Extra‐pair mating in a passerine bird with highly duplicated major histocompatibility complex class II: Preference for the golden mean

Genes of the major histocompatibility complex (MHC) are essential in vertebrate adaptive immunity, and they are highly diverse and duplicated in many lineages. While it is widely established that pathogen‐mediated selection maintains MHC diversity through balancing selection, the role of mate choice in shaping MHC diversity is debated. Here, we investigate female mating preferences for MHC class II (MHCII) in the bluethroat (Luscinia svecica), a passerine bird with high levels of extra‐pair paternity and extremely duplicated MHCII. We genotyped family samples with mixed brood paternity and categorized their MHCII alleles according to their functional properties in peptide binding. Our results strongly indicate that females select extra‐pair males in a nonrandom, self‐matching manner that provides offspring with an allelic repertoire size closer to the population mean, as compared to offspring sired by the social male. This is consistent with a compatible genes model for extra‐pair mate choice where the optimal allelic diversity is intermediate, not maximal. This golden mean presumably reflects a trade‐off between maximizing pathogen recognition benefits and minimizing autoimmunity costs. Our study exemplifies how mate choice can reduce the population variance in individual MHC diversity and exert strong stabilizing selection on the trait. It also supports the hypothesis that extra‐pair mating is adaptive through altered genetic constitution in offspring.     

MHC-based patterns of social and extra-pair mate choice in the Seychelles warbler

The existence and nature of indirect genetic benefits to mate choice remain contentious. Major histocompatibility complex (MHC) genes, which play a vital role in determining pathogen resistance in vertebrates, may be the link between mate choice and the genetic inheritance of vigour in offspring. Studies have shown that MHC-dependent mate choice can occur in mammal and fish species, but little work has focused on the role of the MHC in birds. We tested for MHC-dependent mating patterns in the Seychelles warbler (Acrocephalus sechellensis). There was no influence of MHC class I exon 3 variation on the choice of social mate. However, females were more likely to obtain extra-pair paternity (EPP) when their social mate had low MHC diversity, and the MHC diversity of the extra-pair male was significantly higher than that of the cuckolded male. There was no evidence that females were mating disassortatively, or that they preferred males with an intermediate number of MHC bands. Overall, the results are consistent with the 'good genes' rather than the 'genetic compatibility' hypothesis. As female choice will result in offspring of higher MHC diversity, MHC-dependent EPP may provide indirect benefits in the Seychelles warbler if survival is positively linked to MHC diversity.     

Early arrival is not associated with more extra‐pair fertilizations in a long‐distance migratory bird

When assessing the benefits of early arrival date of migratory birds, a hidden and often ignored component of males’ fitness is the higher chance of early‐arriving birds to obtain extra‐pair fertilizations. Here we investigated how extra‐pair paternity might affect the relationship between male arrival date and number of fertilizations in a model study system, the European pied flycatcher Ficedula hypoleuca. For this purpose, we sampled and genotyped breeding pairs, unpaired males and offspring (including embryos from unhatched eggs when possible) of a Dutch pied flycatcher population. Detailed information on arrival date of males, egg laying date of their social mates and nest success was also recorded. Early‐arriving males had early‐laying females and males with early‐laying females had a higher probability of siring extra‐pair eggs and obtain more fertilizations. However, male arrival date alone did not correlate with the probability to gain extra‐pair paternity and neither to the amount of fertilized eggs. Both early‐ and late‐arriving males had a higher probability of losing paternity in their own nest compared to birds with an intermediate arrival date. Finally, late‐arriving males were more likely to remain unpaired but, interestingly, a few of these birds obtained paternity via extra‐pair copulations. Because earlier arrival date did not lead to more extra‐pair fertilizations and because such relationship seems to be driven mainly by the female's laying date, we conclude that the contribution of extra‐pair paternity to the overall fitness benefits of early male arrival date is relatively small.     

