Sperm Precedence Research Articles

Lifetime inbreeding depression in a leaf beetle.

Ongoing habitat loss and fragmentation result in rapid population size reductions, which can increase the levels of inbreeding. Consequently, many species are threatened by inbreeding depression, a loss of individual fitness following the mating of close relatives. Here, we investigated inbreeding effects on fitness‐related traits throughout the lifetime of the mustard leaf beetle (Phaedon cochleariae) and mechanisms for the avoidance of inbreeding. Previously, we found that these beetles have family‐specific cuticular hydrocarbon profiles, which are likely not used as recognition cue for precopulatory inbreeding avoidance. Thus, we examined whether adult beetles show postcopulatory inbreeding avoidance instead. For this purpose, we determined the larval hatching rate of eggs laid by females mated sequentially with two nonsiblings, two siblings, a nonsibling, and a sibling or vice versa. The beetles suffered from inbreeding depression throughout their entire ontogeny, as evinced by a prolonged larval development, a decreased larval and adult survival and a decreased reproductive output of inbred compared to outbred individuals. The highest larval hatching rates were detected when females were mated with two nonsiblings or first with a sibling and second with a nonsibling. Significantly lower hatching rates were measured in the treatments with a sibling as second male. Thus, the results do not support the existence of postcopulatory inbreeding avoidance in P. cochleariae, but revealed evidence for second male sperm precedence. Consequently, an alternative strategy to avoid inbreeding costs might exist in this beetle, such as a polyandrous mating system, potentially coupled with a specific dispersal behavior.

Testing the direct and genetic benefit hypotheses of polyandry in the wood tiger moth

Polyandry is widespread among many animal taxa, yet the benefits for females are still debated. The two main hypotheses to explain its evolution are the direct benefits and the genetic benefits hypotheses, which are not mutually exclusive. We tested both in the wood tiger moth Arctia plantaginis (Arctiidae) by comparing fitness components in single and multiple mated females. We measured female longevity and number of eggs laid (i.e. direct benefits), as well as offspring hatching success and survival (i.e. genetic benefits). Fitness components did not differ between single and multiple mated females; therefore, there was no evidence to support either direct or genetic benefits hypotheses, or any costs. After paternity analyses, we obtained qualitatively similar results by comparing clutches effectively sired by one male with clutches sired by two males, regardless of the number of times a female mated. We further investigated the proximate mechanisms driving the outcome of paternity patterns. First male precedence, last male precedence, and mixed paternity were present in equal proportions, although there was a trend towards last male sperm precedence in later clutches. Interestingly, in polyandric females, the age of the second male positively affected the number of eggs laid and the number of surviving offspring, indicating an advantage for older males, possibly due to a higher parental investment. We suggest in light of recent theoretical work that the acceptance of more partners in female A. plantaginis may have evolved to ensure fertilization and avoid the risk of virgin death. Why do females mate with multiple males? Here, we investigate the effects of polyandry on female fitness components in the wood tiger moth Arctia plantaginis. We do not find any support for the direct or genetic benefits hypotheses, or any costs of polyandry. We do not find any clear paternity patterns as last male, first male, and mixed paternities are equally present. We suggest that in this species polyandry may have evolved to ensure fertilization and avoid the risk of virgin death.

Last male sperm precedence is modulated by female remating rate in Drosophila melanogaster.

Following multiple matings, sperm from different males compete for fertilization within the female reproductive tract. In many species, this competition results in an unequal sharing of paternity that favors the most recent mate, termed last male sperm precedence (LMSP). Much of our understanding of LMSP comes from studies in Drosophila melanogaster that focus on twice‐mated females with standardized latencies between successive matings. Despite accumulating evidence indicating that females often mate with more than two males and exhibit variation in the latency between matings, the consequences of mating rate on LMSP are poorly understood. Here, we developed a paradigm utilizing D. melanogaster in which females remated at various time intervals with either two or three transgenic males that produce fluorescent sperm (green, red, or blue). This genetic manipulation enables paternity assessment of offspring and male‐specific sperm fate examination in female reproductive tracts. We found that remating latency had no relationship with LMSP in females that mated with two males. However, LMSP was significantly reduced in thrice‐mated females with short remating intervals; coinciding with reduced last‐male sperm storage. Thus, female remating rate influences the relative share of paternity, the overall clutch paternity diversity, and ultimately the acquisition of indirect genetic benefits to potentially maximize female reproductive success.

Extreme fertilization bias towards freshly inseminated sperm in a species exhibiting prolonged female sperm storage.

