Etheostoma xanthovum, the Clarks Darter, is described as a new species endemic to the Clarks River drainage in Kentucky and Tennessee, USA. Etheostoma xanthovum was previously recognized as Etheostoma oophylax based on morphological characters. Subsequent to the description of E. oophylax, molecular phylogenetic analyses consistently resolved specimens from the Clarks River drainage and E. chienense as sister species, which together formed a sister clade to all other sampled populations of E. oophylax. Our analyses of morphological trait data, mitochondrial DNA, and genomic sampling using double digest restriction-site associated DNA sequencing support the distinctiveness of E. xanthovum. Morphologically, E. xanthovum differs slightly from E. oophylax in the modal number of dorsal fin rays (12 versus 11) and in the average number of scale rows around the caudal peduncle (21.8 versus 20.4). Etheostoma xanthovum does not share mitochondrial DNA haplotypes with E. oophylax or E. chienense. Phylogenomic analysis of an average of 28,448 double digest restriction-site associated DNA loci per sampled specimen resolves E. xanthovum and E. chienense as sister species, and assessment of genomic divergence supports the hypothesis that each of these two species represents a distinct and independently evolving lineage. In addition, we report a range extension of E. oophylax in the Obion River drainage, a direct tributary of the Mississippi River.