The direct cost of traumatic secretion transfer in hermaphroditic land snails: individuals stabbed with a love dart decrease lifetime fecundity

Several taxa of simultaneously hermaphroditic land snails exhibit a conspicuous mating behaviour, the so-called shooting of love darts. During mating, such land snail species transfer a specific secretion by stabbing a mating partner''s body with the love dart. It has been shown that sperm donors benefit from this traumatic secretion transfer, because the secretions manipulate the physiology of a sperm recipient and increase the donors'' fertilization success. However, it is unclear whether reception of dart shooting is costly to the recipients. Therefore, the effect of sexual conflict and antagonistic arms races on the evolution of traumatic secretion transfer in land snails is still controversial. To examine this effect, we compared lifetime fecundity and longevity between the individuals that received and did not receive dart shooting from mating partners in Bradybaena pellucida. Our experiments showed that the dart-receiving snails suffered reduction in lifetime fecundity and longevity. These results suggest that the costly mating behaviour, dart shooting, generates conflict between sperm donors and recipients and that sexually antagonistic arms races have contributed to the diversification of the morphological and behavioural traits relevant to dart shooting. Our findings also support theories suggesting a violent escalation of sexual conflict in hermaphroditic animals.     

The darting game in snails and slugs

Love darts are hard 'needles' that many snails and slugs use to pierce their partner during mating. In a few species, darts have been shown to play a role in sperm competition. Two new papers, by Davison et al., and Koene and Schulenburg, might further pique researchers' interest, because they show how the full potential of darts can be tapped for studies of sexual selection in hermaphrodites.     

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.     

Multiple paternity and postcopulatory sexual selection in a hermaphrodite: what influences sperm precedence in the garden snail Helix aspersa?

Sperm competition has been studied in many gonochoric animals but little is known about its occurrence in simultaneous hermaphrodites, especially in land snails. The reproductive behaviour of the land snail Helix aspersa involves several features, like multiple mating, long-term sperm storage and dart-shooting behaviour, which may promote sperm competition. Cryptic female choice may also occur through a spermatheca subdivided into tubules, which potentially allows compartmentalized sperm storage of successive mates. In order to determine the outcome of postcopulatory sexual selection in this species, we designed a cross-breeding experiment where a recipient ('female') mated with two sperm donors ('males'). Mates came from either the same population as the recipient or from a distinct one. To test for the influence a recipient can have on the paternity of its offspring, we excluded the effects of dart shooting by using only virgin snails as sperm donors because they do not shoot any dart before their first copulation. We measured the effects of size of mates as well as time to first and second mating on second mate sperm precedence (P2; established using microsatellite markers). Multiple paternity was detected in 62.5% of clutches and overall there was first-mate sperm precedence with a mean P2 of 0.24. Generalized linear modelling revealed that the best predictors of paternity were the time between matings and the time before first mating. Overall, both first and second mates that copulated quickly got greater parentage, which may suggest that postcopulatory events influence patterns of sperm precedence in the garden snail.     

Random mating with respect to mating status in the simultaneously hermaphroditic land snail Arianta arbustorum

In promiscuous species with sperm storage, males are expected to show a preference for mating with virgin and young females to reduce the risk of sperm competition. In the simultaneous hermaphrodite land snail Arianta arbustorum, sperm production precedes egg production by 2–4 weeks, resulting in a short period of protandric hermaphroditism before shell growth is completed. In natural populations, copulating pairs involving individuals which have not yet completed shell growth (virgins) have been observed. We ran a series of mate-choice experiments to examine whether virgin and nonvirgin (experienced) individuals of A. arbustorum discriminate between virgin and nonvirgin mating partners. We also assessed the number of sperm delivered to partners with different mating status. Neither virgin nor nonvirgin snails showed any preference for mating with a virgin partner. In all test situations mating was random and the number of sperm delivered was not adjusted to the mating status of the partner. Mating success was mainly determined by the activity of the individual. The random mating pattern does not imply random fertilization of eggs because the presence of a sperm-digesting organ and the morphology of the sperm storage organ allow a selective storage and use of sperm in A. arbustorum.     

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.     

