Magister Scientiae (Biodiversity and Conservation Biology) / The last taxonomic treatment of the subtribe Anthosperminae Benth. (Rubiaceae, Rubioideae, Anthospermeae) was in 1986 by Puff., nevertheless, few attempts have been made to resolve the phylogeny and the inter- and infrageneric relationships within the subtribe. The genera Anthospermum L. (39 species) and Nenax Gaertn. (11 species) are considered the most difficult groups to distinguish. Anthospermum species are widely distributed in Sub-Saharan Africa and Madagascar with the highest concentration of taxa in southern Africa, while Nenax species are restricted to southern Africa, in the south-western Cape Floristic Region. The two genera share common morphological and anatomical characters such as the growth form, presence of hairs on the stem, leaf arrangement, presence of petioles, flowers formation, dehiscence and presence of carpophore in fruits. currently combination of characters, woody shrub, needle-like leaves, few-flowered inflorescence and dioecy are considered unique in Nenax. The most recent phylogenetic analysis based on molecular data indicated insights into generic relationships within the two genera and the subtribe Anthosperminae. The present study focussed on expanding the phylogenetic analysis of Anthospermum, Nenax and other genera within the subtribe, as well as assessing the value of selected morphological and anatomical characters for re-assesing generic circumscriptions. Phylogenetic relationships were analysed using Maximum Parsimony, Maximum Likelihood and Bayesian inference, and a Maximum Clade Credibility tree was produced. These analyses were based on both nuclear (ITS, ETS) and plastid (trnL-f, rps16, rpl32) datasets.