolutionary basis of dietary trade-offs.

In variable environments, animals can change their reproductive behaviors and physiology to maximize reproductive returns. Natural environments vary in multifaceted ways, and animals may need to integrate multiple social or physical cues to adopt the most effective behavioral strategy. In a fully factorial 2 × 2 × 2 experiment, we exposed males to three factors: the number of rivals (10 or 30), food availability (present/absent) and mechanical shaking (present/absent). After 60 min of exposure, we recorded the male’s mating latency, copulation duration and the number of offspring produced after a single mating. We also noted the latency of the males partner to remate with a stock male 24 h later. When rival number was increased from 10 per vial to 30 per vial, males sired more offspring. Males also varied their copulation duration and mating latency in response to the number of rivals, but in a condition-dependent manner. In the absence of vortexing, males mated for a shorter time when kept with 30 rivals, but the opposite was observed when males were vortexed. When males were fed and held in groups of 30, they took longer to begin mating compared to the other treatments. Our findings are consistent with the idea that male Drosophila integrate social cues to respond to levels of sperm competition and plastically allocate their ejaculate, but we have demonstrated that they can occur more rapidly (1 h) than previously thought (>24 h). Overall, our data highlight that combinatorial approaches can reveal new relationships between environment and behavior.

Theory predicts that the sex ratio within populations should influence the strength of sexual selection, and sex ratio is often used as a proxy for sexual selection. However, recent studies challenge this relationship. We manipulated adult sex ratios in Drosophila melanogaster to comprehensively investigate the relationship between sex ratio and sexual selection. Consistent with theory, we found stronger sexual selection in males than females and an increased variance in male reproductive success (the opportunity for selection) in male-biased sex ratios. In addition, males faced more intense sperm competition in male-biased sex ratios, although the structure of sexual networks was largely invariant to sex ratio. Despite this, we show that sex ratios did not influence sexual selection in males as measured by the Bateman gradient. We leverage randomized null models to reconcile these results and show that the higher male reproductive variance in male-biased sex ratios may be explained by random chance in mating, rather than competitive mechanisms. Our findings indicate that caution is warranted over the long-standing assumption that sex ratio bias is a good proxy for the strength of sexual selection.

Early life conditions can have long-lasting fitness effects on organisms. In insects, crowding during larval stages impose a significant constraint on adult phenotypes due to increased intraspecific competition for resources, which can modulate males’ success in pre- and post-mating competition in adulthood. Evidence for larval crowding effects on male seminal fluid allocation exists but is limited to a small subset of well-known seminal fluid proteins (Sfps), and often overlooks the interactions between male and female phenotypes. We currently lack a comprehensive understanding of how male and female larval crowding interact to affect production, composition, and transfer of the wider seminal proteome. Here, we manipulated Drosophila melanogaster larval crowding (low versus high) of males and females to generate individuals with large and small body size, respectively. We mated individuals in a fully factorial design, and measured the abundance, composition, and transfer of Sfps. Large males produced Sfps in significantly higher abundances, yet this difference was marginal and not detected when Sfps were analysed individually. Conversely, small males transferred greater quantities of much of their seminal proteome to females than did large males. When analysing proteins individually, 10 Sfps were transferred in significantly higher abundances by small males than large males. Our results suggest that small males invest more on each mating opportunity, potentially as a response of overall fewer mating opportunities due to their reduced size, or due to the larval cues of high population density. This work provides an insight into early life effects on ejaculate allocation in D. melanogaster and sheds light on the physiological and behavioural responses to developmental conditions in insects.

In many species, the order in which males mate with a female explains much of the variation in paternity arising from post-copulatory sexual selection. Research in Drosophila suggests that mating order may account for the majority of the variance in male reproductive success. However, the effects of mating order on paternity bias might not be static but could potentially vary with social or environmental factors. To test this idea, we used an existing dataset, collated from an experiment we previously published (Morimoto et al., PLoS One, 11, 2016, e0154468), with the addition of unpublished data from the same experiment. These previous experiments manipulated larval density in Drosophila melanogaster which generated variation in male and female body size, assembled groups of individuals of different sizes, and measured the mating success and paternity share of focal males. The data presented here provides information on each focal male's mating order and the frequency in which focal males remated with same females (‘repetitive matings’). We combined this information with our previously reported focal male reproductive success to partition variance in paternity into male mating order and repetitive matings across groups that differed in the body size composition of males and females. We found, as expected, that male mating order explained a considerable portion of the variance in male paternity. However, we also found that the impact of male mating order on male paternity was influenced by the body size composition of groups. Specifically, males that tended to mate last had a greater paternity advantage, and displayed lower variance, in groups containing a heterogenous mixture male body sizes than in groups with a single male body size. Repetitive mating only had a minor contribution to the variance in male paternity share across all experiments. Overall, our findings contribute to the growing body of research showing that post-copulatory sexual selection is subject to socio-ecological influences. Abstract Social group composition affects the role of last male sperm precedence in Drosophila.

Dietary lipids (DLs), particularly sterols and fatty acids, are precursors for endogenous lipids that, unusually for macronutrients, shape cellular and organismal function long after ingestion. These functions – cell membrane structure, intracellular signalling, and hormonal activity – vary with the identity of DLs, and scale up to influence health, survival, and reproductive fitness, thereby affecting evolutionary change. Our Ecological Lipidology approach integrates biochemical mechanisms and molecular cell biology into evolution and nutritional ecology. It exposes our need to understand environmental impacts on lipidomes, the lipid specificity of cell functions, and predicts the evolution of lipid-based diet choices. Broad interdisciplinary implications of Ecological Lipidology include food web alterations, species responses to environmental change, as well as sex differences and lifestyle impacts on human nutrition, and opportunities for DL-based therapies.