Influence of Culture Heterogeneity in Cell Surface Charge on Adhesion and Biofilm Formation by
            <i>Enterococcus faecalis</i>

Merode, Annet E. J. van; Mei, Henny C. van der; Busscher, Henk J.; Krom, Bastiaan P.

doi:10.1128/jb.188.7.2421-2426.2006

Cited by 94 publications

(68 citation statements)

References 30 publications

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“…35,36 As an example, in preliminary experiments we have also observed the self-organising process with mesenchymal stem cells (data not shown). Since a washing step might selectively inuence the heterogeneity of surface adhesion, [37][38][39] our assay can also be applied to weakly adherent cells that need more time to settle down. Having to wait several hours for selforganization might seem to be incompatible with highthroughput studies, but commonly used approaches also require 2 to 15 h to allow cells to adhere on the substrate.…”

Section: Resultsmentioning

confidence: 99%

Cellular self-organization on micro-structured surfaces

2014

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Micro-patterned surfaces are frequently used in high-throughput single-cell studies, as they allow one to image isolated cells in defined geometries. Commonly, cells are seeded in excess onto the entire chip, and non-adherent cells are removed from the unpatterned sectors by rinsing. Here, we report on the phenomenon of cellular self-organization, which allows for autonomous positioning of cells on micropatterned surfaces over time. We prepared substrates with a regular lattice of protein-coated adhesion sites surrounded by PLL-g-PEG passivated areas, and studied the time course of cell ordering. After seeding, cells randomly migrate over the passivated surface until they find and permanently attach to adhesion sites. Efficient cellular self-organization was observed for three commonly used cell lines (HuH7, A549, and MDA-MB-436), with occupancy levels typically reaching 40-60% after 3-5 h. The time required for sorting was found to increase with increasing distance between adhesion sites, and is well described by the time-to-capture in a random-search model. Our approach thus paves the way for automated filling of cell arrays, enabling high-throughput single-cell analysis of cell samples without losses.

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Section: Resultsmentioning

confidence: 99%

Cellular self-organization on micro-structured surfaces

2014

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“…Since Atn is important for cellular growth as well as cell lysis, the adherence defect observed after atn deletion could be the result of increased chaining of the mutants (9, 57), leading to a reduction in binding sites or an increase in cell surface charges, causing a lower cell deposition rate. Repulsive interactions between abiotic surfaces and bacteria, as well as between adhered cells and incoming cells, due to surface charges were shown to influence the initial adhesion of bacteria to substrata (73). Alternatively, defects in autolytic processes may result in modification of the cell wall structure and mislocalization of surface proteins required for attachment during biofilm formation.…”

Section: Discussionmentioning

confidence: 99%

Contribution of Autolysin and Sortase A during Enterococcus faecalis DNA-Dependent Biofilm Development

et al. 2009

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Biofilm production is a major attribute of Enterococcus faecalis clinical isolates. Although some factors, such as sortases, autolysin, and extracellular DNA (eDNA), have been associated with E. faecalis biofilm production, the mechanisms underlying the contributions of these factors to this process have not been completely elucidated yet. In this study we define important roles for the major E. faecalis autolysin (Atn), eDNA, and sortase A (SrtA) during the developmental stages of biofilm formation under static and hydrodynamic conditions. Deletion of srtA affects the attachment stage and results in a deficiency in biofilm production. Atn-deficient mutants are delayed in biofilm development due to defects in primary adherence and DNA release, which we show to be particularly important during the accumulative phase for maturation and architectural stability of biofilms. Confocal laser scanning and freeze-dry electron microscopy of biofilms grown under hydrodynamic conditions revealed that E. faecalis produces a DNase I-sensitive fibrous network, which is important for biofilm stability and is absent in atn-deficient mutant biofilms. This study establishes the stage-specific requirements for SrtA and Atn and demonstrates a role for Atn in the pathway leading to DNA release during biofilm development in E. faecalis.Biofilms are bacterial communities encased within an extracellular matrix composed of carbohydrates, proteins, and nucleic acids (10). They are an ideal environment for exchange of genetic materials, such as genes encoding virulence factors and antibiotic resistance determinants, among bacteria within a community (61, 76), while allowing the flow of water and nutrients, as well as ions and various small molecules, to bacteria within the community (8) and providing a protective shield against antibiotics, antimicrobial substances, and phagocytosis (33, 80). Development of a biofilm is a complex multistage process. It is initiated by primary adhesion of the bacteria to a substratum, which is followed by the formation of microcolonies and production of an exopolysaccharide matrix, and it finally culminates with the formation of a three-dimensional (3D) multicellular mature structure (48). Bacterial biofilms are important medically because they have been associated with the pathogenesis of chronic and device-related persistent infections, such as cystic fibrosis, urinary tract infections, and endocarditis (11).Enterococcus faecalis is a gram-positive bacterium that is a commensal of the gastrointestinal tract of healthy individuals. However, it is also an important opportunistic pathogen that is responsible for a wide variety of nosocomial infections, including endocarditis, urinary tract infections, and bacteremia (14,17,23,44). E. faecalis accounts for approximately 65 to 80% of all enterococcal nosocomial infections (16). The ability of E. faecalis to adhere to and develop biofilms on medical devices, such as intravascular and urinary catheters (25,26,36,37,72), is thought to contribute to its pathogenes...