Extra‐pair offspring are less heterozygous than within‐pair offspring in American redstarts Setophaga ruticilla

The vast majority of bird species are socially monogamous; however, extra‐pair paternity is nearly ubiquitous and a number of theories have been proposed to explain the prevalence of this mixed mating strategy. Here, we test the genetic compatibility hypothesis – the idea that females seek extra‐pair copulations with males whose genes are more compatible with her own. For this study, we examined eight years of paternity data (2004–2011) from a Nearctic‐Neotropical migratory bird, the American redstart Setophaga ruticilla, breeding in southeastern Ontario, Canada. We predicted that females paired with genetically similar males (higher relatedness) would be more likely to produce extra‐pair offspring and that extra‐pair offspring would have higher levels of heterozygosity than within‐pair offspring. Alternatively, because this population experiences high levels of immigration, females may produce extra‐pair offspring with more genetically similar males because of the potential for outbreeding depression. Using five highly variable microsatellite markers, we examined patterns of relatedness among social pairs as well as measures of offspring heterozygosity. In contrast to our predictions, we found no difference in relatedness between social pairs where the females produced extra‐pair offspring and social pairs where the females produced only within‐pair offspring. However, extra‐pair offspring were significantly less heterozygous than within‐pair offspring. Together, these findings suggest that females a) are not engaging in extra‐pair fertilizations based on relatedness to their social mate and b) appear to be mating with extra‐pair males that are more genetically similar to themselves. We suggest there may be benefits for females to mate with genetically similar extra‐pair males in highly outbred populations with high rates of immigration, such as for maintaining co‐adapted gene complexes or genes coding for local adaptations.     

High rates of extra‐pair paternity in a socially monogamous beetle with biparental care

1. Nest construction and paternity assurance are predicted to favour biparental care in insects. The horned passalus (Odontotaenius disjunctus) is a socially monogamous beetle with biparental care that breeds in decaying logs. The genetic mating system of the horned passalus was investigated to determine if paternity assurance is likely to drive the evolution or maintenance of paternal care in this system. Parental time budgets were also examined to better understand the types and frequencies of behaviours performed by parents. 2. Genotyping‐by‐sequencing revealed high levels of extra‐pair paternity, with 54.8% of offspring sired by extra‐pair males and 70% of nests containing extra‐pair young. 3. More heterozygous social males were cuckolded less than more homozygous social males. Extra‐pair mating, however, seems unlikely to increase offspring genetic diversity as extra‐pair offspring were not more heterozygous than within‐pair offspring, and average brood heterozygosity did not increase with higher rates of extra‐pair paternity. 4. Behavioural observations demonstrated that parents spent on average 46.5% of their time processing the decaying wood resource for larval offspring. Because resource processing is a by‐product of feeding and provides shareable benefits for all larvae in the brood, this form of paternal care could be favoured despite low paternity.     

Direct benefits and the evolution of female-biased cooperative breeding in Seychelles warblers

Abstract.— Inclusive fitness benefits have been suggested to be a major selective force behind the evolution of cooperative breeding. We investigated the fitness benefits selecting for cooperative breeding in the Seychelles warbler, Acrocephalus sechellensis . A microsatellite-based genotyping method was used to determine the relatedness of subordinates to group offspring in an isolated population of Seychelles warblers. The indirect and direct breeding benefits accruing to individual subordinates were then calculated for every successful breeding event over a three-year period. We show that female subordinates frequently gained parentage and that this, combined with high levels of extra group paternity, resulted in low levels of relatedness between subordinates and non descendent offspring within a territory. Direct breeding benefits were found to be significantly higher than indirect kin benefits for both female and male subordinates. As predicted, female subordinates gained significantly more direct breeding opportunities and therefore higher inclusive fitness benefits by being a subordinate within a group than did males. This may explain why most subordinates in the Seychelles warbler are female.     

Male age and its association with reproductive traits in captive and wild house sparrows

Evolutionary theory predicts that females seek extra‐pair fertilizations from high‐quality males. In socially monogamous bird species, it is often old males that are most successful in extra‐pair fertilizations. Adaptive models of female extra‐pair mate choice suggest that old males may produce offspring of higher genetic quality than young males because they have proven their survivability. However, old males are also more likely to show signs of reproductive senescence, such as reduced sperm quality. To better understand why old males account for a disproportionally large number of extra‐pair offspring and what the consequences of mating with old males are, we compared several sperm traits of both captive and wild house sparrows, Passer domesticus. Sperm morphological traits and cloacal protuberance volume (a proxy for sperm load) of old and young males did not differ substantially. However, old males delivered almost three times more sperm to the female's egg than young males. We discuss the possibility of a post‐copulatory advantage for old over young males and the consequences for females mated with old males.     

