Mechanisms of Phenotypic Rifampicin Tolerance in <i>Mycobacterium tuberculosis</i> Beijing Genotype Strain B0/W148 Revealed by Proteomics

The “successful” Russian clone B0/W148 of <i>Mycobacterium tuberculosis</i> Beijing is well-known for its capacity to develop antibiotic resistance. During treatment, resistant mutants can occur that have inheritable resistance to specific antibiotics. Next to mutations, <i>M. tuberculosis</i> has several mechanisms that increase their tolerance to a variety of antibiotics. Insights in the phenotypic mechanisms that contribute to drug tolerance will increase our understanding of how antibiotic resistance develops in <i>M. tuberculosis</i>. In this study, we examined the (phospho)­proteome dynamics in <i>M. tuberculosis</i> Beijing strain B0/W148 when exposed to a high dose of rifampicin; one of the most potent first-line antibiotics. A total of 2,534 proteins and 191 phosphorylation sites were identified, and revealed the differential regulation of DosR regulon proteins, which are necessary for the development of a dormant phenotype that is less susceptible to antibiotics. By examining independent phenotypic markers of dormancy, we show that persisters of <i>in vitro</i> rifampicin exposure entered a metabolically hypoactive state, which yields rifampicin and other antibiotics largely ineffective. These new insights in the role of protein regulation and post-translational modifications during the initial phase of rifampicin treatment reveal a shortcoming in the antituberculosis regimen that is administered to 8–9 million individuals annually