Asymmetric habitat isolation and sexual isolation predicted by the cost of migration and hybridization generate novel signatures of reinforcing selection

Reinforcement is an evolutionary process whereby increased prezygotic reproductive isolation evolves in response to the cost of hybridization. Despite theory predicting that multiple prezygotic barriers can evolve via reinforcement, most empirical studies examine a single barrier. We test novel predictions for the reinforcement of both habitat isolation and sexual isolation between ecologically divergent lineages under asymmetric migration: the lineage that emigrates more should evolve stronger habitat isolation due to the lower fitness of immigrants in the alternative habitat, while the lineage that receives more immigrants should exhibit stronger sexual isolation due to the lower fitness of hybrids. We found both signatures of reinforcement in two sympatric sister species of gall wasps that are host specific to the southern live oaks, Quercus virginiana and Q. geminata, respectively. Specifically, we observed stronger habitat isolation in the species with higher emigration rates, Belonocnema treatae, and stronger sexual isolation in the species facing more immigrants, B. fossoria. In contrast, comparisons of both species to a third, allopatric, species showed that B. kinseyi exhibited both lower habitat isolation and sexual isolation than the sympatric species, consistent with the classic predictions of reinforcement. Our study provides a rare examination of the interplay of ecology and geography in the evolution of multiple reproductive barriers to gene flow. Given that asymmetric migration between ecologically divergent lineages increasingly appears to be the rule rather than the exception, concomitant asymmetries in the strength of habitat and sexual isolation could be more widespread than currently understood. ### Competing Interest Statement The authors have declared no competing interest.