The storage of sperm by females across successive reproductive cycles is well documented in internal fertilizers, yet the fate of stored sperm when they compete with ‘new’ sperm to fertilize a female's eggs has rarely been considered. This gap in our understanding is likely due to the logistical difficulties of controlling behavioural interactions during or after mating, which in turn may influence how many sperm are inseminated and how stored sperm are ultimately used during successive bouts of sperm competition with freshly inseminated sperm. Here, we use artificial insemination (AI) in guppies (Poecilia reticulata), a polyandrous live-bearing poeciliid fish exhibiting prolonged sperm storage by females, to overcome these challenges. The use of AI enables us to control potential differential maternal effects (e.g. behaviourally mediated cryptic female choice) and specifically test for post-copulatory paternity biases that favour either stored or fresh sperm when they compete to fertilize eggs. Our paternity analyses revealed the almost complete dominance of freshly inseminated sperm over stored sperm, supporting previous studies reporting similar patterns following natural matings across successive brood cycles. However, our use of AI, which excluded behavioural interactions between males and females, most likely generated a far stronger pattern of fresh sperm precedence compared with those reported in previous studies, possibly implicating ‘cryptic' forms of selection by females that may sometimes bolster the success of stored sperm.

Estimating encounter rates as the first step of sexual selection in the lizard Anolis sagrei.

How individuals move through their environment dictates which other individuals they encounter, determining their social and reproductive interactions and the extent to which they experience sexual selection. Specifically, females rarely have the option of mating with all males in a population-they can only choose among the males they encounter. Further, quantifying phenotypic differences between the males that females encounter and those that sire females' offspring lends insight into how social and reproductive interactions shape male phenotypes. We used an explicitly spatio-temporal Markov chain model to estimate the number of potential mates of Anolis sagrei lizards from their movement behaviour, and used genetic paternity assignments to quantify sexual selection on males. Females frequently encountered and mated with multiple males, offering ample opportunity for female mate choice. Sexual selection favoured males that were bigger and moved over larger areas, though the effect of body size cannot be disentangled from last-male precedence. Our approach corroborates some patterns of sexual selection previously hypothesized in anoles based on describing them as territorial, whereas other results, including female multiple mating itself, are at odds with territorial polygyny, offering insight into discrepancies in other taxa between behavioural and genetic descriptions of mating systems.

Modeling strategic sperm allocation: Tailoring the predictions to the species.

Two major challenges exist when empirically testing the predictions of sperm allocation theory. First, the study species must adhere to the assumptions of the model being tested. Unfortunately, the common assumption of sperm allocation models that females mate a maximum of once or twice does not hold for many, if not most, multiply and sequentially mating animals. Second, a model's parameters, which dictate its predictions, must be measured in the study species. Common examples of such parameters, female mating frequency and sperm precedence patterns, are unknown for many species used in empirical tests. Here, we present a broadly applicable model, appropriate for multiply, sequentially mating animals, and test it in three species for which data on all the relevant parameter values are available. The model predicts that relative allocation to virgin females, compared to nonvirgins, depends on the interaction between female mating rate and the sperm precedence pattern: relative allocation to virgins increases with female mating rate under first-male precedence, while the opposite is true under later-male precedence. Our model is moderately successful in predicting actual allocation patterns in the three species, including a cricket in which we measured the parameter values and performed an empirical test of allocation.

How pre- and postcopulatory sexual selection influence male mating decisions in a promiscuous species

When females mate multiply, male reproductive success depends on both pre- and postcopulatory processes, including female choice and sperm competition. However, these processes can favour different mating tactics in males. Here we used the Trinidadian guppy, Poecilia reticulata, system to understand how this conflict is resolved. We asked whether knowledge of recent female mating history leads males to adjust their mating effort with respect to the time devoted to mating activity, and the frequency and the sequence of mating tactics employed. To do this we quantified male mating behaviour in three competitive scenarios: (1) Single, when a focal male arrives near a single female and remains alone with her; (2) First, when a focal male is joined by a rival male; and (3) Second, when a focal male arrives after a rival male. We hypothesized that males adjust their behaviour based on arrival order. If female sequential mate choice is the main process shaping male mating behaviours (favouring First males in guppies), males should avoid competition and invest most when Single. Alternatively, if last-male sperm precedence is the major driver of decision making, males should invest more in mating attempts in the Second scenario. Greatest investment when First implies an intermediate strategy. We found that order of arrival influenced mating decisions with most mating activity during the First rather than the Single and Second scenarios. This result suggests that both pre- and postcopulatory processes influence mating investment, and that individual males make contingent decisions to maximize both mating and fertilization success.

