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First published online December 18, 2003; 10.1104/pp.103.028712

Plant Physiology 134:307-319 (2004)
© 2004 American Society of Plant Biologists

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PLANTS INTERACTING WITH OTHER ORGANISMS

Biocontrol of Bacillus subtilis against Infection of Arabidopsis Roots by Pseudomonas syringae Is Facilitated by Biofilm Formation and Surfactin Production1

Harsh Pal Bais, Ray Fall and Jorge M. Vivanco*

Department of Horticulture and Landscape Architecture (H.P.B., J.M.V.) and Cell and Molecular Biology Program (J.M.V.), Colorado State University, Fort Collins, Colorado 80523-1173; and Department of Chemistry and Biochemistry and Cooperative Institute for Research in Environmental Sciences, University of Colorado, Boulder, Colorado 80309-0215 (R.F.)

Relatively little is known about the exact mechanisms used by Bacillus subtilis in its behavior as a biocontrol agent on plants. Here, we report the development of a sensitive plant infection model demonstrating that the bacterial pathogen Pseudomonas syringae pv tomato DC3000 is capable of infecting Arabidopsis roots both in vitro and in soil. Using this infection model, we demonstrated the biocontrol ability of a wild-type B. subtilis strain 6051 against P. syringae. Arabidopsis root surfaces treated with B. subtilis were analyzed with confocal scanning laser microscopy to reveal a three-dimensional B. subtilis biofilm. It is known that formation of biofilms by B. subtilis is a complex process that includes secretion of surfactin, a lipopeptide antimicrobial agent. To determine the role of surfactin in biocontrol by B. subtilis, we tested a mutant strain, M1, with a deletion in a surfactin synthase gene and, thus, deficient in surfactin production. B. subtilis M1 was ineffective as a biocontrol agent against P. syringae infectivity in Arabidopsis and also failed to form robust biofilms on either roots or inert surfaces. The antibacterial activity of surfactin against P. syringae was determined in both broth and agar cultures and also by live-dead staining methods. Although the minimum inhibitory concentrations determined were relatively high (25 µg mL-1), the levels of the lipopeptide in roots colonized by B. subtilis are likely to be sufficient to kill P. syringae. Our results collectively indicate that upon root colonization, B. subtilis 6051 forms a stable, extensive biofilm and secretes surfactin, which act together to protect plants against attack by pathogenic bacteria.


1 This work was supported by the Colorado State University Agricultural Experiment Station (grant to J.M.V.), by the National Science Foundation-CAREER (grant no. MCB 0093014 to J.M.V.), by the State of Colorado (Invasive Weeds Initiative to J.M.V.),by the Lindbergh Foundation (to J.M.V.), and by the U.S. Department of Energy (grant no. DE-FG03-97ER20274 to R.F.).

Article, publication date, and citation information can be found at www.plantphysiol.org/cgi/doi/10.1104/pp.103.028712.

* Corresponding author; e-mail jvivanco{at}lamar.colostate.edu; fax 970-491-7745.

Received June 17, 2003; returned for revision July 21, 2003; accepted November 3, 2003.




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