Evolutionary trade-offs between testes size and parenting in Neotropical glassfrogs.

In males, large testes size signifies high sperm production and is commonly linked to heightened sperm competition levels. It may also evolve as a response to an elevated risk of sperm depletion due to multiple mating or large clutch sizes. Conversely, weapons, mate or clutch guarding may allow individuals to monopolize mating events and preclude sperm competition, thereby reducing the selection of large testes. Herein, we examined how paternal care, sexual size dimorphism (SSD), weaponry and female fecundity are linked to testes size in glassfrogs. We found that paternal care was associated with a reduction in relative testes size, suggesting an evolutionary trade-off between testes size and parenting. Although females were slightly larger than males and species with paternal care tended to have larger clutches, there was no significant relationship between SSD, clutch size and relative testes size. These findings suggest that the evolution of testes size in glassfrogs is influenced by sperm competition risk, rather than sperm depletion risk. We infer that clutch guarding precludes the risk of fertilization by other males and consequently diminishes selective pressure for larger testes. Our study highlights the prominent role of paternal care in the evolution of testes size in species with external fertilization.

How hibernation in frogs drives brain and reproductive evolution in opposite directions.

Environmental seasonality can promote the evolution of larger brains through cognitive and behavioral flexibility but can also hamper it when temporary food shortage is buffered by stored energy. Multiple hypotheses linking brain evolution with resource acquisition and allocation have been proposed for warm-blooded organisms, but it remains unclear how these extend to cold-blooded taxa whose metabolism is tightly linked to ambient temperature. Here, we integrated these hypotheses across frogs and toads in the context of varying brumation (hibernation) durations and their environmental correlates. We showed that protracted brumation covaried negatively with brain size but positively with reproductive investment, likely in response to brumation-dependent changes in the socio-ecological context and associated selection on different tissues. Our results provide novel insights into resource allocation strategies and possible constraints in trait diversification, which may have important implications for the adaptability of species under sustained environmental change.

The effect of cigarette smoke extract exposure on the size and sexual behaviour of Drosophila melanogaster.

Drosophila melanogaster is a widely used animal model in human diseases and to date it has not been applied to the study of the impact of tobacco use on human sexual function. Hence, this report examines the effects of different concentrations of cigarette smoke extract (CSE) exposure on the size and sexual behavior of D. melanogaster. Wild-type flies were held in vials containing CSE-infused culture media at concentrations of 10%, 25%, and 50% for three days, and their offspring were reared under the same conditions before measuring their body size and mating behavior. CSE exposure during development reduced the tibia length and body mass of emerging adult flies and prolonged the time required for successful courtship copulation success, while courtship behaviors (wing extension, tapping, abdomen bending, attempted copulation) remained largely unchanged. Our findings indicate that CSE exposure negatively affects the development of flies and their subsequent reproductive success. Future experiments should investigate the CSE effect on male female fertility.

Interplay between male quality and male-female compatibility across episodes of sexual selection.

The processes underlying mate choice profoundly influence the dynamics of sexual selection and the evolution of male sexual traits. Consistent preference for certain phenotypes may erode genetic variation in populations through directional selection, whereas divergent preferences (e.g., genetically compatible mates) provide one mechanism to maintain such variation. However, the relative contributions of these processes across episodes of selection remain unknown. Using Drosophila melanogaster, we followed the fate of male genotypes, previously scored for their overall reproductive value and their compatibility with different female genotypes, across pre- and postmating episodes of selection. When pairs of competitor males differed in their intrinsic quality and their compatibility with the female, both factors influenced outcomes from mating success to paternity but to a varying degree between stages. These results add further dimensions to our understanding of how the interactions between genotypes and forms of selection shape reproductive outcomes and ultimately reproductive trait evolution.

Genotype-by-environment interactions influence the composition of the Drosophila seminal proteome.

Ejaculate proteins are key mediators of post-mating sexual selection and sexual conflict, as they can influence both male fertilization success and female reproductive physiology. However, the extent and sources of genetic variation and condition dependence of the ejaculate proteome are largely unknown. Such knowledge could reveal the targets and mechanisms of post-mating selection and inform about the relative costs and allocation of different ejaculate components, each with its own potential fitness consequences. Here, we used liquid chromatography coupled with tandem mass spectrometry to characterize the whole-ejaculate protein composition across 12 isogenic lines of Drosophila melanogaster that were reared on a high- or low-quality diet. We discovered new proteins in the transferred ejaculate and inferred their origin in the male reproductive system. We further found that the ejaculate composition was mainly determined by genotype identity and genotype-specific responses to larval diet, with no clear overall diet effect. Nutrient restriction increased proteolytic protein activity and shifted the balance between reproductive function and RNA metabolism. Our results open new avenues for exploring the intricate role of genotypes and their environment in shaping ejaculate composition, or for studying the functional dynamics and evolutionary potential of the ejaculate in its multivariate complexity.

