During the last decades, cowpox virus, a member of the genus Orthopoxvirus within the Poxviridae family, has appeared as a pathogen in domestic cats, zoo animal species, and humans. At the same time, vaccinia virus, another orthopoxvirus, has been used as a recombinant vaccine vector with foreign genes inserted in the thymidine kinase (TK) gene. By PCR and cycle sequencing, we have determined the nucleotide sequences of the TK gene and the A-type inclusion protein (ATIP) gene of virus isolates from two human cowpox cases in Sweden, as well as a human and a feline case from Norway. We also obtained the corresponding sequences from ectromelia virus (strain Moscow), cowpox virus (strain Brighton) and vaccinia virus (strain Western Reserve). The new virus isolates differed from ectromelia virus and vaccinia virus, and were confirmed to be cowpox virus strains. Isolates originating from the same country had nearly identical TK sequences and fully identical ATIP sequences. They probably represent local geographical strains of cowpox virus.