Exopolysaccharide Production Is Required for Development of Escherichia coli K-12 Biofilm Architecture

Abstract

Although exopolysaccharides (EPSs) are a large component of bacterial biofilms, their contribution to biofilm structure and function has been examined for only a few organisms. In each of these cases EPS has been shown to be required for cellular attachment to abiotic surfaces. Here, we undertook a genetic approach to examine the potential role of colanic acid, an EPS of Escherichia coli K-12, in biofilm formation. Strains either proficient or deficient in colanic acid production were grown and allowed to adhere to abiotic surfaces and were then examined both macroscopically and microscopically. Surprisingly, we found that colanic acid production is not required for surface attachment. Rather, colanic acid is critical for the formation of the complex three-dimensional structure and depth of E. coli biofilms.Bacterial biofilms have been described as sessile bacterial communities that live attached to each other and to surfaces (2,3,4,5,10). In natural settings, many bacterial species live predominantly in these communities, with a smaller portion of the bacterial population subsisting as free-swimming (planktonic) organisms (10). In addition to their abundance in natural environments, biofilms also impinge significantly upon our industrialized world. For example, bacterial biofilms can form on catheters and prostheses and thereby cause persistent, antibiotic-resistant infections (5, 12). Biofilms can also clog pipes (1) and contaminate food in industrial settings (21). However, biofilms can also have beneficial functions, for example, by acting as biocontrol agents by preventing fungal infections in certain plants (9). Given the preponderance of biofilm communities in nature as well as their medical and industrial impact, it is clearly important to understand the molecular mechanisms that govern both the formation and dissolution of these sessile communities.The three-dimensional architecture of a number of singlespecies bacterial biofilms has been previously described (5, 8). The two most generalizable features of these biofilms are microcolonies, composed of cells surrounded by large amounts of exopolysaccharide (EPS), and water-filled channels, which have been hypothesized to promote the influx of nutrients and the efflux of waste products.Previous work with Pseudomonas aeruginosa and with Escherichia coli has shown that EPS (alginate and colanic acid, respectively) synthesis is induced upon attachment of the bacteria to a surface (6,7,17). However, these results have not revealed the role(s) that EPS plays in biofilm formation. Studies with the gram-negative organisms Shewanella putrefaciens and Vibrio cholerae and the gram-positive organism Staphylococcus epidermidis revealed that EPS is required for initial attachment to surfaces (15, 20; D. Newman and R. Kolter, unpublished data). Here, we describe the role of EPS in E. coli biofilm formation and note that this role is dramatically different than that described for S. putrefaciens, V. cholerae, and S. epidermidis.Isolation of an E. coli strain defective in...

Genetic determinants of biofilm development of opaque and translucent Vibrio parahaemolyticus

“…We interpret the inclusion of the cys mutants in this class as the result of a requirement for activated sulphur in the biosynthesis of V. parahaemolyticus CPSA. Although the relative importance of CPSA in biofilm formation is consistent with observations in many organisms Danese et al, 2000;Sutherland, 2001), the fact that CPSA played a central role in both cell types was somewhat surprising in light of prior data demonstrating dramatic differences in the amount of CPSA produced by OP and TR strains (Enos-Berlage and McCarter, 2000). Perhaps TR strains produce sufficient CPSA to foster biofilm development and the additional CPSA produced by OP strains alters the properties and the architecture of the biofilm.…”

“…Thus, consistent with work in many organisms (e.g. Danese et al, 2000), CPS in V. parahaemolyticus appears to play an important role in adherence (measured by crystal violet), and establishment of three-dimensional biofilms (i.e. pellicle formation and microcolony/pillar formation in the submerged coverslip assay).…”

Elevated levels of σ S inhibit biofilm formation in Escherichia coli: a role for the Rcs phosphorelay

“…The activation of the Rcs phosphorelay results in an increase in the expression of the extracellular polysacchar-ide colanic acid (Gottesman et al, 1985). As the expression of colanic acid is required for the maturation of biofilms formed by E. coli, this suggests a positive role for the Rcs phosphorelay in the regulation of biofilm maturation (Danese et al, 2000b). On the other hand, activation of the Rcs phosphorelay has been shown to repress the expression of genes encoding surface adhesins, such as antigen 43 (Ag43) and curli, and it has been reported that overproduction of RcsB (a situation that mimics the activation of the Rcs phosphorelay) inhibits the expression of the flhDC operon, encoding the master regulators of flagella biosynthesis (Ferrières & Clarke, 2003;Francez-Charlot et al, 2003;Vianney et al, 2005).…”

“…Over the past few years, a role for the Rcs phosphorelay in biofilm formation by Escherichia coli has been proposed (Danese et al, 2000b;Ferrières & Clarke, 2003). The Rcs phosphorelay is composed of the sensor kinase, RcsC, the histidine-phosphotransfer (HPt)-containing protein RcsD and the cytoplasmic RcsB response regulator (Huang et al, 2006;.…”

The Pseudomonas aeruginosa oxidative stress regulator OxyR influences production of pyocyanin and rhamnolipids: protective role of pyocyanin

“…Positive cultures are surrounded by a dark blue halo. Overnight cultures were spotted and plates were incubated at 28 uC for 48 h (Danese et al, 2000).…”