Strongly sexually dimorphic forelegs are not more condition-dependent than less dimorphic traits in Drosophila prolongata.

Directional sexual selection drives the evolution of traits that are most closely linked to reproductive success, giving rise to trait exaggeration and sexual dimorphism. Exaggerated structures are often costly and, therefore, thought to be expressed in a condition-dependent manner. Sexual selection theory thus predicts a direct link between directional sexual selection, sexual dimorphism, and sex-specific condition dependence. However, only a handful of studies investigate the relationship between sexual dimorphism and condition dependence. Using 21 genetic lines of Drosophila prolongata, we here compared the degree of sexual dimorphism and sex-specific condition dependence, measured as allometric slopes, in sexually selected and non-sexual traits. Our data revealed male-biased sexual dimorphism in all traits examined, most prominently in the sexually selected forelegs. However, there was no relationship between the degree of sex-specific condition dependence and sexual dimorphism across traits and genetic lines. Our results contradict theoretical predictions and highlight the importance of understanding the role of exaggerated traits in the context of both sexual and natural selection. Supplementary Information: The online version contains supplementary material available at 10.1007/s10682-022-10226-0.

Paternal condition affects offspring reproduction and life history in a sex-specific manner in Drosophila melanogaster.

Nongenetic parental effects can contribute to the adaptation of species to changing environments by circumventing some of the limitations of genetic inheritance. A clearer understanding of the influence of nongenetic inheritance and its potentially sex-specific responses in daughters and sons is needed to better predict the evolutionary trajectories of species. However, whereas nongenetic maternal effects have long been recognized and widely studied, comparatively little is known about corresponding paternal effects. Here, by following 30 isogenic lines of Drosophila melanogaster across two generations, each reared under two dietary regimes in each generation, we tested how protein restriction during larval development of the fathers affects the fitness and health of their daughters and sons. We then quantified genetic and non-genetic paternal, and direct environmental, effects across multiple axes of offspring fitness. Daughters and sons responded differently to their father's developmental history. While isolines differed in mean trait values, their specific responses to protein restriction generally varied little. The sex- and trait-specific responses to paternal effects emphasize the complexity of inter-generational parental effects, which raise important questions about their mode of transmission and adaptive value, including the potential for conflict between the sexes.

Cognition contra camouflage: How the brain mediates predator-driven crypsis evolution.

While crypsis is a prominent antipredator adaptation, the role of the brain in predator-driven evolution remains controversial. Resolving this controversy requires contextualizing the brain with established antipredator traits and predation pressure. We hypothesize that the reduced predation risk through crypsis relaxes predation-driven selection on the brain and provide comparative evidence across 102 Chinese frog species for our hypothesis. Specifically, our phylogenetic path analysis reveals an indirect relationship between predation risk and crypsis that is mediated by brain size. This result suggests that at a low predation risk, frogs can afford to be conspicuous and use their large brain for cognitive predator evasion. This strategy may become less efficient or energetically costlier under higher predation pressure, favoring smaller brains and instead increasing crypsis.

Sperm competition in yellow dung flies: No consistent effect of sperm size.

The male competition for fertilization that results from female multiple mating promotes the evolution of increased sperm numbers and can impact sperm morphology, with theory predicting that longer sperm can at times be advantageous during sperm competition. If so, males with longer sperm should sire more offspring than competitors with shorter sperm. Few studies have directly tested this prediction, and findings are inconsistent. Here we assessed whether longer sperm provide a competitive advantage in the yellow dung fly (Scathophaga stercoraria; Diptera: Scathophagidae). Initially, we let brothers with different temperature-mediated mean sperm lengths compete - thus minimizing confounding effects of genetic background - and found no clear advantage of longer sperm. We then used flies from lines subjected to bidirectional selection on phenoloxidase activity that had shown correlated evolutionary responses in sperm and female spermathecal duct lengths. This experiment also yielded no main effect of sperm size on siring success. Instead, there was a trend for a shorter-sperm advantage, but only when competing in females with longer spermathecal ducts. Our data corroborated many previously reported findings (last-male precedence, effects of copula duration and body size), suggesting our failure to find sperm size effects is not inherently due to our experimental protocols. We conclude that longer sperm are not competitively superior in yellow dung flies under most circumstances, and that, consistent with previous work, in this species competitive fertilization success is primarily determined by the relative numbers of sperm competing.

