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Journal of Antimicrobial Chemotherapy (2003) 52, 598-604
© 2003 The British Society for Antimicrobial Chemotherapy

Biofilm formation by Pseudomonas aeruginosa: role of the C4-HSL cell-to-cell signal and inhibition by azithromycin

Sabine Favre-Bonté*, Thilo Köhler and Christian Van Delden§

Department of Genetics and Microbiology, University of Geneva, CMU, 1, rue Michel-Servet, CH-1211 Geneva 4, Switzerland

Received 6 February 2003; returned 6 April 2003; revised 19 June 2003; accepted 30 June 2003

Objectives: In Pseudomonas aeruginosa, biofilm formation is controlled by a cell-to-cell signalling circuit relying on the secretion of 3-oxo-C12-HSL and C4-HSL. Previous studies suggested that C4-HSL plays no significant role in biofilm formation. However the wild-type PAO1 strain PAO-BI, used as a control in these studies is itself impaired in the production of C4-HSL. We wondered therefore whether the role of C4-HSL in biofilm formation might have been underestimated, and whether azithromycin inhibits biofilm formation by interfering with cell-to-cell signalling.

Methods: We used isogenic mutants of wild-type PAO1 strains PAO-BI and PT5 in a static biofilm model. Biofilm formation was quantified using Crystal Violet staining and exopolysaccharide measurements.

Results: Wild-type strain PAO-BI, as a result of its reduced C4-HSL secretion, produced 40% less biofilm compared with the wild-type PAO1 strain PT5. Using isogenic mutants of strain PT5 we have shown that whereas a lasI mutant (deficient in 3-oxo-C12-HSL) produced similar amounts of biofilm to the wild-type, a rhlI mutant (deficient in C4-HSL) produced 70% less biofilm. In the latter strain, biofilm formation could be restored by addition of exogenous C4-HSL. Azithromycin, known to reduce the production of both 3-oxo-C12-HSL and C4-HSL, inhibited biofilm formation of wild-type PT5 by 45%. This inhibition could be reversed by the addition of both cell-to-cell signals.

Conclusions: Our results indicate that C4-HSL also plays a significant role in biofilm formation. Furthermore, we demonstrate the potential of using cell-to-cell signalling blocking agents such as azithromycin to interfere with biofilm formation.

Keywords: quorum sensing, autoinducers, homoserine-lactones, 3-oxo-C12-HSL

* Present address. UMR CNRS 5557-Ecologie Microbienne, Université Claude Bernard, Lyon 1, France.

§ Corresponding author. Tel: +41-22-702-56-39; Fax: +41-22-702-57-02; E-mail: Christian.VanDelden{at}medecine.unige.ch


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