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Journal of Bacteriology, July 2009, p. 4276-4285, Vol. 191, No. 13
0021-9193/09/$08.00+0     doi:10.1128/JB.00363-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

The Global Consequence of Disruption of the AcrAB-TolC Efflux Pump in Salmonella enterica Includes Reduced Expression of SPI-1 and Other Attributes Required To Infect the Host{triangledown} ,{dagger}

Mark A. Webber,1 Andrew M. Bailey,1 Jessica M. A. Blair,1 Eirwen Morgan,2 Mark P. Stevens,2 Jay C. D. Hinton,3,{ddagger} Al Ivens,4,§ John Wain,4,¶ and Laura J. V. Piddock1*

Antimicrobial Agents Research Group, Division of Immunity and Infection, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom,1 Division of Microbiology, Institute for Animal Health, Compton, Berkshire RG20 7NN, United Kingdom,2 Molecular Microbiology Group, Institute of Food Research, Colney, Norwich, United Kingdom,3 Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge CB10 1SA, United Kingdom4

Received 16 March 2009/ Accepted 22 April 2009

The mechanisms by which RND pumps contribute to pathogenicity are currently not understood. Using the AcrAB-TolC system as a paradigm multidrug-resistant efflux pump and Salmonella enterica serovar Typhimurium as a model pathogen, we have demonstrated that AcrA, AcrB, and TolC are each required for efficient adhesion to and invasion of epithelial cells and macrophages by Salmonella in vitro. In addition, AcrB and TolC are necessary for Salmonella to colonize poultry. Mutants lacking acrA, acrB, or tolC showed differential expression of major operons and proteins involved in pathogenesis. These included chemotaxis and motility genes, including cheWY and flgLMK and 14 Salmonella pathogenicity island (SPI)-1-encoded type III secretion system genes, including sopE, and associated effector proteins. Reverse transcription-PCR confirmed these data for identical mutants in two other S. Typhimurium backgrounds. Western blotting showed reduced production of SipA, SipB, and SipC. The absence of AcrB or TolC also caused widespread repression of chemotaxis and motility genes in these mutants, and for acrB::aph, this was associated with decreased motility. For mutants lacking a functional acrA or acrB gene, the nap and nir operons were repressed, and both mutants grew poorly in anaerobic conditions. All phenotypes were restored to that of the wild type by trans-complementation with the wild-type allele of the respective inactivated gene. These data explain how mutants lacking a component of AcrAB-TolC are attenuated and that this phenotype is a result of decreased expression of numerous genes encoding proteins involved in pathogenicity. The link between antibiotic resistance and pathogenicity establishes the AcrAB-TolC system as fundamental to the biology of Salmonella.

* Corresponding author. Mailing address: Antimicrobial Agents Research Group, Division of Immunity and Infection, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom. Mailing address: Phone: 44 121 414 6966. Fax: 44 121 414 6819. E-mail: l.j.v.piddock{at}bham.ac.uk

{triangledown} Published ahead of print on 1 May 2009.

{dagger} Supplemental material for this article may be found at http://jb.asm.org/.

{ddagger} Present address: Department of Microbiology, the Moyne Institute of Preventive Medicine, Trinity College, Dublin 2, Ireland.

§ Present address: Fios Genomics Ltd., ETTC, King's Buildings, Edinburgh EH9 3JL, United Kingdom.

Present address: Health Protection Agency, Centre for Infections, 61 Colindale Avenue, London NW9 5EQ, United Kingdom.

Journal of Bacteriology, July 2009, p. 4276-4285, Vol. 191, No. 13
0021-9193/09/$08.00+0     doi:10.1128/JB.00363-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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