Sendid, L

Sendid, L. in a dose-dependent decrease in adhesion (50% of the control with a 60-g/ml MAb concentration). In competitive assays -1,2 and -1,2 tetramannosides were the most potent carbohydrate inhibitors, with 50% inhibitory concentrations of 2.58 and 6.99 mM, respectively. Immunolocalization on infected monolayers with MAbs specific for -1,2 and -1,2 oligomannosides showed that these epitopes were shed from the yeast to the enterocyte surface. Taken together, our data indicate that -1,2 and -1,2 oligomannosides are involved in the species are part of the A-419259 commensal flora of the mucosa and skin in humans and other vertebrates. In immunocompromised or intensive-care patients, increased mucosal proliferation secondary to use of broad-spectrum antibiotics, together with reduced host defenses and physical alteration of the mucosal barriers, may result in bloodstream invasion. Altogether, candidemia accounts for 10% of nosocomial bloodstream infections, and is the causative agent in 50 to 70% of disseminated candidiasis (13, 18, 20, 36, 48). Molecular typing methods have shown an overall genetic similarity between strains obtained from blood cultures and colonizing strains obtained from the gastrointestinal tracts of the same patients, confirming endogenous acquisition as the main source of invasive candidiasis (40, 47). On the basis PTCRA of this model, adhesion of the yeasts to the epithelium of the digestive tract is usually a prerequisite for colonization and a critical step in the pathogenesis of invasive candidiasis. Characterization of the adhesins and ligands involved in the with host cell surfaces is usually mediated by the yeast cell wall, a complex and dynamic structure made up of glucan, chitin and mannoproteins (reviewed in reference 6). The outermost layers of the cell wall are made of phosphopeptidomannan (PPM), a polymer of mannose residues and proteins commonly referred to as mannan (3, 6). Mannan has been shown to play a role in adherence (27), immunomodulation (11), and antigenic variability (43). The PPM glycan moiety is composed of O-linked and N-linked oligomannosides. The N-linked part consists of a backbone of -1,6-linked mannopyranose residues with branches A-419259 composed of -1,2- and -1,3-linked mannopyranose models and terminal -1,2 linkages in serotype A (7). Short branches composed of -1,2-linked mannopyranose residues are linked to PPM through phosphodiester bridges in serotypes A and B. These side chains are referred to as the acid-labile fraction of PPM, since they are cleaved by moderate acid treatment (45). -1,2 oligomannosidic chains have also been identified on a 14- to 18-kDa glycolipid, referred to as phospholipomannan (PLM) (46), that is expressed at and shed from the cell wall (25, 39). -1,2 mannosidic linkages are uncommon structures whose presence has been reported in only few bacterial and yeast species (30, A-419259 35). In to the macrophage membrane, at least in part through binding to galectin 3, a member of a family of carbohydrate binding proteins implicated in a variety of biological functions (17, 25). -1,2 oligomannosides also generate protective antibodies (22) and induce cytokine production (26). These unique carbohydrate sequences thus appear to play a key role in the blastospores to Caco-2 cells. Indeed, galectin-3, which binds -1,2 oligomannosides around the macrophage (17), is also expressed in intestinal epithelial cells (1, 8). Moreover, recent studies with a mouse model of candidiasis showed that oral administration of synthetic -1,2 oligomannosides could reduce colonization of the gut, presumably by A-419259 competing with the natural flora for binding to enterocytes (12). We were thus interested in understanding the basis of this phenomenon at the cellular and molecular levels. MATERIALS AND METHODS Growth and preparation of strain for adherence assay. strain VW32 (serotype A) (5) was used throughout this study. This strain, which was originally isolated from a patient with human renal candidiasis, has been employed in prior studies of -mannosidic epitopes (15, 46). Stock cultures were maintained at ?20C on Sabouraud dextrose agar. For adhesion experiments, stock cultures were plated on Sabouraud dextrose agar and incubated at 37C. After 18 to 20 h of culture, the yeasts were recovered, washed with phosphate-buffered saline (PBS), and resuspended in PBS. The yeast concentration was determined by hemocytometer and growth on Sabouraud dextrose agar. For experiments using heat-killed yeasts, blastospores were treated at 100C for 15 min, washed in PBS by centrifugation, and resuspended in PBS. Growth and differentiation of Caco-2 cells. Caco-2 cells were obtained from the American Type.