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1. Populations of a number of sub-tropical stream insect species have been found to show unexpected patterns of genetic variation, with more differences between samples from the same stream than between whole streams or between subcatchments. Many samples also showed deviations from Hardy–Weinberg proportions. It has been proposed that these patterns result from limited numbers of matings contributing to a given stream reach, because adults emerge throughout the year, and low levels of larval drift between reaches. These patterns may be less likely in a northern hemisphere montane species with synchronous emergence of adults and high levels of drift. We tested the hypothesis that patterns of genetic variation in a montane mayfly from the Rocky Mountains, Colorado, would reflect a pattern of ‘isolation by distance’ with samples from the same creek being more similar than samples from different creeks and that deviations from Hardy–Weinberg proportions would be minimal. 2. Based on allozyme variation, the hypothesis of minimal deviations from Hardy–Weinberg proportions was not supported and there was no evidence of isolation by distance. Nevertheless the levels of differentiation among samples from within the same stream were less than those reported for most subtropical species. 3. Results from analysis of a fragment of the cytochrome oxidase gene (subunit 1) revealed contrasting patterns. The levels of genetic differentiation were an order of magnitude higher between streams than among samples within streams. In addition, although there was no significant isolation by distance effect overall, a nested clade analysis provided evidence for restricted gene flow with isolation by distance for some clades. 4. We suggest that these contrasting results may reflect the differences in male and female dispersal patterns. While differentiation at nuclear gene markers (allozymes) give information about both male and female dispersal, mitochondrial DNA markers reflect only female dispersal. We suggest that in this species, female dispersal may be more restricted, perhaps mostly along stream channels, whereas male dispersal is more widespread. An alternative explanation for the different results is the different evolutionary rates of the mitochondrial and nuclear markers. [ABSTRACT FROM AUTHOR]