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The population genetics of aquatic animals in the Florida Everglades may be strongly influ-enced by extinction and colonization dynamics. We combined analyses of allozyme and microsatellite loci to test the hypothesis that two levels of population structure are present for spotted sunfish (Pisces: Centrarchidae: Lepomis punctatus) inhabiting the Everglades. We hypothesized that annual cycles of marsh dry-down increase local-scale genetic vari-ation through a process of local extinction and colonization; we hypothesized that barriers to gene flow by levee/canal systems create a second, regional level of genetic variation. In 1996 and 1997, we sampled spotted sunfish from 11 Everglades sites that were distributed in three regions separated by levees. We documented patterns of genetic variation at 7 poly-morphic allozyme loci and 5 polymorphic microsatellite loci. Most genetic variation was present among local populations, according to both types of genetic markers. Furthermore, samples from marsh sites were heterogeneous, while those from canals were not. These data supported our hypothesis that dry-down events and local population dynamics in th