For species that are habitat specialists or sedentary, population fragmentation may lead to genetic divergence between populations and reduced genetic diversity within populations, with frequent inbreeding. Hundreds of kilometres separate three geographical regions in which small populations of the endangered Eastern Bristlebird, Dasyornis brachypterus, a small, grounddwelling passerine that occurs in fire-prone bushland in eastern Australia, are currently found. Here, we use mitochondrial and microsatellite DNA markers to: (i) assess the sub-specific taxonomy designated to northern range-edge, and central and southern range-edge D. brachypterus, respectively, and (ii) assess levels of standing genetic variation and the degree of genetic subdivision of remnant populations. The phylogenetic relationship among mtDNA haplotypes and their spatial distribution did not support the recognised subspecies boundaries. Populations in different regions were highly genetically differentiated, but in addition, the two largest, neighboring populations (located within the central region and separated by *50 km) were moderately differentiated, and thus are likely closed to migration (microsatellites, F ST = 0.06; mtDNA, F ST = 0.12, H ST = 0.08). Birds within these two populations were genotypically diverse and apparently randomly mating. A long-term plan for the conservation of D. brachypterus's genetic diversity should consider individual populations as separate management units. Moreover, managers should avoid actively mixing birds from different populations or regions, to conserve the genetic integrity of local populations and avoid outbreeding depression, should further translocations be used as a recovery tool for this species.