Body size-dependent gender role in a simultaneous hermaphrodite freshwater snail, Physa acuta

We examined whether gender role in the simultaneous hermaphroditefreshwater snail, Physa acuta, is determined by relative bodysize in a manner predicted by the size-advantage model. We observedthe body-size combinations of pairs in the laboratory by usingfield-collected populations. Smaller individuals tended to playthe "male" role (sperm donor), and larger snails the "female"(sperm recipient). Next, we analyzed the mating behaviors involvedin gender-role decision in snail pairs of three different body-sizecombinations, using "large" and "small" snails. Smaller snailswere more likely to approach the partner as a male in different-sizecombination (large/small), whereas frequent initial approachesas a male and rejection behavior as a female were observed inthe large/large combination. Third, we examined the body sizepreference when a snail can freely choose the partner from twoother individuals of different body sizes (large/large/smallor large/small/small). Small individuals had a significant tendencyto act as the male and positively selected large snails as thefemale partner in both triple combinations. However, the largeindividual acted as both the male and the female with nearlyequal frequency. In the size-differing pairings, copulationsoccurred after fewer male approaches and fewer rejections thanin pairings involving two large snails, suggesting that bodysize difference is one of the behavioral solutions in genderconflict. Clear gender-role switching associated with body sizewas not seen. Smaller snails thus have a tendency to play themale role more frequently but adopt both gender roles when theirbody size is sufficiently large.     

Mating triggers dynamic immune regulations in wood ant queens

Mating can affect female immunity in multiple ways. On the one hand, the immune system may be activated by pathogens transmitted during mating, sperm and seminal proteins, or wounds inflicted by males. On the other hand, immune defences may also be down‐regulated to reallocate resources to reproduction. Ants are interesting models to study post‐mating immune regulation because queens mate early in life, store sperm for many years, and use it until their death many years later, while males typically die after mating. This long‐term commitment between queens and their mates limits the opportunity for sexual conflict but raises the new constraint of long‐term sperm survival. In this study, we examine experimentally the effect of mating on immunity in wood ant queens. Specifically, we compared the phenoloxidase and antibacterial activities of mated and virgin Formica paralugubris queens. Queens had reduced levels of active phenoloxidase after mating, but elevated antibacterial activity 7 days after mating. These results indicate that the process of mating, dealation and ovary activation triggers dynamic patterns of immune regulation in ant queens that probably reflect functional responses to mating and pathogen exposure that are independent of sexual conflict.     

Shell shape and mating behaviour in pulmonate gastropods (Mollusca)

Previous work suggests that low-spired hermaphroditic snails mate face-to-face and have reciprocal sperm exchange, whereas high-spired snails mate by shell mounting and have unilateral sperm exchange. This dichotomy lead others to speculate on the evolution of enigmatic mating behaviours and whole-body enantiomorphy. In the present study, we review the current literature on mating behaviour in pulmonate snails and show that: (1) several pulmonate species show considerable intraspecific variation in mating behaviour; (2) mating position does not predict reciprocity of penis intromission and sperm exchange; (3) dart-shooting may be correlated with reciprocity of sperm exchange but other factors must explain the gain or loss of darts; (4) it is unlikely that the degree of reciprocity is the most important factor in explaining the relationship of whole-body enantiomorphy and shell shape; and (5) the reciprocal intromission of penises does not necessarily involve the reciprocal transfer of sperm. Hence, our survey shows that current ideas on the evolutionary relationship between shell shape and reciprocity with sexual selection (including dart-use) and whole-body enantiomorphy in hermaphroditic snails should be refined. The results obtained demonstrate that our current knowledge on gastropod mating behaviour is too limited to detect general evolutionary trajectories in gastropod mating behaviour and genital anatomy.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 96 , 306–321.     

Functional morphology of the sperm-containing chambers of the sea slug Okenia polycerelloides in the context of sexual selection

Sea slugs are interesting models to study post-copulatory sexual selection in simultaneous hermaphrodites due to the enormous variation of their reproductive systems. However, the knowledge of the functional morphology of their reproductive system is limited to few species, and it is rarely discussed in the context of sexual selection theory. In this study, we investigated the functional morphology of the sperm-containing chambers (i.e., ampulla, seminal receptacle, and bursa copulatrix) of the reproductive system of Okenia polycerelloides (Ortea & Bouchet, 1983), based on light, confocal, and electron microscopy. Although the morphology of the ampulla is similar to other species, indicating that it is a site for autosperm storage, we found some sperm facing the ampullar epithelium, a feature commonly regarded as characteristic of the seminal receptacle of sea slugs. The seminal receptacle of O. polycerelloides showed secretory activity and contained sperm with distribution and orientation suggestive of stratification of allosperm from distinct mating events, a feature that would affect sperm competition. The bursa copulatrix had epithelial cells with secretory and absorptive characteristics, and contained degraded sperm and yolk granules within its lumen. Comparative analyses of the contents of each organ demonstrated that sperm digestion occurs in the bursa copulatrix and affects sperm heads first, changing their morphology from slender and curved to shorter and ellipsoid before complete lysis. Although digestion and absorption of surplus sperm are currently the main hypothesized functions for the bursa copulatrix, its role in cryptic female choice should not be ruled out. The close structural connection between the seminal receptacle and bursa copulatrix, as well as their muscular walls, would enable control over the fate of the sperm received in each mating event, that is, storage or digestion.     

