Sperm competition and reproductive success in the decapod Inachus phalangium (Majidae): a male ghost spider crab that seals off rivals' sperm

Parker's (1970a) hypothesis that the overlap of multiple mating sperm in the female's storage organs promotes sperm competition is tested here for the first time in Crustacea: specifically, the mechanisms and consequences of sperm competition are detailed for the spider crab Inachus phalangium . Females of this species store ejaculates from successive copulations with different males discretely and consecutively in sac-like twin seminal receptacles. During copulation males transfer a large quantity of a sperm-free seminal plasma, followed by the sperm which is stored in small spermatophores and forms a densely-packed sperm packet. It was shown, using ³H-thymidine-labelled ejaculate, that the last male to mate displaces the ejaculate of his predecessors dorsally into the apex of the receptacle. Sperm of previous matings are sealed in with the hardening seminal plasma (sperm gel) and are thus prevented from being used to fertilize eggs, while the last male to mate places his sperm closest to the oviduct and vaginal openings. In experiments using the 'sterile-male' method, sperm from the last male to mate gained all fertilizations in subsequent broods. The seminal plasma forms the sperm gel in ghost spider crabs which is used for displacement of previously stored sperm, whereas various other brachyuran taxa use seminal plasma to produce the sperm plug, which prevents a male's sperm from being displaced.     

Intraspecific variation of P2 value in a coccinellid beetle,Harmonia axyridis

Intraspecific variation in P2 value in a coccinellid beetle (Harmonia axyridis) was investigated. The analytical method by Parker et al. (1990) predicts that sperm-flushing displacement in the spermatheca may exist in the sperm utilization pattern of this species. Long duration of sperm transfer in the second copulation resulted in high fertilization success of the second male. Large male body size itself did not have an advantage in flushing efficiency of the previously stored sperm. However, through long duration of sperm transfer and larger ejaculate, males with large body size gain high fertilization success.     

FEMALE CONTROL OF SPERM TRANSFER AND INTRASPECIFIC VARIATION IN SPERM PRECEDENCE: ANTECEDENTS TO THE EVOLUTION OF A COURTSHIP FOOD GIFT

Manipulation of ejaculates is believed to be an important avenue of female choice throughout the animal kingdom, but evidence of its importance to sexual selection remains scarce. In crickets, such manipulation is manifest in the premature removal of the externally attached spermatophore, which may afford females an important means of postcopulatory mate choice. We tested the hypothesis that premature spermatophore removal contributes significantly to intraspecific variation in sperm precedence by (1) experimentally manipulating spermatophore attachment durations of competing male Gryllodes sigillatus and (2) employing protein electrophoresis to determine the paternity of doubly mated females. The relative spermatophore attachment durations of competing males had a significant influence on male paternity, but the pattern of sperm precedence deviated significantly from the predictions of an ideal lottery. Instead, paternity data and morphological evidence accorded best with a model of partial sperm displacement derived here. Our model is similar to a displacement model of Parker et al. in that sperm of the second male mixes instantaneously with that of the first throughout the displacement process, but the novel feature of our model is that the number of sperm displaced is only a fraction of the number of sperm transferred by the second male. Regardless of the underlying mechanism, female G. sigillatus can clearly alter the paternity of their offspring through their spermatophore-removal behavior, and employ such cryptic choice in favoring larger males and those providing larger courtship food gifts. We discuss how female control of sperm transfer and intraspecific variation in sperm precedence may be important precursors to the evolution of gift giving in insects.     

Linear models for assessing mechanisms of sperm competition: the trouble with transformations

Abstract.— Although sperm competition is a pervasive selective force shaping the reproductive tactics of males, the mechanisms underlying different patterns of sperm precedence remain obscure. Parker et al. (1990) developed a series of linear models designed to identify two of the more basic mechanisms: sperm lotteries and sperm displacement; the models can be tested experimentally by manipulating the relative numbers of sperm transferred by rival males and determining the paternity of offspring. Here we show that tests of the model derived for sperm lotteries can result in misleading inferences about the underlying mechanism of sperm precedence because the required inverse transformations may lead to a violation of fundamental assumptions of linear regression. We show that this problem can be remedied by reformulating the model using the actual numbers of offspring sired by each male, and log-transforming both sides of the resultant equation. Reassessment of data from a previous study (Sakaluk and Eggert 1996) using the corrected version of the model revealed that we should not have excluded a simple sperm lottery as a possible mechanism of sperm competition in decorated crickets, Gryllodes sigillatus .     

