Genetic analysis of the β-lactamases of Mycobacterium tuberculosis and Mycobacterium smegmatis and susceptibility to β-lactam antibiotics

摘要

Mycobacteria produce β-lactamases and are intrinsically resistant to β-lactam antibiotics. In addition to the β-lactamases, cell envelope permeability and variations in certain peptidoglycan biosynthetic enzymes are believed to contribute to β-lactam resistance in these organisms. To allow the study of these additional mechanisms, mutants of the major β-lactamases, BlaC and BlaS, were generated in the pathogenic Mycobacterium tuberculosis strain H37Rv and the model organism Mycobacterium smegmatis strain PM274. The mutants M. tuberculosis PM638 (ΔblaC1) and M. smegmatis PM759 (ΔblaS1) showed an increase in susceptibility to β-lactam antibiotics, as determined by disc diffusion and minimal inhibitory concentration (MIC) assays. The susceptibility of the mutants, as assayed by disc diffusion tests, to penicillin-type β-lactam antibiotics was affected most, compared to the cephalosporin-type β-lactam antibiotics. The M. tuberculosis mutant had no detectable β-lactamase activity, while the M. smegmatis mutant had a residual type 1 β-lactamase activity. We identified a gene, blaE, encoding a putative cephalosporinase in M. smegmatis. A double β-lactamase mutant of M. smegmatis, PM976 (ΔblaS1ΔblaE?:?:?res), had no detectable β-lactamase activity, but its susceptibility to β-lactam antibiotics was not significantly different from that of the ΔblaS1 parental strain, PM759. The mutants generated in this study will help determine the contribution of other β-lactam resistance mechanisms in addition to serving as tools to study the biology of peptidoglycan biosynthesis in these organisms.