Distinguishing Black Bullhead (Ameiurus melas) from Brown Bullhead (A. nebulosus) can be difficult, in part, due to conflicting evidence regarding reliable meristic and morphological characters with which to base identifications. The possibility of hybrids with intermediate traits further complicates diagnosis. Identification guides have typically focused on adults, leaving guidance for juveniles largely undescribed. We compared morphological identifications of Black Bullhead and Brown Bullhead with a cytochrome c oxidase I (COI) barcode and microsatellite assignments. Specifically, we examined: (1) gill-raker count, (2) anal-fin ray count, (3) prominence of pectoral-spine serrae, (4) DNA barcoding using the COI gene, and (5) cluster analysis of nine microsatellite loci. With the exception of anal-fin ray counts, we found a high degree of agreement among all methods. Microsatellite analysis identified two clearly distinct genetic groups, and these groups corresponded to individuals identified as Black Bullhead and Brown Bullhead based on morphology. Although Black Bullhead and Brown Bullhead did not share COI haplotypes (except for one individual), we found greater intraspecific genetic distances among individuals identified as Brown Bullhead than between Brown Bullhead and Black Bullhead, demonstrating that caution is warranted against using COI haplotypes as a barcode to solely distinguish the two species. Pectoral-spine serrae prominence was a useful character to distinguish the two species and worked especially well on individuals <125 mm total length. Gill-raker counts of euthanized fish in the laboratory were appropriate for the identification of all sizes of Black Bullhead and Brown Bullhead (using the lowest number of gill rakers on the first gill arch if asymmetry was observed). Finally, microsatellites did not show evidence of recent hybridization at our sampling sites. We recommend using pectoral-spine serrae prominence to distinguish between Black Bullhead and Brown Bullhead in the field and using gill-raker counts for laboratory based identification.