Resolving higher-level phylogenetic networks with repeated hybridization in a complex of polytypic salamanders (Plethodontidae: Desmognathus)

Repeated hybridization between incipient lineages is a common feature of ecological speciation and ecomorphological diversification. However, computational constraints currently limit our ability to reconstruct network radiations from gene-tree data. Available methods are limited to level-1 networks wherein reticulations do not share edges, and higher-level networks may be non-identifiable in many cases. We present a heuristic method to recover information from higher-level networks across a range of potentially identifiable empirical scenarios, supported by a theorem and success in simulated data. When extrinsic information indicating the location and direction of recent or ancestral hybridization events is available, our method can yield successful estimates of non-level-1 networks, or at least a reduced possible set thereof. We apply this technique to the Pisgah clade of Desmognathus salamanders, which contains four to seven species exhibiting two discrete phenotypes, aquatic "shovel-nosed" and semi-aquatic "black-bellied" forms in the southern Appalachian Mountains of the eastern United States. Phylogenomic data strongly support a single backbone topology with up to five overlapping hybrid edges. These results suggest an unusual mechanism of ecomorphological hybrid speciation, wherein a binary threshold trait causes hybrids to shift between two microhabitat niches, promoting ecological divergence between sympatric hybrids and parentals. This contrasts with other well-known systems in which hybrids exhibit intermediate, novel, or transgressive phenotypes. Geographically proximate populations of both phenotypes exhibit admixture, and at least two black-bellied lineages have been produced via reticulations between shovel-nosed parentals, suggesting complex transmission dynamics. The genetic basis of these phenotypes is unclear and further data are needed to clarify the nature of selection and speciation in the group.