Genetic factors associated with acquired phenotypic drug resistance and its compensatory evolution during tuberculosis treatment

Objectives We elucidated the factors, evolution, and compensation of antimicrobial resistance (AMR) in Mycobacterium tuberculosis (MTB) isolates under dual pressure from the intra-host environment and anti-tuberculosis (anti-TB) drugs. Methods This retrospective case–control study included 337 patients with pulmonary tuberculosis from 15 clinics in Tianjin, China, with phenotypic drug susceptibility testing (DST) results available for at least two time points between January 1, 2009 and December 31, 2016. Patients in the case group exhibited acquired AMR to isoniazid (INH) and/or rifampicin (RIF), while those in the control group lacked acquired AMR. Whole-genome sequencing (WGS) was conducted on 149 serial longitudinal MTB isolates from 46 patients who acquired or reversed phenotypic INH/RIF-resistance during treatment. The genetic basis, associated factors, and intra-host evolution of acquired phenotypic INH/RIF-resistance were elucidated using a combined analysis. Results Anti-TB interruption duration ≥30 d showed association with acquired phenotypic INH/RIF resistance (aOR=2·2, 95%CI: 1·0–5·1) and new rpoB mutations (P = 0·024). MTB evolution was 1·2 (95% CI: 1·02–1·38) single nucleotide polymorphisms per genome per year under dual pressure from the intra-host environment and anti-TB drugs. AMR-associated mutations occurred before phenotypic AMR appearance in cases with acquired phenotypic INH (10/16) and RIF (9/22) resistances. Conclusions Compensatory evolution may promote the fixation of INH/RIF-resistance mutations and affect phenotypic AMR. TB treatment should be adjusted based on gene sequencing results, especially in persistent culture positivity during treatment, which highlight the clinical importance of WGS in identifying reinfection and AMR acquisition before phenotypic DST.