Altered physiology of drug-resistant Mycobacterium tuberculosis restricts adaptation to host-relevant conditions and produces condition-dependent collateral vulnerabilities

ABSTRACT Antibiotic resistance mutations often disrupt essential cellular processes; however, the physiological consequences of these mutations under host-relevant conditions remain poorly understood in many bacterial pathogens. Here, we have used a combination of phenotypic growth assays and metabolomics to investigate how drug resistance affects the metabolism of Mycobacterium tuberculosis and its ability to adapt to different host-relevant conditions. This work shows that (i) the fitness costs of antibiotic resistance in M. tuberculosis are highly variable and can be exacerbated or alleviated depending on the media carbon source, (ii) genetically unrelated resistance mutations have overlapping phenotypes and metabolic dysregulation, (iii) the growth of bedaquiline- and rifampicin-resistant mutants was impaired on host-relevant carbon sources, and (iv) resistance mutants have condition-specific collateral antibiotic susceptibilities, with rifampicin-resistant mutants being hypersusceptible to ATP synthase inhibitors when grown under conditions that require propionate metabolism. Collectively, this study highlights how drug resistance impacts bacterial physiology and places metabolic constraints on the ability of M. tuberculosis to adapt to changes in host-relevant conditions. This work has major implications for our understanding of how resistance evolves within the host context and reinforces the importance of how physiological variation in M. tuberculosis might affect the outcomes of new treatment regimens. IMPORTANCE Mutations that provide antibiotic resistance in bacterial pathogens often disrupt key cellular processes. The effects of these disruptions on cellular physiology are poorly understood, especially within the context of a host environment. In this article, we investigated how antibiotic resistance mutations affect the ability of Mycobacterium tuberculosis , the causative agent of tuberculosis, to grow under host-relevant conditions. Using techniques that characterize growth and metabolism, we demonstrate that antibiotic-resistant M. tuberculosis can have both increased and decreased growth depending on the media carbon source, highlighting how interactions between drug resistance traits and the environment affect the ability of M. tuberculosis to adapt to changing conditions. We also show that unrelated antibiotic-resistant strains share similar metabolic defects, which can be exploited to improve antimicrobial susceptibility. Overall, this work demonstrates that drug resistance reshapes the physiology of M. tuberculosis in growth-dependent ways, having major implications for our understanding of how drug resistance evolves within the host context and for the design of new treatment strategies against antibiotic-resistant M. tuberculosis .