Abstract The giant mud crab, Scylla serrata (Forskål, 1775), is a targeted fishery along its geographic distribution, both for domestic and international markets. Population genetic data, evaluated with genetic markers such as microsatellites, can be used to identify management units (stocks) based on genetic dissimilarities between locations, informing localized management decisions for targeted fisheries. Additionally, microsatellite loci can be used to identify released individuals from stock enhancement programs, while simultaneously monitoring genetic impacts of hatchery-releases on wild populations. We isolated and characterized 16 novel microsatellite markers; fourteen of the markers were further evaluated for population structure among three localities in the US-Affiliated Pacific Islands: Palau, Pohnpei, and Kosrae. The significant genetic differentiation observed between Palau and Kosrae/Pohnpei suggests the newly described markers would be capable of elucidating structure across the broader scope of the geographic distribution of the species. Given the international trade for live mud crab, the evaluation of structure would highlight potential issues regarding the transport of live animals between locations with genetically differentiated stocks. Additionally, a multilocus exclusion probability of 0.999, with as few as three markers, suggests the novel microsatellite markers would be capable of identifying hatchery-sourced individuals as part of a stock enhancement program.