Benefits of polyandry: Molecular evidence from field‐caught dung beetles

When females mate with multiple males, they set the stage for postcopulatory sexual selection via sperm competition and/or cryptic female choice. Surprisingly little is known about the rates of multiple mating by females in the wild, despite the importance of this information in understanding the potential for postcopulatory sexual selection to drive the evolution of reproductive behaviour, morphology and physiology. Dung beetles in the genus Onthophagus have become a laboratory model for studying pre‐ and postcopulatory sexual selection, yet we still lack information about the reproductive behaviour of female dung beetles in natural populations. Here, we develop microsatellite markers for Onthophagus taurus and use them to genotype the offspring of wild‐caught females and to estimate natural rates of multiple mating and patterns of sperm utilization. We found that O. taurus females are highly polyandrous: 88% of females produced clutches sired by at least two males, and 5% produced clutches with as many as five sires. Several females (23%) produced clutches with significant paternity skew, indicating the potential for strong postcopulatory sexual selection in natural populations. There were also strong positive correlations between the number of offspring produced and both number of fathers and paternity skew, which suggests that females benefit from mating polyandrously by inciting postcopulatory mechanisms that bias paternity towards males that can sire more viable offspring. This study evaluates the fitness consequences of polyandry for an insect in the wild and provides strong evidence that female dung beetles benefit from multiple mating under natural conditions.     

Measuring polyandry in wild populations: a case study using promiscuous crickets

Mating rate has important implications for patterns of sexual selection and sexual conflict and hence for issues such as speciation and the maintenance of genetic diversity. Knowledge of natural mating rates can provide insights into the factors driving female mating behaviour. We investigated the level of polyandry in a Spanish population of the field cricket Gryllus bimaculatus using microsatellite markers. Two approaches were employed: (i) genotyping the offspring of wild-caught gravid females to determine the number of males siring the brood and (ii) genotyping sperm stored in the spermathecae of females mated in the wild to estimate the number of mating partners. We compared existing methods for inferring the minimum and probable number of fathers and described a novel probabilistic technique estimating the number of mates by genotyping stored sperm. Using the most conservative allele-counting method, 71% of females produced offspring sired by at least two males (a minimum mean of 2.4 fathers per clutch), and all females had mated to at least two males with minimum mean estimates of 2.7-5.1 mates per female. Our study reveals high levels of polyandry in the wild and suggests that females mate with more males than sire their offspring.     

Polyandry and postcopulatory sexual selection in a wild population

When females mate multiply, postcopulatory sexual selection can occur via sperm competition and cryptic female choice. Although postcopulatory selection has the potential to be a major force in driving evolution, few studies have estimated its strength in natural populations. Likewise, although polyandry is widespread across taxa and is the focus of a growing body of research, estimates of natural female mating rates are still limited in number. Microsatellites can be used to estimate the number of mates represented in females' sperm stores and the number of sires contributing to their offspring, enabling comparisons both of polyandry and of two components of postcopulatory selection: the proportion of males that mate but fail to sire offspring, and the degree of paternity skew among the males that do sire offspring. Here, we estimate the number of mates and sires among wild females in the Hawaiian swordtail cricket Laupala cerasina. We compare these estimates to the actual mating rates and paternity shares we observed in a semi‐natural population. Our results show that postcopulatory sexual selection operates strongly in this species: wild females mated with an average minimum of 3.6 males but used the sperm from only 58% of them. Furthermore, among the males that did sire offspring, paternity was significantly skewed. These patterns were similar to those observed in the field enclosure, where females mated with an average of 5.7 males and used the sperm from 62% of their mates, with paternity significantly skewed among the sires.     

Fitness benefits of polyandry for experienced females

Females often mate with several different males, which may promote sperm competition and increase offspring viability. However, the potential benefits of polyandry remain controversial, particularly in birds where recent reviews have suggested that females gain few genetic benefits from extra‐pair mating. In tree swallows (Tachycineta bicolor), we found that females with prior breeding experience had more sires per brood when paired to genetically similar social mates, and, among experienced females, broods with more sires had higher hatching success. Individual females breeding in two consecutive years also produced broods with more sires when they were more genetically similar to their mate. Thus, experienced females were able to avoid the costs of mating with a genetically similar social mate and realize fitness benefits from mating with a relatively large number of males. This is one of the first studies to show that female breeding experience influences polyandry and female fitness in a natural population of vertebrates. Our results suggest that the benefits of polyandry may only be clear when considering both the number of mates females acquire and their ability to modify the outcome of sexual conflict.     

