Contact zones between divergent lineages of aquatic taxa have been described from the northeastern Pacific Ocean. We surveyed samples of Dolly Varden (Salvelinus malma) from their North American range for variation at 14 microsatellite DNA loci. After accounting for hybridization between Dolly Varden and co-occurring bull trout (Salvelinus confluentus) and Arctic char (Salvelinus alpinus), we found evidence for two genetic lineages of Dolly Varden consistent with the previously recognized subspecies, northern Dolly Varden (S. m. malma) and southern Dolly Varden (S. m. lordii). We documented a contact zone between the two subspecies from the eastern Alaska Peninsula to Cook Inlet, Alaska, where admixture values (i.e., the proportion of the genome estimated to be composed of northern Dolly Varden, QNDV) ranged between QNDV= 0.245 and 0.754 across about 700 ocean kilometres. Populations of Dolly Varden showing low admixture (i.e., less than 5%) were located a minimum of 346 km to the west to 1200 km to the southeast, respectively, from the contact zone. The two lineages of Dolly Varden probably stem from isolation and subsequent divergence in, and dispersal from, distinct northern and southern Pleistocene glacial refugia and substantiate the treatment of S. malma as two subspecies and as at least two designatable units under Canada’s Species at Risk Act.