Novel role of Acinetobacter baumannii RND efflux transporters in mediating decreased susceptibility to biocides
ABSTRACT Biocides and dyes are commonly employed in hospital and laboratory settings. We investigated the biocide susceptibilities of a rapidly emerging pathogen, Acinetobacter baumannii, and the underlying molecular mechanisms, with a primary focus on resistance-nodulation-cell division (RND) efflux systems.
Biocide susceptibilities, efflux and in vitro inactivation profiles were monitored in the presence/absence of efflux pump inhibitors. The RND transporters encoded by adeB and adeJ were detected by PCR; null mutants were constructed in the native host. Expression of adeB and adeJ in clinical isolates was assayed by semi-quantitative RT-PCR.
Susceptibility testing and phenotypic assays demonstrated the role of active efflux in mediating decreased susceptibility to biocides. Inactivation of either the adeB or adeJ transporter gene led to increased susceptibility to biocides. RT-PCR analysis exhibited increased adeB and adeJ expression in clinical isolates.
This is the first study demonstrating the role of efflux pumps in mediating decreased susceptibility to disinfectants and other chemical substrates in A. baumannii.
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ABSTRACT: Among Acinetobacter spp., A. baumannii is the most frequently implicated in nosocomial infections, in particular in intensive care units. It was initially thought that multidrug resistance (MDR) in this species was due mainly to horizontal acquisition of resistance genes. However, it has recently become obvious that increased expression of chromosomal genes for efflux systems plays a major role in MDR. Among the five superfamilies of pumps, resistance-nodulation-division (RND) systems are the most prevalent in multiply resistant A. baumannii. RND pumps typically exhibit a wide substrate range that can include antibiotics, dyes, biocides, detergents, and antiseptics. Overexpression of AdeABC, secondary to mutations in the adeRS genes encoding a two-component regulatory system, constitutes a major mechanism of multiresistance in A. baumannii. AdeIJK, intrinsic to this species, is responsible for natural resistance, but since overexpression above a certain threshold is toxic for the host, its contribution to acquired resistance is minimal. The recently described AdeFGH, probably regulated by a LysR-type transcriptional regulator, also confers multidrug resistance when overexpressed. Non-RND efflux systems, such as CraA, AmvA, AbeM, and AbeS, have also been characterized for A. baumannii, as have AdeXYZ and AdeDE for other Acinetobacter spp. Finally, acquired narrow-spectrum efflux pumps, such as the major facilitator superfamily (MFS) members TetA, TetB, CmlA, and FloR and the small multidrug resistance (SMR) member QacE in Acinetobacter spp., have been detected and are mainly encoded by mobile genetic elements.Antimicrobial Agents and Chemotherapy 12/2010; 55(3):947-53. DOI:10.1128/AAC.01388-10 · 4.45 Impact Factor
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ABSTRACT: To investigate the role of outer membrane proteins in Acinetobacter baumannii resistant to imipenem, 2 strains were procured from the same patient. The imipenem-resistant strain was obtained following a period of imipenem treatment in vivo. The multilocus sequence typing and repetitive extragenic palindromic PCR results indicated that the imipenem-resistant strain originated from the sensitive one. Isoelectric focusing detected no carbapenemases, with neither OXA carbapenemases nor metallo-β-lactamases found. Mass spectrophotometric analysis revealed that 3 outer membrane proteins were expressed differentially in the 2 strains: 2 downregulated proteins (OprD and CarO) and 1 upregulated the 34-kDa efflux pump protein in the resistant strain. A 32-fold decrease in the MIC for imipenem in the presence of Phe-Arg-β-naphthylamide in the same strain indicated a possible involvement of the efflux pump mechanism in its resistance, which was consistent with the findings that the mRNA expression of the 34-kDa efflux pump gene was almost fivefold upregulated in the imipenem-resistant strain compared with that in the imipenem-sensitive strain. Such a significant difference, however, was not found in the expression of AdeB and AdeJ between the 2 strains, and AdeE was not detected. Our results suggested that downregulation of outer membrane proteins in conjunction with efflux pump overexpression might contribute to imipenem resistance induced in vivo in A. baumannii.Chemotherapy 02/2011; 57(1):77-84. DOI:10.1159/000323620 · 1.55 Impact Factor
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ABSTRACT: Traditional antimicrobials are increasingly suffering from the emergence of multidrug resistance among pathogenic microorganisms. To overcome these deficiencies, a range of novel approaches to control microbial infections are under investigation as potential alternative treatments. Multidrug efflux is a key target of these efforts. Efflux mechanisms are broadly recognized as major components of resistance to many classes of chemotherapeutic agents as well as antimicrobials. Efflux occurs due to the activity of membrane transporter proteins widely known as Multidrug Efflux Systems (MES). They are implicated in a variety of physiological roles other than efflux and identifying natural substrates and inhibitors is an active and expanding research discipline. One plausible alternative is the combination of conventional antimicrobial agents/antibiotics with small molecules that block MES known as multidrug efflux pump inhibitors (EPIs). An array of approaches in academic and industrial research settings, varying from high-throughput screening (HTS) ventures to bioassay guided purification and determination, have yielded a number of promising EPIs in a series of pathogenic systems. This synergistic discovery platform has been exploited in translational directions beyond the potentiation of conventional antimicrobial treatments. This venture attempts to highlight different tactical elements of this platform, identifying the need for highly informative and comprehensive EPI-discovery strategies. Advances in assay development genomics, proteomics as well as the accumulation of bioactivity and structural information regarding MES facilitates the basis for a new discovery era. This platform is expanding drastically. A combination of chemogenomics and chemoinformatics approaches will integrate data mining with virtual and physical HTS ventures and populate the chemical-biological interface with a plethora of novel chemotypes. This comprehensive step will expedite the preclinical development of lead EPIs.Current pharmaceutical design 04/2011; 17(13):1291-302. · 3.29 Impact Factor