Prediction and enrichment analyses of the Homo sapiens-Drosophila melanogaster COPD-related orthologs: potential for modeling of human COPD genomic responses with the fruit fly.

The significant similarities in airway epithelial cells between mammals and the fruit fly Drosophila melanogaster have rendered the latter an important model organism for studies of chronic inflammatory lung diseases. Focusing on the chronic obstructive pulmonary disease (COPD), we here mapped human gene orthologs associated with this disease in D. melanogaster to identify functionally equivalent genes for immediate, further screening with the fruit fly model. The DIOPT-DIST tool was accessed for the prediction of the COPD-associated orthologs between humans and Drosophila. Enrichment analyses with respect to pathways of the retrieved functional homologs were performed using the ToppFun and FlyMine tools, identifying 73 unique human genes as well as 438 fruit fly genes. The ToppFun analysis verified that the human gene list is associated with COPD phenotypes. Furthermore, the FlyMine investigation highlighted that the Drosophila genes are functionally connected mainly with the "ABC-family proteins mediated transport" and the "ß-catenin-independent WNT signaling pathway." These results suggest an evolutionarily conserved role toward responses to inhaled toxicants and CO2 in both species. We reason that the predicted orthologous genes should be further studied in the Drosophila models of cigarette smoke-induced COPD.

Sperm depletion in relation to developmental nutrition and genotype in Drosophila melanogaster.

Nutrient limitation during development can restrict the ability of adults to invest in costly fitness traits, and genotypes can vary in their sensitivity to developmental nutrition. However, little is known about how genotype and nutrition affect male ability to maintain ejaculate allocation and achieve fertilization across successive matings. Using 17 isogenic lines of Drosophila melanogaster, we investigated how variation in developmental nutrition affects males' abilities to mate, transfer sperm, and sire offspring when presented with successive virgin females. We found that, with each successive mating, males required longer to initiate copulation, transferred fewer sperm, and sired fewer offspring. Males reared on a low-nutrient diet transferred fewer sperm than those reared on nutritionally superior diets, but the rate at which males depleted their sperm, as well as their reproductive performance, was largely independent of diet. Genotype and the genotype × diet interaction explained little of the variation in these male reproductive traits. Our results show that sperm depletion can occur rapidly and impose substantial fitness costs for D. melanogaster males across multiple genotypes and developmental environments.

Weapons Evolve Faster Than Sperm in Bovids and Cervids.

In polyandrous species, males face reproductive competition both before and after mating. Sexual selection thus shapes the evolution of both pre- and postcopulatory traits, creating competing demands on resource allocation to different reproductive episodes. Traits subject to strong selection exhibit accelerated rates of phenotypic divergence, and examining evolutionary rates may inform us about the relative importance and potential fitness consequences of investing in traits under either pre- or postcopulatory sexual selection. Here, we used a comparative approach to assess evolutionary rates of key competitive traits in two artiodactyl families, bovids (family Bovidae) and cervids (family Cervidae), where male-male competition can occur before and after mating. We quantified and compared evolutionary rates of male weaponry (horns and antlers), body size/mass, testes mass, and sperm morphometrics. We found that weapons evolve faster than sperm dimensions. In contrast, testes and body mass evolve at similar rates. These results suggest strong, but differential, selection on both pre- and postcopulatory traits in bovids and cervids. Furthermore, we documented distinct evolutionary rates among different sperm components, with sperm head and midpiece evolving faster than the flagellum. Finally, we demonstrate that, despite considerable differences in weapon development between bovids and cervids, the overall evolutionary patterns between these families were broadly consistent.

Condition-dependent interaction between mating success and competitive fertilization success in Drosophila melanogaster.

