Different mating systems can strongly affect the extent of genetic diversity and population structure among species. Given the increased effects of genetic drift on reduced population size, theory predicts that species undergoing self-fertilization should have greater population structure than outcrossed species, however demographic dynamics may affect this scenario. The mangrove killifish clade is composed of the two only known examples of self-fertilising species among vertebrates (Kryptolebias marmoratus and K. hermaphroditus). A third species in this clade, K. ocellatus, inhabits mangrove forests in southeast Brazil, however its mating system and patterns of genetic structure have been rarely explored. Here, we examined the genetic structure and phylogeographic patterns of K. ocellatus along its distribution, using mitochondrial DNA and microsatellites to compare its patterns of genetic structure with the predominantly selfing and often syntopic, K. hermaphroditus. Our results indicate that K. ocellatus reproduces mainly by outcrossing across much of its known range, with no current evidence of selfing, despite being an androdioecious species. Our results also reveal a stronger population subdivision in K. ocellatus compared to K. hermaphroditus, contrary to the theoretical predictions based on reproductive biology of the two species. Our findings indicate that, although morphologically similar, K. ocellatus and K. hermaphroditus had remarkably different evolutionary histories when colonising the same mangrove areas in south-eastern Brazil, with other factors (e. g. time of colonisation, dispersal/establishment capacity) having more profound effects on the current population structuring of those species than differences in mating systems.

All details about fish collection, mtDNA amplification and genotyping are described in the main body of the manuscript.