PATTERNS OF SEXUAL ROLES ADOPTED BY THE SCHISTOSOME-VECTOR SNAIL BIOMPHALARIA GLABRATA (PLANORBIDAE)

Relatively little is known about the mating behaviour of hermaphroditefreshwater snails, many of which transmit the Schistosoma trematodesamong humans in developing countries. Knowledge of the breedingbiology of these snails could help in the design of schistosomecontrol programmes, as well as possibly contributing to ourunderstanding of the evolution of simultaneous hermaphroditismin animaL. Here we describe an experiment investigating thepatterns of sexual roles adopted by the Schistosoma mansoni-vectotsnail, Btmphalaria glabrata. During observations on groups offreely interacting snails, no individuals copulated significantlymore often in the male than in the female role, or vice versa.Only one individual showed a pattern of alternating sexual rolesover successive matings that differed significantly from a randomsequence of roles. There was no evidence for reciprocal copulationwith one particular partner, either between consecutive matings(unless they were temporarily isolated from other snails) orbetween non-consecutive matings (separated by copulations withother conspccifics). We discuss these results in the contextof sex allocation and ESS mating strategy theories. ^*Present address (or correspondence 1 G VERNON. Bioscan (UK).Standingford House, Cave Street, St. Clements, Oxford OX4 IBA. (Received 5 October 1995; accepted 6 November 1995)     

The refractory period of female katydids (Orthoptera: Tettigoniidae): sexual conflict over the remating interval?

Female tettigoniids have a refractory period after mating duringwhich they are sexually unreceptive. Because females obtainvaluable nutrients as well as sperm at mating, there is likelyto be sexual conflict over the duration of the refractory period;females should accrue male donations, whereas males should preventtheir mates from accepting additional matings. We examined theeffects of ampulla attachment duration and female diet on theinduction of the refractory period of a zaprochiline tettigoniid.We find that, although the relationship between ampulla attachmentduration and duration of the refractory period holds for femalesmaintained on a high-pollen diet, poorly nourished females onlyshow the typical onset of the refractory period when allowedto consume the spermatophylax. These results suggest that femaleinterests may have a greater influence in determining the outcomeof sexual conflict. Under some circumstances spermatophylaxfeeding may compensate for poor female nutrition leading tothe typical refractory period. Sexual conflict over the rematinginterval could be instrumental in generating larger nutrientinvestments by male tettigoniids. [Behav Ecol 1991;2:276–282]     

Sex peptide receptor is required for the release of stored sperm by mated Drosophila melanogaster females

The storage of sperm in mated females is important for efficient reproduction. After sperm are transferred to females during mating, they need to reach and enter into the site(s) of storage, be maintained viably within storage, and ultimately be released from storage to fertilize eggs. Perturbation of these events can have drastic consequences on fertility. In Drosophila melanogaster, females store sperm for up to 2 weeks after a single mating. For sperm to be released normally from storage, Drosophila females need to receive the seminal fluid protein (SFP) sex peptide (SP) during mating. SP, which binds to sperm in storage, signals through the sex peptide receptor (SPR) to elicit two other effects on mated females: the persistence of egg laying and a reduction in sexual receptivity. However, it is not known whether SPR is also needed to mediate SP’s effect on sperm release. By phenotypic analysis of flies deleted for SPR, and of flies knocked down for SPR, ubiquitously or in specific tissues, we show that SPR is required to mediate SP’s effects on sperm release from storage. We show that SPR expression in ppk⁺ neurons is needed for proper sperm release; these neurons include those that mediate SP’s effect on receptivity and egg laying. However, we find that SPR is also needed in the spermathecal secretory cells of the female reproductive tract for efficient sperm release. Thus, SPR expression is necessary in both the nervous system and in female reproductive tract cells to mediate the release of stored sperm.     

Inter-genomic sexual conflict drives antagonistic coevolution in harvester ants

The reproductive interests of males and females are not always aligned, leading to sexual conflict over parental investment, rate of reproduction and mate choice. Traits that increase the genetic interests of one sex often occur at the expense of the other, selecting for counter-adaptations leading to antagonistic coevolution. Reproductive conflict is not limited to intraspecific interactions; interspecific hybridization can produce pronounced sexual conflict between males and females of different species, but it is unclear whether such conflict can drive sexually antagonistic coevolution between reproductively isolated genomes. We tested for hybridization-driven sexually antagonistic adaptations in queens and males of the socially hybridogenetic ‘J’ lineages of Pogonomyrmex harvester ants, whose mating system promotes hybridization in queens but selects against it in males. We conducted no-choice mating assays to compare patterns of mating behaviour and sperm transfer between inter- and intra-lineage pairings. There was no evidence for mate discrimination on the basis of pair type, and the total quantity of sperm transferred did not differ between intra- and inter-lineage pairs; however, further dissection of the sperm transfer process into distinct mechanistic components revealed significant, and opposing, cryptic manipulation of copulatory investment by both sexes. Males of both lineages increased their rate of sperm transfer to high-fitness intra-lineage mates, with a stronger response in the rarer lineage for whom mating mistakes are the most likely. By contrast, the total duration of copulation for intra-lineage mating pairs was significantly shorter than for inter-lineage crosses, suggesting that queens respond to prevent excessive sperm loading by prematurely terminating copulation. These findings demonstrate that sexual conflict can lead to antagonistic coevolution in both intra-genomic and inter-genomic contexts. Indeed, the resolution of sexual conflict may be a key determinant of the long-term evolutionary potential of host-dependent reproductive strategies, counteracting the inherent instabilities arising from such systems.     

