Butterflies tailor their ejaculate in response to sperm competition risk and intensity

Males of many insects eclose with their entire lifetime sperm supply and have to allocate their ejaculates at mating prudently. In polyandrous species, ejaculates of rival males overlap, creating sperm competition. Recent models suggest that males should increase their ejaculate expenditure when experiencing a high risk of sperm competition. Ejaculate expenditure is also predicted to vary in relation to sperm competition intensity. During high intensity, where several ejaculates compete for fertilization of the female''s eggs, ejaculate expenditure is expected to be reduced. This is because there are diminishing returns of providing more sperm. Additionally, sperm numbers will depend on males'' ability to assess female mating status. We investigate ejaculate allocation in the polyandrous small white butterfly Pieris rapae (Lepidoptera). Males have previously been found to ejaculate more sperm on their second mating when experiencing increased risk of sperm competition. Here we show that males also adjust the number of sperm ejaculated in relation to direct sperm competition. Mated males provide more sperm to females previously mated with mated males (i.e. when competing with many sperm) than to females previously mated to virgin males (competing with few sperm). Virgin males, on the other hand, do not adjust their ejaculate in relation to female mating history, but provide heavier females with more sperm. Although virgin males induce longer non-receptive periods in females than mated males, heavier females remate sooner. Virgin males may be responding to the higher risk of sperm competition by providing more sperm to heavier females. It is clear from this study that males are sensitive to factors affecting sperm competition risk, tailoring their ejaculates as predicted by recent theoretical models.     

Male crickets adjust the viability of their sperm in response to female mating status

Sperm competition theory predicts that males should allocate sperm according to the number of competing ejaculates. Prudent allocation of sperm in response to different levels of sperm competition has been found across a number of taxa; however, some studies suggest that males may not always allocate sperm as expected. Here we examine sperm allocation in the Australian field cricket Teleogryllus oceanicus, using female mating status (virgin, singly mated, or multiply mated) to manipulate male perception of sperm competition risk and intensity. Consistent with theory, we found that male crickets adjust their ejaculates in response to female mating status. However, rather than altering the absolute numbers of sperm transferred to a female, males altered the quality of their sperm. Males ejaculated sperm of low viability (proportion of live vs. dead sperm) when mating with virgins, increased sperm viability when mating with singly mated females, but reduced sperm viability when mating with multiply mated females. Our results show that variation in ejaculate quality can be an important aspect of strategic ejaculation by males and suggest caution in the interpretation of studies in which males do not appear to allocate sperm according to theory.     

Female sperm use and storage between fertilization events drive sperm competition and male ejaculate allocation

Sperm competition theory has traditionally focused on how male allocation responds to female promiscuity, when males compete to fertilize a single clutch of eggs. Here, we develop a model to ask how female sperm use and storage across consecutive reproductive events affect male ejaculate allocation and patterns of mating and paternity. In our model, sperm use (a single parameter under female control) is the main determinant of sperm competition, which alters the effect of female promiscuity on male success and, ultimately, male reproductive allocation. Our theory reproduces the general pattern predicted by existing theory that increased sperm competition favors increased allocation to ejaculates. However, our model predicts a negative correlation between male ejaculate allocation and female promiscuity, challenging the generality of a prevailing expectation of sperm competition theory. Early models assumed that the energetic costs of precopulatory competition and the level of sperm competition are both determined by female promiscuity, which leads to an assumed covariation between these two processes. By modeling precopulatory costs and sperm competition independently, our theoretical framework allows us to examine how male allocation should respond independently to variation in sperm competition and energetic trade‐offs in mating systems that have been overlooked in the past.     

Ejaculate expenditures of male crickets in response to varying risk and intensity of sperm competition: not all species play games

