Constraints on evolution and postcopulatory sexual selection: trade-offs among ejaculate characteristics

Ejaculates function as an integrated unit to ensure male fertility and paternity, can have a complex structure, and can experience multiple episodes of selection. Current studies on the evolution of ejaculates typically focus on phenotypic variation in sperm number, size, or related traits such as testes size as adaptations to postcopulatory male-male competition. However, the evolution of the integrated nature of ejaculate structure and function depends on genetic variation in and covariation between the component parts. Here we report a quantitative genetic study of the components of the ejaculate of the cockroach Nauphoeta cinerea, including those we know to experience postcopulatory sexual selection, in the context of functional integration of ejaculate characters. We use the patterns of genetic variation and covariation to infer how the integration of the functions of the ejaculate constrain and shape its evolution. Ejaculate components were highly variable, showed significant additive genetic variance, and moderate to high evolvability. The level of genetic variation in these characters, despite strong directional or truncating selection, may reflect the integration of multiple episodes of selection that occur in N. cinerea. There were few significant phenotypic correlations, but all the genetic correlations among ejaculate characters were significantly different from zero. The patterns of genetic variation and covariation suggest that there are important trade-offs among individual traits of the ejaculate and that evolution of ejaculate characteristics will not proceed unconstrained. Fully describing the genetic relationships among traits that perform as an integrated unit helps us understand how functional relationships constrain or facilitate the evolution of the complex structure that is the ejaculate.     

Evolution of ejaculates: patterns of phenotypic and genotypic variation and condition dependence in sperm competition traits

Sperm competition is widely recognized as a potent force in evolution, influencing male behavior, morphology, and physiology. Recent game theory analyses have examined how sperm competition can influence the evolution of ejaculate expenditure by males and the morphology of sperm contained within ejaculates. Theoretical analyses rest on the assumption that there is sufficient genetic variance in traits important in sperm competition to allow evolving populations to move to the evolutionarily stable equilibrium. Moreover, patterns of genotypic variation can provide valuable insight into the nature of selection currently acting on traits. However, our knowledge of genetic variance underlying traits important in sperm competition is limited. Here we examine patterns of phenotypic and genotypic variation in four sperm competition traits in the dung beetle Onthophagus taurus. Testis weight, ejaculate volume, and copula duration were found to have high coefficients of additive genetic variation (CV(A)S), which is characteristic of fitness traits and traits subject to sexual selection. Heritabilities were high, and there was some evidence for Y-linked inheritance in testis weight. In contrast, sperm length had a low CV(A), which is characteristic of traits subject to stabilizing selection. Nevertheless, there was little residual variance so that the heritability of sperm length exceeded 1.0. Such a pattern is consistent with Y-linked inheritance in sperm length. Interestingly, we found that testis weight and sperm length were genetically correlated with heritable male condition. This finding holds important implications for potential indirect benefits associated with the evolution of polyandry.     

The evolution of sperm and non-sperm producing organs in male Drosophila

The male ejaculate is made up of two components: sperm and non-sperm. There has been little consideration of how these two basic compartments evolve. If they are subject to trade-offs, theory predicts that when the sperm competition raffle is unfair, when seminal fluid proteins stimulate fecundity and/or when ejaculate components alter fertilization success, there will be differential selection on sperm versus non-sperm ejaculate characteristics. However, the fundamental assumption that there are trade-offs between sperm and non-sperm ejaculate compartments in Drosophila has not yet been tested. To address this, we examined testis (sperm producing) and accessory gland (non-sperm producing) size across 22 species of Drosophila . We also examined how these characters varied with copulation duration, which may represent an additional target for sperm competition. The results showed no evidence of a trade-off between testis length and accessory gland length. Copulation duration correlated negatively with accessory gland length and there was a positive correlation with testis length, but only after correcting for body size. Overall, the results suggest no evidence for gross trade-offs in sperm versus non-sperm compartments across these Drosophila species, and motivate more detailed examination of ejaculate investment patterns.  © 2008 The Linnean Society of London, Biological Journal of the Linnean Society , 2008, 94 , 505–512.     

