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JAC Advance Access published online on November 22, 2005
Journal of Antimicrobial Chemotherapy, doi:10.1093/jac/dki419
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© The Author 2005. 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 August 15, 2005
Revised October 11, 2005
Accepted October 21, 2005

Original article

Interaction of CmeABC and CmeDEF in conferring antimicrobial resistance and maintaining cell viability in Campylobacter jejuni

Masato Akiba 1, Jun Lin 2, Yi-Wen Barton 3, and Qijing Zhang 3 *

1 Department of Microbiology and Preventive Medicine, Iowa State University, Ames, IA 50011, USA; Present address. Department of Safety Research, National Institute of Animal Health, 3-1-5 Kannondai, Tsukuba, Ibaraki 305-0856, Japan
2 Department of Microbiology and Preventive Medicine, Iowa State University, Ames, IA 50011, USA; Present address. Department of Animal Science, The University of Tennessee, 2505 River Drive, Knoxville, TN 37996-4574, USA
3 Department of Microbiology and Preventive Medicine, Iowa State University, Ames, IA 50011, USA

* To whom correspondence should be addressed.
Qijing Zhang, E-mail: zhang123{at}iastate.edu


  Abstract

Objectives: To determine the role of CmeDEF in conferring antimicrobial resistance in Campylobacter jejuni and examine the interaction of CmeABC and CmeDEF in mediating antimicrobial resistance and maintaining cell viability.

Methods: Single and double mutants of cmeF and cmeB were generated in multiple strains using insertional mutagenesis. The mutants were compared with their wild-type strains for antimicrobial susceptibility and growth characteristics. Transcription fusion was used to quantify the expression of cmeDEF and cmeABC. Ethidium bromide (EB) accumulation assay was used to measure the efflux function.

Results: Insertional mutagenesis of the cmeF gene in C. jejuni NCTC 11168 resulted in a 2-fold decrease in the resistance to ampicillin, polymyxin B and EB, whereas the same mutation in C. jejuni 81-176 and 21190 led to a 2-4-fold increase in the resistance to multiple antimicrobials and toxic compounds. The increased resistance in the cmeF mutants of 81-176 and 21190 was associated with the elevated efflux in the mutants. Compared with the cmeB mutant, the cmeF/cmeB double mutants of 81-176 and 21190 showed further decrease in the resistance to various antimicrobials and toxic compounds. Transcription fusion assay indicated that the expression level of cmeF was substantially lower than that of cmeB. Notably, the cmeB/cmeF double mutation, not the single mutations, impaired cell viability in Campylobacter.

Conclusions: CmeDEF interacts with CmeABC in conferring antimicrobial resistance and maintaining cell viability in C. jejuni. CmeABC is the predominant efflux pump in C. jejuni, whereas CmeDEF plays a secondary role in conferring intrinsic resistance to antimicrobials.

Keywords: antibiotics; efflux; mutagenesis.
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