Biochemical Properties of EmeA, a Multidrug Efflux Pump, from Enterococcus faecalis.

ABSTRACT A DNA fragment responsible for resistance to antimicrobial agents was cloned from the chromosomal DNA of Enterococcus faecalis ATCC 29212 by using drug-hypersensitive mutant Escherichia coli KAM32 as a host cell. Cells of E. coli KAM32 harboring a recombinant plasmid (pAEF82) carrying the DNA fragment became resistant to many structurally unrelated antimicrobial agents, such as norfloxacin, ciprofloxacin, doxycycline, acriflavine, 4′,6-diamidino-2-phenylindole, tetraphenylphosphonium chloride, daunorubicin, and doxorubicin. Since the sequence of the whole genome of E. faecalis is known, we sequenced several portions of the DNA insert in plasmid pAEF82 and identified two open reading frames within the insert. We designated the genes efrA and efrB. A search of the deduced amino acid sequences of EfrA and EfrB revealed that they are similar to each other and that they belong to the ATP-binding cassette (ABC) family of multidrug efflux transporters. Transformed E. coli KAM32 cells harboring efrAB showed energy-dependent efflux of acriflavine. The efflux activity was inhibited by reserpine, verapamil, and sodium-o-vanadate, known inhibitors of ABC efflux pumps.

Download Full-text

Topoisomerase mutations and efflux are associated with fluoroquinolone resistance in Enterococcus faecalis

Journal of Medical Microbiology ◽

10.1099/jmm.0.46636-0 ◽

2006 ◽

Vol 55 (10) ◽

pp. 1395-1401 ◽

Cited By ~ 16

Author(s):

Yoshihiro Oyamada ◽

Hideaki Ito ◽

Matsuhisa Inoue ◽

Jun-ichi Yamagishi

Keyword(s):

Enterococcus Faecalis ◽

Efflux Pump ◽

Resistance Mechanisms ◽

Fluoroquinolone Resistance ◽

Multidrug Efflux ◽

Multidrug Efflux Pump ◽

Topoisomerase Iv ◽

Gram Positive Bacteria ◽

Efflux Pump Inhibitors ◽

Resistant Mutants

To understand better the mechanisms of fluoroquinolone resistance in Enterococcus faecalis, fluoroquinolone-resistant mutants isolated from Ent. faecalis ATCC 29212 by stepwise selection with sparfloxacin (SPX) and norfloxacin (NOR) were analysed. The results showed the following. (i) In general, fluoroquinolone-resistance mechanisms in Ent. faecalis are similar to those in other Gram-positive bacteria, such as Staphylococcus aureus and Streptococcus pneumoniae, namely, mutants with amino acid changes in both GyrA and ParC exhibited high fluoroquinolone resistance, and single GyrA mutants and a single ParC mutant were more resistant to SPX and NOR, respectively, than the parent strain, indicating that the primary targets of SPX and NOR in Ent. faecalis are DNA gyrase and topoisomerase IV, respectively. (ii) Alterations in GyrB (ΔKGA, residues 395–397) and ParE (Glu-459 to Lys) were associated with fluoroquinolone resistance in some mutants. Moreover, the facts that the NOR MIC, but not the SPX MIC, decreased in the presence of multidrug efflux pump inhibitors, that NOR accumulation decreased in the cells, and that the EmeA mRNA expression level did not change, strongly suggested that a NorA-like efflux pump, rather than EmeA, was involved in resistance to NOR.

Download Full-text

Functional Cloning and Expression of emeA, and Characterization of EmeA, a Multidrug Efflux Pump from Enterococcus faecalis

Biological and Pharmaceutical Bulletin ◽

10.1248/bpb.26.266 ◽

2003 ◽

Vol 26 (2) ◽

pp. 266-270 ◽

Cited By ~ 27

Author(s):

Eun-Woo Lee ◽

Jing Chen ◽

Md. Nazmul Huda ◽

Teruo Kuroda ◽

Tohru Mizushima ◽

...

Keyword(s):

Enterococcus Faecalis ◽

Efflux Pump ◽

Cloning And Expression ◽

Multidrug Efflux ◽

Multidrug Efflux Pump

Download Full-text

Faculty Opinions recommendation of Structure of the AcrAB-TolC multidrug efflux pump.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.718356663.793495679 ◽

2014 ◽

Author(s):

Paul Savage

Keyword(s):

Efflux Pump ◽

Multidrug Efflux ◽

Multidrug Efflux Pump

Download Full-text

Faculty Opinions recommendation of Molecular basis for inhibition of AcrB multidrug efflux pump by novel and powerful pyranopyridine derivatives.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.726218324.793516072 ◽

2016 ◽

Author(s):

Herschel Wade

Keyword(s):

