Background: The need for enhancing the productivity of fisheries in Africa triggered the introduction of non-native fish, causing dramatic changes to local species. In East Africa, the extensive translocation of Nile tilapia (Oreochromis niloticus) is one of the major factors in this respect. Using 40 microsatellite loci with SSR-GBS techniques, we amplified a total of 664 individuals to investigate the genetic structure of O. niloticus from East Africa in comparison to Ethiopian and Burkina Faso populations.Results: All three African regions were characterized by independent gene-pools, however, the Ethiopian population from Lake Tana was genetically more divergent (F st = 2.1) than expected suggesting that it might be a different sub-species. In East Africa, the genetic structure was congruent with both geographical location and anthropogenic activities (Isolation By Distance for East Africa, R 2 = 0.67 and Uganda, R 2 = 0.24). O. niloticus from Lake Turkana (Kenya) was isolated, while in Uganda, despite populations being rather similar to each other, two main natural catchments were able to be defined. We show that these two groups contributed to the gene-pool of different non-native populations. Moreover, admixture and possible hybridization with other tilapiine species may have contributed to the genetic divergence found in some populations such as Lake Victoria. We detected other factors that might be affecting Nile tilapia genetic variation. For example, most of the populations have gone through a reduction in genetic diversity, which can be a consequence of bottleneck (G-W, < 0.5) caused by overfishing, genetic erosion due to fragmentation or founder effect resulting from stocking activities. Conclusions: The anthropogenic activities particularly in the East African O. niloticus translocations, promoted artificial admixture among Nile Tilapia populations. Translocations may also have triggered hybridization with the native congenerics, which needs to be further studied. These events may contribute to outbreeding depression and hence compromising the sustainability of the species in the region.