<p>Habitat fragmentation in urban environments has the potential to increase deleterious effects of isolation and genetic drift for species occupying urban parks and green spaces. Ant species vary in urban tolerance, but the impacts of urbanization on ant population genetics are unclear. In this study, we evaluated how urbanization in the Philadelphia, USA metropolitan area has affected population genetics of six ant species: three species in the <i>Aphaenogaster rudis</i> group, all urban avoider woodland ants; <i>Camponotus pennsylvanicus</i>, an urban utilizer species which can inhabit edge habitat; and <i>Nylanderia flavipes</i>, an invasive urban dweller species. Individuals were collected from nine urban parks and at five points in nearby contiguous forest and genotyped at four polymorphic microsatellite loci. Both <i>C. pennsylvanicus</i> and <i>N. flavipes</i> occurred in urban and forest habitats, but of the three <i>Aphaenogaster</i> species, two occupied primarily urban parks while the third was restricted to forests. Allelic richness did not differ for any species between urbanized and forested regions. <i>Aphaenogaster</i> in urban parks showed very weak urbanization-induced genetic isolation, which may reflect high population densities and/or effective dispersal mechanisms. In contrast, <i>C. pennsylvanicus</i> population genetic structure significantly correlated with the urban landscape, while the invasive <i>N. flavipes</i> displayed significantly increased genetic differentiation in forested habitats. Altogether, these results show that genetic response to urbanization in ants varies depending on species and context, with moderate predictability based on aptitude for the urban environment.</p>