Dietary restriction during development can affect adult body size and condition. In many species, larger (high-condition) males gain higher mating success through male-male competition and female choice, and female condition can affect the extent of both female mate choice and male investment in courtship or ejaculates. However, few studies have examined the joint effects and interplay of male and female condition during both the pre- and the postcopulatory phases of sexual selection. We therefore manipulated the larval diet of male and female Drosophila melanogaster to study how body size variation in both sexes biases competitive outcomes at different reproductive stages, from mating to paternity. We did not find a difference in mate preference or mating latency between females of different conditions, nor any interaction between male and female conditions. However, large males were more successful in gaining matings, but only when in direct competition, whereas mating latencies were shorter for low-condition males in noncompetitive settings. Small males also transferred more sperm to nonvirgin females, displaced a larger proportion of resident sperm, and achieved higher paternity shares per mating than large males. In agreement with existing theory, we suggest that small males might partially compensate for their low mating success by strategically investing in larger sperm numbers and potentially other, unmeasured ejaculate traits, when they do have a mating opportunity.

The Drosophila septate junctions beyond barrier function: Review of the literature, prediction of human orthologs of the SJ-related proteins and identification of protein domain families.

The involvement of Septate Junctions (SJs) in critical cellular functions that extend beyond their role as diffusion barriers in the epithelia and the nervous system has made the fruit fly an ideal model for the study of human diseases associated with impaired Tight Junction (TJ) function. In this study, we summarized current knowledge of the Drosophila melanogaster SJ-related proteins, focusing on their unconventional functions. Additionally, we sought to identify human orthologs of the corresponding genes as well as protein domain families. The systematic literature search was performed in PubMed and Scopus databases using relevant key terms. Orthologs were predicted using the DIOPT tool and aligned protein regions were determined from the Pfam database. 3-D models of the smooth SJ proteins were built on the Phyre2 and DMPFold protein structure prediction servers. A total of 30 proteins were identified as relatives to the SJ cellular structure. Key roles of these proteins, mainly in the regulation of morphogenetic events and cellular signalling, were highlighted. The investigation of protein domain families revealed that the SJ-related proteins contain conserved domains that are required not only for cell-cell interactions and cell polarity but also for cellular signalling and immunity. DIOPT analysis of orthologs identified novel human genes as putative functional homologs of the fruit fly SJ genes. A gap in our knowledge was identified regarding the domains that occur in the proteins encoded by eight SJ-associated genes. Future investigation of these domains is needed to provide functional information.

How female × male and male × male interactions influence competitive fertilization in Drosophila melanogaster.

How males and females contribute to joint reproductive success has been a long-standing question in sexual selection. Under postcopulatory sexual selection, paternity success is predicted to derive from complex interactions among females engaging in cryptic female choice and males engaging in sperm competition. Such interactions have been identified as potential sources of genetic variation in sexually selected traits but are also expected to inhibit trait diversification. To date, studies of interactions between females and competing males have focused almost exclusively on genotypes and not phenotypic variation in sexually selected traits. Here, we characterize within- and between-sex interactions in Drosophila melanogaster using isogenic lines with heritable variation in both male and female traits known to influence competitive fertilization. We confirmed, and expanded on, previously reported genotypic interactions within and between the sexes, and showed that several reproductive events, including sperm transfer, female sperm ejection, and sperm storage, were explained by two- and three-way interactions among sex-specific phenotypes. We also documented complex interactions between the lengths of competing males' sperm and the female seminal receptacle, which are known to have experienced rapid female-male co-diversification. Our results highlight the nonindependence of sperm competition and cryptic female choice and demonstrate that complex interactions between the sexes do not limit the ability of multivariate systems to respond to directional sexual selection.

How sperm competition shapes the evolution of testes and sperm: a meta-analysis.

Females of many species mate with multiple males, thereby inciting competition among ejaculates from rival males for fertilization. In response to increasing sperm competition, males are predicted to enhance their investment in sperm production. This prediction is so widespread that testes size (correcting for body size) is commonly used as a proxy of sperm competition, even in the absence of any other information about a species' reproductive behaviour. By contrast, a debate about whether sperm competition selects for smaller or larger sperm has persisted for nearly three decades, with empirical studies demonstrating every possible response. Here, we synthesize nearly 40 years of sperm competition research in a meta-analytical framework to determine how the evolution of sperm number (i.e. testes size) and sperm size (i.e. sperm head, midpiece, flagellum and total length) is influenced by varying levels of sperm competition across species. Our findings support the long-held assumption that higher levels of sperm competition are associated with relatively larger testes. We also find clear evidence that sperm competition is associated with increases in all components of sperm length. We discuss these results in the context of different theoretical predictions and general patterns in the breeding biology and selective environment of sperm. This article is part of the theme issue 'Fifty years of sperm competition'.

Relative Brain Size Is Predicted by the Intensity of Intrasexual Competition in Frogs.