Consequences of sperm displacement for female dung flies,Scatophaga stercoraria

Displacement of stored sperm during copulation occurs in many insects. This process provides direct benefits for males via increased fertilization success, but the fitness consequences of sperm displacement for females are less clear. Here we investigate potential benefits of sperm displacement for female yellow dung flies, Scatophaga stercoraria. We find no evidence that female dung flies gain direct benefits from displacement of previously stored sperm in terms of increased fertility or fecundity. There was no difference in the relative survival rate, development time, size or fluctuating asymmetry of offspring produced by females that had previously stored sperm displaced before oviposition and those that did not. Females using previously stored sperm to fertilize their eggs produced significantly higher ratios of male to female offspring. These novel findings have important implications for understanding the evolutionary dynamics of male–female interactions in sperm competition.     

Optimal copula duration in yellow dung flies: ejaculatory duct dimensions and size-dependent sperm displacement

We aim to interpret sperm displacement in relation to male size in the yellow dung fly, Scatophaga stercoraria, and to compare the general properties of indirect and direct size-dependent sperm displacement in insects. We examine the hypothesis that male size-dependent sperm displacement in dung flies can be explained by size-dependent increases in the ejaculatory apparatus, allowing greater sperm flow rates in larger males. We expect sperm flow rates to be proportional to the diameter of the aedeagus duct to the power x, where x lies between 2 and 3. We test this hypothesis using a simulation model of indirect sperm displacement that has been developed to accommodate recent observations on sperm transfer, in which sperm flow from the male into the female bursa and are then transferred to the spermathecae by movements of the female tract. The indirect model approximates to the pattern of size-related sperm displacement, with scaling power 3 giving a better fit than power 2. Copula duration shows a male size-dependent decrease in this species. We apply the indirect model of sperm displacement, in conjunction with parameters obtained from field and laboratory data, to predict size-dependent changes in optimal copula duration from the male perspective. This model concurs with the observations by predicting a size-dependent decline in optimal copula duration, as did an earlier model in which displacement was direct (new sperm displace previously stored sperm directly from the sperm stores). Our new approach gives a better fit than the earlier direct model. Thus, both results (displacement rates and copula duration) can be explained by size-dependent changes in the ejaculatory apparatus of the male with the female's exchange rate of sperm (from bursa to spermathecae) remaining constant with respect to male size, although we discuss the possibility that this female process may accelerate with increased male size. In general, where the sperm input rate is around the same magnitude as the exchange rate, indirect displacement will be dependent on the size of the male, as in dung flies, but this dependency is lost if the input rate is very high relative to the exchange rate across the entire range of male size. Size-dependent displacement should always apply for males with direct displacement.     

Functional morphology of the female reproductive system of a crab with highly extensible seminal receptacles and extreme sperm storage capacity

We studied the functional morphology of the female reproductive system of the purple stone crab Danielethus crenulatus . The most remarkable feature is the relative storage capacity and extensibility of the seminal receptacles. These receptacles are a pair of simple sacs that lack internal structures dividing the internal lumen. Differences in seminal receptacle size and contents are accompanied by conspicuous changes in receptacle lining at a tissue level. Full seminal receptacles contain discrete sperm masses formed by hardened fluid and densely packed spermatophores. Different sperm masses are likely from different mates and their stratified disposition within the seminal receptacles is compatible with rival sperm displacement and last sperm precedence. Additionally, the anatomical structure of the vulva and vagina suggest active female control over copula. We discuss our results in the general context of sperm storage in brachyurans and the implications for the mating system of this species.     

INDIVIDUAL VARIATION IN SPERM COMPETITION SUCCESS OF YELLOW DUNG FLIES,SCATOPHAGA STERCORARIA

Intraspecific variation in the proportion of offspring sired by the second male to mate with a female (P₂) is an aspect of sperm competition that has received little attention. We examined variation in the sperm competition success of individual male dung flies, Scatophaga stercoraria. In unmanipulated matings, copula duration was dependent on male size with smaller males copulating for longer. A principal component analysis was used to generate uncorrelated scores based on a male's size and copula duration. Using these scores demonstrated that P₂ values were dependent both on the relative size and copula durations of competing males. When copula duration was held constant, the success of an individual male increased as his body size, relative to the first male, increased. We interrupted copulations of “large” and “small” second males and fitted the resultant P₂ values to a linear model of sperm competition with unequal ejaculates. The data fit well to a model of sperm displacement in which sperm mix quickly on introduction to the sperm stores. Furthermore, they show that “large” males have a greater rate of sperm displacement than “small” males. The levels of prey availability during testis maturation may influence a male's success in sperm competition although his immediate mating history does not. We show why an understanding of variation in sperm competition success is important for understanding the mechanisms and evolutionary significance of sperm competition.     

