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Applied and Environmental Microbiology, March 2006, p. 2064-2069, Vol. 72, No. 3
0099-2240/06/$08.00+0     doi:10.1128/AEM.72.3.2064-2069.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Chelator-Induced Dispersal and Killing of Pseudomonas aeruginosa Cells in a Biofilm{dagger}

Ehud Banin,1 Keith M. Brady,2 and E. Peter Greenberg1*

Department of Microbiology, School of Medicine, University of Washington, Seattle, Washington 98195-7242,1 Department of Microbiology, Roy and Lucille Carver College of Medicine, University of Iowa, Iowa City, Iowa 522422

Received 2 November 2005/ Accepted 20 December 2005

Biofilms consist of groups of bacteria attached to surfaces and encased in a hydrated polymeric matrix. Bacteria in biofilms are more resistant to the immune system and to antibiotics than their free-living planktonic counterparts. Thus, biofilm-related infections are persistent and often show recurrent symptoms. The metal chelator EDTA is known to have activity against biofilms of gram-positive bacteria such as Staphylococcus aureus. EDTA can also kill planktonic cells of Proteobacteria like Pseudomonas aeruginosa. In this study we demonstrate that EDTA is a potent P. aeruginosa biofilm disrupter. In Tris buffer, EDTA treatment of P. aeruginosa biofilms results in 1,000-fold greater killing than treatment with the P. aeruginosa antibiotic gentamicin. Furthermore, a combination of EDTA and gentamicin results in complete killing of biofilm cells. P. aeruginosa biofilms can form structured mushroom-like entities when grown under flow on a glass surface. Time lapse confocal scanning laser microscopy shows that EDTA causes a dispersal of P. aeruginosa cells from biofilms and killing of biofilm cells within the mushroom-like structures. An examination of the influence of several divalent cations on the antibiofilm activity of EDTA indicates that magnesium, calcium, and iron protect P. aeruginosa biofilms against EDTA treatment. Our results are consistent with a mechanism whereby EDTA causes detachment and killing of biofilm cells.

* Corresponding author. Mailing address: Box 357242, Department of Microbiology, University of Washington School of Medicine, Seattle, WA 98195-7242. Phone: (206) 616-2881. Fax: (206) 616-2938. E-mail: epgreen{at}u.washington.edu.

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

Applied and Environmental Microbiology, March 2006, p. 2064-2069, Vol. 72, No. 3
0099-2240/06/$08.00+0     doi:10.1128/AEM.72.3.2064-2069.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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