Costs incurred in the manufacture of ejaculates may constrainthe number of sperm that males can produce, so males shouldshow some economy in their allocation of sperm across multiplematings. In species in which females mate with multiple malesand are capable of storing sperm for extended periods, spermallocation of males should be tailored to the risk of spermcompetition. Recent game theory predicts that males shouldtransfer the least sperm when there are no other rivals, andthe most sperm when only one other rival is likely to inseminatethe female. However, as the numbers of competitors increasesbeyond two, the models predict a corresponding decrease in ejaculate expenditure. We tested these predictions in three cricket species, Gryllodes sigillatus, Gryllus veletis, and Gryllus texensis, assessing the sperm allocation of males held under three levelsof apparent interrival competition: no rivals, one rival andsix rivals. Sperm allocation of G. veletis varied accordingto theory: males increased their sperm allocation with an increasedrisk of sperm competition (no rivals vs. one), but decreasedtheir allocation with an increased intensity of sperm competition(one rival vs. six). Sperm allocation of male G. texensis showedno significant response to the density of rivals, and spermallocation in G. sigillatus was influenced by an unexpectedinteraction between treatment density and the order in whichmales experienced the three treatments. The observed interspecificvariation in facultative sperm allocation may be due to interspecificdifferences in population density, rearing environment, orfemale mating behavior.     

Discriminating Males and Unpredictable Females: Males Bias Sperm Allocation in Favor of Virgin Females

When both sexes mate with multiple partners, theory predicts that males should adjust their investment in ejaculates in response to the risk and/or intensity of sperm competition. Here, we demonstrate that, in the harlequin beetle riding pseudoscorpion, Cordylochernes scorpioides, males use cues deposited on females by previous males to distinguish between virgin, once‐mated, and multiply‐mated females and adjust sperm allocation accordingly. Sperm number declined in direct proportion to the number of previous males, with virgin females receiving nearly three times more sperm than females exposed to three previous males. Given the lack of first‐male sperm precedence in C. scorpioides, this pattern is not consistent with current sperm competition models and appears best explained by a significant risk of wasting ejaculates on deceitful, mated females. In C. scorpioides, males transfer sperm indirectly to females via a stalked spermatophore deposited on the substrate. Mated females often feign sexual receptivity and cooperate throughout mating, only to reject the sperm packet produced by the male. While indirect sperm transfer facilitates a high level of female deceit and control, females of many species are able to influence the number and fate of sperm transferred during copulation and are likely to conceal their sexual unreceptivity to minimize male retaliation. If males cannot accurately assess female receptivity, increased risk of sperm rejection by mated females could outweigh the risk of sperm competition and favor greater sperm allocation to virgin females.     

Theoretical influence of female mating status and remating propensity on male sperm allocation patterns

Theoretical models predict that males should allocate more sperm in matings where the immediate risk of sperm competition is high. It has therefore often been argued that males should invest less sperm in matings with virgin females compared with matings with already mated females. However, with relatively polyandrous females, high sperm competition risk will covary with high sperm competition intensity leading to more unpredictable conditions, as high competition intensity should favour smaller ejaculates. With the use of a genetic algorithm, we found that males should allocate more sperm in matings with virgin females when female mating frequency is relatively high, whereas low remating rates will select for higher effort in matings with nonvirgin females. At higher remating rates, first male sperm precedence favours larger ejaculates in matings with virgin females and second male precedence favours the reverse. These results shed some light on several findings that have been difficult to explain adaptively by the hitherto developed theory on sperm allocation.     

SPERM COMPETITION GAMES: A GENERAL MODEL FOR PRECOPULATORY MALE–MALE COMPETITION

Reproductive males face a trade‐off between expenditure on precopulatory male–male competition—increasing the number of females that they secure as mates—and sperm competition—increasing their fertilization success with those females. Previous sperm allocation models have focused on scramble competition in which males compete by searching for mates and the number of matings rises linearly with precopulatory expenditure. However, recent studies have emphasized contest competition involving precopulatory expenditure on armaments, where winning contests may be highly dependent on marginal increases in relative armament level. Here, we develop a general model of sperm allocation that allows us to examine the effect of all forms of precopulatory competition on sperm allocation patterns. The model predicts that sperm allocation decreases if either the “mate‐competition loading,”a, or the number of males competing for each mating, M, increases. Other predictions remain unchanged from previous models: (i) expenditure per ejaculate should increase and then decrease, and (ii) total postcopulatory expenditure should increase, as the level of sperm competition increases. A negative correlation between a and M is biologically plausible, and may buffer deviations from the previous models. There is some support for our predictions from comparative analyses across dung beetle species and frog populations.     

