scholarly journals Relevant Role of Fibronectin-Binding Proteins in Staphylococcus aureus Biofilm-Associated Foreign-Body Infections

2009 ◽  
Vol 77 (9) ◽  
pp. 3978-3991 ◽  
Author(s):  
Marta Vergara-Irigaray ◽  
Jaione Valle ◽  
Nekane Merino ◽  
Cristina Latasa ◽  
Begoña García ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Binding Proteins ◽  
Surface Protein ◽  
Multicopy Plasmid ◽  
Chronic Infections ◽  
Implanted Medical Devices ◽  
Relevant Role ◽  
Fibronectin Binding ◽  
Biofilm Development ◽  
Biofilm Matrix

ABSTRACT Staphylococcus aureus can establish chronic infections on implanted medical devices due to its capacity to form biofilms. Analysis of the factors that assemble cells into a biofilm has revealed the occurrence of strains that produce either a polysaccharide intercellular adhesin/poly-N-acetylglucosamine (PIA/PNAG) exopolysaccharide- or a protein-dependent biofilm. Examination of the influence of matrix nature on the biofilm capacities of embedded bacteria has remained elusive, because a natural strain that readily converts between a polysaccharide- and a protein-based biofilm has not been studied. Here, we have investigated the clinical methicillin (meticillin)-resistant Staphylococcus aureus strain 132, which is able to alternate between a proteinaceous and an exopolysaccharidic biofilm matrix, depending on environmental conditions. Systematic disruption of each member of the LPXTG surface protein family identified fibronectin-binding proteins (FnBPs) as components of a proteinaceous biofilm formed in Trypticase soy broth-glucose, whereas a PIA/PNAG-dependent biofilm was produced under osmotic stress conditions. The induction of FnBP levels due to a spontaneous agr deficiency present in strain 132 and the activation of a LexA-dependent SOS response or FnBP overexpression from a multicopy plasmid enhanced biofilm development, suggesting a direct relationship between the FnBP levels and the strength of the multicellular phenotype. Scanning electron microscopy revealed that cells growing in the FnBP-mediated biofilm formed highly dense aggregates without any detectable extracellular matrix, whereas cells in a PIA/PNAG-dependent biofilm were embedded in an abundant extracellular material. Finally, studies of the contribution of each type of biofilm matrix to subcutaneous catheter colonization revealed that an FnBP mutant displayed a significantly lower capacity to develop biofilm on implanted catheters than the isogenic PIA/PNAG-deficient mutant.

scholarly journals Protein A-Mediated Multicellular Behavior in Staphylococcus aureus

2008 ◽  
Vol 191 (3) ◽  
pp. 832-843 ◽  
Author(s):  
Nekane Merino ◽  
Alejandro Toledo-Arana ◽  
Marta Vergara-Irigaray ◽  
Jaione Valle ◽  
Cristina Solano ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Biofilm Formation ◽  
Protein A ◽  
Surface Proteins ◽  
Growth Media ◽  
Dependent Manner ◽  
Chronic Infections ◽  
The Matrix ◽  
Implanted Medical Devices ◽  
Biofilm Development

ABSTRACT The capacity of Staphylococcus aureus to form biofilms on host tissues and implanted medical devices is one of the major virulence traits underlying persistent and chronic infections. The matrix in which S. aureus cells are encased in a biofilm often consists of the polysaccharide intercellular adhesin (PIA) or poly-N-acetyl glucosamine (PNAG). However, surface proteins capable of promoting biofilm development in the absence of PIA/PNAG exopolysaccharide have been described. Here, we used two-dimensional nano-liquid chromatography and mass spectrometry to investigate the composition of a proteinaceous biofilm matrix and identified protein A (spa) as an essential component of the biofilm; protein A induced bacterial aggregation in liquid medium and biofilm formation under standing and flow conditions. Exogenous addition of synthetic protein A or supernatants containing secreted protein A to growth media induced biofilm development, indicating that protein A can promote biofilm development without being covalently anchored to the cell wall. Protein A-mediated biofilm formation was completely inhibited in a dose-dependent manner by addition of serum, purified immunoglobulin G, or anti-protein A-specific antibodies. A murine model of subcutaneous catheter infection unveiled a significant role for protein A in the development of biofilm-associated infections, as the amount of protein A-deficient bacteria recovered from the catheter was significantly lower than that of wild-type bacteria when both strains were used to coinfect the implanted medical device. Our results suggest a novel role for protein A complementary to its known capacity to interact with multiple immunologically important eukaryotic receptors.

