Evolutionary Implications of Reproductive Tract Microbiota in the Polygynandrous Red Junglefowl (Gallus gallus)

Host-associated microbiota are temporally unstable—novel bacteria are acquired from the environment and exchanged between hosts. In the current literature, hypotheses have emerged about the potential for host social behaviour to influence its gut microbiota, while the gut bacteria may affect host social decision making. A similar, yet under-explored evolutionary interdependence may also occur between host mating behaviour and its reproductive microbiota. For example, reproductive bacteria may partake in host reproductive decisions, play a role in sexual selection and conflict, and determine host socio- sexual network structure. How bacterial communities in the reproductive tract relate to host evolution may be more evident in non-human hosts. We therefore investigated this association by characterising the microbiota in the reproductive tracts of both female and male red junglefowl (Gallus gallus)—the main wild ancestor of the domestic chicken (G. domesticus)—that mates frequently and with multiple partners (polygynandry) over the breeding season. By carrying out 16S rRNA gene sequencing on functionally-distinct reproductive tract samples from both sexes, ejaculates from males, and cloacal wipes from sexually-interacting social groups, we show that both females and males harbour bacteria across the continuum of their internal reproductive tracts, and that these assemblages are distinct from the gut microbiota. We find evidence for spatial structure in female reproductive tract microbiota, while male reproductive tract and ejaculate microbiota are more homogenous, suggesting a potential for sexual conflict between females and males. We also report that reproductive tract bacteria are sexually transmitted in social groups of red junglefowl, and that individuals that fail to secure a partner diverge in their cloacal microbiota from the rest of the mating population. Combined, our findings highlight the importance of exploring bacterial communities in the context of host evolution, and spark hypotheses about the sex-specific costs and benefits of exchanging bacteria during mating.