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Data from 10 microsatellite DNA loci were used to describe the genetic structure of the two extant species (Cyprinodon macularius and C. eremus) of the endangered Desert Pupfish complex of southwestern United States and northwestern Mexico. Variation at microsatellite loci was significantly correlated (Mantel test) with that of previous mtDNA results, both for the complex and for the relatively wide-ranging C. macularius alone. Both species showed unusually high levels of microsatellite diversity for non-marine fish (H (e) = 0.84-0.93; A(R) = 11.9-17.0). There was evidence (R (ST) > F (ST)) that the two extant populations of C. eremus have been isolated sufficiently long for mutation to contribute significantly to genetic divergence, whereas divergence among the nine assayed populations of C. macularius could be attributed to genetic drift alone. Correspondingly, 10% of the diversity in C. eremus was attributable to differences between the two populations, whereas, for C. macularius, only 2.7% was attributable to among-population variation. Within C. macularius, a small (0.8%), but statistically significant, portion was attributable to differences between populations in the Salton Sea area and those on the lower Colorado River delta. The two populations of C. eremus and five groups of populations of C. macularius are recommended as management units for conservation genetics management of the two species.