Alternative reproductive strategies explain asymmetries in reproductive isolation and reinforcement in two Ischnura damselfly species.

Theoretical and empirical studies of reinforcement have advanced our understanding of speciation, yet its role in polymorphic species remains understudied. Because morphs differ in behavior, morphology, and reproductive strategies, reinforcement may act unevenly among them, generating asymmetric reproductive isolation. We tested this prediction in the polymorphic damselflies Ischnura elegans and Ischnura graellsii, in which female morphs adopt alternative reproductive strategies. These species form two independent hybrid zones where reinforcement has strengthened mechanical isolation and driven reproductive character displacement in mating-related structures. We quantified five reproductive barriers across female morphs to evaluate how color polymorphism interacts with reinforcement. We found clear asymmetry between morphs: gynochrome females of both species showed reinforced mechanical isolation, whereas androchromes did not, consistent with their contrasting reproductive strategies. Additionally, gametic barriers evolved in opposite directions between species. Fertility isolation was reinforced in I. elegans, while oviposition and fertility barriers relaxed in I. graellsii, but symmetrically between female morphs, likely reflecting gene flow and purging of incompatibilities. Reinforcement strengthened reproductive isolation in a morph-specific manner, as pre-existing differences between female morphs influenced the likelihood of heterospecific mating and therefore the strength of selection against hybridization. Future work should examine whether these asymmetric dynamics generate cascading effects within species and contribute to morph-level diversification.