Age‐dependent changes in infidelity in Seychelles warblers

Extra‐pair paternity (EPP) is often linked to male age in socially monogamous vertebrates; that is, older males are more likely to gain EPP and less likely to be cuckolded. However, whether this occurs because males improve at gaining paternity as they grow older, or because “higher quality” males that live longer are preferred by females, has rarely been tested, despite being central to our understanding of the evolutionary drivers of female infidelity. Moreover, how extra‐pair reproduction changes with age within females has received even less attention. Using 18 years of longitudinal data from an individually marked population of Seychelles warblers (Acrocephalus sechellensis), we found considerable within‐individual changes in extra‐pair reproduction in both sexes: an early‐life increase and a late‐life decline. Furthermore, males were cuckolded less as they aged. Our results indicate that in this species age‐related patterns of extra‐pair reproduction are determined by within‐individual changes with age, rather than differences among individuals in longevity. These results challenge the hypothesis—based on longevity reflecting intrinsic quality—that the association between male age and EPP is due to females seeking high‐quality paternal genes for offspring. Importantly, EPP accounted for up to half of male reproductive success, emphasizing the male fitness benefits of this reproductive strategy. Finally, the occurrence of post‐peak declines in extra‐pair reproduction provides explicit evidence of senescence in infidelity in both males and females.     

Variation in parent–offspring kinship in socially monogamous systems with extra‐pair reproduction and inbreeding

Female extra‐pair reproduction in socially monogamous systems is predicted to cause cuckolded socially‐paired males to conditionally reduce paternal care, causing selection against extra‐pair reproduction and underlying polyandry. However, existing models and empirical studies have not explicitly considered that cuckolded males might be related to their socially‐paired female and/or to her extra‐pair mate, and therefore be related to extra‐pair offspring that they did not sire but could rear. Selection against paternal care, and hence against extra‐pair reproduction, might then be weakened. We derive metrics that quantify allele‐sharing between within‐pair and extra‐pair offspring and their mother and her socially‐paired male in terms of coefficients of kinship and inbreeding. We use song sparrow (Melospiza melodia) paternity and pedigree data to quantify these metrics, and thereby quantify the joint effects of extra‐pair reproduction and inbreeding on a brood's total allelic value to its socially‐paired parents. Cuckolded male song sparrows were almost always detectably related to extra‐pair offspring they reared. Consequently, although brood allelic value decreased substantially following female extra‐pair reproduction, this decrease was reduced by within‐pair and extra‐pair reproduction among relatives. Such complex variation in kinship within nuclear families should be incorporated into models considering coevolutionary dynamics of extra‐pair reproduction, parental care, and inbreeding.     

Extra‐pair mating in a socially monogamous and paternal mouth‐brooding cardinalfish

Many vertebrates form monogamous pairs to mate and care for their offspring. However, genetic tools have increasingly shown that offspring often arise from matings outside of the monogamous pair bond. Social monogamy is relatively common in coral reef fishes, but there have been few studies that have confirmed monogamy or extra‐pair reproduction, either for males or for females. Here, long‐term observations and genetic tools were applied to examine the parentage of embryos in a paternally mouth‐brooding cardinalfish, Sphaeramia nematoptera. Paternal care in fishes, such as mouth‐brooding, is thought to be associated with a high degree of confidence in paternity. Two years of observations confirmed that S. nematoptera form long‐term pair bonds within larger groups. However, genetic parentage revealed extra‐pair mating by both sexes. Of 105 broods analysed from 64 males, 30.1% were mothered by a female that was not the partner and 11.5% of broods included eggs from two females. Despite the high paternal investment associated with mouth‐brooding, 7.6% of broods were fertilized by two males. Extra‐pair matings appeared to be opportunistic encounters with individuals from outside the immediate group. We argue that while pair formation contributes to group cohesion, both males and females can maximize lifetime reproductive success by taking advantage of extra‐pair mating opportunities.     

