We report the genetic structure, relatedness and mating structure of a population of the endangered giant anteater Myrmecophaga tridactyla Linnaeus, 1758 in the Emas National Park, Brazil, based on variability at five microsatellite loci. Additionally, we addressed the hypothesis that the M. tridactyla population studied has low levels of polymorphism and high levels of inbreeding and relatedness and that animals with overlapping home range are highly related. All five microsatellite loci displayed low levels of polymorphism and of expected and observed heterozygosity. The low level of polymorphism and high inbreeding showed by the population studied may be the outcome of high mortality and reduction in population size due to recurrent fire events in the Emas National Park, as reported in 1994. The reduction in population size may have led to a higher frequency of mating between closely related animals, augmented by the isolation of the population in the park because of the expansion of agricultural land and fragmentation of the Cerrado environment. The natural history of M. tridactyla and the phylopatric (sex-biased dispersal) behavior of females should increase the effects of isolation and bottlenecking, decreasing gene flow and increasing inbreeding. However, the low levels of polymorphism found in this population may simply be due to the natural history and evolution of M. tridactyla as reported for other species. The genetic structure and dynamics of this population needs to be investigated more profoundly in order to provide sound data for the design of conservation strategies for M. tridactyla in the Emas National Park.