Polyandry and paternity skew in natural and experimental populations of Drosophila serrata

Many species engage in polyandry, resulting in the potential for sexual selection to continue post-copulation through sperm competition and/or cryptic female choice. The relative importance of pre- vs. post-copulatory processes remains unknown for most species despite this information being fundamental for understanding the evolutionary consequences of sexual selection. The Australian fruit fly Drosophila serrata has become a prominent model system for studying precopulatory sexual selection, such as mating preferences and their influence on the evolution of sexually selected traits. Here, we investigated polyandry and the potential for post-copulatory sexual selection in this species using indirect paternity analysis. We genotyped 21 wild-caught and 19 laboratory-reared mothers and their offspring (a total of 787 flies) at six microsatellite loci and found extensive polyandry, with all broods surveyed having at least two sires. Female remating rates were higher than in other Drosophila surveyed to date and no significant differences were found between laboratory and field populations. Additionally, we found evidence for biased sperm usage in several broods of D. serrata . Paternity skew occurred more frequently in broods from the field population than the laboratory one, suggesting differences between the two environments in the level of post-copulatory sexual selection. Our data suggest that D. serrata represents a promising system for studying the interaction between pre- and post-copulatory sexual selection in driving the evolution of sexually selected phenotypes.     

Female multiple mating in wild and laboratory populations of the two-spot ladybird, Adalia bipunctata

Female mating rate is an important variable for understanding the role of females in the evolution of mating systems. Polyandry influences patterns of sexual selection and has implications for sexual conflict over mating, as well as for wider issues such as patterns of gene flow and levels of genetic diversity. Despite this, remarkably few studies of insects have provided detailed estimates of polyandry in the wild. Here we combine behavioural and molecular genetic data to assess female mating frequency in wild populations of the two-spot ladybird, Adalia bipunctata (Coleoptera: Coccinellidae). We also explore patterns of sperm use in a controlled laboratory environment to examine how sperm from multiple males is used over time by females, to link mating with fertilization. We confirm that females are highly polyandrous in the wild, both in terms of population mating rates (approximately 20% of the population found in copula at any given time) and the number of males siring offspring in a single clutch (three to four males, on average). These patterns are consistent across two study populations. Patterns of sperm use in the laboratory show that the number of mates does not exceed the number of fathers, suggesting that females have little postcopulatory influence on paternity. Instead, longer copulations result in higher paternity for males, probably due to the transfer of larger numbers of sperm in multiple spermatophores. Our results emphasize the importance of combining field and laboratory data to explore mating rates in the wild.     

High opportunity for postcopulatory sexual selection under field conditions

In polygamous systems, male fitness is determined not only by mating success but also by fertilization success. Despite the growing interest over the past several decades in postcopulatory sexual selection, its relative importance compared to precopulatory sexual selection remains a subject of debate. Here, we use extensive behavioral observations of a seminatural population of Hawaiian swordtail crickets, Laupala cerasina, and molecular paternity assignment to measure the opportunities for pre‐ and postcopulatory selection. Because postcopulatory selection has the potential to operate at multiple stages, we also separately attribute its effects to factors specific to mating events versus factors specific to males. We find that variance in postcopulatory success is over four times as great as variance in precopulatory success, with most of it unexplained by male mating order or the number of nuptial gifts given. Surprisingly, we also find that male singing effort is under postcopulatory selection, suggesting that males who sing more frequently also have more competitive ejaculates. Our results are consistent with the hypothesis that high polyandry levels promote greater relative postcopulatory selection. They also highlight the need for detailed behavioral observations under conditions as natural as possible when measuring mating and reproductive success.     

