Genetic variation is expected to be influenced by the interaction between reproductive mode and dispersal traits on the one hand, and environmental and habitat setting affecting establishment success on the other. We evaluated how environmental/habitat setting affect population genetic variation (i.e. variation in genetic diversity and structure) when regulated by contrasting dispersal traits. We used fungus-specific microsatellite markers to examine genetic diversity and structure of two closely related epiphytic lichen fungi that differ in their primary reproductive mode: Nephroma laevigatum (sexually reproducing, N = 191, ten microsatellites) and N. parile (asexually, N = 182, twelve microsatellites), along a steep climatic gradient in Scotland. Despite their reproductive differences, we found a high proportion of clones in both species and a background pattern of genetic structure related to climatic gradients. We also demonstrated that woodland connectivity, rather than geographic distance, explained genetic diversity in both species. Environmental/habitat setting, modulated by the reproductive mode of the species, affects genetic diversity and structure, but the putative dissimilarity in their reproductive mode is less important than has been previously assumed. We reinforce the importance of protecting highly connected populations, positioned along a gradient capturing the segregation of gene pool differences in response to climatic variation.