Knowledge of genetic diversity and population structuring represents a key component for the conservation of endangered species, especially where translocations and re-introduction operations are integral tools for population management. The blue duck (Hymenolaimus malacorhynchos) is a threatened riverine specialist that is endemic to New Zealand. Populations from the North and South Island form two distinct mitochondrial lineages, which currently necessitate separate conservation management. Here we examine the patterns of variability at 11 microsatellite loci and mitochondrial control region data to assess the range-wide genetic diversity and population structure of blue duck. Our data suggest that North and South Island blue duck populations likely diverged in the late Pleistocene with very limited gene flow, strongly reinforcing the current management strategy to avoid translocation between islands. Genetic diversity within both islands follows a pattern of isolation by distance with relatively high levels of gene flow among populations, likely driven by male-juvenile dispersal. The overall genetic diversity in blue duck is low and effective population size is small. These data will provide important information for conservation management of this species.