Inducement and Reversal of Tetracycline Resistance in
            <i>Escherichia coli</i>
            K-12 and Expression of Proton Gradient-Dependent Multidrug Efflux Pump Genes

Viveiros, Miguel; Jesus, Ana Rita; Brito, Marisa P. de; Leandro, Clara; Martins, Marta; Ordway, Diane; Molnar, Ana Maria; Molnár, Joséph; Amaral, Leonard

doi:10.1128/aac.49.8.3578-3582.2005

Cited by 111 publications

(161 citation statements)

References 36 publications

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“…Our data are consistent with those of Olliver et al (22), who also found PA␤N-mediated inhibition of AcrEF activity in Salmonella enterica serovar Typhimurium, but inconsistent with those of Viveiros et al (37) and Amaral et al (36), who concluded that PA␤N is an AcrB-specific inhibitor. A reason for this discrepancy could be that their conclusion was based on the lack of a PA␤N effect on tetracycline resistance resulting from transient, tetracycline-induced expression of AcrEF and other pump proteins in the absence of AcrAB (37). We successfully used PA␤N in the NPN efflux assay and showed, for the first time, that it is an effective and potent inhibitor of both AcrAB-and AcrEF-mediated NPN efflux.…”

Section: Discussionmentioning

confidence: 99%

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Importance of Real-Time Assays To Distinguish Multidrug Efflux Pump-Inhibiting and Outer Membrane-Destabilizing Activities in Escherichia coli

et al. 2015

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The constitutively expressed AcrAB multidrug efflux system of Escherichia coli shows a high degree of homology with the normally silent AcrEF system. Exposure of a strain with acrAB deleted to antibiotic selection pressure frequently leads to the insertion sequence-mediated activation of the homologous AcrEF system. In this study, we used strains constitutively expressing either AcrAB or AcrEF from their normal chromosomal locations to resolve a controversy about whether phenylalanylarginine ␤-naphthylamide (PA␤N) inhibits the activities of AcrAB and AcrEF and/or acts synergistically with antibiotics by destabilizing the outer membrane permeability barrier. Real-time efflux assays allowed a clear distinction between the efflux pump-inhibiting activity of PA␤N and the outer membrane-destabilizing action of polymyxin B nonapeptide (PMXBN). When added in equal amounts, PA␤N, but not PMXBN, strongly inhibited the efflux activities of both AcrAB and AcrEF pumps. In contrast, when outer membrane destabilization was assessed by the nitrocefin hydrolysis assay, PMXBN exerted a much greater damaging effect than PA␤N. Strong action of PA␤N in inhibiting efflux activity compared to its weak action in destabilizing the outer membrane permeability barrier suggests that PA␤N acts mainly by inhibiting efflux pumps. We concluded that at low concentrations, PA␤N acts specifically as an inhibitor of both AcrAB and AcrEF efflux pumps; however, at high concentrations, PA␤N in the efflux-proficient background not only inhibits efflux pump activity but also destabilizes the membrane. The effects of PA␤N on membrane integrity are compounded in cells unable to extrude PA␤N. IMPORTANCEThe increase in multidrug-resistant bacterial pathogens at an alarming rate has accelerated the need for implementation of better antimicrobial stewardship, discovery of new antibiotics, and deeper understanding of the mechanism of drug resistance. The work carried out in this study highlights the importance of employing real-time fluorescence-based assays in differentiating multidrug efflux-inhibitory and outer membrane-destabilizing activities of antibacterial compounds. Multidrug resistance among human bacterial pathogens remains a grave social concern. Numerous strategies have been proposed to curtail the rampant increase in multidrug resistance among human pathogens, ranging from the effective integration of pharmacokinetic and pharmacodynamic parameters and implementation of antimicrobial stewardship (1) to the development of novel antibiotics based on either an existing or a novel chemical scaffold, exploitation of new cellular targets, and directly tackling the cellular mechanisms that confer multidrug resistance (2). Efflux of antibiotics from the cell is one of the common mechanisms of antibiotic resistance in bacteria, with resistance developing when the rate of drug efflux across the membrane exceeds that of drug influx (3).Bacterial genomes encode several membrane-bound multidrug efflux systems (4, 5). These systems are usually under t...