scholarly journals A Novel Staphylococcus aureus Biofilm Phenotype Mediated by the Fibronectin-Binding Proteins, FnBPA and FnBPB

2008 ◽  
Vol 190 (11) ◽  
pp. 3835-3850 ◽  
Author(s):  
Eoghan O'Neill ◽  
Clarissa Pozzi ◽  
Patrick Houston ◽  
Hilary Humphreys ◽  
D. Ashley Robinson ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Biofilm Formation ◽  
Binding Proteins ◽  
Clinical Problem ◽  
Growth Conditions ◽  
Surface Proteins ◽  
Fibrinogen Binding ◽  
Fibronectin Binding ◽  
Mrsa Strains ◽  
Biofilm Development

ABSTRACT Device-associated infections involving biofilm remain a persistent clinical problem. We recently reported that four methicillin-resistant Staphylococcus aureus (MRSA) strains formed biofilm independently of the icaADBC-encoded exopolysaccharide. Here, we report that MRSA biofilm development was promoted under mildly acidic growth conditions triggered by the addition of glucose to the growth medium. Loss of sortase, which anchors LPXTG-containing proteins to peptidoglycan, reduced the MRSA biofilm phenotype. Furthermore introduction of mutations in fnbA and fnbB, which encode the LPXTG-anchored multifunctional fibrinogen and fibronectin-binding proteins, FnBPA and FnBPB, reduced biofilm formation by several MRSA strains. However, these mutations had no effect on biofilm formation by methicillin-sensitive S. aureus strains. FnBP-promoted biofilm occurred at the level of intercellular accumulation and not primary attachment. Mutation of fnbA or fnbB alone did not substantially affect biofilm, and expression of either gene alone from a complementing plasmid in fnbA fnbB mutants restored biofilm formation. FnBP-promoted biofilm was dependent on the integrity of SarA but not through effects on fnbA or fnbB transcription. Using plasmid constructs lacking regions of FnBPA to complement an fnbAB mutant revealed that the A domain alone and not the domain required for fibronectin binding could promote biofilm. Additionally, an A-domain N304A substitution that abolished fibrinogen binding did not affect biofilm. These data identify a novel S. aureus biofilm phenotype promoted by FnBPA and FnBPB which is apparently independent of the known ligand-binding activities of these multifunctional surface proteins.

scholarly journals Fibrinogen Activates the Capture of Human Plasminogen by Staphylococcal Fibronectin-Binding Proteins

mBio ◽  
2017 ◽  
Vol 8 (5) ◽  
Author(s):  
Philippe Herman-Bausier ◽  
Giampiero Pietrocola ◽  
Timothy J. Foster ◽  
Pietro Speziale ◽  
Yves F. Dufrêne
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Surface ◽  
Molecular Mechanism ◽  
Conformational Changes ◽  
Binding Proteins ◽  
Biological Function ◽  
Surface Protein ◽  
Activation Mechanism ◽  
Human Plasminogen ◽  
Fibronectin Binding

