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JAC Advance Access published online on June 6, 2006
Journal of Antimicrobial Chemotherapy, doi:10.1093/jac/dkl202
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© The Author 2006. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org
Received December 16, 2005
Revised April 16, 2006
Accepted April 26, 2006

Original article

Bacteroides fragilis BmeABC efflux systems additively confer intrinsic antimicrobial resistance

Lilian Pumbwe 1, Ohmi Ueda 2, Fuminobu Yoshimura 3, Abraham Chang 4, Rachel L. Smith 4, and Hannah M. Wexler 1 *

1 Greater Los Angeles Veterans Administration Healthcare Systems, Los Angeles, CA, USA; Department of Medicine, University of California, Los Angeles, CA, USA
2 Department of Oral Microbiology, Matsumoto Dental University, Shiojiri, Japan
3 Department of Microbiology, School of Dentistry, Aichi-Gakuin University, Nagoya, Japan
4 Greater Los Angeles Veterans Administration Healthcare Systems, Los Angeles, CA, USA

* To whom correspondence should be addressed.
Hannah M. Wexler, E-mail: hwexler{at}ucla.edu


  Abstract

Objectives: To determine the prevalence of expression and function(s) of Bacteroides fragilis RND family efflux transport systems (bmeABC1-16).

Methods: The mRNA transcripts of bmeB efflux pump genes were detected in a wild-type strain ADB77 by RT-PCR and expression in different strains was quantified by comparative quantitative real-time RT-PCR. In order to determine independent or additive functions, BmeB 1, 3, 12 and 15 (the first efflux pumps identified) were deleted as singles, doubles, triples or quadruples by the double cross-over technique with pADB242 and antimicrobial susceptibility was assayed by the spiral gradient endpoint technique.

Results: All efflux pumps except bmeB9 were expressed in the wild-type parental strain. Susceptibility to {beta}-lactams, fluoroquinolones, ethidium bromide, SDS and triclosan was increased in ADB77{Delta}bmeB3 (up to 3-fold) and ADB77{Delta}bmeB1{Delta}bmeB3{Delta}bmeB12 (up to 5-fold). Expression of bmeB9 was increased and that of bmeB11 repressed in the latter deletant. A quadruple deletant (ADB77{Delta}bmeB1{Delta}bmeB3{Delta}bmeB12{Delta}bmeB15) had similar changes as well as a 2-fold increase in expression of bmeB16 and norfloxacin resistance. Expression of bmeB3 was increased in two triple deletants ADB77{Delta}bmeB1{Delta}bmeB12{Delta}bmeB15-type I (2-fold) and ADB77{Delta}bmeB1{Delta}bmeB12{Delta}bmeB15-type II (5.8-fold). Antimicrobial MICs were also increased in the latter deletant; ampicillin (2.6-fold), cefoperazone (3.4-fold), cefoxitin (1.8-fold), tetracycline (36.4-fold), SDS (1.7-fold) and triclosan (2-fold).

Conclusions: These data demonstrate that constitutive bmeB expression is prevalent in B. fragilis. At least seven BmeB efflux pumps are functional in transporting antimicrobials and have overlapping substrate profiles, and at least four confer intrinsic resistance.

Keywords: membrane proteins; co-expression; susceptibility.
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