Do male secondary sexual characters signal ejaculate quality? A meta‐analysis

There are two reasons why researchers are interested in the phenotypic relationship between the expression of male secondary sexual characters (SSCs) and ‘ejaculate quality’ (defined as sperm/ejaculate traits that are widely assumed to increase female fertility and/or sperm competitiveness). First, if the relationship is positive then females could gain a direct benefit by choosing more attractive males for fertility assurance reasons (‘the phenotype‐linked fertility’ hypothesis). Second, there is much interest in the direction of the correlation between traits favoured by pre‐copulatory sexual selection (i.e. affecting mating success) and those favoured by post‐copulatory sexual selection (i.e. increasing sperm competitiveness). If the relationship is negative this could lead to the two forms of selection counteracting each other. Theory predicts that the direction of the relationship could be either positive or negative depending on the underlying genetic variance and covariance in each trait, the extent of variation among males in condition (resources available to allocate to reproductive traits), and variation among males in the cost or rate of mating. We conducted a meta‐analysis to determine the average relationship between the expression of behavioural and morphological male secondary sexual characters and four assays of ejaculate quality (sperm number, viability, swimming speed and size). Regardless of how the data were partitioned the mean relationship was consistently positive, but always statistically non‐significant. The only exception was that secondary sexual character expression was weakly but significantly positively correlated with sperm viability (r = 0.07, P < 0.05). There was no significant difference in the strength or direction of the relationship between behavioural and morphological SSCs, nor among relationships using the four ejaculate quality assays. The implications of our findings are discussed.     

Distinct evolutionary patterns of morphometric sperm traits in passerine birds

The striking diversity of sperm shape across the animal kingdom is still poorly understood. Postcopulatory sexual selection is an important factor driving the evolution of sperm size and shape. Interestingly, morphometric sperm traits, such as the length of the head, midpiece and flagellum, exhibit a strong positive phenotypic correlation across species. Here we used recently developed comparative methods to investigate how such phenotypic correlations between morphometric sperm traits may evolve. We compare allometric relationships and evolutionary trajectories of three morphometric sperm traits (length of head, midpiece and flagellum) in passerine birds. We show that these traits exhibit strong phenotypic correlations but that allometry varies across families. In addition, the evolutionary trajectories of the midpiece and flagellum are similar while the trajectory for head length differs. We discuss our findings in the light of three scenarios accounting for correlated trait evolution: (i) genetic correlation; (ii) concerted response to selection acting simultaneously on different traits; and (iii) phenotypic correlation between traits driven by mechanistic constraints owing to selection on sperm performance. Our results suggest that concerted response to selection is the most likely explanation for the phenotypic correlation between morphometric sperm traits.     

Intraspecific variation in ejaculate traits of the northern watersnake (Nerodia sipedon)

Sperm competition occurs when the sperm of more than one male compete to fertilize the eggs of a female. In reptiles, sperm competition is particularly prevalent and is an important agent of sexual selection in males. Spermatogenesis in reptiles can be energetically expensive, suggesting that there may be costs to producing high-quality ejaculates. The northern watersnake Nerodia sipedon has a mating system characterized by aggregations in which a single female mates with multiple males, resulting in high levels of multiple paternities. Under these circumstances sperm competition is likely important, and selection should favour sperm and ejaculate traits that enhance a male's reproductive success. In this study, we examined intraspecific variation in ejaculate quality (sperm length, motility, sperm density, spermactocrit) in male watersnakes and determined whether ejaculate traits varied with body size and condition, using both size-corrected mass and haematocrit as indices of condition. We found large variation among males in all these traits, except for sperm length. Although there was significant variation in sperm length among males, the majority of variation in sperm length occurred within rather than among individuals. Males with high haematocrit had sperm that were less variable with respect to length, and large males produced ejaculates that were less concentrated with respect to sperm than small males. The lack of condition dependence of most ejaculate traits is consistent with previous studies that indicate that male reproductive effort in this species is generally not energy limited, perhaps because of opportunistic foraging during the mating season.     

