Restriction site mapping of mitochondrial DNA with 23 restriction endonucleases was used to examine the genetic and phylogenetic relationships of populations of D. silvestris and D. heteroneura from the island of Hawaii. Two morphological races of D. silvestris are known on the island of Hawaii. One has three bristle rows on the tibia of the foreleg and is found on the east side of the island. The other is found on the west side of the island and has the ancestral bristle row character of two rows on the tibia of the foreleg. All D. heteroneura have the ancestral bristle row character state. We demonstrate that mtDNA restriction site analysis can also differentiate the two D. silvestris races, and that the two bristle row D. silvestris are more closely related to D. heteroneura than they are to their three bristle row conspecifics using both distance and character state analysis. Our study (which uses six base recognition restriction endonucleases) is not sensitive enough to determine the phylogenetic relationships of populations within either of the D. silvestris lineages.