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Applied and Environmental Microbiology, May 2009, p. 2861-2868, Vol. 75, No. 9
0099-2240/09/$08.00+0     doi:10.1128/AEM.01317-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

DNA as an Adhesin: Bacillus cereus Requires Extracellular DNA To Form Biofilms{triangledown} ,{dagger}

Sébastien Vilain,1,3 Jakobus M. Pretorius,2 Jacques Theron,2 and Volker S. Brözel1*

Department of Biology and Microbiology, South Dakota State University, Brookings, South Dakota 57007,1 Department of Microbiology and Plant Pathology, University of Pretoria, Pretoria 0002, South Africa,2 Laboratoire de Biotechnologie des Protéines Recombinantes à Visée Santé (EA4135), ESTBB, Université Victor Segalen Bordeaux 2, 33076 Bordeaux Cedex, France3

Received 12 June 2008/ Accepted 15 February 2009

The soil saprophyte Bacillus cereus forms biofilms at solid-liquid interfaces. The composition of the extracellular polymeric matrix is not known, but biofilms of other bacteria are encased in polysaccharides, protein, and also extracellular DNA (eDNA). A Tn917 screen for strains impaired in biofilm formation at a solid-liquid interface yielded several mutants. Three mutants deficient in the purine biosynthesis genes purA, purC, and purL were biofilm impaired, but they grew planktonically like the wild type in Luria-Bertani broth. Biofilm populations had higher purA, purC, and purL transcript ratios than planktonic cultures, as measured by real-time PCR. Laser scanning confocal microscopy (LSCM) of BacLight-stained samples indicated that there were nucleic acids in the cell-associated matrix. This eDNA could be mobilized off the biofilm into an agarose gel matrix through electrophoresis, and it was a substrate for DNase. Glass surfaces exposed to exponentially growing populations acquired a DNA-containing conditioning film, as indicated by LSCM. Planktonic exponential-phase cells released DNA into an agarose gel matrix through electrophoresis, while stationary-phase populations did not do this. DNase treatment of planktonic exponential-phase populations rendered cells more susceptible than control populations to the DNA-interacting antibiotic actinomycin D. Exponential-phase purA cells did not contain detectable eDNA, nor did they convey a DNA-containing conditioning film to the glass surface. These results indicate that exponential-phase cells of B. cereus ATCC 14579 are decorated with eDNA and that biofilm formation requires DNA as part of the extracellular polymeric matrix.

* Corresponding author. Mailing address: Department of Biology and Microbiology, South Dakota State University, Brookings, SD 57007. Phone: (605) 688-5624. Fax: (605) 688-6483. E-mail: volker.brozel{at}sdstate.edu

{triangledown} Published ahead of print on 27 February 2009.

{dagger} Journal series publication 3626 from the South Dakota Agricultural Experiment Station.

Applied and Environmental Microbiology, May 2009, p. 2861-2868, Vol. 75, No. 9
0099-2240/09/$08.00+0     doi:10.1128/AEM.01317-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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