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Applied and Environmental Microbiology, May 2007, p. 2777-2786, Vol. 73, No. 9
0099-2240/07/$08.00+0     doi:10.1128/AEM.00984-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Two Host-Induced Ralstonia solanacearum Genes, acrA and dinF, Encode Multidrug Efflux Pumps and Contribute to Bacterial Wilt Virulence ^,

Darby G. Brown, Jill K. Swanson, and Caitilyn Allen^*

University of Wisconsin—Madison, Department of Plant Pathology, Madison, Wisconsin 53706

Received 26 April 2006/ Accepted 19 February 2007

Multidrug efflux pumps (MDRs) are hypothesized to protect pathogenic bacteria from toxic host defense compounds. We created mutations in the Ralstonia solanacearum acrA and dinF genes, which encode putative MDRs in the broad-host-range plant pathogen. Both mutations reduced the ability of R. solanacearum to grow in the presence of various toxic compounds, including antibiotics, phytoalexins, and detergents. Both acrAB and dinF mutants were significantly less virulent on the tomato plant than the wild-type strain. Complementation restored near-wild-type levels of virulence to both mutants. Addition of either dinF or acrAB to Escherichia coli MDR mutants KAM3 and KAM32 restored the resistance of these strains to several toxins, demonstrating that the R. solanacearum genes can function heterologously to complement known MDR mutations. Toxic and DNA-damaging compounds induced expression of acrA and dinF, as did growth in both susceptible and resistant tomato plants. Carbon limitation also increased expression of acrA and dinF, while the stress-related sigma factor RpoS was required at a high cell density (>10⁷ CFU/ml) to obtain wild-type levels of acrA expression both in minimal medium and in planta. The type III secretion system regulator HrpB negatively regulated dinF expression in culture at high cell densities. Together, these results show that acrAB and dinF encode MDRs in R. solanacearum and that they contribute to the overall aggressiveness of this phytopathogen, probably by protecting the bacterium from the toxic effects of host antimicrobial compounds.

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* Corresponding author. Mailing address: Department of Plant Pathology, University of Wisconsin—Madison, 1630 Linden Dr., Madison, WI 53706. Phone: (608) 262-9578. Fax: (608) 263-2626. E-mail: cza{at}plantpath.wisc.edu

Published ahead of print on 2 March 2007.

Supplemental material for this article may be found at http://aem.asm.org/.

Present address: School of Biological Sciences, University of Auckland, Private Bag 92019, Auckland 1020, New Zealand.

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Applied and Environmental Microbiology, May 2007, p. 2777-2786, Vol. 73, No. 9
0099-2240/07/$08.00+0     doi:10.1128/AEM.00984-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

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