Freshwater crayfish of the genus Euastacus are among Australia’s most threatened freshwater taxa, yet the distributions and population status of many species remain poorly understood. The Fitzroy Falls spiny crayfish (Euastacus dharawalus) is listed as critically endangered and was previously only known to occur in one creek located in the Southern Highlands of NSW, Australia. Here, we investigated the distribution, demographic characteristics and genetic structure of newly discovered E. dharawalus populations across the Southern Highlands. Surveys were conducted across the Nepean, Wingecarribee and Burke River catchments. Catch-per-unit-effort was used to compare relative abundance among sites, while size structure, sex ratio and body condition were assessed to characterise population demographics. Genome-wide single nucleotide polymorphisms (SNPs) were used to validate species identity and assess genetic structure among populations in the Nepean, Wingecarribee and Wildes Meadow Creek catchments. Euastacus dharawalus was detected throughout the Nepean and Wingecarribee catchments but was absent from the Burke catchment, where E. spinifer was present at all surveyed sites. Newly discovered populations exhibited broad size distributions and no pronounced demographic imbalance. SNP analyses confirmed the identity of the newly discovered populations as E. dharawalus and revealed substantial genetic differentiation among the three catchments. These findings substantially expand the known distribution of E. dharawalus and highlight the importance of protecting genetically distinct populations across its range. Identifying these previously unknown populations provides critical information for conservation, recovery planning and water management to support the long-term persistence of this critically endangered species.