SPERM MORPHOLOGY AND VELOCITY ARE GENETICALLY CODETERMINED IN THE ZEBRA FINCH

Sperm morphology (size and shape) and sperm velocity are both positively associated with fertilization success, and are expected to be under strong selection. Until recently, evidence for a link between sperm morphology and velocity was lacking, but recent comparative studies have shown that species with high levels of sperm competition have evolved long and fast sperm. It is therefore surprising that evidence for a phenotypic or genetic relationship between length and velocity within species is equivocal, even though sperm competition is played out in the intraspecific arena. Here, we first show that sperm velocity is positively phenotypically correlated with measures of sperm length in the zebra finch Taeniopygia guttata . Second, by using the quantitative genetic "animal model" on a dataset from a multigenerational-pedigreed population, we show that sperm velocity is heritable, and positively genetically correlated to a number of heritable components of sperm length. Therefore, selection for faster sperm will simultaneously lead to the evolution of longer sperm (and vice versa). Our results provide, for the first time, a clear phenotypic and genetic link between sperm length and velocity, which has broad implications for understanding how recently described macroevolutionary patterns in sperm traits have evolved.     

Intraspecific variation in sperm length is negatively related to sperm competition in passerine birds

Spermatozoa are among the most diversified cells in the animal kingdom, but the underlying evolutionary forces affecting intraspecific variation in sperm morphology are poorly understood. It has been hypothesized that sperm competition is a potent selection pressure on sperm variation within species. Here, we examine intraspecific variation in total sperm length of 22 wild passerine bird species (21 genera, 11 families) in relation to the risk of sperm competition, as expressed by the frequency of extrapair paternity and relative testis size. We demonstrate, by using phylogenetic comparative methods, that between-male variation in sperm length within species is closely and negatively linked to the risk of sperm competition. This relationship was even stronger when only considering species in which data on sperm length and extrapair paternity originated from the same populations. Intramale variation in sperm length within species was also negatively, although nonsignificantly, related to sperm competition risk. Our findings suggest that postcopulatory sexual selection is a powerful evolutionary force reducing the intraspecific phenotypic variation in sperm-size traits, potentially driving the diversification of sperm morphology across populations and species.     

Sperm traits negatively covary with size and asymmetry of a secondary sexual trait in a freshwater crayfish

In species where females mate promiscuously, the reproductive success of males depends both on their ability to acquire mates (pre-copulatory sexual selection) and ability of their ejaculates to outcompete those of other males (post-copulatory sexual selection). Sperm competition theory predicts a negative relationship between investment in body traits favouring mate acquisition (secondary sexual characters, SSCs) and investment in ejaculate size or quality, due to the inherent costs of sperm production. In contrast, the phenotype-linked fertility hypothesis posits that male fertilizing efficiency is reliably reflected by the phenotypic expression of male SSCs, allowing females to obtain direct benefits by selecting more ornamented males as copulation partners. In this study, we investigated the relationships between male SSCs and size and quality (viability and longevity) of ejaculates allocated to females in mating trials of the freshwater crayfish Austropotamobius italicus. We showed that the relative size of male weapons, the chelae, was negatively related to ejaculate size, and that chelae asymmetry, resulting from regeneration of lost chelipeds, negatively covaried with sperm longevity. Moreover, males allocated more viable sperm to mates from their own rather than different stream of origin. Our findings thus suggest that, according to sperm competition theory, pre-copulatory sexual selection for large weapons used in male fighting may counteract post-copulatory sperm competition in this crayfish species, and that investment in cheliped regeneration may impair ejaculate quality.     

Causes of male sexual trait divergence in introduced populations of guppies

Males from different populations of the same species often differ in their sexually selected traits. Variation in sexually selected traits can be attributed to sexual selection if phenotypic divergence matches the direction of sexual selection gradients among populations. However, phenotypic divergence of sexually selected traits may also be influenced by other factors, such as natural selection and genetic constraints. Here, we document differences in male sexual traits among six introduced Australian populations of guppies and untangle the forces driving divergence in these sexually selected traits. Using an experimental approach, we found that male size, area of orange coloration, number of sperm per ejaculate and linear sexual selection gradients for male traits differed among populations. Within populations, a large mismatch between the direction of selection and male traits suggests that constraints may be important in preventing male traits from evolving in the direction of selection. Among populations, however, variation in sexual selection explained more than half of the differences in trait variation, suggesting that, despite within‐population constraints, sexual selection has contributed to population divergence of male traits. Differences in sexual traits were also associated with predation risk and neutral genetic distance. Our study highlights the importance of sexual selection in trait divergence in introduced populations, despite the presence of constraining factors such as predation risk and evolutionary history.     