Correction to: Female remating decisions and a shorter inter-mating interval diminish last-male sperm precedence

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Do Sperm Really Compete and Do Eggs Ever Have a Choice? Adult Distribution and Gamete Mixing Influence Sexual Selection, Sexual Conflict, and the Evolution of Gamete Recognition Proteins in the Sea.

The evolution of gametic compatibility and the effectiveness of compatibility, within and across species, depend on whether sperm from different males directly compete for an egg and whether eggs ever have a choice. Direct sperm competition and egg choice depend on whether sperm from different males arrive at an egg in the brief interval between first sperm contact and fertilization. Although this process may be relevant for all sexually reproducing organisms, it is most easily examined in aquatic external fertilizers. When sperm are released into the sea, packets of seawater at the spatial scale relevant to single eggs might contain sperm from only one male, eliminating the potential for direct sperm competition and egg choice. Field experiments and a simple heuristic model examining the degree of sperm mixing for the sea urchin Strongylocentrotus franciscanus indicate that degree of competitive fertilization depends on density and distribution of competing males and that the nature of this competition influences whether males with high- or low-affinity gamete recognition protein genotypes have higher reproductive success. These results provide a potential explanation for the generation and maintenance of variation in gamete recognition proteins and why effectiveness of conspecific sperm precedence can be density dependent.

Evolution of external female genital mutilation: why do males harm their mates?

Sperm competition may select for male reproductive traits that influence female mating or oviposition rate. These traits may induce fitness costs to the female; however, they may be costly for the males as well as any decrease in female fitness also affects male fitness. Male adaptations to sperm competition manipulate females by altering not only female behaviour or physiology, but also female morphology. In orb-weaving spiders, mating may entail mutilation of external structures of the female genitalia, which prevents genital coupling with subsequent males. Here, we present a game theoretical model showing that external female genital mutilation is favoured even under relatively high costs of mutilation, and that it is favoured by a high number of mate encounters per female and last-male sperm precedence.

Last-male sperm precedence in Rhynchophorus ferrugineus (Olivier): observations in laboratory mating experiments with irradiated males.

The Red Palm Weevil (RPW) Rhynchophorus ferrugineus (Olivier 1790) is an invasive pest from southeastern Asia and Melanesia that in the last 30 years has spread widely in the Middle East and Mediterranean Basin. Its stem-boring larvae cause great damage to several palm species of the Arecaceae family, many of which are economically important for agricultural and ornamental purposes. Therefore, great attention has recently been focused in studying this species to identify sustainable and effective eradication strategies, such as sterile insect technique (SIT). The rapid spread of RPW is associated with its high reproductive success. To evaluate the suitability of a SIT strategy, particular physiological and behavioral aspects of RPW reproduction, such as the presence of polyandry and post-copulatory sperm selection mechanisms, were investigated. To determine paternity of progeny from multiply mated females, double-crossing experiments were carried out confining individual females with either a wild-type male or a γ-irradiated male (Co-60). Fecundity and fertility of females were scored to evaluate post-copulatory sperm selection. Results showed that progeny were almost exclusively produced by the sperm of the second male, suggesting that a last-male sperm precedence is expressed at high levels in this species, and providing interesting insights for an area-wide RPW management strategy such as the SIT.

Artificial insemination unveils a first-male fertilization advantage in the guppy

Several factors are involved in determining the outcome of sperm competition. In addition to sperm number, sperm quality and male phenotype, insemination order is often associated with skewed paternity share. Patterns of sperm precedence can be produced by the mechanics of sperm storage and fertilization, or by active processes under male or female control. However, as males and females always interact during copulation, it is difficult to identify the mechanism responsible. The Trinidadian guppy, Poecilia reticulata, is a polyandric species characterized by last-male sperm precedence in natural matings. During such matings, females allow attractive males to inseminate more sperm by controlling copulation duration. We used artificial insemination to clarify the extent to which female control of sperm transfer influences the observed pattern of sperm precedence in this species. This technique allowed us to experimentally manipulate the number of sperm transferred and the timing of insemination. We found a significant first-male fertilization advantage. This advantage, however, declined as the time between insemination and parturition increased. Presumably, the anatomy and the physiology of the female genital tract favour egg fertilization by the first ejaculate inseminated, whereas sperm mixing is likely to be responsible for the reduction in first-male advantage associated with longer insemination–parturition intervals. Our results suggest that the last-male precedence detected after two consecutive natural matings is caused by cryptic female preference for attractive males associated with a female trading-up strategy (i.e. the second male is more frequently more attractive than the first male), rather than by insemination order per se. As the pattern of sperm precedence has important consequences for male reproductive strategies (for example mate guarding and male mate choice copying), unravelling its dynamic represents an important contribution to understanding the sexual behaviour of this model species.

