Genomic and phenotypic divergence informs translocation strategies for an endangered freshwater fish.

Translocation, the movement of organisms for conservation purposes, can result in unintended introgression if genetic material flows between populations in new ways. The Bluemask Darter Etheostoma akatulo is a federally endangered species of freshwater fish inhabiting the Caney Fork River system and three of its tributaries (Collins River, Rocky River, and Cane Creek) in Tennessee. The current conservation strategy for Bluemask Darters involves translocating the progeny of broodstock from the Collins River (in the west) to the Calfkiller River (in the east) where the species had been extirpated. In this study, we use ddRAD sequence data from across the extant range to assess this translocation strategy in light of population structure, phylogeny, and demography. We also include museum specimen data to assess morphological variation among extant and extirpated populations. Our analyses reveal substantial genetic and phenotypic disparities between a western population in the Collins River and an eastern population encompassing the Rocky River, Cane Creek, and upper Caney Fork, the two of which shared common ancestry more than 100,000 years ago. Furthermore, morphological analyses classify 12 of 13 Calfkiller River specimens with phenotypes consistent with the eastern population. These results suggest that current translocations perturb the evolutionary boundaries between two delimited populations. Instead, we suggest that repopulating the Calfkiller River using juveniles from the Rocky River could balance conflicting signatures of demography, diversity, and divergence. Beyond conservation, the microgeographic structure of Bluemask Darter populations adds another puzzle to the phylogeography of the hyperdiverse freshwater fishes in eastern North America.