A novel approach combining the Calgary Biofilm Device and Phenotype MicroArray for the characterization of the chemical sensitivity of bacterial biofilms

Abstract

A rapid method for screening the metabolic susceptibility of biofilms to toxic compounds was developed by combining the Calgary Biofilm Device (MBEC device) and Phenotype MicroArray (PM) technology. The method was developed using Pseudomonas alcaliphila 34, a Cr(VI)-hyper-resistant bacterium, as the test organism. P. alcaliphila produced a robust biofilm after incubation for 16 h, reaching the maximum value after incubation for 24 h (9.4 × 10⁶ ± 3.3 × 10⁶ CFU peg⁻¹). In order to detect the metabolic activity of cells in the biofilm, dye E (5×) and menadione sodium bisulphate (100 μM) were selected for redox detection chemistry, because they produced a high colorimetric yield in response to bacterial metabolism (340.4 ± 6.9 Omnilog Arbitrary Units). This combined approach, which avoids the limitations of traditional plate counts, was validated by testing the susceptibility of P. alcaliphila biofilm to 22 toxic compounds. For each compound the concentration level that significantly lowered the metabolic activity of the biofilm was identified. Chemical sensitivity analysis of the planktonic culture was also performed, allowing comparison of the metabolic susceptibility patterns of biofilm and planktonic cultures.

Keywords:

• Phenotype MicroArray
• Calgary Biofilm Device
• biofilm
• planktonic culture
• Pseudomonas
• chemical sensitivity analysis

Acknowledgments

This work was supported by the Italian Ministero dell'Istruzione, dell'Università e della Ricerca (MIUR) (PRIN, 2008). The authors thank Barry R. Bochner for the critical revision of the manuscript.