Interspersed repetitive DNA sequences are unlikely to be parasitic

It is suggested that sperm competition (competition between the sperm from two or more males over the fertilization of ova) may account for the fact that sperm are so small and so numerous. In the entire absence of sperm competition, selection may favour an increase in sperm size so that the sperm contributes nutriment to the subsequent viability and success of the zygote. However, an extremely low incidence of sperm competition is adequate to prevent sperm size increasing. Vertebrate sperm should remain at minimal size provided that double matings (one female mated by two males) occur more often than about 4 times the ratio of sperm size: ovum size. The classical theory that sperm are small simply because of the difficulties of ensuring that ova do get fertilized may also explain sperm size, and both effects (sperm competition and ensuring fertilization) are likely to contribute to the stability of anisogamy. Large numbers of sperm can be produced because sperm are tiny and the optimal allocation of reproductive reserves to ejaculates is not trivially small even when double matings are rather rare. It is suggested that of its total mating effort, a male vertebrate should spend a fraction on sperm that is roughly equivalent to a quarter of the probability of double mating.     

Do Male Cook Strait Giant Weta Prudently Allocate Sperm?

Sperm production is costly and so males are expected to prudently allocate sperm to matings in a manner that maximizes their fitness. Sperm competition hypotheses predict that when facing increased sperm competition risk males should increase their investment in ejaculates. In contrast, when facing high future mating opportunities, males are expected to decrease their sperm investment in the current mating. This is because males should keep in reserve an amount of sperm proportional to their expected future mating opportunities. We experimentally tested whether male Cook Strait giant weta (Anostostomatidae: Orthoptera: Deinacrida rugosa) phenotypically adjust their investment in ejaculates in relation to their perceived risk of sperm competition and future mating opportunities. D. rugosa is a large flightless orthopteran insect in which males pass multiple spermatophores to females during a day-long mating bout. Contrary to expectation, we found that low female availability (i.e. increased sperm competition risk) had no effect on male resource allocation to sperm (i.e. number of spermatophores) compared to controls whereas, contrary to expectation, males experiencing high female availability increased their ejaculate investment by transferring significantly more spermatophores to their mates. Our results might be a consequence of males being insensitive to increased presence of rival males, reducing their allocation to sperm under increasingly risky circumstances, or due to females prolonging copulations when their perceived future mating opportunities are low.     

Male crickets adjust ejaculate quality with both risk and intensity of sperm competition

Sperm competition theory predicts that males should increase their expenditure on the ejaculate with increasing risk of sperm competition, but decrease their expenditure with increasing intensity. There is accumulating evidence for sperm competition theory, based on examinations of testes size and/or the numbers of sperm ejaculated. However, recent studies suggest that ejaculate quality can also be subject to selection by sperm competition. We used experimental manipulations of the risk and intensity of sperm competition in the cricket, Teleogryllus oceanicus. We found that males produced ejaculates with a greater percentage of live sperm when they had encountered a rival male prior to mating. However, when mating with a female that presented a high intensity of sperm competition, males did not respond to risk, but produced ejaculates with a reduced percentage of live sperm. Our data suggest that males exhibit a fine-tuned hierarchy of responses to these cues of sperm competition.     

Sperm competition and ejaculate economics

Sperm competition was identified in 1970 as a pervasive selective force in post‐copulatory sexual selection that occurs when the ejaculates of different males compete to fertilise a given set of ova. Since then, sperm competition has been much studied both empirically and theoretically. Because sperm competition often favours large ejaculates, an important challenge has been to understand the evolution of strategies through which males invest in sperm production and economise sperm allocation to maximise reproductive success under competitive conditions. Sperm competition mechanisms vary greatly, depending on many factors including the level of sperm competition, space constraints in the sperm competition arena, male mating roles, and female influences on sperm utilisation. Consequently, theoretical models of ejaculate economics are complex and varied, often with apparently conflicting predictions. The goal of this review is to synthesise the theoretical basis of ejaculate economics under sperm competition, aiming to provide empiricists with categorised model assumptions and predictions. We show that apparent contradictions between older and newer models can often be reconciled and there is considerable consensus in the predictions generated by different models. We also discuss qualitative empirical support for some of these predictions, and detail quantitative matches between predictions and observations that exist in the yellow dung fly. We argue that ejaculate economic theory represents a powerful heuristic to explain the diversity in ejaculate traits at multiple levels: across species, across males and within individual males. Future progress requires greater understanding of sperm competition mechanisms, quantification of trade‐offs between ejaculate allocation and numbers of matings gained, further knowledge of mechanisms of female sperm selection and their associated costs, further investigation of non‐sperm ejaculate effects, and theoretical integration of pre‐ and post‐copulatory episodes of sexual selection.     

