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EfrAB, an ABC multidrug efflux pump in Enterococcus faecalis.

Eun-Woo Lee1, M Nazmul Huda, Teruo Kuroda

  • 1Department of Microbiology, Faculty of Pharmaceutical Sciences, Okayama University, Tsushima, Okayama, 700-8530, Japan.

Antimicrobial Agents and Chemotherapy
|November 26, 2003
PubMed
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Researchers cloned a DNA fragment from Enterococcus faecalis conferring broad antimicrobial resistance in E. coli. This fragment contains two genes, efrA and efrB, encoding ATP-binding cassette (ABC) multidrug efflux transporters.

Area of Science:

  • Microbiology
  • Molecular Biology
  • Genetics

Background:

  • Multidrug resistance (MDR) is a significant challenge in healthcare.
  • Efflux pumps are a major mechanism contributing to MDR in bacteria.
  • Identifying novel efflux pump genes is crucial for developing strategies to combat MDR.

Purpose of the Study:

  • To clone and characterize a DNA fragment from Enterococcus faecalis conferring resistance to multiple antimicrobial agents.
  • To identify the specific genes responsible for this resistance phenotype.
  • To elucidate the mechanism of action of the identified resistance genes.

Main Methods:

  • Cloning of a DNA fragment from Enterococcus faecalis ATCC 29212 into a drug-hypersensitive Escherichia coli KAM32 host.
  • Transformation of E. coli with recombinant plasmid pAEF82 and assessment of antimicrobial resistance.

Related Experiment Videos

  • Sequencing of the DNA insert in pAEF82 to identify open reading frames (ORFs).
  • Bioinformatic analysis of deduced amino acid sequences to identify gene families.
  • Functional analysis of transformed E. coli for acriflavine efflux and inhibition studies.

    • Main Results:

    • A DNA fragment in plasmid pAEF82 conferred resistance to structurally unrelated antimicrobial agents including fluoroquinolones, tetracyclines, and intercalating dyes.
    • Two ORFs, designated efrA and efrB, were identified within the DNA insert.
    • EfrA and EfrB showed similarity to each other and belonged to the ATP-binding cassette (ABC) transporter family.
    • Transformed E. coli expressing efrAB exhibited energy-dependent efflux of acriflavine, inhibited by known ABC pump inhibitors (reserpine, verapamil, sodium-o-vanadate).

    Conclusions:

    • The cloned DNA fragment contains the efrAB genes, which encode a novel ABC multidrug efflux transporter system in Enterococcus faecalis.
    • This efflux system confers resistance to a broad spectrum of antimicrobial agents.
    • Understanding the function of EfrAB provides insights into MDR mechanisms and potential targets for antimicrobial drug development.