Binding of the fibronectin-binding protein FnBPA fromStaphylococcus aureus to the human protein fibronectin has previously been implicated in the development of infective endocarditis, specifically in the processes of platelet activation and invasion of the endothelium. We recently proposed a model for binding of fibronectin to FnBPA in which the bacterial protein contains 11 potential binding sites (FnBPA-1 to FnBPA-11), each composed of motifs that bind to consecutive fibronectin type 1 modules in the N-terminal domain of fibronectin. Here we show that six of the 11 sites bind with dissociation constants in the nanomolar range; other sites bind more weakly. The high affinity binding sites include FnBPA-1, the sequence of which had previously been thought to be encompassed by the fibrinogen-binding A domain of FnBPA. Both the number and sequence conservation of the type-1 module binding motifs appears to be important for high affinity binding. The in vivo relevance of the in vitro binding studies is confirmed by the presence of antibodies in patients with S. aureus infections that specifically recognize complexes of these six high affinity repeats with fibronectin.Staphylococcus aureus is one of the most important bacterial pathogens to affect humans. Clinical manifestations of infection range from superficial skin infections (1) to life-threatening conditions, such as endocarditis (2, 3) and difficult to treat infections of the bones and joints (4, 5). The increasing virulence and antibiotic resistance exhibited by this major source of both community and hospital-acquired infection presents an urgent challenge. Furthering our understanding of the mechanism by which staphylococcal pathogenesis occurs is thus imperative for the development of novel therapeutic and preventative strategies.Since attachment to host tissue is a critical early step in infection, one particular group of targets for intervention is the microbial surface components recognizing adhesive matrix molecules (MSCRAMM) 4 family of surface-expressed adhesins (6, 7). These proteins exploit extracellular matrix proteins, such as fibronectin (Fn), using them as a bridge between the bacterial cell surface and host cell receptors that effect downstream signaling (8, 9). Although classically regarded as an exclusively extracellular pathogen, S. aureus has been shown to adhere to and invade several host cell types (10 -14), and the Fn-binding subfamily of MSCRAMMs (FnBPs) appears to be involved in this process (15). There is an emerging view that S. aureus can exist intracellularly, hijacking and invading host cells to establish persistence (16). Conceivably, this mechanism could facilitate rapid and effective bloodstream dissemination while allowing the bacterium to evade antibiotics and host immune surveillance, an apposite theory given the prevalence of bacterial metastasis (3) and infection relapse in staphylococcal disease (17).Fn is a large glycoprotein present in a soluble form in human plasma and other body fluids and in an insoluble form in...

Staphylococcus aureus is responsible for a wide range of infections, including soft tissue infections and potentially fatal bacteremias. The primary niche for S. aureus in humans is the nares, and nasal carriage is a documented risk factor for staphylococcal infection. Previous studies with rodent models of nasal colonization have implicated capsule and teichoic acid as staphylococcal surface factors that promote colonization. In this study, a mouse model of nasal colonization was utilized to demonstrate that S. aureus mutants that lack clumping factor A, collagen binding protein, Staphylococcus aureus causes a diverse spectrum of severe infections in humans, including bacteremia, endocarditis, and osteomyelitis, as well as skin and soft tissue infections. Notorious for decades as a major source of nosocomial infections, S. aureus has recently taken on a new role in causing an escalating number of community-acquired infections. To offset the problems of antibiotic-resistant S. aureus strains, preventive measures (e.g., immunization) should be explored as a complement to existing therapeutic approaches aimed at controlling this bacterial pathogen.Humans are a reservoir for S. aureus, and the nose is the principal site of staphylococcal colonization. Approximately 20% of people persistently carry S. aureus in the anterior nares, ϳ60% are intermittent carriers, and ϳ20% are noncarriers (19). Nasal carriage is a known risk factor for staphylococcal infection in a number of clinical settings (51). Certain patient populations that show higher rates of S. aureus nasal colonization have an increased risk of staphylococcal infection. These populations include patients with diabetes, eczema, and human immunodeficiency virus infection, individuals receiving continuous ambulatory peritoneal dialysis or hemodialysis, and injection drug users (19). Moreover, patients in hospitals or individuals living in crowded conditions often show higherthan-normal rates of S. aureus nasal colonization. The source of ϳ80% of S. aureus bacteremias is endogenous since infecting bacteria have been shown by genotypic analysis to be identical to organisms recovered from the nasal mucosa (48, 53). These observations support an approach in which systemic S. aureus infections are prevented by eliminating or reducing nasal carriage.One approach commonly used to reduce S. aureus carriage in individuals at risk for staphylococcal infection involves topical treatment with a nasal ointment containing the antibiotic mupirocin. Eradication of nasal carriage with topical mupirocin has been correlated with a reduction in the incidence of S. aureus infection in some patient populations (20, 45), but not in others (40,54). Whereas mupirocin is effective in decolonizing nasal carriers, recolonization often occurs from extranasal carriage sites (52). Of further concern is the emergence of mupirocin resistance in S. aureus (31, 46). The utility of