SPERM COMPETITION AND THE EVOLUTION OF SEMINAL FLUID COMPOSITION

Male ejaculates include large amounts of seminal fluid proteins (Sfps) that influence male sperm competitive success. In spite of their diverse proximate functions, Sfps involved in sperm competition increase male fitness in one of three ways: (1) “avoidance” proteins help males avoid sperm competition, (2) “defense” proteins help males defend their sperm from displacement by the female's subsequent mate, and (3) “offense” proteins aid males in displacing sperm of preceding males. Here, we present a population genetic model of the evolution of allocation of finite resources by males to the three kinds of Sfps. We analyze the influence of relative efficiencies of different Sfps, of plasticity in resource allocation, and of differences in viability costs of Sfps. We find that in absence of plasticity or different viability costs, equal investment in defense and offense Sfps evolves, irrespective of their relative efficiency. In all cases, males evolve to invest more in avoidance when avoidance proteins are increasingly efficient, and when offense is more efficient than defense. Differences in viability costs result in lower investment in costly proteins, whereas plasticity has complex effects, influencing both the optimal seminal fluid composition and maintenance of variation in investment in these proteins across populations.     

Condition-dependent ejaculate size and composition in a ladybird beetle

Sexually selected male ejaculate traits are expected to depend on the resource state of males. Theory predicts that males in good condition will produce larger ejaculates, but that ejaculate composition will depend on the relative production costs of ejaculate components and the risk of sperm competition experienced by low- and high-condition males. Under some conditions, when low condition leads to poorer performance in sperm competition, males in low condition may produce ejaculates with higher sperm content relative to their total ejaculate and may even transfer more sperm than high-condition males in an absolute sense. Previous studies in insects have shown that males in good condition transfer larger ejaculates or more sperm, but it has not been clear whether increased sperm content represents a shift in allocation or simply a larger ejaculate, and thus the condition dependence of ejaculate composition has been largely untested. We examined condition dependence in ejaculate by manipulating adult male condition in a ladybird beetle (Adalia bipunctata) in which males transfer three distinct ejaculate components during mating: sperm, non-sperm ejaculate retained within the female reproductive tract, and a spermatophore capsule that females eject and ingest following mating. We found that high condition males indeed transferred larger ejaculates, potentially achieved by an increased rate of ejaculate transfer, and allocated less to sperm compared with low-condition males. Low-condition males transferred ejaculates with absolutely and proportionally more sperm. This study provides the first experimental evidence for a condition-dependent shift in ejaculate composition.     

Releasing small ejaculates slowly increases per‐gamete fertilization success in an external fertilizer: Galeolaria caespitosa (Polychaeta: Serpulidae)

The idea that male reproductive strategies evolve primarily in response to sperm competition is almost axiomatic in evolutionary biology. However, externally fertilizing species, especially broadcast spawners, represent a large and taxonomically diverse group that have long challenged predictions from sperm competition theory—broadcast spawning males often release sperm slowly, with weak resource‐dependent allocation to ejaculates despite massive investment in gonads. One possible explanation for these counter‐intuitive patterns is that male broadcast spawners experience strong natural selection from the external environment during sperm dispersal. Using a manipulative experiment, we examine how male reproductive success in the absence of sperm competition varies with ejaculate size and rate of sperm release, in the broadcast spawning marine invertebrate Galeolaria caespitosa (Polychaeta: Serpulidae). We find that the benefits of Fast or Slow sperm release depend strongly on ejaculate size, but also that the per‐gamete fertilization rate decreases precipitously with ejaculate size. Overall, these results suggest that, if males can facultatively adjust ejaculate size, they should slowly release small amounts of sperm. Recent theory for broadcast spawners predicts that sperm competition can also select for Slow release rates. Taken together, our results and theory suggest that selection often favours Slow ejaculate release rates whether males experience sperm competition or not.     

