Mitochondrial DNA sequences and 5 microsatellite DNA loci were used to assess the genetic structure of Micropterus punctulatus (spotted bass) in the upper Red, Ouachita, and Arkansas river basins. Results for 318 spotted bass from 14 localities indicated that extant populations in East Cache Creek, which potentially supported the nominal subspecies M. punctulatus wichitae, are either introductions or they are weakly divergent native populations. Both mtDNA and micro-satellite DNA indicated that populations of M. punctulatus in the Ouachita River basin are more similar to those in the Arkansas River basin than to those in the Red River basin. This conflicts with a previous Pleistocene model for the aquatic biogeography of the region. Microsatellites showed no evidence of genetic introgression by M. dolomieu; mtDNA was not divergent between the 2 species, possibly because of ancient introgression.