Evidence for specific binding of uncapacitated boar spermatozoa to porcine zonae pellucidae in vitro

Washed ejaculated boar sperm and sperm from the cauda epididymis bind to the zona pellucida of fixed porcine eggs in large numbers. Sperm incubated in the presence of dextran sulfate (8 K daltons or 500 K daltons) or fucoidan and then washed no longer bind to eggs. Other acid carbohydrates (heparin, chondroitin sulfates, inositol hexasulfate, carboxymethylcellulose) fail to block sperm-egg binding even when added directly to sperm-egg suspensions. Seminal plasma and the seminal vesicle secretion contain basic proteins which bind tightly to sperm and bind reversibly to eggs preventing sperm from binding to eggs. When dextran sulfate or fucoidan are mixed with the vesicular secretion, from which seminal plasma basic proteins originate (Hunt et al., '83), the secretion loses the capacity to prevent sperm from binding to eggs; this suggests that seminal vesicle proteins can bind to the same site on zonae as do sperm and thus seminal plasma may modify sperm-egg interactions. Corpus and cauda epididymal sperm also bind in large numbers to the zona pellucida of isolated eggs but high concentrations of caput sperm, which exhibit high motility in the presence of caffeine, bind only in few numbers. Thus a component that enhances sperm-zona binding is apparently formed on the plasma membranes of uncapacitated sperm during passage through the epididymis. This finding, and an earlier observation that antibodies raised against uncapacitated sperm plasma membranes block sperm-egg binding in vivo (Peterson et al., '83) suggest that this component may be involved in sperm zona interaction in vivo.     

Purification and partial characterization of the 17 kDa sperm coating protein from boar seminal plasma.

Sperm coating proteins of 16, 17, and 19 kDa have been purified from boar seminal plasma. The 17 kDa protein has been identified as an antigen recognized by monoclonal antibody ACR.3 and is thus identical to low molecular mass zona pellucida binding protein from boar spermatozoa (Moos et al., 1990). The 17 and 19 kDa proteins are glycosylated and tend to form hetero-complexes. The 17 kDa ACR.3 antigen is sequentially released from the sperm cell surface during capacitation and, after induction of the acrosome reaction, the 16 kDa form was also observed. Immunocytochemical studies on boar reproductive tissues have suggested that the seminal vesicle epithelium may be the source of these proteins.     

Female remating, sperm competition and sexual selection in Drosophila

Female remating is fundamental to evolutionary biology as it determines the pattern of sexual selection and sexual conflict. Remating in females is an important component of Drosophila mating systems because it affects sperm usage patterns and sexual selection. Remating is common in females of many species of Drosophila in both natural and laboratory populations. It has been reported in many insect species and also in vertebrates. Female remating is a prerequisite for sperm competition between males, and the consequences of this competition, such as sperm precedence or sperm displacement, have been reported for many species of Drosophila. Female remating is dependent on the amount of sperm stored, the male seminal fluid components, nutrition, the quantity of eggs laid, experimental design and density of flies in laboratory. Remating by a female is an insurance against male sterility and sub-fertility and increases genetic heterogeneity of female offspring. Remating gives greater female productivity in many species of Drosophila. We examined female remating with respect to sperm competition and sexual selection in Drosophila and addressed the possible benefits for females. We also reviewed the role of accessory gland fluid in remating, costs associated with remating, the genetic basis of female remating and some possible mechanisms of sperm competition in the light of last male sperm priority and paternity assurance in Drosophila and other insects. We also suggest future areas of research.     