Male scorpionflies assess the amount of rival sperm transferred by females' previous mates

Theory predicts that when sperm compete numerically, selection will favor males who vary the number of sperm they transfer with the immediate level of sperm competition. In this study, I measured male mating investment in response to both female mating status (virgin vs. mated) and the number of foreign sperm stored by females in a previous mating in the scorpionfly Panorpa cognata. Female sperm storage was manipulated by interrupting copulations at different time points. Female mating status did not significantly influence male mating investment, but resource-limited males invested strategically in relation to the amount of sperm stored by females in a previous mating. I found continuously decreasing male investment in response to increasing amounts of competing sperm. These results demonstrate an unprecedented male ability to assess the number of sperm stored by females. As a result, males are capable of an extraordinarily fine-tuned reaction to the intensity of sperm competition.     

Sperm competition affects male behaviour and sperm output in the rainbow darter

Rainbow darters, Etheostoma caeruleum, are promiscuous fish with moderate rates of group spawning (between one and five males may simultaneously mate with one female). In this study, I examined male sperm output and male willingness to spawn under different levels of sperm competition intensity. One male and one female were allowed to spawn in an aquarium where they had visual and olfactory access to one of four treatments: four males, one male, zero males, or one female. Theory predicts that males should reduce sperm output when there are more than the average number of males at a group spawning (four-male treatment) and should increase sperm output when there are fewer than average males at a group spawning (one-male treatment). Mean sperm output did not differ among treatments. However, males released more sperm when spawning in the presence of competing males (four-male and one-male treatments pooled) than when spawning in the absence of competing males (zero-male and one-female treatments pooled). Males were also most likely to forego spawning opportunities when sperm competition intensity was high. Furthermore, male willingness to spawn was size dependent. Large males were more likely to forego spawning opportunities under high sperm competition intensity. Large males may be better off waiting for future spawning opportunities when there is a lower potential for sperm competition intensity.     

Strategic allocation of ejaculates by male Adélie penguins

Sperm competition theory suggests that males should strategically allocate sperm to those females that will bring them the best possible genetic returns. Although males of a number of species of insects and fishes have been shown to allocate sperm strategically, we provide, to our knowledge, the first evidence that an avian species is also capable of allocating ejaculates. Male Adélie penguins (Pygoscelis adeliae) are more likely to transfer sperm during extra-pair copulations (EPCs) than during pair copulations. We investigated the question of how males allocate ejaculates within the constraints of limited sperm availability and found (i) that males that engaged in EPC attempts ejaculated less often when copulating with their social partner than males that made no EPC attempts, and (ii) that there was no difference between males that were involved in failed EPC attempts and those that were involved in successful EPCs in the proportion of copulations that resulted in sperm transfer. These results indicate that males achieve strategic allocation of sperm within the constraints of limited sperm availability by withholding ejaculates from their social partners.     

Sperm competition games: a comparison of loaded raffle models and their biological implications

Our main aim is to compare the additive model, due to Mesterton-Gibbons, and the multiplicative model, due to Parker, of sperm allocation under sperm competition, when other influences are treated in the same way. We first review these (and other) models and their foundations, leading to a generalization of the multiplicative model. Sperm is assumed to cost energy, and this constraint is incorporated differently in the two models. These give the same results in the random-roles situation when the males occupy roles (of first and second to mate) randomly: the number of sperm ejaculated in the favoured role is greater than that in the disfavoured role by an amount that depends on the effect of sperm limitation (i.e. the probability that there is insufficient sperm to ensure full fertility). If the latter is negligible, or the fertilization raffle fair, this difference is zero, as Parker found originally. In the constant roles situation (where males of a particular type always occupy the same role) the predictions differ: the additive model has the same predictions as in the random roles case, but the multiplicative model predicts that males of the type occupying the favoured role ejaculate less than males of the type occupying the disfavoured role, in accord with Parker's original conclusion. The fitnesses of the two types of male can be calculated in the multiplicative model: the fitness of the favoured male is usually higher, even if he has to expend more energy in "finding" a female, e.g. through fighting, etc. These conclusions relate to inter-male behaviour (i.e. of different male types), as distinct from intra-male behaviour (i.e. of a given male when in different roles). We analyse situations in which one male type has some probability of acting in its less usual role: calculations with varying amounts of sperm limitation are presented. It is found that the presence of a male of a different type has an effect on intra-male ejaculate behaviour, which also depends critically on the role usually occupied. We conclude that the multiplicative model is the more accurate model and provides more information. Some experimental data on sperm numbers are used to find the effects of sperm limitation. For species which conform to the loaded raffle model, sperm limitation typically has small or negligible effects: in this case, we argue that empiricists should look for equal ejaculates in the two roles when studying random role situations; when roles are occupied non-randomly average sperm expenditure should be greater by male types typically occupying the disfavoured role, but within a male type, expenditure should be greater in the role it typically occupies.     

