The intercellular adhesin involved in biofilm accumulation of Staphylococcus epidermidis is a linear beta-1,6-linked glucosaminoglycan: purification and structural analysis

Abstract

The primary attachment to polymer surfaces followed by accumulation in multilayered cell clusters leads to biofilm production of Staphylococcus epidermidis, which is thought to contribute to virulence in biomaterialrelated infections. We purified a specific polysaccharide antigen of biofilm-producing S. epidermidis 1457 and RP62A, which was recently shown to have a function in the accumulative phase of biofilm production by mediating intercellular adhesion (D. Mack, M. Nedelmann, A. Krokotsch, A. Schwarzkopf, J. Heesemann, and R. Laufs, Infect. Immun. 62:3244-3253, 1994). Following Sephadex G-200 gel filtration, this antigen was separated by Q-Sepharose chromatography into a major polysaccharide, polysaccharide I (>80%), which did not bind to Q-Sepharose, and a minor polysaccharide, polysaccharide II (<20%), which was moderately anionic. As shown by chemical analyses and nuclear magnetic resonance spectroscopy, polysaccharide I is a linear homoglycan of at least 130 ␤-1,6-linked 2-deoxy-2-amino-D-glucopyranosyl residues. On average, 80 to 85% of them are N acetylated; the rest are non-N-acetylating and positively charged. Chain cleavage by deamination with HNO 2 revealed a more or less random distribution of the non-N-acetylated glucosaminyl residues, with some prevalence of glucosaminyl-rich sequences. Cation-exchange chromatography separated molecular species whose content of non-N-acetylated glucosaminyl residues varied between 2 and 26%. Polysaccharide II is structurally related to polysaccharide I but has a lower content of non-N-acetylated Dglucosaminyl residues and contains phosphate and ester-linked succinate, rendering it anionic. Enzyme-linked immunosorbent assay inhibition with various monosaccharides revealed the ␤-anomeric form and the acetylated amino group of the D-glucosaminyl residues as important for reactivity with the specific antiserum. The unbranched polysaccharide structure favors long-range contacts and interactions between polysaccharide strands and the cell wall and/or lectin-like proteins, leading to intercellular adhesion and biofilm accumulation. The structure of the polysaccharide is, so far, considered to be unique and, according to its function, is referred to as S. epidermidis polysaccharide intercellular adhesin (PIA).At present, coagulase-negative staphylococci, mostly Staphylococcus epidermidis, represent the most frequent cause by far of nosocomial sepsis and are the most prominent organisms responsible for infections associated with implanted biomaterials like intravascular catheters, peritoneal dialysis catheters, cerebrospinal fluid shunts, prosthetic heart valves, and prosthetic joints, resulting in substantial morbidity and mortality (25,37,54,56).By scanning electron microscopy, coagulase-negative staphylococci were shown to colonize intravascular catheters in large adherent biofilms composed of multilayered cell clusters embedded in an amorphous extracellular material, which is composed of exopolysaccharides referred to as slime or glycocalyx (13,16,28,45,51). In vitro...

Staphylococcus epidermidisBacteremia Induces Brain Injury in Neonatal Mice via Toll-like Receptor 2-Dependent and -Independent Pathways

“…WT S. epidermidis 1457, a clinical strain obtained from an adult patient with a central venous catheter infection [21], was a kind gift from Dr Michael Otto's laboratory. For the preparation of inocula, 500 mL of bacterial stock was added to 30 mL of trypticase soy broth in a 125 mL baffled flask.…”

Biofilm formation in invasive Staphylococcus aureus isolates is associated with the clonal lineage

“…The best-described mechanism of biofilm development in S. aureus involves the extracellular molecule polysaccharide intercellular adhesin or poly-Nacetylglucosamine (PIA/PNAG) (Mack et al, 1996). PIA/PNAG synthesis is regulated by the intercellular adhesion (ica) locus (Cramton et al, 1999).…”

Biofilm exopolysaccharides: a strong and sticky framework

“…Inorganic residues, such as phosphate or rarely sulphate, may also confer polyanionic status (Sutherland, 1990). A very few EPS may even be polycationic, as exemplified by the adhesive polymer obtained from strains of Staphylococcus epidermidis strains associated with biofilms (Mack et al, 1996). The composition and structure of the polysaccharides determines their primary conformation.…”