Multiple paternity in a natural population of a wild tobacco fly, Bactrocera cacuminata (Diptera: Tephritidae), assessed by microsatellite DNA markers

Mating frequency has important implications for patterns of sexual selection and sexual conflict and hence for issues such as speciation and the maintenance of genetic diversity. Knowledge of natural mating patterns can also lead to more effective control of pest tephritid species, in which suppression programmes, such as the sterile insect technique (SIT) are employed. Multiple mating by females may compromise the success of SIT. We investigated the level of polyandry and sperm utilization in a Brisbane field population of the tropical fruit fly, Bactrocera cacuminata (Hering), using seven polymorphic microsatellite loci. The offspring of 22 wild-caught gravid females were genotyped to determine the number of males siring each brood and paternity skew, using the programs gerud and scare. Our data showed that 22.7% of females produced offspring sired by at least two males. The mean number of mates per female was 1.72. Paternal contributions of double-sired broods were skewed with the most successful male having sired between 76.9% and 87.5% of the offspring. These results have implications for SIT, because the level of remating we have identified would indicate that wild females could mate with one or more resident fertile males.     

Pre‐ and postcopulatory sexual selection favor aggressive,young males in polyandrous groups of red junglefowl

A challenge in evolutionary biology is to understand the operation of sexual selection on males in polyandrous groups, where sexual selection occurs before and after mating. Here, we combine fine‐grained behavioral information (>41,000 interactions) with molecular parentage data to study sexual selection in replicated, age‐structured groups of polyandrous red junglefowl, Gallus gallus. Male reproductive success was determined by the number of females mated (precopulatory sexual selection) and his paternity share, which was driven by the polyandry of his female partners (postcopulatory sexual selection). Pre‐ and postcopulatory components of male reproductive success covaried positively; males with high mating success also had high paternity share. Two male phenotypes affected male pre‐ and postcopulatory performance: average aggressiveness toward rival males and age. Aggressive males mated with more females and more often with individual females, resulting in higher sexual exclusivity. Similarly, younger males mated with more females and more often with individual females, suffering less intense sperm competition than older males. Older males had a lower paternity share even allowing for their limited sexual exclusivity, indicating they may produce less competitive ejaculates. These results show that—in these populations—postcopulatory sexual selection reinforces precopulatory sexual selection, consistently promoting younger and more aggressive males.     

Genetic parentage assessment in the crayfish Orconectes placidus,a high-fecundity invertebrate with extended maternal brood care

Microsatellite data have recently been introduced in the context of genetic maternity and paternity assignments in high-fecundity fish species with single-parent-tended broods. Here we extend such analyses to an aquatic invertebrate, the crayfish Orconectes placidus, in which gravid females carry large numbers of offspring. Genetic parentage analyses of more than 900 progeny from 15 wild crayfish broods revealed that gravid females were invariably the exclusive dams of the offspring they tended (i.e. there was no allomaternal care), and that most of the females had mated with multiple (usually two) males who contributed sometimes highly skewed numbers of offspring to a brood. Within any multiply sired brood, the unhatched eggs (or the hatched juveniles) from different fathers were randomly distributed across the mother's brood space. All of these genetic findings are discussed in the light of observations on the mating behaviours and reproductive biology of crayfish.     

Multiple Mating,Paternity and Complex Fertilisation Patterns in the Chokka Squid Loligo reynaudii

Polyandry is widespread and influences patterns of sexual selection, with implications for sexual conflict over mating. Assessing sperm precedence patterns is a first step towards understanding sperm competition within a female and elucidating the roles of male- and female-controlled factors. In this study behavioural field data and genetic data were combined to investigate polyandry in the chokka squid Loligo reynaudii. Microsatellite DNA-based paternity analysis revealed multiple paternity to be the norm, with 79% of broods sired by at least two males. Genetic data also determined that the male who was guarding the female at the moment of sampling was a sire in 81% of the families tested, highlighting mate guarding as a successful male tactic with postcopulatory benefits linked to sperm deposition site giving privileged access to extruded egg strings. As females lay multiple eggs in capsules (egg strings) wherein their position is not altered during maturation it is possible to describe the spatial / temporal sequence of fertilisation / sperm precedence There were four different patterns of fertilisation found among the tested egg strings: 1) unique sire; 2) dominant sire, with one or more rare sires; 3) randomly mixed paternity (two or more sires); and 4) a distinct switch in paternity occurring along the egg string. The latter pattern cannot be explained by a random use of stored sperm, and suggests postcopulatory female sperm choice. Collectively the data indicate multiple levels of male- and female-controlled influences on sperm precedence, and highlights squid as interesting models to study the interplay between sexual and natural selection.     

