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The native North American fire ants (SolenopsisWestwood) comprise a difficult group taxonomically that has undergone multiple revisions in the past century yet remains in a state of taxonomic uncertainty. In the present study, we utilised a set of 59 microsatellite markers analysed in 238 specimens to conduct the first robust genetic analysis of the four nominal species. Our approach used a variety of methods to test operational criteria commonly employed in species delimitation, including genotypic clustering, reproductive isolation/cohesion and monophyly. We conclude from our results that the recognised North American fire ant species represent evolutionarily independent entities and, moreover, we confirm the presumed sister status of the desert fire ants, S. aureaWheeler and S. amblychilaWheeler. However, the presence of at least two genetically divergent populations within the nominal species boundaries, including a western form of S. xyloni and a distinct population of S. aurea endemic to the Salton Trough, suggests that the current taxonomy does not fully capture the species-level diversity in this group. Our study provides the molecular foundation for future integrated studies of the taxonomy and evolution of this scientifically and economically important group of insects.