Mycobacterium tuberculosis complex Lineage 8 causes drug resistant pulmonary tuberculosis also in South Kivu, Eastern Democratic Republic of Congo

Background Lineage 8 (L8) occupies a unique phylogenetic position as a sister clade to all other known Mycobacterium tuberculosis complex (MTBC) lineages. Although the initial identification of L8 strains was limited to Rwanda and Uganda, the complete geographical distribution of this lineage remains undetermined due to limited genomic surveillance across eastern and central Africa. Methods We performed whole genome sequencing (WGS) on MTBC isolates from rifampicin-resistant tuberculosis (TB) patients’ sputum in Bukavu, South Kivu province, eastern Democratic Republic of Congo (DRC). Lineage assignment was performed using TBProfiler with classification based on single nucleotide polymporphisms (SNPs). Phylogenetic reconstruction was conducted using maximum likelihood analysis against publicly available L8 reference genomes from Rwanda and Uganda. Drug-resistance profiles were determined through both genotypic prediction from WGS data and phenotypic drug-susceptibility testing. Clinical and epidemiologic data were extracted from programmatic records. Results We identified three clinical isolates belonging to MTBC L8 among fifteen sequenced isolates collected in Bukavu between January and December 2023 enriched for rifampicin resistant TB as determined by Xpert Ultra. Phylogenetic analysis confirmed placement within the L8 clade while revealing unique SNP profiles distinct from previously published Rwandan and Ugandan L8 strains by 16–20 to >200 SNPs, respectively, consistent with local diversification rather than recent cross-border importation. Ancestral genomic features characteristic of L8 were preserved, including the intact cobF region and complete pks8 gene. Drug-resistance prediction from genomic data indicated multidrug-resistant and pre-extensively-resistant TB. Conclusions This report extends the known geographic distribution of MTBC L8 to eastern DRC, supporting the hypothesis that the African Great Lakes region harbors under-characterized ancestral diversity of the tubercle bacillus. Systematic genomic surveillance across the DRC-Uganda-Rwanda-Burundi corridor, including rifampicin-susceptible isolates, is needed to determine whether L8 represents a rare relict lineage or an under-detected endemic variant circulating through regional transmission networks.