The black bullhead, Ameiurus melas of the family Ictaluridae, is a freshwater fish native to North America that was introduced throughout Europe in the late 1800s. Using mitochondrial DNA (mtDNA) as a genetic marker, the present study investigates the genetic structure and historical demography of A. melas in North America. MtDNA-based phylogenetic analyses revealed the existence of two distinct lineages (A and B) of A. melas. While lineage A clustered with the previously reported sequences of A. melas, lineage B emerged as a unique clade like other species of the genus Ameiurus. Individuals belonging to lineage B were mostly from the south central region of the United States, the region that never glaciated during the last Ice Age. Results of a fossil-based molecular clock analysis suggest that lineage A and the paraphyletic lineage B of A. melas diverged from their respective common ancestors approximately 3 (±0.2) and 15.9 (±1.3) million years ago. Lineage B could possibly be a hybrid species, possessing the mtDNA haplotype of its maternal parent, an Ameiurus species that has gone extinct. While lineage A showed evidence of population expansion, lineage B did not show any evidence of population expansion, but rather is comprised of geographically structured populations.