Nestmate recognition is of prime importance in maintaining ant colony integration and organization. Monogyne red imported fire ant, Solenopsis invicta Buren, colonies are highly territorial and aggressive toward non-nestmate conspecific workers. In contrast, workers from polygyne nests in the United States show no aggression toward workers from other conspecific colonies (polygyne or monogyne). Nests within a polygyne population form a ‘supercolony,’ with free exchange of workers and food between nests. The difference in conspecific nestmate recognition is a major distinguishing feature of the two S. invicta forms in the United States. We report here the discovery of an exception to this dichotomy. High levels of worker aggression are released by the introduction of newly mated queens into both polygyne and monogyne colonies. This suggests that nestmate recognition involving female sexuals does not follow the same mechanism used to explain nestmate recognition behavior between workers.