Pathogen burden,co‐infection and major histocompatibility complex variability in the European badger (Meles meles)

Pathogen‐mediated selection is thought to maintain the extreme diversity in the major histocompatibility complex (MHC) genes, operating through the heterozygote advantage, rare‐allele advantage and fluctuating selection mechanisms. Heterozygote advantage (i.e. recognizing and binding a wider range of antigens than homozygotes) is expected to be more detectable when multiple pathogens are considered simultaneously. Here, we test whether MHC diversity in a wild population of European badgers (Meles meles) is driven by pathogen‐mediated selection. We examined individual prevalence (infected or not), infection intensity and co‐infection of 13 pathogens from a range of taxa and examined their relationships with MHC class I and class II variability. This population has a variable, but relatively low, number of MHC alleles and is infected by a variety of naturally occurring pathogens, making it very suitable for the investigation of MHC–pathogen relationships. We found associations between pathogen infections and specific MHC haplotypes and alleles. Co‐infection status was not correlated with MHC heterozygosity, but there was evidence of heterozygote advantage against individual pathogen infections. This suggests that rare‐allele advantages and/or fluctuating selection, and heterozygote advantage are probably the selective forces shaping MHC diversity in this species. We show stronger evidence for MHC associations with infection intensity than for prevalence and conclude that examining both pathogen prevalence and infection intensity is important. Moreover, examination of a large number and diversity of pathogens, and both MHC class I and II genes (which have different functions), provide an improved understanding of the mechanisms driving MHC diversity.     

High rates of infidelity in the Grey Fantail Rhipidura albiscapa suggest that testis size may be a better correlate of extra‐pair paternity than sexual dimorphism

The Grey Fantail Rhipidura albiscapa is a socially monogamous passerine endemic to Australia. Behavioural and morphological clues point to opposing conclusions as to its breeding system; sexual monomorphism and monochrome colorations suggest monogamy, whereas relatively large testes and a prominent cloacal protruberance are more indicative of multiple mating and sperm competition. We used five highly variable microsatellite loci to investigate the genetic breeding system of this species. Paternity was assigned to 49 of 69 (71%) offspring tested and the overall rate of partner infidelity was high, with 55% of offspring being sired by an extra‐pair male and 64% of all clutches containing extra‐pair young. This puts the Grey Fantail amongst the most promiscuous socially monogamous species yet studied. Where extra‐pair fathers were identified, these were invariably in neighbouring territories, and although larger males did not gain more paternities overall, extra‐pair offspring tended to be fathered by larger males than expected by chance. We interpret our findings in light of some of the potential costs and benefits associated with extra‐pair paternity.     

Lack of evidence for improved immune response of extra‐pair nestlings in collared flycatcher Ficedula albicollis

Extra‐pair paternity is common in many socially monogamous bird species. Increasing evidence suggests that extra‐pair copulations are female‐driven, but benefits for females mating outside social pair‐bonds are still poorly understood. The most influential explanation, “good genes” hypothesis, states that females mated socially with low quality males, engage in extra‐pair copulations to obtain genetic benefits for their progeny. According to this model, enhanced performance of extra‐pair offspring is expected. Here, based on 4‐year study of collared flycatcher Ficedula albicollis, we compared the condition of extra‐pair and within‐pair young. We found no difference in immune response and body size between maternal half‐siblings raised in the same nests. Additionally sex ratio was not biased among extra‐pair nestlings, and paternity was not associated with hatching rank. Our results failed to reveal “good genes” effects in the studied population. These effects might be hard to detect, but other hypotheses should also be studied more thoroughly in the future.     