SummaryThe fibronectin binding protein, FnBPA, is a multifunctional microbial surface component recognizing adhesive matrix molecule (MSCRAMM) that promotes bacterial adherence to immobilized fibrinogen and elastin via the N-terminal A domain. The binding site for fibrinogen and elastin was localized to subdomains N2N3. A three-dimensional structural model of FnBPA was created based on the known crystal structure of the domains N2N3 of clumping factor A (ClfA). The role of individual residues in the putative ligand binding trench was examined by testing the affinity of mutants for fibrinogen and elastin. Two residues (N304 and F306) were crucial for binding both ligands and are in the equivalent positions to residues known to be important for fibrinogen binding by ClfA. A peptide comprising the C-terminus of the g-chain of fibrinogen and a monoclonal anti-rAFnBPA antibody were potent inhibitors of the FnBPA-elastin interaction. This suggests that FnBPA binds to fibrinogen and elastin in a similar manner. Amino acid sequence divergence of 26.5% occurred between the A domains of FnBPA from strains 8325-4 and P1. Most variant residues were predicted to be located on the surface of domains N2N3 while few occurred in the putative ligand binding trench and the latching peptide explaining limited immunocross reactivity while ligand binding activity is conserved.

Staphylococcus epidermidis is a ubiquitous human skin commensal that has emerged as a major cause of foreign-body infections. Eleven genes encoding putative cell-wall-anchored proteins were identified by computer analysis of the publicly available S. epidermidis unfinished genomic sequence. Four genes encode previously described proteins (Aap, Bhp, SdrF and SdrG), while the remaining seven have not been characterized. Analysis of primary sequences of the Staphylococcus epidermidis surface (Ses) proteins indicates that they have a structural organization similar to the previously described cell-wall-anchored proteins from S. aureus and other Gram-positive cocci. However, not all of the Ses proteins are direct homologues of the S. aureus proteins. Secondary and tertiary structure predictions suggest that most of the Ses proteins are composed of several contiguous subdomains, and that the majority of these predicted subdomains are folded into β-rich structures. PCR analysis indicates that certain genes may be found more frequently in disease isolates compared to strains isolated from healthy skin. Patients recovering from S. epidermidis infections had higher antibody titres against some Ses proteins, implying that these proteins are expressed during human infection. Western blot analyses of early-logarithmic and late-stationary in vitro cultures suggest that different regulatory mechanisms control the expression of the Ses proteins.

Osteointegrative dental implants are widely used in implantology for their well-known excellent performance once implanted in the host. Remarkable bacterial colonization along the transgingival region may result in a progressive loss of adhesion at gum-implant interface and an increase of the bone area exposed to pathogens. This phenomenon may negatively effect the osteointegration process and cause, in the most severe cases, implant failure. The presence of bacteria at implant site affect the growth of new bone tissue and consequently, the achievement of a mechanically stable bone-implant interface, key parameters for a suitable implant osteointegration. In the present work, a novel surface treatment has been developed and optimized in order to convert the amorphous titanium oxide in a crystalline layer enriched in anatase capable of providing not only antibacterial properties but also of stimulating the precipitation of apatite when placed in simulated body fluid. The collected data have shown that the tested treatment results in a crystalline anatase-type titanium oxide layer able to provide a remarkable decrease in bacterial attachment without negatively effecting cell metabolic activity. In conclusion, the surface modification treatment analyzed in the present study might be an elegant way to reduce the risk of bacterial adhesion and increase the lifetime of the transgingival component in the osteointegrated dental implant.