The Eastern-Japanese common toad, Bufo japonicus formosus, is distributed from southern Hokkaido to Kinki district of northeastern Honshu, Japan. However, Hokkaido populations are suspected to be nonnative and derived from relatively recent artificial introduction from Honshu. To clarify the origin of Hokkaido populations, we analyzed sequence variations of the mitochondrial cytochrome b gene in the toads from Hokkaido and compared the sequence data from previous studies. We obtained ten haplotypes from six sites on Hokkaido. Samples from the sites located along Ishikari river system, Asahikawa, Ebetsu, and Ishikari cities, had six haplotypes. Only one haplotype was obtained in Sapporo city and it was genetically far from Ishikari river's ones. Hakodate and Muroran cities samples had one and two haplotypes, respectively. All ten haplotypes from Hokkaido were genetically identical or close to haplotype data from Kanto district or Shizuoka Prefecture, mid-eastern Honshu, far from Hokkaido. Therefore, the toad populations of Hokkaido were considered to be nonnative and multiple introductions from eastern Honshu to Hokkaido were implied.