Sperm in competition: not playing by the numbers

The outcome of sperm competition is mediated largely by the relative numbers of sperm from competing males. However, substantial variation in features of sperm morphology and behaviour, such as length, longevity and motility, exists and researchers have suggested that this variation functions in postcopulatory sexual selection. Recent studies have determined the effect of these sperm-quality traits on fertilization success and a synthesis of this literature reveals that they are important in both sperm competition and cryptic female choice. To understand how postcopulatory sexual selection influences sperm traits, future research should determine sex-specific interactions that influence paternity, identify genetic correlations between ejaculate characters, quantify the relative costs of producing different sperm traits, and test assumptions of models of sperm quality evolution. Such research will shed light on what evolutionary pressures are responsible for the diversity in sperm morphometry and behaviour.     

Genetic and environmental sources of covariance among internal reproductive traits in the yellow dung fly

Substantial inter- and intraspecific variation is found in reproductive traits, but the evolutionary implications of this variation remain unclear. One hypothesis is that natural selection favours female reproductive morphology that allows females to control mating and fertilization and that diverse male reproductive traits arise as counter adaptations to subvert this control. Such co-evolution predicts the establishment of genetic correlations between male and female reproductive traits that closely interact during mating. Therefore, we measured phenotypic and genetic correlations between male and female reproductive tract characteristics in the yellow dung fly, Scathophaga stercoraria (Diptera: Scathophagidae), using a nested half-sib breeding experiment. We found significant heritabilities for the size of most reproductive tract traits investigated in both females (spermathecae and their ducts, accessory glands and their ducts) and males (testis size but not sperm length). Within the sexes, phenotypic and genetic correlations were mostly nil or positive, suggesting functional integration of or condition-dependent investment in internal reproductive traits. Negative intrasexual genetic correlations, potentially suggestive of resource allocation trade-offs, were not evident. Intersexual genetic correlations were mostly positive, reflecting expected allometries between male and female morphologies. Most interestingly, testis size correlated positively with female accessory gland size and duct length, potentially indicative of a co-evolutionary arms race. We discuss these and alternative explanations for these patterns of genetic covariance.     

Quantitative genetic evidence that males trade attractiveness for ejaculate quality in guppies

Polyandry, where females mate with multiple males, means that a male''s reproductive success will depend both on his ability to acquire mates and the ability of his sperm to compete effectively for fertilizations. But, how do males partition their reproductive investment between these two episodes of selection? Theory predicts that increases in ejaculate investment will come at a cost to investment in other reproductive traits. Although evidence revealing such trade-offs is accumulating, we know little about their genetic basis. Here, I report patterns of genetic (co)variation for a range of traits subject to pre- and post-copulatory sexual selection in the guppy Poecilia reticulata, a promiscuous livebearing fish in which males alternate between courtship and sneak matings to obtain copulations. The analyses of genetic variation and covariation for these behaviours revealed a strong genetic predisposition for one tactic over the other. Both mating tactics were also strongly genetically integrated with the level of sexual ornamentation and ejaculate quality. Males that predominantly performed sneak matings were less ornamented but had faster swimming sperm than those that predominantly used courtship. These patterns of genetic variation and covariation reveal potential evolutionary constraints on the direction of selection of pre- and post-copulatory traits, and support sperm competition theory by revealing a trade-off between sexual attractiveness and investment in ejaculates.     