Malaria infection status predicts extra‐pair paternity in the blue tit

Extra‐pair matings comprise a common reproductive strategy among socially monogamous bird species. However, it remains unclear why females decide to mate with extra‐pair males. Indirect benefits in terms of improving offspring genetic quality are usually invoked to explain this phenomenon. Parasite resistance genes are often considered as a female target of seeking extra‐pair matings, but the direct test of this hypothesis is generally lacking. Here, we report on a relationship between the status of infection with malaria parasites (Plasmodium and Haemoproteus) and occurrence of extra‐pair paternity in a wild population of the blue tit Cyanistes caeruleus inhabiting Gotland (Sweden). We found that the probability of extra‐pair paternity is significantly related to the infection status of social parents. Infected males showed higher probability of being cuckolded than uninfected ones. However, this was observed only among males mated to uninfected females. Thus, avian malaria may potentially contribute to explanation of extra‐pair mating behaviour.     

Family‐assisted inference of the genetic architecture of major histocompatibility complex variation

With their direct link to individual fitness, genes of the major histocompatibility complex (MHC) are a popular system to study the evolution of adaptive genetic diversity. However, owing to the highly dynamic evolution of the MHC region, the isolation, characterization and genotyping of MHC genes remain a major challenge. While high‐throughput sequencing technologies now provide unprecedented resolution of the high allelic diversity observed at the MHC, in many species, it remains unclear (i) how alleles are distributed among MHC loci, (ii) whether MHC loci are linked or segregate independently and (iii) how much copy number variation (CNV) can be observed for MHC genes in natural populations. Here, we show that the study of allele segregation patterns within families can provide significant insights in this context. We sequenced two MHC class I (MHC‐I) loci in 1267 European barn owls (Tyto alba), including 590 offspring from 130 families using Illumina MiSeq technology. Coupled with a high per‐individual sequencing coverage (~3000×), the study of allele segregation patterns within families provided information on three aspects of the architecture of MHC‐I variation in barn owls: (i) extensive sharing of alleles among loci, (ii) strong linkage of MHC‐I loci indicating tandem architecture and (iii) the presence of CNV in the barn owl MHC‐I. We conclude that the additional information that can be gained from high‐coverage amplicon sequencing by investigating allele segregation patterns in families not only helps improving the accuracy of MHC genotyping, but also contributes towards enhanced analyses in the context of MHC evolutionary ecology.     

Increased extra‐pair paternity in broods of aging males and enhanced recruitment of extra‐pair young in a migratory bird

Despite keen interest in extra‐pair mating in birds, its adaptive significance remains unresolved. Here, we use a multi‐year dataset to test whether traits of a female's social mate influence her propensity to produce extra‐pair offspring in a population of house wrens, and whether producing extra‐pair young has consequences for a female's fitness through effects on offspring survival. Females were most likely to produce extra‐pair offspring when paired with old males and when paired with males on poor‐quality territories, although this latter effect was marginally nonsignificant. Among offspring, the cutaneous immunity of within‐pair young decreased as the age of their sires increased, but cutaneous immunity of extra‐pair young was not affected by the age of their extra‐pair sires or by the age of the males rearing them. Extra‐pair offspring were more likely than within‐pair offspring to return as breeding adults to the local population, with extra‐pair sons being more likely to return as a breeder for multiple years. Our findings support the hypothesis that females produce extra‐pair offspring to enhance their inclusive fitness beyond what they are capable of given the male with which they are socially paired.     

Characterization of MHC-I in the blue tit (Cyanistes caeruleus) reveals low levels of genetic diversity and trans-population evolution across European populations

The major histcompatibility complex (MHC) is a vital component of the adaptive immune system in all vertebrates. This study is the first to characterize MHC class I (MHC-I) in blue tits (Cyanistes caeruleus), and we use MHC-I exon 3 sequence data from individuals originating from three locations across Europe: Spain, the Netherlands to Sweden. Our phylogeny of the 17 blue tit MHC-I alleles contains one allele cluster with low nucleotide diversity compared to the remaining more diverse alleles. We found a significant evidence for balancing selection in the peptide-binding region in the diverse allele group only. No separation according to geographic location was found in the phylogeny of alleles. Although the number of MHC-I loci of the blue tit is comparable to that of other passerine species, the nucleotide diversity of MHC-I appears to be much lower than that of other passerine species, including the closely related great tit (Parus major) and the severely inbred Seychelles warbler (Acrocephalus sechellensis). We believe that this initial MHC-I characterization in blue tits provides an important step towards understanding the mechanisms shaping MHC-I diversity in natural populations.     

