What's on the Outside Matters: The Role of the Extracellular Polymeric Substance of Gram-negative Biofilms in Evading Host Immunity and as a Target for Therapeutic Intervention

Gunn, John S.; Bakaletz, Lauren O.; Wozniak, Daniel J.

doi:10.1074/jbc.r115.707547

Cited by 146 publications

(134 citation statements)

References 102 publications

(98 reference statements)

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“…The primary function of the matrix in human infections is protection of the bacterial community against the hazards of external (e.g., antibiotics) and internal (e.g., innate immune system) factors (6). While these general functions are associated with most microbial biofilms, the individual ECM components often possess unique properties for the bacterial community and with regard to the host (7).…”

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confidence: 99%

“…Additionally, biofilm formation on and invasion of the gallbladder epithelium aid in the establishment and maintenance of carriage (11). Salmonella biofilm ECM components have been determined and include proteins (curli fimbriae and BapA), polysaccharides (extracellular DNA, cellulose, O-antigen capsule, colanic acid [in nontyphoidal serovars], and Vi antigen [ViAg] [in serovars Typhi, Dublin, and Paratyphi C]) (7,(13)(14)(15)(16)(17). Individual mutations in these ECM factors have been tested by various groups for their roles in biofilm formation, and some have also been examined for phenotypes in vertebrate models of infection (18)(19)(20)(21)(22)(23)(24).…”

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Salmonella Extracellular Matrix Components Influence Biofilm Formation and Gallbladder Colonization

et al. 2016

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Salmonella enterica serovar Typhi, the causative agent of typhoid fever in humans, forms biofilms encapsulated by an extracellular matrix (ECM). Biofilms facilitate colonization and persistent infection in gallbladders of humans and mouse models of chronic carriage. Individual roles of matrix components have not been completely elucidated in vitro or in vivo. To examine individual functions, strains of Salmonella enterica serovar Typhimurium, the murine model of S. Typhi, in which various ECM genes were deleted or added, were created to examine biofilm formation, colonization, and persistence in the gallbladder. Studies show that curli contributes most significantly to biofilm formation. Expression of Vi antigen decreased biofilm formation in vitro and virulence and bacterial survival in vivo without altering the examined gallbladder pro-or anti-inflammatory cytokines. Oppositely, loss of all ECM components (⌬wcaM ⌬csgA ⌬yihO ⌬bcsE) increased virulence and bacterial survival in vivo and reduced gallbladder interleukin-10 (IL-10) levels. Colanic acid and curli mutants had the largest defects in biofilm-forming ability and contributed most significantly to the virulence increase of the ⌬wcaM ⌬csgA ⌬yihO ⌬bcsE mutant strain. While the ⌬wcaM ⌬csgA ⌬yihO ⌬bcsE mutant was not altered in resistance to complement or growth in macrophages, it attached and invaded macrophages better than the wild-type (WT) strain. These data suggest that ECM components have various levels of importance in biofilm formation and gallbladder colonization and that the ECM diminishes disseminated disease in our model, perhaps by reducing cell attachment/invasion and dampening inflammation by maintaining/inducing IL-10 production. Understanding how ECM components aid acute disease and persistence could lead to improvements in therapeutic treatment of typhoid fever patients.

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Salmonella Extracellular Matrix Components Influence Biofilm Formation and Gallbladder Colonization

et al. 2016

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Here we show that thread-like eDNA fill the space between bacteria throughout Pseudomonas aeruginosa prototypical strain PA01 biofilm (Figure 1).

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confidence: 87%

“…Extracellular polymeric substances (EPS) are biofilm self-produced matrices that provide a protective shed for biofilm bacteria [1]. eDNA represents an important structural and functional component of EPS.…”

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STEM Observation of eDNA as a Dominant Component of EPS in Pseudomonas aeruginosa Biofilm

et al. 2018

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Extracellular polymeric substances (EPS) are biofilm self-produced matrices that provide a protective shed for biofilm bacteria [1]. eDNA represents an important structural and functional component of EPS.Here we show that thread-like eDNA fill the space between bacteria throughout Pseudomonas aeruginosa prototypical strain PA01 biofilm (Figure 1).Biofilm structures have been investigated by diverse microscopic methods, including fluorescence light microscopy, atomic force microscopy, transmission electron microscopy, and scanning electron microscopy [2][3][4][5]. Among them, confocal laser scanning microscopy is a powerful tool often used to study biofilm spatial structures. However, the resolution of confocal laser scanning microscopy limits its application to ultrastructural study of EPS. Most observation of eDNA and other EPS in biofilm are limited by resolution. In this work scanning transmission electron microscopy (STEM) is applied to the structural study of in vitro P. aeruginosa PA01 biofilm.The biofilms were cultured on polycarbonate membrane in agar plate at 37°C for 48 hours. Biofilm discs were chemically fixed with 2.5% glutaraldehyde and 2% paraformaldehyde in 0.15M cacodylate buffer, and then en bloc stained with 2% reduced osmium tetroxide. After dehydration and infiltration, samples were embedded in 100% durcupan resin and incubated at 65°C for 2 days. The resin embedded biofilm discs were processed to 90nm ultra-fine sections by using a Reichert-Jung Ultracut E ultramicrotome (Leica, Wetzlar, Germany). The thin sections were picked up with a loop and put on 400 mesh copper grids. The grids were air dried and then coated with amorphous carbon (t=3nm) on both sides. Electron microscopy data were collected in STEM mode on a Tecnai F20 S/TEM (Thermo Fisher Scientific, Hillsboro) operating at 200kV.In P. aeruginosa PA01biofilm, eDNA was reported as the major component of EPS [6]. Our results showed that thread-like EPS presented throughout the PA01 biofilm, with especially large concentrations in the region close to the culture base (Figure 1). Recently, Turnbull et al. reported that explosive lysis of P. aeruginosa contributed significantly to the eDNA contents of biofilm [7]. Our images also indicated the thread-like structures had morphological similarity to the bacterial internal content (Figure 1), suggesting that the thread-like structures are DNA.P. aeruginosa biofilm is frequently associated with chronic wound infection. Using STEM imaging techniques, we were able to gain an insight into the structures of P. aeruginosa PA01 biofilm. The structure provided the biofilm architecture for better understanding the biofilm EPS organization, which is likely to serve as structure basis for further therapeutic intervention [8].