Competition over mates is a powerful force shaping trait evolution. For instance, better cognitive abilities may be beneficial in male-male competition and thus be selected for by intrasexual selection. Alternatively, investment in physical attributes favoring male performance in competition for mates may lower the resources available for brain development, and more intense male mate competition would coincide with smaller brains. To date, only indirect evidence for such relationships exists, and most studies are heavily biased toward primates and other homoeothermic vertebrates. We tested the association between male brain size (relative to body size) and male-male competition across N=30 species of Chinese anurans. Three indicators of the intensity of male mate competition-operational sex ratio (OSR), spawning-site density, and male forelimb muscle mass-were positively associated with relative brain size, whereas the absolute spawning group size was not. The relationship with the OSR and male forelimb muscle mass was stronger for the male than for the female brains. Taken together, our findings suggest that the increased cognitive abilities of larger brains are beneficial in male-male competition. This study adds taxonomic breadth to the mounting evidence for a prominent role of sexual selection in vertebrate brain evolution.

Fitness consequences of the combined effects of veterinary and agricultural pesticides on a non-target insect.

Pesticides and veterinary products that are globally used in farming against pests and parasites are known to impact non-target beneficial organisms. While most studies have tested the lethal and sub-lethal effects of single chemicals, species are exposed to multiple contaminants that might interact and exacerbate the toxic responses of life-history fitness components. Here we experimentally tested an ecotoxicological scenario that is likely to be widespread in nature, with non-target dung communities being exposed both to cattle parasiticides during the larval stage and to agricultural insecticides during their adult life. We assessed the independent and combined consumptive effects of varying ivermectin and spinosad concentration on juvenile life-history and adult reproductive traits of the widespread yellow dung fly (Scathophaga stercoraria; Diptera: Scathophagidae). Larval exposure to ivermectin prolonged development time and reduced egg-to-adult survival, body size, and the magnitude of the male-biased sexual size dimorphism. The consumption by the predatory adult flies of spinosad-contaminated prey showed an additional, independent (from ivermectin) negative effect on female clutch size, and subsequent egg hatching success, but not on the body size and sexual size dimorphism of their surviving offspring. However, there were interactive synergistic effects of both contaminants on offspring emergence and body size. Our results document adverse effects of the combination of different chemicals on fitness components of a dung insect, highlighting transgenerational effects of adult exposure to contaminants for their offspring. These findings suggest that ecotoxicological tests should consider the combination of different contaminants for more accurate eco-assessments.

Sperm quality and quantity evolve through different selective processes in the Phasianidae.

Sperm competition is often considered the primary selective force underlying the rapid and diversifying evolution of ejaculate traits. Yet, several recent studies have drawn attention to other forms of selection with the potential of exceeding the effects of sperm competition. Since ejaculates are complex, multivariate traits, it seems plausible that different ejaculate components vary in their responses to different selective pressures. Such information, however, is generally lacking as individual ejaculate traits tend to be studied in isolation. Here, we studied the macroevolutionary patterns of ejaculate volume, sperm number, sperm length and the proportion of viable normal sperm in response to varying levels of sperm competition, body size and the duration of female sperm storage in pheasants and allies (Phasianidae). Ejaculate volume, sperm number and sperm viability were all relatively higher in polygamous than in monogamous mating systems. However, whereas ejaculate volume additionally covaried with body size, sperm number instead increased with the female sperm-storage duration, in conjunction with a decrease in sperm length. Overall, our results revealed important details on how different forms of selection can jointly shape ejaculates as complex, composite traits.

Sexual ornaments but not weapons trade off against testes size in primates.

Males must partition their limited reproductive investments between traits that promote access to females (sexual ornaments and weapons) and traits that enhance fertilization success, such as testes and ejaculates. Recent studies show that if the most weaponized males can monopolize access to females through contest competition, thereby reducing the risk of sperm competition, they tend to invest less in sperm production. However, how males invest in sexual ornaments relative to sperm production remains less clear. If male ornaments serve as badges of status, with high-ranking males attaining near-exclusive access to females, similar to monopolizing females through combat, their expression should also covary negatively with investment in post-mating traits. In a comparative study across primates, which exhibit considerable diversification in sexual ornamentation, male weaponry and testes size, we found relative testes size to decrease with sexual ornaments but increase with canine size. These contrasting evolutionary trajectories might be driven by differential selection, functional constraints or temporal patterns of metabolic investment between the different types of sexual traits. Importantly, however, our results indicate that the theory of relative investments between weapons and testes in the context of monopolizing females can extend to male ornaments.