The causes and consequences of historical contingencies in the evolution of extensively drug-resistant Mycobacterium tuberculosis

Tuberculosis (TB) remains a major global health crisis, surpassing HIV/AIDS and malaria in fatalities, with highest mortality from Multidrug-resistant TB (MDR-TB). A new WHO-endorsed regimen for MDR-TB, combining the drugs bedaquiline, pretomanid, linezolid, and moxifloxacin (BPaLM), has been used in Georgia since 2021. Yet, emerging resistance against these drugs jeopardizes BPaLM effectiveness, with unclear evolutionary drivers. The burden of MDR-TB are often driven by few highly transmissible strains. In Georgia, we showed that 30% of the MDR-TB cases were incarceration-related. We hypothesize that prisons facilitated the emergence and spread of these MDR-outbreak strains, and that resistance to BPaLM depends on their evolutionary histories. We will take advantage of our collection of >20,000 clinical isolates from Georgia covering three decades. We aim to i) define the role of prisons in the emergence of resistance in Georgia, ii) explore the evolutionary history, genomic and phenotypic features of the dominating MDR-outbreak strains, iii) study the effect of historical contingencies and assess risk factors for BPaLM resistance. We will include a prospective patient cohort, and apply quantitative drug susceptibility testing, long-read genome sequencing, RNAseq and proteomics. This project will generate fundamental insights into the role of historical contingencies in bacterial adaptation, and guide diagnostics and treatment strategies for this lethal disease.