Polyandry in dragon lizards: inbred paternal genotypes sire fewer offspring

Multiple mating in female animals is something of a paradox because it can either be risky (e.g., higher probability of disease transmission, social costs) or provide substantial fitness benefits (e.g., genetic bet hedging whereby the likelihood of reproductive failure is lowered). The genetic relatedness of parental units, particularly in lizards, has rarely been studied in the wild. Here, we examined levels of multiple paternity in Australia's largest agamid lizard, the eastern water dragon (Intellagama lesueurii), and determined whether male reproductive success is best explained by its heterozygosity coefficient or the extent to which it is related to the mother. Female polyandry was the norm: 2/22 clutches (9.2%) were sired by three or more fathers, 17/22 (77.2%) were sired by two fathers, and only 3/22 (13.6%) clutches were sired by one father. Moreover, we reconstructed the paternal genotypes for 18 known mother–offspring clutches and found no evidence that females were favoring less related males or that less related males had higher fitness. However, males with greater heterozygosity sired more offspring. While the postcopulatory mechanisms underlying this pattern are not understood, female water dragons likely represent another example of reproduction through cryptic means (sperm selection/sperm competition) in a lizard, and through which they may ameliorate the effects of male‐driven precopulatory sexual selection.     

EXPERIMENTAL EVOLUTION OF SPERM QUALITY VIA POSTCOPULATORY SEXUAL SELECTION IN HOUSE MICE

Individuals of many species copulate with multiple mates (polygamy). Multiple mating by females (polyandry) promotes sperm competition, which has broad implications for the evolution of the ejaculate. Multigenerational studies of polygamous insects have shown that the removal of sexual selection has profound fitness consequences for females, and can lead to an evolutionary divergence in ejaculate traits. However, the evolutionary implications of polygamous mating across successive generations have not before been demonstrated in a vertebrate. By manipulating the mating system we were able to reinstate postcopulatory sexual selection in a house mouse population that had a long history of enforced monogamy. Following eight generations of selection, we performed sperm quality assays on males from both the polygamous and monogamous selection lines. We applied a principal component analysis to summarize the variation among 12 correlated sperm traits, and found that males evolving under sperm competition had significantly larger scores on the first axis of variation, reflecting greater numbers of epididymal sperm and increased sperm motility, compared to males from lines under relaxed selection. Moreover, we found a correlated response in the size of litters born to females in lines subject to sperm competition. Thus, we present significant evidence that sperm competition has profound fitness consequences for both male and female house mice.     

Evolutionary causes and consequences of sequential polyandry in anuran amphibians

Among anuran amphibians (frogs and toads), there are two types of polyandry: simultaneous polyandry, where sperm from multiple males compete to fertilize eggs, and sequential polyandry, where eggs from a single female are fertilized by multiple males in a series of temporally separate mating events, and sperm competition is absent. Here we review the occurrence of sequential polyandry in anuran amphibians, outline theoretical explanations for the evolution of this mating system and discuss potential evolutionary implications. Sequential polyandry has been reported in a limited number of anurans, but its widespread taxonomic and geographic distribution suggests it may be common. There have been no empirical studies that have explicitly investigated the evolutionary consequences of sequential polyandry in anurans, but species with this mating pattern share an array of behavioural, morphological and physiological characteristics, suggesting that there has been common sexual selection on their reproductive system. Sequential polyandry may have a number of adaptive benefits, including spreading the risk of brood failure in unpredictable environments, insuring against male infertility, or providing genetic benefits, either through good genes, intrinsic compatibility or genetic diversity effects. Anurans with sequential polyandry provide untapped opportunities for innovative research approaches that will contribute significantly to understanding anuran evolution and also, more broadly, to the development of sexual‐selection and life‐history theory.     

