Emergence of bedaquiline-resistant tuberculosis and of multidrug-resistant and extensively drug-resistant Mycobacterium tuberculosis strains with rpoB Ile491Phe mutation not detected by Xpert MTB/RIF in Mozambique: a retrospective observational study

Background In 2021, an estimated 4800 people developed rifampicin-resistant tuberculosis in Mozambique, 75% of which went undiagnosed. Detailed molecular data on rifampicin-resistant and multidrug-resistant (MDR) tuberculosis are not available. Here, we aimed at gaining precise data on the determinants of rifampicin-resistant and MDR tuberculosis in Mozambique. Methods In this retrospective observational study, we performed whole-genome sequencing of 704 rifampicin-resistant Mycobacterium tuberculosis complex (Mtbc) strains submitted to the National Tuberculosis Reference Laboratory (NTRL) in Maputo, Mozambique, between 2015 and 2021. Phylogenetic strain classification, genomic resistance prediction, and cluster analysis were performed. Findings Between Jan 1, 2015, and July 31, 2021, 2606 Mtbc isolates with an isoniazid or rifampicin resistance were identified in the NTRL biobank, of which, 1483 (56·9%) were from men, 1114 (42·7%) from women, and nine (0·4%) were unknown. Genome-based drug-resistant prediction classified 704 Mtbc strains as rifampicin resistant. 628 (89%) of the 704 Mtbc strains were classified MDR; of those, 146 (23%) were pre-extensively drug resistant (pre-XDR; additional fluoroquinolone resistance), and 24 (4%) extensively drug resistant (XDR; combined fluoroquinolone and bedaquiline resistance). Overall, 61 (9%) of 704 strains revealed resistance to bedaquiline: five (7%) of 76 rifampicin resistant plus bedaquiline resistant, 32 (7%) of 458 MDR plus bedaquiline resistant, and 24 (100%) of 24 XDR. Prevalence of bedaquiline resistance increased from 3% in 2016 to 14% in 2021. The cluster rate (12 single-nucleotide polymorphism threshold) was 42% for rifampicin-resistant strains, 78% for MDR strains, 94% for pre-XDR strains, and 96% for XDR Mtbc strains. 31 (4%) of 704 Mtbc strains, belonging to a diagnostic escape outbreak strain previously described in Eswatini (group_56), had an rpoB Ile491Phe mutation which is not detected by Xpert MTB/RIF (no other rpoB mutation). Of these, 23 (74%) showed additional resistance to bedaquiline, 13 (42%) had bedaquiline and fluoroquinolone resistance, and two (6%) were bedaquiline, fluoroquinolone, and delamanid resistant. Interpretation Pre-XDR resistance is highly prevalent among MDR Mtbc strains in Mozambique and so is bedaquiline resistance; and the frequency of bedaquiline resistance quadrupled over time and was found even in Mtbc strains without fluoroquinolone resistance. Importantly, strains with Ile491Phe mutation were frequent, accounting for 31% (n=10) of MDR plus bedaquiline-resistant strains and 54% (n=13) of XDR Mtbc strains. Given the current diagnostic algorithms and treatment regimens, both the emergence of rifampicin resistance due to Ile491Phe and bedaquiline resistance might jeopardise MDR tuberculosis prevention and care unless sequencing-based technology is rolled out. The potential cross border spread of diagnostic escape strains needs further investigation. Funding The German Ministry of Health through the Seq_MDRTB-Net project, the Deutsche Forschungsgemeinschaft under Germany's Excellence Strategy Precision Medicine in Inflammation and the Research Training Group 2501 TransEvo, the Leibniz Science Campus Evolutionary Medicine of the Lung, and the German Ministry of Education and Research via the German Center for Infection Research.