Abstract: Ilex integra is an insular endemic and endangered species that is distributed only in East Asia. Here we used inter-simple sequence repeat (ISSR) markers to investigate the genetic structure of six populations (five natural populations and one cultivated population) of the species from Zhoushan Archipelago, China. A total of 78 discernible loci were obtained from 57 individuals of all sampled populations using nine primers. Of these loci, 57.7% were polymorphic (PPL=57.7%). Both the average percentage of polymorphic loci (PPL=42.1%) and Nei’s genetic diversity (H_E=0.153), estimated by POPGENE, indicated that I. integra had lower levels of genetic diversity than those reported in many other insular plant species. Our results also showed that populations of I. integra were highly differentiated (G_st=0.316, F_st=0.295，S=0.304). Geographic isolation may be a significant factor to the genetic differentiation among these populations. In addition, a significant correlation was found between genetic distance and geographic distance (r=0.649, P<0.05). UPGMA analysis revealed that Putuo Island population had a close relationship with Zhujiajian Island populations, and the cultivated populations on Zhoushan Island might have originated from Taohua Island. Based on the above findings, we propose that conservation management of I. integra should include protection of natural habitats occupied by all five wild populations and preservation of germplasm in bo-tanical gardens from multiple seed sources.