Nile tilapia (Oreochromis niloticus) is one of the most important species in Tanzania for inland fisheries and aquaculture. Although indigenous to the country, it is only naturally distributed within the margins of Lake Tanganyika and peripheral water bodies. The widespread distribution across other parts of the country is a consequence of introductions that started in the 1950s. We investigated the population genetic structure of Nile tilapia across Tanzania using nuclear microsatellite markers, and compared the head and body morphology of populations using geometric morphometric analyses. We found the Lake Tanganyika population to be genetically distinct from the introduced populations. However, there were no clear morphological differences in head and body shape that distinguished the Lake Tanganyika population from the others. We conclude that the Lake Tanganyika population of Nile tilapia represents a unique genetic resource within the country. We suggest that Nile tilapia aquaculture within the Lake Tanganyika catchment should be restricted to the indigenous strain.