Genetic diversity in over-exploited and threatened species has important, but often neglected, implications for conservation. In South Africa, the bark of the regionally endangered pepper-bark tree, Warburgia salutaris, is extensively harvested for subsistence medicine. We developed 11 polymorphic microsatellite markers to examine the genetic diversity and structure among 227 W. salutaris individuals sampled from across much of its range in South Africa’s Kruger National Park (KNP) and 31 trees growing in local nurseries. We also tested the cross-amplification success of these markers in two closely related species, Warburgia elongata (IUCN listed as Endangered) and W. stuhlmannii (IUCN listed as Vulnerable). Contrary to our expectations, we found evidence of clonality and for high genetic diversity in all wild KNP sub-populations of W. salutaris. We identified between 114 and 149 unique multi-locus genotypes in the 258 individuals included in our analyses. Bayesian assignment tests identified three distinct genetic clusters that are represented by the sampled KNP sub-population of W. salutaris, with minimal geographic structure and some admixture. Three genetic clusters were also identified among all sampled individuals, i.e., KNP and nursery trees. Nursery W. salutaris germplasm comprised unique genotypes with similar levels of observed heterozygosity to the wild population. High levels of clonality and reported self-incompatibility, along with short distance pollination, likely explain the lack of recent sexual recruitment (i.e., reduced fruit set and lack of seedlings) in the wild population. These findings will be useful to identify sub-populations where genets may be sampled to maximize genetic diversity within subpopulations for restoration throughout KNP.