Pleistocene speciation and glacial refugia in a temperate freshwater biodiversity hotspot

Among temperate regions of the world, freshwater fish species richness is highest in the Central Highlands of eastern North America. Historical biogeographic and phylogeographic researchers have investigated mechanisms driving this exceptional diversity, yet the role of major climatic events, like Pleistocene glaciation, is incompletely characterized. In this study, we analyze genomic DNA sequence data sampled from populations of the widely distributed Gilt Darter, Percina evides Jordan & Copeland 1877, to reconstruct pre-glacial drainage patterns and assess the impact of Pleistocene glaciation on generating the high species diversity of eastern North American freshwater fishes. Phylogenomics, population genomic analyses, and evaluation of morphology delimit four species currently classified as P. evides. These species likely diverged via allopatric speciation among the disjunct regions of the Central Highlands driven by the onset of Pleistocene glaciation. Divergence times among newly delimited species of the Gilt Darter complex are congruent with the onset of glaciation, periods of river incision and aggradation, and river network rearrangement. The discovery of new species in the Percina evides complex and their phylogenetic relationships highlight how Pleistocene glaciation and glacial refugia contributed to the remarkable temperate freshwater biodiversity hotspot of the Central Highlands.