Kinship among group members has long been recognized as a main factor promoting the evolution of sociality and reproductive altruism, yet some ants have an extraordinary social organization, called unicoloniality, whereby individuals mix freely among physically separated nests. This type of social organization is not only a key attribute responsible for the ecological dominance of these ants, but also an evolutionary paradox because relatedness between nestmates is effectively zero. Recently, it has been proposed that, in the Argentine ant, unicoloniality is a derived trait that evolved after its introduction into new habitats. Here we test this basic assumption by conducting a detailed genetic analysis of four native and six introduced populations with five to 15 microsatellite loci and one mitochondrial gene. In contrast to the assumption that native populations consist of family-based colonies with related individuals who are aggressive toward members of other colonies, we found that native populations also form supercolonies, and are effectively unicolonial. Moreover, just as in introduced populations, the relatedness between nestmates is not distinguishable from zero in these native range supercolonies. Genetic differentiation between native supercolonies was very high for both nuclear and mitochondrial markers, indicating extremely limited gene flow between supercolonies. The only important difference between the native and introduced populations was that supercolonies were several orders of magnitude smaller in the native range (25–500 m). This size difference has important consequences for our understanding of the evolution and stability of unicolonial structures because the relatively small size of supercolonies in the native range implies that competition can occur between supercolonies, which can act as a break on the spread of selfish mutants by eliminating supercolonies harboring them.