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Chemical Genetic Interaction Profiling Reveals Determinants of Intrinsic Antibiotic Resistance in Mycobacterium tuberculosis.

ABSTRACT: Chemotherapy for tuberculosis (TB) is lengthy and could benefit from synergistic adjuvant therapeutics that enhance current and novel drug regimens. To identify genetic determinants of intrinsic antibiotic susceptibility in Mycobacterium tuberculosis, we applied a chemical genetic interaction (CGI) profiling approach. We screened a saturated transposon mutant library and identified mutants that exhibit altered fitness in the presence of partially inhibitory concentrations of rifampin, ethambutol, isoniazid, vancomycin, and meropenem, antibiotics with diverse mechanisms of action. This screen identified the M. tuberculosis cell envelope to be a major determinant of antibiotic susceptibility but did not yield mutants whose increase in susceptibility was due to transposon insertions in genes encoding efflux pumps. Intrinsic antibiotic resistance determinants affecting resistance to multiple antibiotics included the peptidoglycan-arabinogalactan ligase Lcp1, the mycolic acid synthase MmaA4, the protein translocase SecA2, the mannosyltransferase PimE, the cell envelope-associated protease CaeA/Hip1, and FecB, a putative iron dicitrate-binding protein. Characterization of a deletion mutant confirmed FecB to be involved in the intrinsic resistance to every antibiotic analyzed. In contrast to its predicted function, FecB was dispensable for growth in low-iron medium and instead functioned as a critical mediator of envelope integrity.

SUBMITTER: Xu W

PROVIDER: S-EPMC5700314 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Chemical Genetic Interaction Profiling Reveals Determinants of Intrinsic Antibiotic Resistance in Mycobacterium tuberculosis.

Xu Weizhen W DeJesus Michael A MA Rücker Nadine N Engelhart Curtis A CA Wright Meredith G MG Healy Claire C Lin Kan K Wang Ruojun R Park Sae Woong SW Ioerger Thomas R TR Schnappinger Dirk D Ehrt Sabine S

Antimicrobial agents and chemotherapy 20171122 12

Chemotherapy for tuberculosis (TB) is lengthy and could benefit from synergistic adjuvant therapeutics that enhance current and novel drug regimens. To identify genetic determinants of intrinsic antibiotic susceptibility in <i>Mycobacterium tuberculosis</i>, we applied a chemical genetic interaction (CGI) profiling approach. We screened a saturated transposon mutant library and identified mutants that exhibit altered fitness in the presence of partially inhibitory concentrations of rifampin, eth ...[more]

PMID: 28893793

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Project description:Pyrazinamide (PZA) is a prodrug that is converted to pyrazinoic acid by the enzyme pyrazinamidase, encoded by the pncA gene in Mycobacterium tuberculosis. Molecular identification of mutations in pncA offers the potential for rapid detection of pyrazinamide resistance (PZA(r)). However, the genetic variants are highly variable and scattered over the full length of pncA, complicating the development of a molecular test. We performed a large multicenter study assessing pncA sequence variations in 1,950 clinical isolates, including 1,142 multidrug-resistant (MDR) strains and 483 fully susceptible strains. The results of pncA sequencing were correlated with phenotype, enzymatic activity, and structural and phylogenetic data. We identified 280 genetic variants which were divided into four classes: (i) very high confidence resistance mutations that were found only in PZA(r) strains (85%), (ii) high-confidence resistance mutations found in more than 70% of PZA(r) strains, (iii) mutations with an unclear role found in less than 70% of PZA(r) strains, and (iv) mutations not associated with phenotypic resistance (10%). Any future molecular diagnostic assay should be able to target and identify at least the very high and high-confidence genetic variant markers of PZA(r); the diagnostic accuracy of such an assay would be in the range of 89.5 to 98.8%. Importance: Conventional phenotypic testing for pyrazinamide resistance in Mycobacterium tuberculosis is technically challenging and often unreliable. The development of a molecular assay for detecting pyrazinamide resistance would be a breakthrough, directly overcoming both the limitations of conventional testing and its related biosafety issues. Although the main mechanism of pyrazinamide resistance involves mutations inactivating the pncA enzyme, the highly diverse genetic variants scattered over the full length of the pncA gene and the lack of a reliable phenotypic gold standard hamper the development of molecular diagnostic assays. By analyzing a large number of strains collected worldwide, we have classified the different genetic variants based on their predictive value for resistance which should lead to more rapid diagnostic tests. This would assist clinicians in improving treatment regimens for patients.

Dataset Information

Chemical Genetic Interaction Profiling Reveals Determinants of Intrinsic Antibiotic Resistance in Mycobacterium tuberculosis.

Publications

Chemical Genetic Interaction Profiling Reveals Determinants of Intrinsic Antibiotic Resistance in Mycobacterium tuberculosis.

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