NorA mediated efflux in Staphylococcus epidermidis

I Couto, SS Costa, M Viveiros, Constança Pomba

Research output: Contribution to conferencePoster

Abstract

Staphylococcus epidermidisis a colonizer of the human skin and a common nosocomial pathogen that is often resilient to antibiotic therapeutics. Moreover, antibiotic resistant strains are frequently isolated. In Staphylococcus aureus, resistance to antibiotics and biocides can be mediated by multidrug efflux pumps (EP) and efflux is now recognized as a relevant mechanism in the development of resistance. In opposition, very little is known about efflux-mediated resistance in S. epidermidis. The aim of this work is to expand our knowledge on this topic by exposing a susceptible strain to ethidium bromide (EtBr), a surrogate marker for efflux activity.The reference strain S. epidermidisATCC12228, with a minimum inhibitory concentration (MIC) of EtBr of 0.25 mg/L, was subjected to a step-wise exposure to EtBr. After the exposure process, an EtBr-adapted strain was obtained (ATCC12228_EtBr). Strains ATCC12228 and the EtBr-adapted counterpart, ATCC12228_EtBr, were evaluated regarding antibiotic and biocide susceptibility by determination of MICs. Efflux activity was assessed by real-time fluorometry and MIC re-determination in the presence of efflux inhibitors. Finally, the expression levels of homologs of the S. aureusEP genes norA, norB, norCand of the regulator mgrA, as well as of a putative MATE transporter were analyzed by RT-qPCR.Strain ATCC12228_EtBr displayed an EtBr MIC of 32 mg/L (64-fold higher), together with reduced susceptibility to biocides (quaternary ammonium compounds and chlorhexidine), as well as low-level phenotypic resistance to the fluoroquinolones ciprofloxacin (MIC of 4 mg/L) and norfloxacin (MIC of 16 mg/L). This phenotype was accompanied by an increase in the overall efflux activity relatively to the parental strain as determined by fluorometric EtBr efflux assays. In addition, all efflux inhibitors tested (thioridazine, chlorpromazine, reserpine and verapamil) were able to significantly reduce the MICs of EtBr,fluoroquinolones and selected biocides. RT-qPCR gene expression analysis revealed an overexpression of only the norAgene in ATCC12228_EtBr relatively to ATCC12228.This work demonstrates that efflux mediated by the native EP NorA is a contributor to antibiotic and biocide resistance in S. epidermidisand highlights the need of further studies to fully ascertain its role in S. epidermidisand other staphylococci of interest.
Original languageEnglish
Pages95
Number of pages1
Publication statusPublished - 19 Apr 2017
Event4th National Congress of Tropical Medicine - Lisbon, Portugal
Duration: 19 Apr 201721 Apr 2017

Conference

Conference4th National Congress of Tropical Medicine
Abbreviated title4th CNMT
CountryPortugal
CityLisbon
Period19/04/1721/04/17

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