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Journal of Bacteriology, July 2004, p. 4492-4501, Vol. 186, No. 14
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.14.4492-4501.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Phosphorus Limitation Enhances Biofilm Formation of the Plant Pathogen Agrobacterium tumefaciens through the PhoR-PhoB Regulatory System

Thomas Danhorn,1 Morten Hentzer,2 Michael Givskov,2 Matthew R. Parsek,3 and Clay Fuqua1*

Department of Biology, Indiana University, Bloomington, Indiana 47405,1 Biocentrum-DTU, Technical University of Denmark, DK-2800 Lyngby, Denmark,2 Department of Microbiology, University of Iowa, Iowa City, Iowa 522423

Received 11 November 2003/ Accepted 26 February 2004

The plant pathogen Agrobacterium tumefaciens forms architecturally complex biofilms on inert surfaces. Adherence of A. tumefaciens C58 was significantly enhanced under phosphate limitation compared to phosphate-replete conditions, despite slower overall growth under low-phosphate conditions. Replacement of Pi with sn-glycerol-3-phosphate and 2-aminoethylphosphonate yielded similar results. The increase in surface interactions under phosphate limitation was observed in both static culture and continuous-culture flow cells. Statistical analysis of confocal micrographs obtained from the flow cell biofilms revealed that phosphate limitation increased both the overall attached biomass and the surface coverage, whereas the maximum thickness of the biofilm was not affected. Functions encoded on the two large plasmids of A. tumefaciens C58, pTiC58 and pAtC58, were not required for the observed phosphate effect. The phosphate concentration at which increased attachment was observed triggered the phosphate limitation response, controlled in many bacteria by the two-component regulatory system PhoR-PhoB. The A. tumefaciens phoB and phoR orthologues could only be disrupted in the presence of plasmid-borne copies of the genes, suggesting that this regulatory system might be essential. Expression of the A. tumefaciens phoB gene from a tightly regulated inducible promoter, however, correlated with the amount of biofilm under both phosphate-limiting and nonlimiting conditions, demonstrating that components of the Pho regulon influence A. tumefaciens surface interactions.

* Corresponding author. Mailing address: Department of Biology, 1001 E. 3rd St., Jordan Hall 142, Indiana University, Bloomington, IN 47405-1847. Phone: (812) 856-6005. Fax: (812) 855-6705. E-mail: cfuqua{at}bio.indiana.edu.

Journal of Bacteriology, July 2004, p. 4492-4501, Vol. 186, No. 14
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.14.4492-4501.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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