Human activities have a major impact on habitat connectivity and landscape structure. In this context, it is critical to better understand animal movements and gene flow to develop appropriate conservation and land management measures. It is also important to better understand difference between sexes in space use and spatial scale of dispersal. We studied the common adder (Vipera berus), an elusive snake species with low mobility that is facing a substantial decline in Europe. A systematic sampling was carried out to clarify the dispersal pattern at a fine spatial scale (10 × 7 km 2 ) in a rural landscape with both semi-natural (preserved heathlands, hedgerow networks) and degraded (crops, roads) habitats. Based on 280 captured adults and using 11 microsatellite markers, we detected no marked genetic differentiation, however, we detected relatively strong isolation-by-distance (IBD). Under IBD, we quantified a low neighborhood size (N s ≈ 50) associated with limited natal dispersal (σ ≤ 1 km). We detected sex-biased dispersal in favor of males, but the pattern was dependent on the spatial scale considered. Our results also suggest that there is higher genetic diversity in the preserved habitat, notably among males. Overall, our study underlines the importance of sex variation in dispersal, and the spatial scale of landscape effects. This contrast between sexes should be considered to improve functional connectivity at fine spatial scales for reptile conservation.