Journal
JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART A
Volume 70A, Issue 2, Pages 274-282Publisher
WILEY
DOI: 10.1002/jbm.a.30077
Keywords
biofilm; green fluorescent protein; confocal laser scanning microscopy; Escherichia coli; curli
Funding
- NIBIB NIH HHS [EB-00279] Funding Source: Medline
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We developed a novel method of evaluating biofilm architecture on a synthetic material using green fluorescent protein-expressing Escherichia coli and red fluorescence staining of exopolysaccharides. Confocal laser scanning microscopy observation revealed the time course of the change in the in situ three-dimensional structural features of biofilm on a polyurethane film without structural destruction: initially adhered cells are grown to form cellular aggregates and secrete exopolysaccharides. These cells were spottily distributed on the surface at an early incubation time but fused to form a vertically grown biofilm with incubation time. Fluorescence intensity, which is a measure of the number of cells, determined using a fluorometer and biofilm thickness determined from confocal laser scanning microscopy vertical images were found to be effective for quantification of time-dependent growth of biofilms. The curli (surface-located fibers specifically binding to fibronectin and laminin)-producing Escherichia coli strain, YMel, significantly proliferated on fibronectin-coated polyurethane, whereas the curli-deficient isogenic mutant, YMel-1, did not. The understanding of biofilm architecture in molecular and morphological events and new fluorescence microscopic techniques may help in the logical surface design of biomaterials with a high antibacterial potential. (C) 2004 Wiley Periodicals, Inc.
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