Variation in Sperm Displacement and Its Association with Accessory Gland Protein Loci in Drosophila Melanogaster

Genes that influence mating and/or fertilization success may be targets for strong natural selection. If females remate frequently relative to the duration of sperm storage and rate of sperm use, sperm displacement may be an important component of male reproductive success. Although it has long been known that mutant laboratory stocks of Drosophila differ in sperm displacement, the magnitude of the naturally occurring genetic variation in this character has not been systematically quantified. Here we report the results of a screen for variation in sperm displacement among 152 lines of Drosophilia melanogaster that were made homozygous for second and/or third chromosomes recovered from natural populations. Sperm displacement was assayed by scoring the progeny of cn;bw females that had been mated sequentially to cn;bw and tested males in either order. Highly significant differences were seen in both the ability to displace sperm that is resident in the female's reproductive tract and in the ability to resist displacement by subsequent sperm. Most lines exhibited nearly complete displacement, having nearly all progeny sired by the second male, but several lines had as few as half the progeny fathered by the second male. Lines that were identified in the screen for naturally occurring variation in sperm displacement were also characterized for single-strand conformation polymorphisms (SSCP) at seven accessory gland protein (Acp) genes, Glucose dehydrogenase (Gld), and Esterase-6 (Est-6). Acp genes encode proteins that are in some cases known to be transmitted to the female in the seminal fluid and are likely candidates for genes that might mediate the phenomenon of sperm displacement. Significant associations were found between particular Acp alleles at four different loci (Acp26Aa/Ab, Acp29B, Acp36DE and Acp53E) and the ability of males to resist displacement by subsequent sperm. There was no correlation between the ability to displace resident sperm and the ability to resist being displaced by subsequent sperm. This lack of correlation, and the association of Acp alleles with resisting subsequent sperm only, suggests that different mechanisms mediate the two components of sperm displacement.     

Comparison of models for flow induced deformation of soft biological tissue

The behaviour of a deformable porous medium during the flow of fluid under a pressure difference is examined for both infinitesimal and finite deformations. Models for both cases are solved for the problem of steady one-dimensional compression and compared with experimental data from Parker et al. (J. appl. Mech. 54, 794-800, 1987) for a polyurethane sponge. The purpose of this study is to identify a simple model which agrees qualitatively with these published results. To relate the stress relations for biological tissues to the data for polymer sponges (Parker et al., 1987) a translation of 1.1 kPa was introduced. This allows for some structural differences between the two media. It was found that the infinitesimal models were adequate up to 20% strain, but significant divergence occurred for higher strains. A finite deformation model with the permeability depending exponentially on the strain gave the most consistent results and required the fitting of only two parameters.     

Seminal fluid transferrin as an index of gonadal function in men

Seminal fluid transferrin concentrations of proven fertile donors and normozoospermic patients were significantly higher (P less than 0.001) than those in other groups examined. There were no significant differences in the transferrin values among vasectomized, azoospermic and very severe oligozoospermic subjects. Values were also similar in patients affected by secretory or excretory azoospermia. Regression analysis showed a positive correlation (P less than 0.001, r = 0.72) between seminal fluid transferrin concentrations and sperm density. A negative correlation (P less than 0.02, r = 0.28) existed between circulating FSH and seminal fluid transferrin concentrations. There were no significant differences between seminal fluid transferrin and the percentages of abnormal sperm cells or immature seminal line elements. These results indicate that the nature of seminiferous tubule dysfunction can be precisely defined by examining seminal fluid transferrin in combination with other biological values usually used to explore testicular function.     

Evidence for biased use of sperm sources in wild female giant cuttlefish (Sepia apama)

In species where females store sperm from their mates prior to fertilization, sperm competition is particularly probable. Female Sepia apama are polyandrous and have access to sperm from packages (spermatangia) deposited by males onto their buccal area during mating and to sperm stored in internal sperm-storage organs (receptacles) located below the beak. Here, we describe the structure of the sperm stores in the female's buccal area, use microsatellite DNA analyses to determine the genetic diversity of stored sperm and combine these data with offspring genotypes to determine the storage location of paternal sperm. The number of male genotypes represented in the sperm receptacles was significantly lower than that found among the spermatangia. Estimation of the volumes of sperm contained in the receptacles and the spermatangia were statistically comparable; however, paternal sperm were more likely to have come from spermatangia than from the sperm receptacles. These results confirm a genetic polyandrous mating system in this species and suggest that fertilization pattern with respect to the sperm stores used is not random.     

Mated Drosophila melanogaster females require a seminal fluid protein, Acp36DE, to store sperm efficiently.

