Bordetellae are gram-negative bacteria that colonize the respiratory tracts of animals and humans. We and others have recently shown that these bacteria are capable of living as sessile communities known as biofilms on a number of abiotic surfaces. During the biofilm mode of existence, bacteria produce one or more extracellular polymeric substances that function, in part, to hold the cells together and to a surface. There is little information on either the constituents of the biofilm matrix or the genetic basis of biofilm development by Bordetella spp. By utilizing immunoblot assays and by enzymatic hydrolysis using dispersin B (DspB), a glycosyl hydrolase that specifically cleaves the polysaccharide poly-␤-1,6-N-acetyl-D-glucosamine (poly-␤-1,6-GlcNAc), we provide evidence for the production of poly-␤-1,6-GlcNAc by various Bordetella species (Bordetella bronchiseptica, B. pertussis, and B. parapertussis) and its role in their biofilm development. We have investigated the role of a Bordetella locus, here designated bpsABCD, in biofilm formation. The bps (Bordetella polysaccharide) locus is homologous to several bacterial loci that are required for the production of poly-␤-1,6-GlcNAc and have been implicated in bacterial biofilm formation. By utilizing multiple microscopic techniques to analyze biofilm formation under both static and hydrodynamic conditions, we demonstrate that the bps locus, although not essential at the initial stages of biofilm formation, contributes to the stability and the maintenance of the complex architecture of Bordetella biofilms.In contrast to the planktonic or free swimming mode of existence in laboratory settings, bacteria predominantly form surface-attached communities known as biofilms in their natural habitats. Biofilms are commonly defined as highly structured communities of cells that are encased in a self-produced polymeric organic matrix (7). Biofilms are increasingly being recognized as important contributors to chronic bacterial diseases (8,19,48). Biofilms provide protection from both the innate and the adaptive components of the immune system, and bacteria in biofilms are extremely recalcitrant to antibiotic therapy and other antimicrobial agents (15,20,31,40).We are studying the members of the bacterial genus Bordetella as a model system in order to understand how bacteria adapt to cope with the selective pressures inside mammalian hosts. Given the demonstrated importance of biofilms in contributing to bacterial persistence, we are interested in characterizing the role that biofilms play in Bordetella physiology and the infectious cycle within hosts. Bordetellae are small, aerobic, gram-negative coccobacilli that colonize the respiratory tracts of humans and animals (42). Bordetella pertussis, the human pathogen, results in the disease known as whooping cough, while B. bronchiseptica mainly infects animals and causes a variety of respiratory diseases (42, 64). B. parapertussis strains can be divided into two genetically distinct types: those that infect humans, causing a...

The majority of Bordetella sp. virulence determinants are regulated by the BvgAS signal transduction system. BvgAS mediates the control of multiple phenotypic phases and a spectrum of gene expression profiles specific to each phase in response to incremental changes in the concentrations of environmental signals. Studies highlighting the critical role of this signaling circuitry in the Bordetella infectious cycle have focused on planktonically growing bacterial cells. It is becoming increasingly clear that the major mode of bacterial existence in the environment and within the body is a surface-attached state known as a biofilm. Biofilms are defined as consortia of sessile microorganisms that are embedded in a matrix. During routine growth of Bordetella under agitating conditions, we noticed the formation of a bacterial ring at the air-liquid interface of the culture tubes. We show here that this surface adherence property reflects the ability of these organisms to form biofilms. Our data demonstrate that the BvgAS locus regulates biofilm development in Bordetella. The results reported in this study suggest that the Bvg-mediated control in biofilm development is exerted at later time points after the initial attachment of bacteria to the different surfaces. Additionally, we show that these biofilms are highly tolerant of a number of antimicrobials, including the ones that are currently recommended for treatment of veterinary and human infections caused by Bordetella spp. Finally, we discuss the significance of the biofilm lifestyle mode as a potential contributor to persistent infections.

Two analogues of the marine natural product bromoageliferin have been synthesized and subsequently assayed for the ability to inhibit the formation of Pseudomonas aeruginosa biofilms. Both compounds inhibited the ability of Pseudomonas aeruginosa strains PAO1 and PA14 to form biofilms; however, they displayed differential toxicity to planktonic bacteria. The cis analogue, CAGE, was found to possess greater toxicity than the trans analogue, TAGE. These compounds have no structural homology to any known inhibitors of Pseudomonas aeruginosa biofilms.