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Section: Discussionmentioning

confidence: 99%

“…AcrAB and AcrEF share high amino acid sequence identity (66.49% and 77.56%, respectively) and thus are expected to respond very similarly. It was therefore somewhat surprising that two reports concluded that PA␤N is an AcrB-specific inhibitor (36,37).…”

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confidence: 99%

Importance of Real-Time Assays To Distinguish Multidrug Efflux Pump-Inhibiting and Outer Membrane-Destabilizing Activities in Escherichia coli

et al. 2015

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“…The respective MICs of the various compounds against the bacteria were determined using a rapid XTT colorimetric assay [14]. Briefly, samples were first dissolved in dimethyl sulphoxide (DMSO)/MHB.…”

Section: Determination Of Bacterial Susceptibilitymentioning

confidence: 99%

Antibacterial activity of some natural products against bacteria expressing a multidrug-resistant phenotype

Kuete

Alibert²,

Eyong

et al. 2011

International Journal of Antimicrobial Agents

142

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“…In E. coli, AcrAB-TolC is upregulated in response to different signals, such as aromatic weak acids (salicylate), superoxides (generated by paraquat), bile salts, fatty acids (decanoate), and tetracycline (49). These toxic compounds activate the transcription of global regulators, and they cause upregulation of the AcrAB-TolC efflux pump (11,40,45,48).…”

Section: Discussionmentioning

confidence: 99%

Effect of Transcriptional Activators SoxS, RobA, and RamA on Expression of Multidrug Efflux Pump AcrAB-TolC in Enterobacter cloacae

Pérez

Poza

Aranda

et al. 2012

Antimicrob Agents Chemother

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Control of membrane permeability is a key step in regulating the intracellular concentration of antibiotics. Efflux pumps confer innate resistance to a wide range of toxic compounds such as antibiotics, dyes, detergents, and disinfectants in members of the Enterobacteriaceae. The AcrAB-TolC efflux pump is involved in multidrug resistance in Enterobacter cloacae. However, the underlying mechanism that regulates the system in this microorganism remains unknown. In Escherichia coli, the transcription of acrAB is upregulated under global stress conditions by proteins such as MarA, SoxS, and Rob. In the present study, two clinical isolates of E. cloacae, EcDC64 (a multidrug-resistant strain overexpressing the AcrAB-TolC efflux pump) and Jc194 (a strain with a basal AcrAB-TolC expression level), were used to determine whether similar global stress responses operate in E. cloacae and also to establish the molecular mechanisms underlying this response. A decrease in susceptibility to erythromycin, tetracycline, telithromycin, ciprofloxacin, and chloramphenicol was observed in clinical isolate Jc194 and, to a lesser extent in EcDC64, in the presence of salicylate, decanoate, tetracycline, and paraquat. Increased expression of the acrAB promoter in the presence of the above-described conditions was observed by flow cytometry and reverse transcription-PCR, by using a reporter fusion protein (green fluorescent protein). The expression level of the AcrAB promoter decreased in E. cloacae EcDC64 derivates deficient in SoxS, RobA, and RamA. Accordingly, the expression level of the AcrAB promoter was higher in E. cloacae Jc194 strains overproducing SoxS, RobA, and RamA. Overall, the data showed that SoxS, RobA, and RamA regulators were associated with the upregulation of acrAB, thus conferring antimicrobial resistance as well as a stress response in E. cloacae. In summary, the regulatory proteins SoxS, RobA, and RamA were cloned and sequenced for the first time in this species. The involvement of these proteins in conferring antimicrobial resistance through upregulation of acrAB was demonstrated in E. cloacae.

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Inducement and Reversal of Tetracycline Resistance in Escherichia coli K-12 and Expression of Proton Gradient-Dependent Multidrug Efflux Pump Genes

Cited by 111 publications

References 36 publications

Importance of Real-Time Assays To Distinguish Multidrug Efflux Pump-Inhibiting and Outer Membrane-Destabilizing Activities in Escherichia coli

Importance of Real-Time Assays To Distinguish Multidrug Efflux Pump-Inhibiting and Outer Membrane-Destabilizing Activities in Escherichia coli

Antibacterial activity of some natural products against bacteria expressing a multidrug-resistant phenotype

Effect of Transcriptional Activators SoxS, RobA, and RamA on Expression of Multidrug Efflux Pump AcrAB-TolC in Enterobacter cloacae

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