ABSTRACT Invasive bacterial pathogens can capture host plasminogen (Plg) and allow it to form plasmin. This process is of medical importance as surface-bound plasmin promotes bacterial spread by cleaving tissue components and favors immune evasion by degrading opsonins. In Staphylococcus aureus, Plg binding is in part mediated by cell surface fibronectin-binding proteins (FnBPs), but the underlying molecular mechanism is not known. Here, we use single-cell and single-molecule techniques to demonstrate that FnBPs capture Plg by a sophisticated activation mechanism involving fibrinogen (Fg), another ligand found in the blood. We show that while FnBPs bind to Plg through weak (∼200-pN) molecular bonds, direct interaction of the adhesins with Fg through the high-affinity dock, lock, and latch mechanism dramatically increases the strength of the FnBP-Plg bond (up to ∼2,000 pN). Our results point to a new model in which the binding of Fg triggers major conformational changes in the FnBP protein, resulting in the buried Plg-binding domains being projected and exposed away from the cell surface, thereby promoting strong interactions with Plg. This study demonstrated a previously unidentified role for a ligand-binding interaction by a staphylococcal cell surface protein, i.e., changing the protein orientation to activate a cryptic biological function. IMPORTANCE Staphylococcus aureus captures human plasminogen (Plg) via cell wall fibronectin-binding proteins (FnBPs), but the underlying molecular mechanism is not known. Here we show that the forces involved in the interaction between Plg and FnBPs on the S. aureus surface are weak. However, we discovered that binding of fibrinogen to FnBPs dramatically strengthens the FnBP-Plg bond, therefore revealing an unanticipated role for Fg in the capture of Plg by S. aureus. These experiments favor a model where Fg-induced conformational changes in FnBPs promote their interaction with Plg. This work uncovers a previously undescribed activation mechanism for a staphylococcal surface protein, whereby ligand-binding elicits a cryptic biological function. IMPORTANCE Staphylococcus aureus captures human plasminogen (Plg) via cell wall fibronectin-binding proteins (FnBPs), but the underlying molecular mechanism is not known. Here we show that the forces involved in the interaction between Plg and FnBPs on the S. aureus surface are weak. However, we discovered that binding of fibrinogen to FnBPs dramatically strengthens the FnBP-Plg bond, therefore revealing an unanticipated role for Fg in the capture of Plg by S. aureus. These experiments favor a model where Fg-induced conformational changes in FnBPs promote their interaction with Plg. This work uncovers a previously undescribed activation mechanism for a staphylococcal surface protein, whereby ligand-binding elicits a cryptic biological function.

scholarly journals Increased Virulence of a Fibronectin-Binding Protein Mutant of Staphylococcus aureus in a Rat Model of Pneumonia

2002 ◽  
Vol 70 (7) ◽  
pp. 3865-3873 ◽  
Author(s):  
Mary C. McElroy ◽  
David J. Cain ◽  
Christine Tyrrell ◽  
Timothy J. Foster ◽  
Christopher Haslett
Keyword(s):  
Staphylococcus Aureus ◽  
Epithelial Cells ◽  
Rat Model ◽  
Binding Proteins ◽  
Binding Protein ◽  
Alveolar Epithelial Cells ◽  
Alveolar Epithelial ◽  
Fibronectin Binding Protein ◽  
Fibronectin Binding

ABSTRACT Fibronectin-binding proteins mediate Staphylococcus aureus internalization into nonphagocytic cells in vitro. We have investigated whether fibronectin-binding proteins are virulence factors in the pathogenesis of pneumonia by using S. aureus strain 8325-4 and isogenic mutants in which fibronectin-binding proteins were either deleted (DU5883) or overexpressed [DU5883(pFnBPA4)]. We first demonstrated that fibronectin-binding proteins mediate S. aureus internalization into alveolar epithelial cells in vitro and that S. aureus internalization into alveolar epithelial cells requires actin rearrangement and protein kinase activity. Second, we established a rat model of S. aureus-induced pneumonia and measured lung injury and bacterial survival at 24 and 96 h postinoculation. S. aureus growth and the extent of lung injury were both increased in rats inoculated with the deletion mutant (DU5883) in comparison with rats inoculated with the wild-type (8325-4) and the fibronectin-binding protein-overexpressing strain DU5883(pFnBPA4) at 24 h postinfection. Morphological evaluation of infected lungs at the light and electron microscopic levels demonstrated that S. aureus was present within neutrophils from both 8325-4- and DU5883-inoculated lungs. Our data suggest that fibronectin-binding protein-mediated internalization into alveolar epithelial cells is not a virulence mechanism in a rat model of pneumonia. Instead, our data suggest that fibronectin-binding proteins decrease the virulence of S. aureus in pneumonia.

scholarly journals Increased expression of fibronectin-binding proteins by fluoroquinolone-resistant Staphylococcus aureus exposed to subinhibitory levels of ciprofloxacin.