Mated females of many animal species store sperm. Sperm storage profoundly influences the number, timing, and paternity of the female's progeny. To investigate mechanisms for sperm storage in Drosophila melanogaster, we generated and analyzed mutations in Acp36DE. Acp36DE is a male seminal fluid protein whose localization in mated females suggested a role in sperm storage. We report that male-derived Acp36DE is essential for efficient sperm storage by females. Acp36DE(1) (null) mutant males produced and transferred normal amounts of sperm and seminal fluid proteins. However, mates of Acp36DE(1) males stored only 15% as many sperm and produced 10% as many adult progeny as control-mated females. Moreover, without Acp36DE, mated females failed to maintain an elevated egg-laying rate and decreased receptivity, behaviors whose persistence (but not initiation) normally depends on the presence of stored sperm. Previous studies suggested that a barrier in the oviduct confines sperm and Acp36DE to a limited area near the storage organs. We show that Acp36DE is not required for barrier formation, but both Acp36DE and the barrier are required for maximal sperm storage. Acp36DE associates tightly with sperm. Our results indicate that Acp36DE is essential for the initial storage of sperm, and that it may also influence the arrangement and retention of stored sperm.     

Optimal copula duration in yellow dung flies: effects of female size and egg content

We examine data on copula duration in dung flies, Scatophaga stercoraria, in relation to female phenotype. We use a marginal value theorem approach based on the plausible mechanisms of sperm competition to predict the effect of female variation on optimal copula duration, t *, from the male perspective. Future fertilizations are expected to have a trivial effect on t * with fully gravid females, but an increasing relative effect on t * towards completion of oviposition. t * is expected to be affected by female size because of variation in (1) a female's egg content, which increases the maximum egg gain available from a mating, and (2) the female reproductive tract, which affects the rate at which sperm are displaced. In fully gravid females, t * was not dependent on egg number variation, but showed a positive relation with egg content in females that had laid a varying proportion of their mature egg load at the time of mating, and were therefore not fully gravid. Our models predict that if a male can estimate egg content only by the distension of a female's abdomen, t * should increase in a similar way to that seen with 'take-over' females. We predict t * for fully gravid females by assuming that males can monitor female size. The data showed that sperm displacement rate decreased, and average egg content increased, with female size. Under two models for a sperm displacement mechanism, one (which assumes indirect displacement at a rate proportional to the increase in spermathecal volume) predicts the observed relation between t * and female size almost exactly. Small males copulated for longer than large males (as predicted and reported previously). Copyright 1999 The Association for the Study of Animal Behaviour.     

Sperm competition and the evolution of ejaculate composition

We present a model of sperm competition that incorporates both sperm and nonsperm parts of the ejaculate. Our primary focus is on determining how ejaculate composition and size evolves as a function of the effects of seminal fluid on male reproductive success and as a function of asymmetry in sperm usage by females. The model predicts that different patterns of investment in sperm and seminal products are expected to evolve as a function of the bias in sperm usage by females. It also predicts the evolution of distinct patterns in ejaculate composition depending on the function of seminal fluid. In the discussion, we highlight a number of potential approaches for testing the theory that we develop.     

Male mating rate is constrained by seminal fluid availability in bedbugs, Cimex lectularius

Sexual selection, differences in reproductive success between individuals, continues beyond acquiring a mating partner and affects ejaculate size and composition (sperm competition). Sperm and seminal fluid have very different roles in sperm competition but both components encompass production costs for the male. Theoretical models predict that males should spend ejaculate components prudently and differently for sperm and seminal fluid but empirical evidence for independent variation of sperm number and seminal fluid volume is scarce. It is also largely unknown how sperm and seminal fluid variation affect future mating rate. In bedbugs we developed a protocol to examine the role of seminal fluids in ejaculate allocation and its effect on future male mating rate. Using age-related changes in sperm and seminal fluid volume we estimated the lowest capacity at which mating activity started. We then showed that sexually active males allocate 12% of their sperm and 19% of their seminal fluid volume per mating and predicted that males would be depleted of seminal fluid but not of sperm. We tested (and confirmed) this prediction empirically. Finally, the slightly faster replenishment of seminal fluid compared to sperm did not outweigh the faster decrease during mating. Our results suggest that male mating rate can be constrained by the availability of seminal fluids. Our protocol might be applicable to a range of other organisms. We discuss the idea that economic considerations in sexual conflict research might benefit from distinguishing between costs and benefits that are ejaculate dose-dependent and those that are frequency-dependent on the mating rate per se.     

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.