Polyandry facilitates postcopulatory inbreeding avoidance in house mice

The avoidance of genetic incompatibilities between parental genotypes has been proposed to account for the evolution of polyandry. An extension of this hypothesis suggests polyandry may provide an opportunity for females to avoid the cost of inbreeding by exploiting postcopulatory mechanisms that bias paternity toward unrelated male genotypes. Here we test the inbreeding avoidance hypothesis in house mice by experimentally manipulating genetic compatibility via matings between siblings and nonsiblings. We observed little difference in reproductive success between females mated to two siblings or females mated to two nonsiblings. Females mated to both a sibling and a nonsibling tended to have a lower litter survival, but only when the first male to mate was a sibling. Microsatellite data revealed that paternity was biased toward nonsiblings when a female mated with both a sibling and a nonsibling. Unlike previous studies of invertebrates, paternity bias toward the sibling male was independent of mating sequence. We provide one of the first empirical demonstrations that polyandry facilitates postcopulatory sexual selection in a vertebrate. We discuss this result in relation to the possibility of selective fertilization of ova based on major histocompatibility complex (MHC) haploid expression of sperm.     

Molecular Evidence for High Frequency of Multiple Paternity in a Freshwater Shrimp Species Caridina ensifera

Background

Molecular genetic analyses of parentage provide insights into mating systems. Although there are 22,000 members in Malacostraca, not much has been known about mating systems in Malacostraca. The freshwater shrimp Caridina ensifera blue, is a new species belonging to Malacostraca which was discovered recently in Sulawesi, Indonesia. Due to its small body size and low fecundity, this species is an ideal species to study the occurrence and frequency of multiple paternity and to understand of how the low fecundity species persist and evolve.

Methodology/Principal Findings

In this study, we developed four polymorphic microsatellites from C. ensifera and applied them to investigate the occurrence and frequency of multiple paternity in 20 C. ensifera broods caught from Lake Matano, Sulawesi. By genotyping the mother and all offspring from each brood we discovered multiple paternity in all 20 broods. In most of the 20 broods, fathers contributed skewed numbers of offspring and there was an apparent inverse correlation between reproductive success of sires and their relatedness to mothers.

Conclusions/Significance

Our results in combination with recent reports on multiple paternity in crayfish, crab and lobster species suggests that multiple paternity is common in Malacostraca. Skewed contribution of fathers to the numbers of offspring and inverse correlation between reproductive success of sires and their relatedness to mothers suggest that sperm competition occurred and/or pre- and postcopulatory female choice happen, which may be important for avoiding the occurrence of inbreeding and optimize genetic variation in offspring and for persistence and evolution of low fecundity species.     

An integrative mating system assessment of a nonmodel,economically important Pacific rockfish (Sebastes melanops) reveals nonterritorial polygamy and conservation implications for a large species flock

Characterizing the mating systems of long‐lived, economically important Pacific rockfishes comprising the viviparous Sebastes species flock is crucial for their conservation. However, direct assignment of mating success to sires is precluded by open, offshore populations and high female fecundity. We addressed this challenge by integrating paternity‐assigned mating success of females with the adult sex ratio (ASR) of the population, male evolutionary responses to receptive females, and reproductive life history traits—in the framework of sexual selection theory—to assess the mating system of Sebastes melanops. Microsatellite parentage analysis of 17 pregnant females, 1,256 of their progeny, and 106 adults from the population yielded one to four sires per brood, a mean of two sires, and a female mate frequency distribution with a truncated normal (random) pattern. The 11 multiple paternity broods all contained higher median allele richness than the six single paternity broods (Wilcoxon test: W = 0, p < .001), despite similar levels of average heterozygosity. By sampling sperm and alleles from different males, polyandrous females gain opportunities to enhance their sperm supply and to lower the cost of mating with genetically incompatible males through reproductive compensation. A mean of two mates per mated female with a variance of one, an ASR = 1.2 females per male, and the expected population mean of 2.4 mates for mated males (and the estimated 35 unavailable sires), fits polygamous male mate frequency distributions that distinguish polygynandry and polyandrogyny mating systems, that is, variations of polygamy, but not polyandry. Inference for polygamy is consistent with weak premating sexual selection on males, expected in mid‐water, schooling S. melanops, owing to polyandrous mating, moderately aggregated receptive females, an even ASR, and no territories and nests used for reproduction. Each of these characteristics facilitates more mating males and erodes conspicuous sexual dimorphism. Evaluation of male evolutionary responses of demersal congeners that express reproductively territorial behavior revealed they have more potential mechanisms for producing premating sexual selection, greater variation in reproductive success, and a reduced breeding effective population size of adults and annual effective size of a cohort, compared to S. melanops modeled with two mates per adult. Such divergence in behavior and mating system by territorial species may differentially lower their per capita birth rates, subsequent population growth, and slow their recovery from exploitation.     

