Spiders are a prominent group of predatory arthropods in most terrestrial ecosystems, and are especially diverse in the Savanna Biome of South Africa. Despite this, there is a considerable paucity of information on spider assemblages in particular microhabitats, including tree bark. The spiders associated with the bark of Vachellia xanthophloea (fever trees) were studied at five wetland sites in the Ndumo Game Reserve, South Africa. The influence of various seasonal, habitat and phenological effects on spider assemblages was investigated. In total, 8341 spiders were collected, of which only 2726 were adults (32.7 %), representing 25 families and 108 species. While total spider abundance and adult species richness did not differ significantly between sites, significant differences were found in adult abundance and in overall assemblages between sites. There were also no significant seasonal effects on spider assemblages and guild composition. Only eight of the species collected can be considered exclusive bark-dwelling spiders, while similar proportions are facultative bark-dwellers (43.5 %) or accidental bark-dwellers (49.1 %). This study demonstrates that targeted sampling of specialised microhabitats can yield a significant proportion of the spider diversity in a particular locality, and can contribute to understanding patterns of biodiversity in conserved areas.