Feral rabbit populations in Australia have generally been managed using localized control procedures. While these procedures may result in local extinctions, persistence of populations will depend on the probability of recolonization. Genetic markers developed using temperature gradient gel electrophoresis (TGGE) combined with heteroduplex analysis (HA) of mitochondrial DNA (mtDNA) were used to characterize the degree of subdivision and extent of gene flow within and among rabbit populations distributed over large distances (up to 1000 km) in southern Queensland (QLD) and north-west New South Wales (NSW), Australia. TGGE analyses revealed significant heterogeneity in mtDNA control region haplotype frequencies. From heterogeneity χ2 tests, it was evident that the differentiation observed was largely attributable to five sites which were located in the semiarid eastern region, whereas haplotype frequencies were homogeneous throughout the arid western region. These results suggest that there are independent population systems within the study area. The extent of gene flow among local populations within each system is related to the spatial configuration of acceptable habitat patches and the persistence of the populations is determined by the probability of recolonization following local extinction. These data suggest that to provide better overall control of rabbit populations, different management strategies may be necessary in arid and semiarid ecosystems. In arid south-west QLD and north-west NSW, where extensive gene flow occurs over large distances, rabbit populations should be managed at a regional level. In semiarid eastern QLD, where gene flow is restricted and populations are more isolated, localized control procedures may provide effective short-term relief. These results indicate that in nonequilibrium systems with patchy distribution of individuals, the interpretation of migration rate from estimates of gene flow obtained using existing genetic models must include an understanding of the spatial and temporal scales over which population processes operate.