Reversed sexual conflict in a promiscuous antelope

A general tenet of sexual conflict theory is that males have higher optimum mating rates than do females and therefore should be more persistent when it comes to mating. However, in promiscuous species, females might benefit from high mating rates as a result of increased conception probability with favored males, whereas favored males benefit from mating selectively because of sperm depletion. When this results in higher optimum mating rates for females than for males, there is potential for reversed sexual conflicts between persistent females and resistant males. Here I report evidence of such a reversed sexual conflict in a promiscuous antelope, the African topi. Rather than mating randomly, favored males prefer to balance mating investment equally between females as predicted by strategic sperm allocation theory. Females, however, enhance their probability of mating with favored males through aggression toward mating pairs. Supporting the idea that aggressive females thereby harass males to mate at a rate that is suboptimal from the males' perspective, males become increasingly likely to counterattack aggressive females with whom they have already mated disproportionately, and such male counterattacks are associated with refusal to mate with the aggressive females. This study points to reversed sexual conflict as a more significant evolutionary force in promiscuous mammals than previously thought; however, such conflicts probably often go unnoticed because males, in contrast to females, can avoid mating without conspicuous resistance.     

Effects of two seminal fluid transcripts on post‐mating behaviour in the simultaneously hermaphroditic flatworm Macrostomum lignano

The seminal fluid proteins (SFPs) transferred to mating partners along with sperm often play crucial roles in mediating post‐mating sexual selection. One way in which sperm donors can maximize their own reproductive success is by modifying the partner's (sperm recipient's) post‐copulatory behaviour to prevent or delay re‐mating, thereby decreasing the likelihood or intensity of sperm competition. Here, we adopted a quantitative genetic approach combining gene expression and behavioural data to identify candidates that could mediate such a response in the simultaneously hermaphroditic flatworm Macrostomum lignano. We identified two putative SFPs—Mlig‐pro46 and Mlig‐pro63—linked to both mating frequency and ‘suck’ frequency, a distinctive behaviour, in which, upon ejaculate receipt, the worm places its pharynx over its female genital opening and apparently attempts to remove the received ejaculate. We, therefore, performed a manipulative experiment using RNA interference‐induced knockdown to ask how the loss of Mlig‐pro46 and Mlig‐pro63 expression, singly and in combination, affects mating frequency, partner suck propensity and sperm competitive ability. None of the knockdown treatments impacted strongly on the mating frequency or sperm competitive ability, but knockdown of Mlig‐pro63 resulted in a significantly decreased suck propensity of mating partners. This suggests that Mlig‐pro63 may normally act as a cue in the ejaculate to trigger recipient suck behaviour and—given that other proteins in the ejaculate have the opposite effect—could be one component of an ongoing arms race between donors and recipients over the control of ejaculate fate. However, the adaptive significance of Mlig‐pro46 and Mlig‐pro63 from a donor perspective remains enigmatic.     

Mate sampling and the sexual conflict over mating in seaweed flies

The order in which females encounter, or sample, males in apopulation may have important consequences for mate choice,with the information gathered about males influencing boththe preference function and degree of choosiness of females.Sexual selection may be affected as a result. Sampling of particularsubsets of males may be a crucial component of individual variation in mate preferences within populations. However, the sequencein which males are sampled may also be important in specieswithout traditional, active mate choice, such as when sexualselection involves sexual conflict over mating. This wouldoccur if the likelihood of a female mating with a male of acertain phenotype changes as a result of previous encounters.We examined the effects of encountering males differing inbody size, a sexually selected phenotype, in the seaweed flyCoelopa frigida. Sexual selection occurs in this species asa result of a sexual conflict over mating. We show that theoutcome of the sexual conflict is independent of the orderin which males are encountered by female seaweed flies, withthe overall mating advantage to large males being unaffected.In addition, we explored female preference functions and evaluatethe heterogeneity in female willingness to mate. We suggestthat consideration of mate sampling theory is valuable whenexamining mate choice in species in which sexual selectionis driven by sexual conflict.