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. 2025 Feb 11:16:1535160.
doi: 10.3389/fmicb.2025.1535160. eCollection 2025.

Whole genomic analysis uncovers high genetic diversity of rifampicin-resistant Mycobacterium tuberculosis strains in Botswana

Tuelo Mogashoa  1   2 , Johannes Loubser  2 , Ontlametse T Choga  1   3 , Justice Tresor Ngom  2 , Wonderful T Choga  1   3 , Mpaphi B Mbulawa  4 , Tuduetso Molefi  5 , One Stephen  4 , Topo Makhondo  5 , Kedumetse Seru  1 , Patience Motshosi  1 , Boitumelo Zuze  1 , Joseph Makhema  1 , Rosemary M Musonda  1 , Dimpho Otukile  6 , Chawangwa Modongo  6 , Botshelo T Kgwaadira  7 , Keabetswe Fane  7 , Simani Gaseitsiwe  1 , Rob M Warren  2 , Sikhulile Moyo #  1   8   9   10 , Anzaan Dippenaar #  11 , Elizabeth M Streicher #  2
Affiliations

Whole genomic analysis uncovers high genetic diversity of rifampicin-resistant Mycobacterium tuberculosis strains in Botswana

Tuelo Mogashoa et al. Front Microbiol. .

Abstract

Background: The emergence of drug-resistant Mycobacterium tuberculosis (M. tb) strains remains a threat to tuberculosis (TB) prevention and care. Understanding the drug resistance profiles of circulating strains is crucial for effective TB control. This study aimed to describe the genetic diversity of rifampicin-resistant M. tb strains circulating in Botswana using whole genome sequencing (WGS).

Methods: This study included 202 stored M. tb isolates from people diagnosed with rifampicin-resistant TB (RR-TB) between January 2016 and June 2023. Genomic DNA was extracted using the cetyltrimethylammonium bromide (CTAB) method. Library preparation was performed using the Illumina DNA prep kit following the manufacturer's instructions. Sequencing was done on Illumina NextSeq2000. TBProfiler software was used to identify known M. tb lineages and drug resistance profiles. Statistical analyses were performed on STATA version 18.

Results: WGS analysis revealed multidrug resistance (57.9%: 95% CI; 50.7-64.8), Pre-XDR (16.8%, 95% CI: 11.9-22.7), RR-TB (20.2%: 95% CI: 14.98-26.5), and HR-TB (0.5%, 95% CI; 0.01-2.7). We identified a high genetic diversity with three predominant lineages: lineage 4 (60.9%, 95% CI; 53.8-67.7), lineage 1 (22.8%: 95% CI; 17.2-29.2), and lineage 2 (13.9%, 95% CI: 9.4-19.4). The most frequently observed drug resistance mutations for rifampicin, isoniazid, ethambutol, streptomycin, pyrazinamide, and fluoroquinolones were rpoB S450L (28.6%), katG S315T (60.5%), embA_c.-29_-28delCT, embB Q497R (31.7%), rrs_n.517C>T (47.1%), pncA_c.375_389delCGATGAGGTCGATGT (36.0%) and gyrA A90V (79.4%), respectively. No bedaquiline and delamanid resistance-associated mutations were detected.

Conclusions: This study highlights the high genetic diversity of M. tb strains, with a predominance of lineage 4 among people with RR-TB in Botswana. It provides valuable insights into the genetic diversity of rifampicin-resistant M. tb strains circulating in Botswana.

Keywords: Botswana; genetic diversity; rifampicin resistance; tuberculosis; whole genome sequencing.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Maximum-likelihood phylogeny of 202 rifampicin-resistant Mycobacterium tuberculosis isolates. The color-coded annotations include year of collection, health district, main lineage and drug resistance profiles. The black and white circles indicate the genotypic resistance profiles to 18 antibiotics (black and white represent resistance and susceptibility to the specific antibiotics, respectively).
See this image and copyright information in PMC

References

    1. Barilar I., Battaglia S., Borroni E., Brandao A. P., Brankin A., Cabibbe A. M., et al. . (2024). Quantitative measurement of antibiotic resistance in Mycobacterium tuberculosis reveals genetic determinants of resistance and susceptibility in a target gene approach. Nat. Commun. 15:488. 10.1038/s41467-023-44325-5 - DOI - PMC - PubMed
    1. Carey A. F., Rock J. M., Krieger I. V., Chase M. R., Fernandez-Suarez M., Gagneux S., et al. . (2018). TnSeq of Mycobacterium tuberculosis clinical isolates reveals strain-specific antibiotic liabilities. PLoS Pathog. 14:e1006939. 10.1371/journal.ppat.1006939 - DOI - PMC - PubMed
    1. Centner C. M., Munir R., Tagliani E., Rieß F., Brown P., Hayes C., et al. . (2024). Reflex Xpert MTB/XDR testing of residual rifampicin-resistant specimens: a clinical laboratory-based diagnostic accuracy and feasibility study in South Africa. Open Forum Infect. Dis. 11:ofae437. 10.1093/ofid/ofae437 - DOI - PMC - PubMed
    1. Chihota V. N., Niehaus A., Streicher E. M., Wang X., Sampson S. L., Mason P., et al. . (2018). Geospatial distribution of Mycobacterium tuberculosis genotypes in Africa. PLoS ONE 13:e0200632. 10.1371/journal.pone.0200632 - DOI - PMC - PubMed
    1. Chisompola N. K., Streicher E. M., Dippenaar A., Whitfield M. G., Tembo M., Mwanza S., et al. . (2021). Drug resistant tuberculosis cases from the Copperbelt province and Northern regions of Zambia: genetic diversity, demographic and clinical characteristics. Tuberculosis 130:102122. 10.1016/j.tube.2021.102122 - DOI - PubMed

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