Pervasive gene flow despite strong and varied reproductive barriers in swordtails.

UNLABELLED: One of the mechanisms that can lead to the formation of new species occurs through the evolution of reproductive barriers. However, recent research has demonstrated that hybridization has been pervasive across the tree of life even in the presence of strong barriers. Swordtail fishes (genus Xiphophorus ) are an emerging model system for studying the interface between these barriers and hybridization. We document overlapping mechanisms that act as barriers between closely related species, X. birchmanni and X. cortezi , by combining genomic sequencing from natural hybrid populations, artificial crosses, behavioral assays, sperm performance, and developmental studies. We show that strong assortative mating plays a key role in maintaining subpopulations with distinct ancestry in natural hybrid populations. Lab experiments demonstrate that artificial F 1 crosses experience dysfunction: crosses with X. birchmanni females were largely inviable and crosses with X. cortezi females had a heavily skewed sex ratio. Using F 2 hybrids we identify several genomic regions that strongly impact hybrid viability. Strikingly, two of these regions underlie genetic incompatibilities in hybrids between X. birchmanni and its sister species X. malinche . Our results demonstrate that ancient hybridization has played a role in the origin of this shared genetic incompatibility. Moreover, ancestry mismatch at these incompatible regions has remarkably similar consequences for phenotypes and hybrid survival in X. cortezi Ô X. birchmanni hybrids as in X. malinche Ô X. birchmanni hybrids. Our findings identify varied reproductive barriers that shape genetic exchange between naturally hybridizing species and highlight the complex evolutionary outcomes of hybridization.

SIGNIFICANCE STATEMENT: Biologists are fascinated by how the diverse species we see on Earth have arisen and been maintained. One driver of this process is the evolution of reproductive barriers between species. Despite the commonality of these barriers, many species still exchange genes through a process called hybridization. Here, we show that related species can have a striking array of reproductive barriers-from genetic interactions that harm hybrids to mate preferences that reduce hybridization in the first place. However, we also find that genetic exchange between these species is very common, and may itself play an important role in the evolution of reproductive barriers. Together, our work highlights the complex web of interactions that impact the origin and persistence of distinct species.