Despite having winged queens, female dispersal in the monogynous ant Cataglyphis cursor is likely to be restricted because colonies reproduce by fission. We investigated the pattern of population genetic structure of this species using eight microsatellite markers and a mitochondrial DNA (mtDNA) sequence, in order to examine the extent of female and nuclear gene flow in two types of habitat. Sampling was carried out at a large spatial scale (16 sites from 2.5 to 120 km apart) as well as at a fine spatial scale (two 4.5-km transects, one in each habitat type). The strong spatial clustering of mtDNA observed at the fine spatial scale strongly supported a restricted effective female dispersal. In agreement, patterns of the mtDNA haplotypes observed at large and fine spatial scales suggested that new sites are colonized by nearby sites. Isolation by distance and significant nuclear genetic structure have been detected at all the spatial scales investigated. The level of local genetic differentiation for mitochondrial marker was 15 times higher than for the nuclear markers, suggesting differences in dispersal pattern between the two sexes. However, male gene flow was not sufficient to prevent significant nuclear genetic differentiation even at short distances (500 m). Isolation-by-distance patterns differed between the two habitat types, with a linear decrease of genetic similarities with distance observed only in the more continuous of the two habitats. Finally, despite these low dispersal capacities and the potential use of parthenogenesis to produce new queens, no signs of reduction of nuclear genetic diversity was detected in C. cursor populations.