Molecular Basis ◽

Efflux Pump ◽

Multidrug Efflux ◽

Multidrug Efflux Pump

Download Full-text

Faculty Opinions recommendation of Biased partitioning of the multidrug efflux pump AcrAB-TolC underlies long-lived phenotypic heterogeneity.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.727537002.793532571 ◽

2017 ◽

Author(s):

Chris Whitfield

Keyword(s):

Efflux Pump ◽

Phenotypic Heterogeneity ◽

Multidrug Efflux ◽

Multidrug Efflux Pump

Download Full-text

Molecular Insights into an Antibiotic Enhancer Action of New Morpholine-Containing 5-Arylideneimidazolones in the Fight against MDR Bacteria

International Journal of Molecular Sciences ◽

10.3390/ijms22042062 ◽

2021 ◽

Vol 22 (4) ◽

pp. 2062

Author(s):

Aneta Kaczor ◽

Karolina Witek ◽

Sabina Podlewska ◽

Veronique Sinou ◽

Joanna Czekajewska ◽

...

Keyword(s):

Molecular Modelling ◽

Efflux Pump ◽

Gram Negative Bacteria ◽

Potential Mechanism ◽

Multidrug Efflux ◽

Aromatic Rings ◽

Allosteric Site ◽

Multidrug Efflux Pump ◽

Dual Action

In the search for an effective strategy to overcome antimicrobial resistance, a series of new morpholine-containing 5-arylideneimidazolones differing within either the amine moiety or at position five of imidazolones was explored as potential antibiotic adjuvants against Gram-positive and Gram-negative bacteria. Compounds (7–23) were tested for oxacillin adjuvant properties in the Methicillin-susceptible S. aureus (MSSA) strain ATCC 25923 and Methicillin-resistant S. aureus MRSA 19449. Compounds 14–16 were tested additionally in combination with various antibiotics. Molecular modelling was performed to assess potential mechanism of action. Microdilution and real-time efflux (RTE) assays were carried out in strains of K. aerogenes to determine the potential of compounds 7–23 to block the multidrug efflux pump AcrAB-TolC. Drug-like properties were determined experimentally. Two compounds (10, 15) containing non-condensed aromatic rings, significantly reduced oxacillin MICs in MRSA 19449, while 15 additionally enhanced the effectiveness of ampicillin. Results of molecular modelling confirmed the interaction with the allosteric site of PBP2a as a probable MDR-reversing mechanism. In RTE, the compounds inhibited AcrAB-TolC even to 90% (19). The 4-phenylbenzylidene derivative (15) demonstrated significant MDR-reversal “dual action” for β-lactam antibiotics in MRSA and inhibited AcrAB-TolC in K. aerogenes. 15 displayed also satisfied solubility and safety towards CYP3A4 in vitro.

Download Full-text

Correlation between the Antibiotic Resistance Genes and Susceptibility to Antibiotics among the Carbapenem-Resistant Gram-Negative Pathogens

Antibiotics ◽

10.3390/antibiotics10030255 ◽

2021 ◽

Vol 10 (3) ◽

pp. 255

Author(s):

Salma M. Abdelaziz ◽

Khaled M. Aboshanab ◽

Ibrahim S. Yahia ◽

Mahmoud A. Yassien ◽

Nadia A. Hassouna

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Efflux Pump ◽

Antibiotic Resistance Genes ◽

Multiple Drug ◽

P Value ◽

Multidrug Efflux ◽

Multidrug Efflux Pump ◽

Gram Negative ◽

Carbapenem Resistant

In this study, the correlation between the antibiotic resistance genes and antibiotic susceptibility among the carbapenem-resistant Gram-negative pathogens (CRGNPs) recovered from patients diagnosed with acute pneumonia in Egypt was found. A total of 194 isolates including Klebsiella pneumoniae (89; 46%), Escherichia coli (47; 24%) and Pseudomonas aeruginosa (58; 30%) were recovered. Of these, 34 (18%) isolates were multiple drug resistant (MDR) and carbapenem resistant. For the K. pneumoniae MDR isolates (n = 22), blaNDM (14; 64%) was the most prevalent carbapenemase, followed by blaOXA-48 (11; 50%) and blaVIM (4; 18%). A significant association (p value < 0.05) was observed between the multidrug efflux pump (AcrA) and resistance to β-lactams and the aminoglycoside acetyl transferase gene (aac-6’-Ib) gene and resistance to ciprofloxacin, azithromycin and β-lactams (except for aztreonam). For P. aeruginosa, a significant association was noticed between the presence of the blaSHV gene and the multidrug efflux pump (MexA) and resistance to fluoroquinolones, amikacin, tobramycin, co-trimoxazole and β-lactams and between the aac-6’-Ib gene and resistance to aminoglycosides. All P. aeruginosa isolates (100%) harbored the MexAB-OprM multidrug efflux pump while 86% of the K. pneumoniae isolates harbored the AcrAB-TolC pump. Our results are of great medical importance for the guidance of healthcare practitioners for effective antibiotic prescription.

Download Full-text