Occurrence of extra‐pair paternity is connected to social male's MHC‐variability in the scarlet rosefinch Carpodacus erythrinus

Genes of the Major Histocompatibility Complex (MHC) represent an essential component of the vertebrate acquired immune system. In the last decades, the role of MHC genes in mate choice has been subject of particular scientific interest. However, results of studies dealing with this topic in different species are equivocal and mechanisms conducting MHC‐based mate choice are still puzzling. We investigated the impact of MHC class I variability on within‐pair and extra‐pair fertilisation success in a wild population of a socially monogamous passerine bird with considerable rates of extra‐pair paternity, the scarlet rosefinch Carpodacus erythrinus. We found some support for the ‘good‐genes‐as‐heterozygosity model’, as social males of high MHC‐heterozygosity were cheated by their females less frequently than less MHC‐heterozygous males. However, cuckolding males were not more MHC‐heterozygous than the cheated social males, nor were extra‐pair young more MHC‐heterozygous than within‐pair young. We did not find any evidence for mating preferences according to the complementarity model.     

Genetic covariance between components of male reproductive success: within‐pair vs. extra‐pair paternity in song sparrows

The evolutionary trajectories of reproductive systems, including both male and female multiple mating and hence polygyny and polyandry, are expected to depend on the additive genetic variances and covariances in and among components of male reproductive success achieved through different reproductive tactics. However, genetic covariances among key components of male reproductive success have not been estimated in wild populations. We used comprehensive paternity data from socially monogamous but genetically polygynandrous song sparrows (Melospiza melodia) to estimate additive genetic variance and covariance in the total number of offspring a male sired per year outside his social pairings (i.e. his total extra‐pair reproductive success achieved through multiple mating) and his liability to sire offspring produced by his socially paired female (i.e. his success in defending within‐pair paternity). Both components of male fitness showed nonzero additive genetic variance, and the estimated genetic covariance was positive, implying that males with high additive genetic value for extra‐pair reproduction also have high additive genetic propensity to sire their socially paired female's offspring. There was consequently no evidence of a genetic or phenotypic trade‐off between male within‐pair paternity success and extra‐pair reproductive success. Such positive genetic covariance might be expected to facilitate ongoing evolution of polygyny and could also shape the ongoing evolution of polyandry through indirect selection.     

Avian β-defensin variation in bottlenecked populations: the Seychelles warbler and other congeners

β-defensins are important components of the vertebrate innate immune system responsible for encoding a variety of anti-microbial peptides. Pathogen-mediated selection is thought to act on immune genes and potentially maintain allelic variation in the face of genetic drift. The Seychelles warbler, Acrocephalus sechellensis, is an endemic passerine that underwent a recent bottleneck in its last remaining population, resulting in a considerable reduction in genome-wide variation. We genotyped avian β-defensin (AvBD) genes in contemporary (2000–2008) and museum samples (1876–1940) of the Seychelles warbler to investigate whether immunogenetic variation was lost through this bottleneck, and examined AvBD variation across four other Acrocephalus species with varying demographic histories. No variation was detected at four of the six AvBD loci screened in the post-bottleneck population of Seychelles warbler, but two silent nucleotide polymorphisms were identified at AvBD8 and one potentially functional amino-acid variation was observed at AvBD11. Variation in the Seychelles warbler was significantly lower than in the mainland migratory congeneric species investigated, but it similar to that found in other bottlenecked species. In addition, screening AvBD7 in 15 museum specimens of Seychelles warblers sampled prior to the bottleneck (1877–1905) revealed that this locus possessed two alleles previously, compared to the single allele in the contemporary population. Overall, the results show that little AvBD variation remains in the Seychelles warbler, probably as a result of having low AvBD diversity historically rather than the loss of variation due to drift associated with past demographic history. Given the limited pathogen fauna, this lack of variation at the AvBD loci may currently not pose a problem for this isolate population of Seychelles warblers, but it may be detrimental to the species’ long-term survival if new pathogens reach the population in the future.