A comparison of nuclear and cytoplasmic genetic effects on sperm competitiveness and female remating in a seed beetle

It is widely assumed that male sperm competitiveness evolves adaptively. However, recent studies have found a cytoplasmic genetic component to phenotypic variation in some sperm traits presumed important in sperm competition. As cytoplasmic genes are maternally transmitted, they cannot respond to selection on sperm and this constraint may affect the scope in which sperm competitiveness can evolve adaptively. We examined nuclear and cytoplasmic genetic contributions to sperm competitiveness, using populations of Callosobruchus maculatus carrying orthogonal combinations of nuclear and cytoplasmic lineages. Our design also enabled us to examine genetic contributions to female remating. We found that sperm competitiveness and remating are primarily encoded by nuclear genes. In particular, a male's sperm competitiveness phenotype was contingent on an interaction between the competing male genotypes. Furthermore, cytoplasmic effects were detected on remating but not sperm competitiveness, suggesting that cytoplasmic genes do not generally play a profound evolutionary role in sperm competition.     

RELATIONSHIPS BETWEEN SPERM LENGTH AND SPEED DIFFER AMONG THREE INTERNALLY AND THREE EXTERNALLY FERTILIZING SPECIES

It is often assumed that longer sperm, by virtue of their increased swimming speed, have a fertilization advantage over shorter sperm when in competition to fertilize eggs. However, there is surprisingly little evidence for a positive correlation between sperm length and speed. Here we use an approach that accounts for within‐male variation in sperm traits to examine the relationships between sperm length and sperm speed across a broad range of species, including three internally fertilizing species and three externally fertilizing species. Our results reveal that correlations between sperm size and speed are indeed present and possibly more common than currently thought. However, the direction of the correlations between sperm length and speed, which are more prevalent within a male's ejaculate than among males, were influenced by fertilization mode in contrasting and unexpected ways. Broadly, the patterns revealed that in externally fertilizing species sperm with longer flagellum and shorter heads relative to their flagellum swam faster, whereas in internally fertilizing species sperm with shorter flagellum and longer heads relative to their flagellum swam faster. We discuss these results in light of sperm competition theory and contrast the intraspecific patterns observed in this study with macroevolutionary patterns of sperm evolution reported elsewhere.     

Variation in sperm morphology among Afrotropical sunbirds

Birds show considerable variation in sperm morphology. Closely related species and subspecies can show diagnostic differences in sperm size. There is also variation in sperm size among males within a population, and recent evidence from passerine birds suggests that the coefficient of inter‐male variation in sperm length is negatively associated with the level of sperm competition. Here we examined patterns of inter‐ and intra‐specific variation in sperm length in 12 species of sunbird (Nectariniidae) from Nigeria and Cameroon, a group for which such information is extremely limited. We found significant variation among species in sperm total length, with mean values ranging from 74 μm to 116 μm, placing these species within the short to medium sperm length range for passerine birds. Most of this variation was explained by the length of the midpiece, which contains the fused mitochondria and is an important structure for sperm energetics. Relative midpiece length was negatively correlated with the coefficient of inter‐male variation in sperm total length across species, suggesting that sperm competition may have selected for greater midpiece length in this group. We also mapped sperm lengths onto a time‐calibrated phylogeny and found support for a phylogenetic signal in all sperm length components, except head length. A test of various evolutionary or tree transformation models gave strongest support for the Brownian motion model for all sperm components, i.e. divergences were best predicted by the phylogenetic distance between lineages. The coefficients of inter‐male variation in sperm total length indicate that sperm competition is high but variable among sunbird species, as is the case with passerine birds at large.     

Variability in sperm form and function in the context of sperm competition risk in two Tupinambis lizards

In polyandrous species, sperm morphometry and sperm velocity are under strong sexual selection. Although several hypotheses have been proposed to explain the role of sperm competition in sperm trait variation, this aspect is still poorly understood. It has been suggested that an increase in sperm competition pressure could reduce sperm size variation or produce a diversity of sperm to maximize male fertilization success. We aim at elucidating the variability of sperm morphometric traits and velocity in two Tupinambis lizards in the context of sperm competition risk. Sperm traits showed substantial variation at all levels examined: between species, among males within species, and within the ejaculate of individual males. Sperm velocity was found to be positively correlated with flagellum: midpiece ratio, with relatively longer flagella associated with faster sperm. Our results document high variability in sperm form and function in lizards.     

