The Australian freshwater crocodile (Crocodylus johnstoni) is endemic to the northern mainland tropics of Australia and is widespread across the Kimberley region in the northwest Australia. Currently, there is limited understanding of the genetic structure and diversity of these populations, which impacts on our ability to evaluate the conservation status of the species. Population genetic analyses of 173 freshwater crocodiles from the Ord River, Fitzroy River, and Lennard River basins were conducted using single-nucleotide polymorphisms (SNPs). After filtering, 1185 SNPs were retained for downstream population genetic analysis. STRUCTURE and principal component analyses identified three clusters consistent with the three river basins. Population differentiation gave an FST of 0.15 between western and eastern Kimberley and the pairwise FST range was 0.06–0.18 among the three river basins. Assignment tests identified three migration events between the adjacent Fitzroy River and Lennard River basins, which may be explained by possible overland movement across these river basins. The population structure found here indicates that delimitation of management units should be based on river basins with the proximity of adjacent river basins taken into consideration when gene flow exists. Estimates of effective population size showing a low ratio of effective population size to census size in Lake Argyle may raise the concern of future monitoring in this area. Further population genetic studies across the species’ full range are required to better understand the extent of river basins acting as discrete population units, gene flow, population dynamics, and demographic history.