MALE CONTEST COMPETITION AND THE COEVOLUTION OF WEAPONRY AND TESTES IN PINNIPEDS

Male reproductive success is influenced by competitive interactions during precopulatory and postcopulatory selective episodes. Consequently, males can gain reproductive advantages during precopulatory contest competition by investing in weaponry and during postcopulatory sperm competition by investing in ejaculates. However, recent theory predicts male expenditure on weaponry and ejaculates should be subject to a trade‐off, and should vary under increasing risk and intensity of sperm competition. Here, we provide the first comparative analysis of the prediction that expenditure on weaponry should be negatively associated with expenditure on testes mass. Specifically, we assess how sexual selection influences the evolution of primary and secondary sexual traits among pinnipeds (seals, sea lions, and walruses). Using recently developed comparative methods, we demonstrate that sexual selection promotes rapid divergence in body mass, sexual size dimorphism (SSD), and genital morphology. We then show that genital length appears to be positively associated with the strength of postcopulatory sexual selection. However, subsequent analyses reveal that both genital length and testes mass are negatively associated with investment in precopulatory weaponry. Thus, our results are congruent with recent theoretical predictions of contest‐based sperm competition models. We discuss the possible role of trade‐offs and allometry in influencing patterns of reproductive trait evolution in pinnipeds.     

Sperm competition games: sperm selection by females

We analyse a co-evolutionary sexual conflict game, in which males compete for fertilizations (sperm competition) and females operate sperm selection against unfavourable ejaculates (cryptic female choice). For simplicity, each female mates with two males per reproductive event, and the competing ejaculates are of two types, favourable (having high viability or success) or unfavourable (where progeny are less successful). Over evolutionary time, females can increase their level of sperm selection (measured as the proportion of unfavourable sperm eliminated) by paying a fecundity cost. Males can regulate sperm allocations depending on whether they will be favoured or disfavoured, but increasing sperm allocation reduces their mating rate. The resolution of this game depends on whether males are equal, or unequal. Males could be equal: each is favoured with probability, p, reflecting the proportion of females in the population that favour his ejaculate (the 'random-roles' model); different males are favoured by different sets of females. Alternatively, males could be unequal: given males are perceived consistently by all females as two distinct types, favoured and disfavoured, where p is now the frequency of the favoured male type in the population (the 'constant-types' model). In both cases, the evolutionarily stable strategy (ESS) is for females initially to increase sperm selection from zero as the viability of offspring from unfavourable ejaculates falls below that of favourable ejaculates. But in the random-roles model, sperm selection decreases again towards zero as the unfavourable ejaculates become disastrous (i.e. as their progeny viability decreases towards zero). This occurs because males avoid expenditure in unfavourable matings, to conserve sperm for matings in the favoured role where their offspring have high viability, thus allowing females to relax sperm selection. If sperm selection is costly to females, ESS sperm selection is high across a region of intermediate viabilities. If it is uncostly, there is no ESS in this region unless sperm limitation (i.e. some eggs fail to be fertilized because sperm numbers are too low) is included into the model. In the constant-types model, no relaxation of sperm selection occurs at very low viabilities of disfavoured male progeny. If sperm selection is sufficiently costly, ESS sperm selection increases as progeny viability decreases down towards zero; but if it is uncostly, there is no ESS at the lowest viabilities, and unlike the random-roles model, this cannot be stabilized by including sperm limitation. Sperm allocations in the ESS regions differ between the two models. With random roles, males always allocate more sperm in the favoured role. With constant types, the male type that is favoured allocates less sperm than the disfavoured type. These results suggests that empiricists studying cryptic female choice and sperm allocation patterns need to determine whether sperm selection is applied differently, or consistently, on given males by different females in the same population.