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Background and aims  –    Brachystegia    is a species-rich tree genus found in tropical Africa and a typical element of Miombo woodlands, a widely distributed subtype of the Zambezian savanna. Plastid DNA was shown to be largely uninformative to assess species phylogenetic relationships due to widespread chloroplast capture among species. Here, we aim to assess the capacity of nuclear ribosomal DNA ( rDNA ) to clarify the phylogeny of    Brachystegia    species while accounting for intra-individual site polymorphisms ( 2ISPs ), which are often present in  rDNA  and potentially phylogenetically informative.Material and methods  – Genome skimming sequencing on 47 samples representing 27 of the 29 currently recognized    Brachystegia    species, allowed us to retrieve complete nuclear ribosomal cistrons encoding for 18S, 5.8S, and 25S rRNA genes (35S  rDNA ). We reconstructed the    Brachystegia    phylogeny using Maximum Likelihood methods based on the standard substitution model or integrating  2ISPs  (GENOTYPE implementation in RAxML-NG). We additionally tested the effect of partitioning the data (one partition for  rDNA  genes and one for the  ITS1 + ITS2 ). We also conducted network inferences (Neighbor-Net splits graph), as a strict bifurcative approach might not properly model topological uncertainty at shallow phylogenetic depth.Key results  –  2ISPs -aware and standard phylogenetic reconstructions are largely congruent. We identified several well-supported main clades clarifying the species relationships, including two clades of Miombo woodlands species. Miombo Group A includes species with ovoid to globose axillary dormant buds, while Miombo Group B species have flattened ones. Two morphologically close    Brachystegia    species (   B.   kennedyi    and    B.   leonensis   ) found in Guineo-Congolian rain forests form also a robustly supported clade.  2ISPs  coding allowed to identify an additional Guineo-Congolian clade (   B.   eurycoma    and    B.   nigerica   ). Ribosomal DNA therefore proves more useful to explore the generic phylogeny than plastid DNA but the species relationships within and among the main clades remain poorly resolved, probably due to recent diversification and/or recurrent hybridization, so that the diversification of    Brachystegia    remains to be more properly characterised.Conclusion  – Nuclear and plastid phylogenetic reconstructions of    Brachystegia    species are discordant. Even if not well-resolved,  rDNA  phylograms and networks are characterized by taxonomic sorting, while we observe a strictly geographic sorting in the plastid dataset. Most of the species’ relationships remain to be characterized using additional nuclear markers combined with in-depth morphological investigations.

Nuclear ribosomal phylogeny of Brachystegia : new markers for new insights about rain forests and Miombo woodlands evolution