Egg‐induced changes to sperm phenotypes shape patterns of multivariate selection on ejaculates

Sperm cells exhibit extraordinary phenotypic diversity and rapid rates of evolution, yet the adaptive value of most sperm traits remains equivocal. Recent findings suggest that to understand how selection targets ejaculates, we must recognize that female‐imposed physiological conditions often alter sperm phenotypes. These phenotypic changes may influence the relationships among sperm traits and their association with fitness. Here, we show that chemical substances released by eggs (known to modify sperm physiology and behaviour) alter patterns of selection on a suite of sperm traits in the mussel Mytilus galloprovincialis. We use multivariate selection analyses to characterize linear and nonlinear selection acting on sperm traits in (a) seawater alone and (b) seawater containing egg‐derived chemicals (egg water). Our analyses revealed that nonlinear selection on canonical axes of multiple traits (notably sperm velocity, sperm linearity and percentage of motile sperm) was the most important form of selection overall, but importantly these patterns were only evident when sperm phenotypes were measured in egg water. These findings reveal the subtle way that females can alter patterns of selection, with the implication that overlooking environmentally moderated changes to sperm, may result in erroneous interpretations of how selection targets phenotypic (co)variation in sperm traits.     

Density‐dependent patterns of multivariate selection on sperm motility and morphology in a broadcast spawning mussel

Sperm cells exhibit extraordinary phenotypic variation, both among taxa and within individual species, yet our understanding of the adaptive value of sperm trait variation across multiple contexts is incomplete. For species without the opportunity to choose mating partners, such as sessile broadcast spawning invertebrates, fertilization depends on gamete interactions, which in turn can be strongly influenced by local environmental conditions that alter the concentration of sperm and eggs. However, the way in which such environmental factors impact phenotypic selection on functional gamete traits remains unclear in most systems. Here, we analyze patterns of linear and nonlinear multivariate selection under experimentally altered local sperm densities (densities within the capture zone of eggs) on a range of functionally important sperm traits in the broadcast spawning marine mussel, Mytilus galloprovincialis. Specifically, we assay components of sperm motility and morphology across two fertilization environments that simulate either sperm limitation (when there are too few sperm to fertilize all available eggs), or sperm saturation (when there are many more sperm than required for fertilization, and the risk of polyspermy and embryonic failure is heightened). Our findings reveal that the strength, form, and targets of selection on sperm depend on the prevailing fertilization environment. In particular, our analyses revealed multiple significant axes of nonlinear selection on sperm motility traits under sperm limitation, but only significant negative directional selection on flagellum length under sperm saturation. These findings highlight the importance of local sperm densities in driving the adaptation of sperm phenotypes, particularly those related to sperm motility, in broadcast spawning invertebrates.     

Sperm morphology and evidence for sperm competition among parrots

Sperm competition is an important component of post‐copulatory sexual selection that has shaped the evolution of sperm morphology. Previous studies have reported that sperm competition has a concurrently directional and stabilizing effect on sperm size. For example, bird species that show higher levels of extrapair paternity and larger testes (proxies for the intensity of sperm competition) have longer sperm and lower coefficients of variation in sperm length, both within and between males. For this reason, these sperm traits have been proposed as indexes to estimate the level of sperm competition in species for which other measures are not available. The relationship between sperm competition and sperm morphology has been explored mostly for bird species that breed in temperate zones, with the main focus on passerine birds. We measured sperm morphology in 62 parrot species that breed mainly in the tropics and related variation in sperm length to life‐history traits potentially indicative of the level of sperm competition. We showed that sperm length negatively correlated with the within‐male coefficient of variation in sperm length and positively with testes mass. We also showed that sperm is longer in sexually dichromatic and in gregarious species. Our results support the general validity of the hypothesis that sperm competition drives variation in sperm morphology. Our analyses suggest that post‐copulatory sexual selection is also important in tropical species, with more intense sperm competition among sexually dichromatic species and among species that breed at higher densities.