1997 ◽  
Vol 41 (5) ◽  
pp. 906-913 ◽  
Author(s):  
C Bisognano ◽  
P E Vaudaux ◽  
D P Lew ◽  
E Y Ng ◽  
D C Hooper
Keyword(s):  
Staphylococcus Aureus ◽  
Bacterial Adhesion ◽  
Binding Proteins ◽  
Solid Phase ◽  
Fluoroquinolone Resistance ◽  
Adhesion Assay ◽  
Topoisomerase Iv ◽  
Resistance Determinants ◽  
Fibronectin Binding ◽  
Resistant Mutants

Bacterial adhesion, which plays an important role in Staphylococcus aureus colonization and infection, may be altered by the presence of antibiotics or/and antibiotic resistance determinants. This study evaluated the effect of fluoroquinolone resistance determinants on S. aureus adhesion to solid-phase fibronectin, which is specifically mediated by two surface-located fibronectin-binding proteins. Five isogenic mutants, derived from strain NCTC 8325 and expressing various levels of quinolone resistance, were tested in an in vitro bacterial adhesion assay with polymethylmethacrylate coverslips coated with increasing amounts of fibronectin. These strains contained single or combined mutations in the three major loci contributing to fluoroquinolone resistance, namely, grlA, gyrA, and flqB, which code for altered topoisomerase IV, DNA gyrase, and increased norA-mediated efflux of fluoroquinolones, respectively. Adhesion characteristics of the different quinolone-resistant mutants grown in the absence of fluoroquinolone showed only minor differences from those of parental strains. However, more important changes in adhesion were exhibited by mutants highly resistant to quinolones following their exponential growth in the presence of one-quarter MIC of ciprofloxacin. Increased bacterial adhesion of the highly quinolone-resistant mutants, which contained combined mutations in grlA and gyrA, was associated with and explained by the overexpression of their fibronectin-binding proteins as assessed by Western ligand affinity blotting. These findings contradict the notion that subinhibitory concentrations of antibiotics generally decrease the expression of virulence factors by S. aureus. Perhaps the increased adhesion of S. aureus strains highly resistant to fluoroquinolones contributes in part to that emergence in clinical settings.

scholarly journals Genetic Evidence for an Alternative Citrate-Dependent Biofilm Formation Pathway in Staphylococcus aureus That Is Dependent on Fibronectin Binding Proteins and the GraRS Two-Component Regulatory System

2008 ◽  
Vol 76 (6) ◽  
pp. 2469-2477 ◽  
Author(s):  
Robert M. Q. Shanks ◽  
Michael A. Meehl ◽  
Kimberly M. Brothers ◽  
Raquel M. Martinez ◽  
Niles P. Donegan ◽  
...  
Keyword(s):  
Gene Expression ◽  
Staphylococcus Aureus ◽  
Biofilm Formation ◽  
Binding Proteins ◽  
Regulatory System ◽  
Tca Cycle ◽  
Formation Pathway ◽  
Two Component ◽  
Fibronectin Binding ◽  
Tca Cycle Intermediates

ABSTRACT We reported previously that low concentrations of sodium citrate strongly promote biofilm formation by Staphylococcus aureus laboratory strains and clinical isolates. Here, we show that citrate promotes biofilm formation via stimulating both cell-to-surface and cell-to-cell interactions. Citrate-stimulated biofilm formation is independent of the ica locus, and in fact, citrate represses polysaccharide adhesin production. We show that fibronectin binding proteins FnbA and FnbB and the global regulator SarA, which positively regulates fnbA and fnbB gene expression, are required for citrate's positive effects on biofilm formation, and citrate also stimulates fnbA and fnbB gene expression. Biofilm formation is also stimulated by several other tricarboxylic acid (TCA) cycle intermediates in an FnbA-dependent fashion. While aconitase contributes to biofilm formation in the absence of TCA cycle intermediates, it is not required for biofilm stimulation by these compounds. Furthermore, the GraRS two-component regulator and the GraRS-regulated efflux pump VraFG, identified for their roles in intermediate vancomycin resistance, are required for citrate-stimulated cell-to-cell interactions, but the GraRS regulatory system does not impact the expression of the fnbA and fnbB genes. Our data suggest that distinct genetic factors are required for the early steps in citrate-stimulated biofilm formation. Given the role of FnbA/FnbB and SarA in virulence in vivo and the lack of a role for ica-mediated biofilm formation in S. aureus catheter models of infection, we propose that the citrate-stimulated biofilm formation pathway may represent a clinically relevant pathway for the formation of these bacterial communities on medical implants.

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