Characterization of rifampicin-resistant Mycobacterium tuberculosis in Khyber Pakhtunkhwa, Pakistan

Anwar Sheed Khan,Sajjad Ahmad,Otavio Cabral-Marques,Hazir Rahman,Taj Ali Khan,Sajid Ali,Shulin Zhang,Muhammad Qasim,Taane G Clark,Dong-Qing Wei,Jody E Phelan,Muhammad Tahir Khan,Gary Napier,Susana Campino

doi:10.1038/s41598-021-93501-4

Abstract

Tuberculosis (TB), caused by Mycobacterium tuberculosis, is endemic in Pakistan. Resistance to both firstline rifampicin and isoniazid drugs (multidrug-resistant TB; MDR-TB) is hampering disease control. Rifampicin resistance is attributed to rpoB gene mutations, but rpoA and rpoC loci may also be involved. To characterise underlying rifampicin resistance mutations in the TB endemic province of Khyber Pakhtunkhwa, we sequenced 51 M. tuberculosis isolates collected between 2016 and 2019; predominantly, MDR-TB (n = 44; 86.3%) and lineage 3 (n = 30, 58.8%) strains. We found that known mutations in rpoB (e.g. S405L), katG (e.g. S315T), or inhA promoter loci explain the MDR-TB. There were 24 unique mutations in rpoA, rpoB, and rpoC genes, including four previously unreported. Five instances of within-host resistance diversity were observed, where two were a mixture of MDR-TB strains containing mutations in rpoB, katG, and the inhA promoter region, as well as compensatory mutations in rpoC. Heteroresistance was observed in two isolates with a single lineage. Such complexity may reflect the high transmission nature of the Khyber Pakhtunkhwa setting. Our study reinforces the need to apply sequencing approaches to capture the full-extent of MDR-TB genetic diversity, to understand transmission, and to inform TB control activities in the highly endemic setting of Pakistan.

Highlights

Tuberculosis (TB), caused by Mycobacterium tuberculosis (MTB), affects ~ 10 million people annually[1]
A TB control program was launched at Hayatabad Medical Complex Peshawar to monitor disease incidence and perform drug susceptibility testing (DST) to assess levels of RR-TB and MDR-TB
Of the fifty-one M. tuberculosis isolates, the majority were from lineage 3 (30, 58.8%), and were sourced from female patients (n = 29, 56.9%) and previously treated cases (n = 30, 58.8%) (Table 1)

Summary

Introduction

Tuberculosis (TB), caused by Mycobacterium tuberculosis (MTB), affects ~ 10 million people annually[1]. Khyber Pakhtunkhwa is the third largest province of Pakistan, with an area of 74,521 k m2 and a population size of approximately 30.5 million individuals. It has a history of armed conflicts, and TB control has been a neglected area of public health. Secondary mutations in the rpoA or rpoC genes can mitigate the initial fitness cost caused by a rpoB mutation[6,7]. By sequencing fifty-one M. tuberculosis, we aimed to summarise the diversity and potential role of rpoB, rpoA, and rpoC mutations in the high MDR incidence setting of Khyber Pakhtunkhwa, to inform diagnostics tools for TB control and implementation. The analysis revealed some mixed MDR infections and heteroresistance, reflecting the complex dynamics of the high transmission setting in Pakistan, and reinforcing the need for robust diagnostic tools

Objectives

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Results

Conclusion