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ABSTRACT The increasing clinical significance of Mycobacterium abscessus is owed to its innate high-level, broad-spectrum resistance to antibiotics and therefore rapidly evolves as an important human pathogen. This warrants the identification of novel targets for aiding the discovery of new drugs or drug combinations to treat M. abscessus infections. This study is inspired by the drug-hypersensitive profile of a mutant M. abscessus (U14) with transposon insertion in MAB_1915. We validated the role of MAB_1915 in intrinsic drug resistance in M. abscessus by constructing a selectable marker-free in-frame deletion in MAB_1915 and complementing the mutant with the same or extended version of the gene and then followed by drug susceptibility testing. Judging by the putative function of MAB_1915, cell envelope permeability was studied by ethidium bromide accumulation assay and susceptibility testing against dyes and detergents. In this study, we established genetic evidence of the role of MAB_1915 in intrinsic resistance to rifampicin, rifabutin, linezolid, clarithromycin, vancomycin, and bedaquiline. Disruption of MAB_1915 has also been observed to cause a significant increase in cell envelope permeability in M. abscessus. Restoration of resistance is observed to depend on at least 27 base pairs upstream of the coding DNA sequence of MAB_1915. MAB_1915 could therefore be associated with cell envelope permeability, and hence its role in intrinsic resistance to multiple drugs in M. abscessus, which presents it as a novel target for future development of effective antimicrobials to overcome intrinsic drug resistance in M. abscessus.IMPORTANCEThis study reports the role of a putative fadD (MAB_1915) in innate resistance to multiple drugs by M. abscessus, hence identifying MAB_1915 as a valuable target and providing a baseline for further mechanistic studies and development of effective antimicrobials to check the high level of intrinsic resistance in this pathogen.

BACKGROUND Ionic liquids (ILs) have become increasingly attractive as alternatives for organic solvents. In this study, the influences of ILs on the cellular growth rate, cellular activity, cortisone acetate (CA) conversion, CA solubility, and cell membrane permeability of Arthrobacter simplex CPCC 140451 ( ASP) were examined. RESULTS Results showed that ILs with ≥3 carbon atoms in their side chains increased CA conversion, CA solubility, and cell membrane permeability of ASP and universally inhibited the cellular growth rate and activity, with ILs with [PF6]-anion exhibiting the most severe inhibition activity. Inhibition was also more serious using ILs with longer alkyl side chains and at higher concentrations. CA conversion, CA solubility, and leakage of cell inclusions showed higher increase under ILs with [PF6]-anion than those of ILs with [BF4]-anion and lower increase for ILs with longer side chains. CONCLUSION This study improved the understanding of the influence and primary selection of imidazolium-based ILs when applied in steroid biotransformation with Arthrobacter simplex whole cells. The data provided a foundation for the selection of suitable ILs for various applications. © 2017 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Hydroxypropyl-β-cyclodextrin (HP-β-CD) enhances steroid 1-dehydrogenation biotransformation by Arthrobacter simplex. In this work, HP-β-CD-induced improvement of A. simplex CPCC 140451 cell envelope permeability which had positive effects on the steroid bioconversion was confirmed by a comparative investigation which showed a lower dehydrogenase activity and higher cell permeability of the cells after being incubated with HP-β-CD. Atomic force microscopy and transmission electron microscopy micrographs showed that HP-β-CD altered the size, sharpness, and surface structure of the cell envelope. The analysis of lipid composition revealed that the proportion of extractable lipids decreased and the fatty acids profile was considerably altered. The contents of unsaturated fatty acids and long-chain fatty acids were reduced by 11.77 and 14.98 %, respectively. The total leakage of protein level increased to 8 %. Proteins belonging to the ATP-binding cassette superfamily and major facilitator superfamily were observed outside the cell. These alterations can explain the change of permeability on the molecular level under HP-β-CD treatment. Results showed the material basis and mechanisms underlying the cellular changes, thus most likely contributing to the conversion rate in addition to cyclodextrins known effects on substrate solubility. [ABSTRACT FROM AUTHOR]

The opportunistic pathogen Pseudomonas aeruginosa is a leading cause of nosocomial infections. Its relatively impermeable outer membrane (OM) limits antibiotic entry, and a chromosomally encoded AmpC β-lactamase inactivates β-lactam antibiotics. AmpC expression is linked to peptidoglycan (PG) recycling, and soluble (sLT) or membrane-bound (mLT) lytic transglycosylases are responsible for generating the anhydromuropeptides that induce AmpC expression. Thus, inhibition of LT activity could reduce AmpC-mediated β-lactam resistance in P. aeruginosa. Here, we characterized single and combination LT mutants. Strains lacking SltB1 or MltB had increased β-lactam minimum inhibitory concentrations (MICs) compared to wild type, while only loss of Slt decreased MICs. An sltB1 mltB double mutant had elevated β-lactam MICs compared to either the sltB1 or mltB single mutants (96 vs. 32 μg/mL cefotaxime), without changes to AmpC levels. Time-kill assays with β-lactams suggested that increased MIC correlated with a slower rate of autolysis in the sltB1 mltB mutant - an antisuicide phenotype. Strains lacking multiple mLTs were more sensitive to β-lactams and up to 16-fold more sensitive to vancomycin, normally incapable of crossing the OM. Multi-mLT mutants were also sensitive to bile salts and osmotic stress, and were hyperbiofilm formers, all phenotypes consistent with cell envelope compromise. Complementation with genes encoding inactive forms of the enzymes - or alternatively, overexpression of Braun's lipoprotein - reversed the mutants' cell envelope damage phenotypes, suggesting that mLTs help to stabilize the OM. We conclude that P. aeruginosa mLTs contribute physically to cell envelope stability, and that Slt is the preferred target for future development of LT inhibitors that could synergize with β-lactams., (© 2015 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)