Female remating decisions and a shorter inter-mating interval diminish last-male sperm precedence

Highly variable within and across species, patterns of sperm use not only are often driven by post-copulatory sexual selection but can also be impacted by experimental design. In investigations of paternity bias using competitive double matings, the inter-mating interval is a temporal factor that can affect sperm use patterns if the first male’s sperm is used or lost at an appreciable rate between matings or if its viability or relative competitiveness is influenced by the time since ejaculation. Rapid loss of first-male sperm within the female after mating has been established in the seed beetle (Callosobruchus maculatus), a model system in sperm competition studies. However, our understanding of sperm precedence in this species, which disproportionately favors the last (second) male to mate, is based on long inter-mating intervals. Here, I determine the effect of a shortened inter-mating interval on second-male paternity (P 2) and, importantly, the extent to which females in this species are willing to remate immediately. I find that P 2 is significantly reduced when females remate immediately than when they remate 24 or 48 h after the first mating and that immediate remating is common, indicating that there is a substantial potential for female remating decisions to influence the intensity of sperm competition within species. To understand the variation in inter-mating intervals from a female perspective, I further identify key differences between females that did and did not remate at three inter-mating intervals (0, 24, and 48 h after the initial mating) and discuss potential mechanisms for the observed variation in female refractoriness. Females often mate with multiple males, and in insects it is common for the last male to mate to obtain the greatest share of paternity. One possible explanation for this bias towards last-male sperm use is the passive loss of sperm from the first male with time since mating. Here, I examine the effect of different intervals between matings in a seed beetle species to determine if first-male paternity is increased when females remate sooner. My results show that first males do achieve higher paternity when females remate sooner, which is more common than expected in this species. I also show the differences between females that do and do not remate, which can help explain potential causes for variation in female remating behavior. Together, these results show the significant impact that female remating decisions can have on male competition and, ultimately, their reproductive success.

An insect anti-antiaphrodisiac.

Passive mechanisms of mate guarding are used by males to promote sperm precedence with little cost, but these tactics can be disadvantageous for their mates and other males. Mated females of the plant bug Lygus hesperus are rendered temporarily unattractive by seminal fluids containing myristyl acetate and geranylgeranyl acetate. These antiaphrodisiac pheromones are gradually released from the female's gonopore, declining until they no longer suppress male courtship. Because starting quantities of these compounds can vary widely, the repellant signal becomes less reliable over time. Evidence was found of a complimentary mechanism that more accurately conveys female mating status. Once inside the female, geranylgeranyl acetate is progressively converted to geranylgeraniol then externalized. Geranylgeraniol counteracts the antiaphrodisiac effect despite having no inherent attractant properties of its own. This is the first evidence for such an anti-antiaphrodisiac pheromone, adding a new element to the communication mechanisms regulating reproductive behaviors.

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.

Male spiders reduce pre- and postmating sexual investment in response to sperm competition risk

The interplay between pre- and postmating responses to intrasexual competition remains enigmatic. Sperm competition models often assume a trade-off between pre- and postmating traits that enhance mate acquisition and fertilization success, respectively. However, when males court females through food donations (i.e., nuptial gifts), pre- and postmating responses may be aligned, as nuptial gifts have the dual function of facilitating both mate acquisition and sperm transfer. In the spider Pisaura mirabilis, nuptial gifts consist of silk-wrapped prey. We tested whether males respond to a competitor by altering: 1) premating investment in the gift, 2) postmating sperm investment, and 3) whether pre- and postmating responses are coupled and respond to competition in the same direction or not. Under competition risk males silk-wrapped their gifts for significantly shorter time and transferred less sperm to females, pointing to a reduction of both pre- and postmating responses. Because silk is not a target of female choice, reducing gift construction may speed up mate acquisition. In accordance with models of sperm allocation, perceived high levels of competition and/or sperm priority patterns may explain the reduced patterns of sperm transfer found in our study. Overall, our findings suggest that in competitive environments pre- and postmating traits are coupled and respond in the same direction.