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“…Furthermore, for both E. faecium and E. faecalis, isolates were found that carried the esp gene but that failed to produce biofilms. Esp-independent biofilm formation by E. faecalis is probably mediated by multiple additional factors, like GelE (12,16,25), BopD (13), the fsr locus (12), the bee locus (37), and heterogeneity in surface charge (43). The observations that biofilms produced by E. faecalis are much thicker than those produced by E. faecium and that the biofilm development of E. faecalis is much more sensitive to the growth medium used (29) may indicate that in E. faecalis, additional factors aside from Esp are involved.…”

Section: Discussionmentioning

confidence: 99%

“…In E. faecalis, several other factors have been associated with biofilm development, like the sugar-binding transcriptional regulator BopD (13), the quorum-sensing locus fsr (12), heterogeneity in surface charge (43), the bee locus (37), and the secreted metalloprotease GelE (12,16,25). Yet absence of a correlation between gelatinase and biofilm development has also been found (6,21,43).…”

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confidence: 99%

Growth Condition-Dependent Esp Expression by Enterococcus faecium Affects Initial Adherence and Biofilm Formation

et al. 2007

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A genetic subpopulation of Enterococcus faecium, called clonal complex 17 (CC-17), is strongly associated with hospital outbreaks and invasive infections. Most CC-17 strains contain a putative pathogenicity island encoding the E. faecium variant of enterococcal surface protein (Esp). Western blotting, flow cytometric analyses, and electron microscopy showed that Esp is expressed and exposed on the surface of E. faecium, though Esp expression and surface exposure are highly varied among different strains. Furthermore, Esp expression depends on growth conditions like temperature and anaerobioses. When grown at 37°C, five of six esp-positive E. faecium strains showed significantly increased levels of surface-exposed Esp compared to bacteria grown at 21°C, which was confirmed at the transcriptional level by real-time PCR. In addition, a significant increase in surface-exposed Esp was found in half of these strains when grown at 37°C under anaerobic conditions compared to the level in bacteria grown under aerobic conditions. Finally, amounts of surface-exposed Esp correlated with initial adherence to polystyrene (R 2 ‫؍‬ 0.7146) and biofilm formation (R 2 ‫؍‬ 0.7535). Polystyrene adherence was competitively inhibited by soluble recombinant N-terminal Esp. This study demonstrates that Esp expression on the surface of E. faecium (i) varies consistently between strains, (ii) is growth condition dependent, and (iii) is quantitatively correlated with initial adherence and biofilm formation. These data indicate that E. faecium senses and responds to changing environmental conditions, which might play a role in the early stages of infection when bacteria transit from oxygen-rich conditions at room temperature to anaerobic conditions at body temperature. In addition, variation of surface exposure may explain the contrasting findings reported on the role of Esp in biofilm formation.

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Influence of Culture Heterogeneity in Cell Surface Charge on Adhesion and Biofilm Formation by Enterococcus faecalis

Cited by 94 publications

References 30 publications

Cellular self-organization on micro-structured surfaces

Cellular self-organization on micro-structured surfaces

Contribution of Autolysin and Sortase A during Enterococcus faecalis DNA-Dependent Biofilm Development

Growth Condition-Dependent Esp Expression by Enterococcus faecium Affects Initial Adherence and Biofilm Formation

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