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“…eDNA provides structural stability to the matrix, acts as a sink for antimicrobial peptides, protects resident bacteria from the host immune response, provides a source of "common goods" for the resident bacteria, acts as a universal structural material conducive to microbial community architecture, and facilitates the uptake of genetic material between bacterial species via a process known as horizontal gene transfer (5,(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). Members of the DNABII family of DNA-binding proteins, integration host factor (IHF) and histone-like protein (HU), are known for their strong structural influences on DNA intracellularly (18-24), and are also critical to the stability of the lattice-like 3D DNA structure found in biofilm matrices extracellularly (13,(25)(26)(27)(28)(29)(30)(31). To date, the presence of bacterial eDNA within biofilms has been attributed to release via various mechanisms, including cell lysis (autolysis or phagemediated) (12,32,33), or active secretion through type IV secretion systems (T4SSs) (34,35).…”

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NontypeableHaemophilus influenzaereleases DNA and DNABII proteins via a T4SS-like complex and ComE of the type IV pilus machinery

Jurcisek

Brockman

Novotny

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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Biofilms formed by nontypeable Haemophilus influenzae (NTHI) are central to the chronicity, recurrence, and resistance to treatment of multiple human respiratory tract diseases including otitis media, chronic rhinosinusitis, and exacerbations of both cystic fibrosis and chronic obstructive pulmonary disease. Extracellular DNA (eDNA) and associated DNABII proteins are essential to the overall architecture and structural integrity of biofilms formed by NTHI and all other bacterial pathogens tested to date. Although cell lysis and outermembrane vesicle extrusion are possible means by which these canonically intracellular components might be released into the extracellular environment for incorporation into the biofilm matrix, we hypothesized that NTHI additionally used a mechanism of active DNA release. Herein, we describe a mechanism whereby DNA and associated DNABII proteins transit from the bacterial cytoplasm to the periplasm via an inner-membrane pore complex (TraC and TraG) with homology to type IV secretion-like systems. These components exit the bacterial cell through the ComE pore through which the NTHI type IV pilus is expressed. The described mechanism is independent of explosive cell lysis or cell death, and the release of DNA is confined to a discrete subpolar location, which suggests a novel form of DNA release from viable NTHI. Identification of the mechanisms and determination of the kinetics by which critical biofilm matrix-stabilizing components are released will aid in the design of novel biofilmtargeted therapeutic and preventative strategies for diseases caused by NTHI and many other human pathogens known to integrate eDNA and DNABII proteins into their biofilm matrix.eDNA | Tra | secretin | IHF | HU B iofilms contribute substantially to the chronic and recurrent nature of many bacterial diseases of humans and animals, including those of the airway, urogenital tract, skin, and oral cavity (1-3). Bacteria within a biofilm are protected from environmental stresses by a self-produced extracellular matrix, called the extrapolymeric substance (EPS), whose composition varies greatly depending upon the microbes that produce it and the environment in which it is formed. Despite vast compositional variability, the EPS is typically composed of a complex mixture of lipopolysaccharides, proteins, lipids, glycolipids, and nucleic acids (4). Extracellular DNA (eDNA) is a major constituent of the EPS formed by multiple human pathogens (5-7) and serves diverse roles within the bacterial biofilm. eDNA provides structural stability to the matrix, acts as a sink for antimicrobial peptides, protects resident bacteria from the host immune response, provides a source of "common goods" for the resident bacteria, acts as a universal structural material conducive to microbial community architecture, and facilitates the uptake of genetic material between bacterial species via a process known as horizontal gene transfer (5,(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). Members of the DNABII family of DNA-binding proteins, integ...

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What's on the Outside Matters: The Role of the Extracellular Polymeric Substance of Gram-negative Biofilms in Evading Host Immunity and as a Target for Therapeutic Intervention

Cited by 146 publications

References 102 publications

Salmonella Extracellular Matrix Components Influence Biofilm Formation and Gallbladder Colonization

Salmonella Extracellular Matrix Components Influence Biofilm Formation and Gallbladder Colonization

STEM Observation of eDNA as a Dominant Component of EPS in Pseudomonas aeruginosa Biofilm

NontypeableHaemophilus influenzaereleases DNA and DNABII proteins via a T4SS-like complex and ComE of the type IV pilus machinery

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