Paternity following multiple mating in ladybird Harmonia axyridis

We investigated factors which may determine realised paternity following multiple mating in a promiscuous ladybird Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae), namely the mating order and ability of males of different morphs to fertilize females. In experiments using males of two colour morphs, we found that the first male had a reproductive advantage over the second male (sperm precedence P2 = 0.38). Males of the melanic morph spectabilis had a reproductive advantage over males of the semi-melanic morph axyridis (P2 = 0.35). Analysis of paternity using microsatellites applied to young larvae, which avoids possible differential mortality of progeny, confirmed the precedence of the first male and higher paternity rate of colour morphs that are more frequent in the wild. Hatchability of eggs increased from 65% after the first mating to 85% after the second mating delayed by five days. Average duration of mating was 157 min and did not differ between the first and second mating and between the colour morphs of males.

Male spider mites use chemical cues, but not the female mating interval, to choose between mates.

The choice of the partner an individual will mate with is expected to strongly impact its fitness. Hence, natural selection has favoured the evolution of cues to distinguish among mates that will provide different fitness benefits to the individual that is choosing. In species with first-malesperm precedence, this is particularly important for males, as mating with mated females will result in no offspring. In the spider mite Tetranychus urticae only the first mating is effective, except if the interval between first and second copulations is shorter than 24h. In line with this, males prefer to mate with virgin over mated females. They do not, however, choose between females that have mated at different time intervals. Here, we tested which type of cues males use to distinguish between females with different mating status (virgin versus mated). To do so, we firstly confirmed that males prefer virgins over mated females and that they do not select females on the basis of their age or mating interval. Next, we tested whether contact and volatile compounds or chemical trails affected male discrimination between mated and virgin females, by systematically varying the exposure of males to these cues. We found that volatile compounds and chemical trails were sufficient to induce discrimination between virgin and mated females in males. Direct contact with females, however, does not seem to play a role in this discrimination. The composition of such chemical cues (trails and volatiles) remains to be identified.

Impact of cryptic female choice on insemination success: Larger sized and longer copulating male squid ejaculate more, but females influence insemination success by removing spermatangia.

In polyandrous mating systems, sperm competition and cryptic female choice (CFC) are well recognized as postcopulatory evolutionary forces. However, it remains challenging to separate CFC from sperm competition and to estimate how much CFC influences insemination success because those processes usually occur inside the female's body. The Japanese pygmy squid, Idiosepius paradoxus, is an ideal species in which to separate CFC from sperm competition because sperm transfer by the male and sperm displacement by the female can be observed directly at an external location on the female's body. Here, we counted the number of spermatangia transferred to, removed from, and remaining on the female body during single copulation episodes. We measured behavioral and morphological characteristics of the male, such as duration of copulation and body size. Although males with larger body size and longer copulation time were capable of transferring larger amounts of sperm, females preferentially eliminated sperm from males with larger body size and shorter copulation time by spermatangia removal; thus, CFC could attenuate sperm precedence by larger males, whereas it reinforces sperm precedence by males with longer copulation time. Genetic paternity analysis revealed that fertilisation success for each male was correlated with remaining sperm volume that is adjusted by females after copulation.

Males Can Benefit from Sexual Cannibalism Facilitated by Self-Sacrifice

In a number of species, males are cannibalized by females after mating (reviewed in [1, 2]), and some males actually appear to facilitate their own cannibalism (reviewed in [3]). Such self-sacrifice can evolve if being eaten sufficiently enhances either fertilization success (mating effort) or offspring number or fitness (paternal effort). While there is some support for the mating-effort hypothesis, few studies have found support for paternal effort. We used two experiments to test the paternal-effort hypothesis in the dark fishing spider, Dolomedes tenebrosus. Males of this species provide themselves as a material contribution: they spontaneously die during copulation and are subsequently eaten by females. In support of the paternal-effort predictions, when females were allowed to consume their mating partner, we found large and significant increases in (1) the number, (2) the size, and (3) the survivorship of the offspring. Similar benefits were not seen when females were allowed to consume a cricket in lieu of a male, suggesting that it is the consumption of themale's body per se that is responsible for thesefitness benefits. Together, our results suggest thatD.tenebrosus males can benefit from self-sacrifice behavior through paternal effort. Such behavior may be particularly likely to evolve when high rates of postcopulatory cannibalism trap males into investing in their first mate instead of investing in acquiring additional matings and/or if strong first-male sperm precedence reduces the benefits of both investing in additional matings and paternity protection.