Historical events have had a great impact on the biogeography of fishes of western North America. We examined the genetic variation of the speckled dace (Rhinichthys osculus) to determine the effects on this species of extensive hydrological changes during the last 10 million years in the Bonneville and Lahontan basins of the Great Basin and the upper Snake River Basin. Eight hundred sixty-nine base pairs of the mitochondrial gene cytochrome b were sequenced from 97 individuals representing 22 populations within these 3 basins, as well as from 2 individuals of longnose dace (Rhinichthys cataractae) that served as outgroups. Additionally, 13 speckled dace sequences representing 3 Bonneville populations were used from GenBank. Phylogenetic relationships were reconstructed using maximum parsimony and maximum likelihood criteria. Analysis of molecular variance was used to determine population structure and to estimate the amount of gene flow across the community boundaries. Three distinct clades were reconstructed representing the Lahontan Basin, the northern Bonneville and upper Snake River basins, and the southern Bonneville Basin. Additionally, most of the population structuring was explained by variation among basins (65.33%) Speckled dace demonstrated high genetic variation. As hypothesized, the northern and southern Bonneville specimens formed separate clades; however, the southern Bonneville clade was basal to a sister clade formed by the northern Bonneville/upper Snake River and Lahontan clades. These relationships indicate that Pliocene connections between the Snake, Lahontan, and Bonneville drainages, rather than more recent Pleistocene connections, best explain population structuring in speckled dace.