Mitochondrial DNA (mtDNA) variations were investigated among eighteen species in the Drosophila montium subgroup. The phylogenetic relationships of eighteen species in the D. montium subgroup were deduced by the fragment patterns of mtDNA using fourteen restriction endonucleases for the analysis. Eighteen species could be divided into three complexes, the kikkawai complex (6 species), the jambulina complex (4 species) and the auraria complex (8 species). Based on the number of shared fragment patterns, the kikkawai complex and the jambulina complex were closely related and the auraria complex was aparted from the other two. The phylogeny of eighteen species in the D. montium subgroup based on the mtDNA data under the consideration was constructed with the data such as reproductive isolation and two-dimensional electrophoresis. The genetic mechanism of speciation in the D. montium subgroup was discussed.