A comparison of DNA sequences from one Danish (Laesø) and four Swedish strains of Apis mellifera mellifera (the black honeybee), A. m. ligustica (the Italian honeybee), A. m. carnica, and A. m. anatoliaca was performed. The sequences were produced using PCR amplification and direct sequencing of two mtDNA sequences common to all species. The two segments tested were: 1) a 897 bp region of the mt CO-I gene, and 2) a 434-881 bp region of the last part of the mt CO-I gene, the tRNAleu gene, an intergenic region and a small section in the beginning of the CO-II gene. The comparisons of the 897 bp region of the mt CO-I gene showed that two haplotypes with a difference of 15 bp were observed - one haplotype comprising the black honeybee and one all the other subspecies. The alignments of the CO-I / CO-II regions indicate that the strains of the black honeybee could be divided into two types in the studied region: one with 685 bp comprised the Danish strain (Laesø) and the strain from Stockholm's SkærgÅrd, and the other with 881 bp included Swedish strains from UmeÅ, Västerbotten and Jamtland. All the other subspecies, which in the studied region differed from each other only in a few base substitutions, have 434 bp. The length differences are mainly due to duplications in the investigated region. The study shows that the different subspecies can be identified on the basis of mtDNA. The phylogenetic analyses showed that the black honeybee, which can be further divided into different strains on basis of the number of duplications, seems to stand in an isolated position - originating presumably from a long termed isolation during and after the last glaciation.