Extracellular DNA Required for Bacterial Biofilm Formation

Whitchurch, Cynthia B.; Tolker‐Nielsen, Tim; Ragas, Paula Cornelia; Mattick, John S.

doi:10.1126/science.295.5559.1487

Cited by 1,829 publications

(1,674 citation statements)

References 8 publications

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“…Commonly, EPS provides ϳ85% vol/vol of the space occupied by a biofilm (but this figure widely varies) and the remainder (ϳ15%) consists of bacteria. EPS is not simply made up of so-called exopolysaccharides (since it is much more complex) and includes heterogeneous combinations of polysaccharides, proteins, and minor amounts of lipids, nucleic acids, and other polymers (11,64,68). In fact, larger tangible biomaterials such as outer membrane vesicles, flagella, phages, pili, and debris from lysed cells are also present in variable amounts ( Fig.…”

Section: Resultsmentioning

confidence: 99%

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High-Resolution Visualization ofPseudomonas aeruginosaPAO1 Biofilms by Freeze-Substitution Transmission Electron Microscopy

Hunter

Beveridge

2005

J Bacteriol

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High-pressure freeze-substitution and transmission electron microscopy have been used for high-resolution imaging of the natural structure of a gram-negative biofilm. Unlike more conventional embedding techniques, this method confirms many of the observations seen by confocal microscopy but with finer structural detail. It further reveals that there is a structural complexity to biofilms at both the cellular and extracellular matrix levels that has not been seen before. Different domains of healthy and lysed cells exist randomly dispersed within a single biofilm as well as different structural organizations of exopolymers. Particulate matter is suspended within this network of fibers and appears to be an integral part of the exopolymeric substance (EPS). O-side chains extending from the outer membrane are integrated into EPS polymers so as to form a continuum. Together, the results support the concept of physical microenvironments within biofilms and show a complexity that was hitherto unknown.

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Section: Resultsmentioning

confidence: 99%

“…Accordingly, most staining of this PAO1 polymeric EPS must be due to the KDO moiety. It also must be remembered, though, that other highly acidic polymers may also be present, such as DNA (11,52,68).…”

mentioning

confidence: 99%

High-Resolution Visualization ofPseudomonas aeruginosaPAO1 Biofilms by Freeze-Substitution Transmission Electron Microscopy

Hunter

Beveridge

2005

J Bacteriol

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“…eDNA has been subsequently shown to contribute to biofilm formation by clinical P. aeruginosa isolates as well as by a variety of bacterial species, including Staphylococcus epidermidis, through analyses of biofilm formation by lysis-defective mutants and of DNA removal from the biofilm matrix (67,117,133,146,150). The contribution of eDNA to attachment and biofilm formation, however, appears to be temporal: experiments utilizing DNase I have suggested that cells in young PAO1 biofilms are held together by eDNA, whereas the cells in more-mature PAO1 biofilms are held together primarily by components other than eDNA (98,166). Subsequent microscopic investigations of mature P. aeruginosa biofilms stained with nucleic acid stains suggested a structural function of eDNA specific to the formation of mushroom-shaped microcolonies (5).…”

Section: Being Sticky Is the Keymentioning

confidence: 99%

Sticky Situations: Key Components That Control Bacterial Surface Attachment

Petrova

Sauer

2012

Journal of Bacteriology

319

251

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“…On the other hand, results using wild-type P. aeruginosa PAO1 and PA14 showed that alginate was not associated with biofilms (72) and that a second polysaccharide likely encoded by a separate polysaccharide biosynthetic gene cluster may provide the extracellular matrix material for those cells (18,19,29). In another report, using P. aeruginosa PAO1, DNA was suggested to be the extracellular matrix material, since the addition of DNase to biofilms dispersed the bacteria (64). Many bacterial extracellular polysaccharides, including alginate, contain negatively charged uronic acid residues.…”

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confidence: 98%

Calcium-Induced Virulence Factors Associated with the Extracellular Matrix of MucoidPseudomonas aeruginosaBiofilms

Sarkisova¹,

Patrauchan

Berglund³

et al. 2005

J Bacteriol

197

173

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Pseudomonas aeruginosa colonizes the pulmonary tissue of patients with cystic fibrosis (CF), leading to biofilm-associated infections. The pulmonary fluid of CF patients usually contains elevated concentrations of cations and may contain the P. aeruginosa redox-active pigment pyocyanin, which is known to disrupt calcium homeostasis of host cells. Since divalent cations are important bridging ions for bacterial polysaccharides and since they may play regulatory roles in bacterial gene expression, we investigated the effect of calcium ions on the extracellular matrix constituents of P. aeruginosa biofilms. For mucoid strain P. aeruginosa FRD1, calcium addition (1.0 and 10 mM as CaCl 2 ) resulted in biofilms that were at least 10-fold thicker than biofilms without added calcium. Scanning confocal laser microscopy showed increased spacing between cells for the thick biofilms, and Fourier transform infrared spectroscopy revealed that the material between cells is primarily alginate. An algD transcriptional reporter demonstrated that calcium addition caused an eightfold increase in alg gene expression in FRD1 biofilms. Calcium addition also resulted in increased amounts of three extracellular proteases (AprA, LasB, and PrpL). Immunoblots of the biofilm extracellular material established that AprA was harbored within the biofilm extracellular matrix. An aprA deletion mutation and a mutation in gene for a putative P. aeruginosa calmodulin-like protein did not significantly affect calcium-induced biofilm structure. Two-dimensional gel electrophoresis showed increased amounts of phenazine biosynthetic proteins in FRD1 biofilms and in calcium-amended planktonic cultures. Spectrochemical analyses showed that the calcium addition causes a three-to fivefold increase in pyocyanin production. These results demonstrate that calcium addition affects the structure and extracellular matrix composition of mucoid P. aeruginosa biofilms, through increased expression and stability of bacterial extracellular products. The calcium-induced extracellular matrix of mucoid P. aeruginosa consists primarily of the virulence factor alginate and also harbors extracellular proteases and perhaps pyocyanin, a biomolecule that may further disrupt cellular calcium levels.

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Extracellular DNA Required for Bacterial Biofilm Formation

Cited by 1,829 publications

References 8 publications

High-Resolution Visualization ofPseudomonas aeruginosaPAO1 Biofilms by Freeze-Substitution Transmission Electron Microscopy

High-Resolution Visualization ofPseudomonas aeruginosaPAO1 Biofilms by Freeze-Substitution Transmission Electron Microscopy

Sticky Situations: Key Components That Control Bacterial Surface Attachment

Calcium-Induced Virulence Factors Associated with the Extracellular Matrix of MucoidPseudomonas aeruginosaBiofilms

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