Polyandry as a mediator of sexual selection before and after mating

The Darwin–Bateman paradigm recognizes competition among males for access to multiple mates as the main driver of sexual selection. Increasingly, however, females are also being found to benefit from multiple mating so that polyandry can generate competition among females for access to multiple males, and impose sexual selection on female traits that influence their mating success. Polyandry can reduce a male''s ability to monopolize females, and thus weaken male focused sexual selection. Perhaps the most important effect of polyandry on males arises because of sperm competition and cryptic female choice. Polyandry favours increased male ejaculate expenditure that can affect sexual selection on males by reducing their potential reproductive rate. Moreover, sexual selection after mating can ameliorate or exaggerate sexual selection before mating. Currently, estimates of sexual selection intensity rely heavily on measures of male mating success, but polyandry now raises serious questions over the validity of such approaches. Future work must take into account both pre- and post-copulatory episodes of selection. A change in focus from the products of sexual selection expected in males, to less obvious traits in females, such as sensory perception, is likely to reveal a greater role of sexual selection in female evolution.     

Breaking the rules: sex roles and genetic mating system of the pheasant coucal

Generally in birds, the classic sex roles of male competition and female choice result in females providing most offspring care while males face uncertain parentage. In less than 5% of species, however, reversed courtship sex roles lead to predominantly male care and low extra-pair paternity. These role-reversed species usually have reversed sexual size dimorphism and polyandry, confirming that sexual selection acts most strongly on the sex with the smaller parental investment and accordingly higher potential reproductive rate. We used parentage analyses and observations from three field seasons to establish the social and genetic mating system of pheasant coucals, Centropus phasianinus, a tropical nesting cuckoo, where males are much smaller than females and provide most parental care. Pheasant coucals are socially monogamous and in this study males produced about 80% of calls in the dawn chorus, implying greater male sexual competition. Despite the substantial male investments, extra-pair paternity was unusually high for a socially monogamous, duetting species. Using two or more mismatches to determine extra-pair parentage, we found that 11 of 59 young (18.6%) in 10 of 21 broods (47.6%) were not sired by their putative father. Male incubation, starting early in the laying sequence, may give the female opportunity and reason to seek these extra-pair copulations. Monogamy, rather than the polyandry and sex-role reversal typical of its congener, C. grillii, may be the result of the large territory size, which could prevent females from monopolising multiple males. The pheasant coucal’s exceptional combination of classic sex-roles and male-biased care for extra-pair young is hard to reconcile with current sexual selection theory, but may represent an intermediate stage in the evolution of polyandry or an evolutionary remnant of polyandry.     

Alternative mating tactics and extreme male dimorphism in fig wasps

The dimorphisms in morphology and behaviour of male fig wasps are among the most extreme in the animal kingdom, and offer excellent opportunities to test the predictions of certain sexual selection models. Winged males resemble their conspecific females closely, but wingless males are so divergent in form that they have repeatedly been classified into different taxa. Wingless males mate within their natal fig fruits, whereas winged males disperse to mate. Individual species may have winged males, wingless males or both morphs. A key hypothesis proposes that sexual selection on male mating opportunities favours winged males in species with small broods and wingless males in species with large broods. Using data from 114 species in 33 genera, we show that both simple and formal comparative analyses support the correlated evolution of large brood size and male winglessness. Theoretical models further predict that, in male dimorphic species, the proportion of winged males should equal (in cases without local mate competition) or exceed (in cases with local mate competition) the proportion of females developing in fig fruits without wingless males. These predictions are met by eight out of nine male dimorphic species studied. Taken together, the patterns across all species, and between different male dimorphic species, strongly support sexual selection on mating opportunities as the major determinant of male morph ratios in fig wasps.