Efflux-mediated response of Staphylococcus aureus exposed to ethidium bromide

Abstract

Exposure of S. aureus to quaternary compounds such as ethidium bromide results in decreased susceptibility of the organism to a wide variety of compounds, including quinolones and biocides through an efflux-mediated response, which for strain ATCC 25923 is mainly NorA-mediated. This altered expression may result from alterations in the norA promoter region.

Molecular Characterization of Fluoroquinolone Resistance of Methicillin –Resistant Clinical Staphylococcus aureus Isolates from Rawalpindi, Pakistan

“…Therefore, a V371I mutation probably exhibited no change in the expression of norA, norB, norC, and mdeA. In conclusion, our findings not only correlate with the earlier studies [6,8,10,12,26] but also describe two novel point mutations in the norA gene and the deletion of nucleotide "A" upstream of a putative fur-binding box and their possible role in the regulation of efflux pump genes norA, norB, norC, and mdeA. The role of these mutations in efflux pump-mediated FQ resistance S. aureus is quite evident and will be useful in potential development of effective EPIs for better mitigation and treatment strategies of multidrug-resistant pathogens in hospitals.…”

“…Additional mutations resulting in V615I, S437P, and D646Y substitutions were also observed. In addition to the mutations in gyrase and topoisomerase genes, the contribution of efflux pumps that confer resistance to multiple antimicrobial agents in S. aureus has also been reported [6,18,24,27]. We observed the presence of efflux pump activity in all S. aureus isolates by an increased EtBr uptake and a reduction of antibiotic MICs in the presence of efflux pump inhibitors, CCCP and RES, as reported earlier [8,24,30].…”

“…All Rights Reserved 4 | P a g e Volume 2, Issue 2 and the norA genes was carried out in a GeneAmp PCR system (Bio-Rad, Hercules, CA, USA) using previously published cycling conditions [6,18,31] as summarized in supplementary table S1. The PCR reactions were performed in a total volume of 50 μl, which contained 1.6 mM each deoxynucleoside triphosphate (Applied Biosystems, Foster City, CA, USA), 20 pmol of each primer (Table S1), 1 mM MgCl 2 , 0.5 U of AmpliTaq Gold Enzyme (Applied Biosystems), and 0.5 μl of genomic DNA (~ 500 ng of DNA) in 1X buffer.…”

“…Real-time PCR was performed in triplicate using the CFX96 Real-time PCR System (Bio-Rad) and the primers previously published [6,18,31] and listed in supplementary table 1. The PCR reactions were carried out in a 20 l volume using Applied Bioscience's SYBR Green 2X Master mix kit according to the manufacturer's instructions.…”

“…Other than mutations in the QRDR hotspots, the presence of efficient efflux pumps, which extrude drugs or other noxious agents from these bacterial cells, enhances resistance to FQ [6,8,25]. Mutations within the promoter region of the multidrug efflux pump gene norA have been shown to directly affect the over-expression of the norA efflux pump in S. aureus.…”