Transcriptional Profiling of in a Two Species Biofilm with .

Biofilms on silicone rubber voice prostheses are the major cause for frequent failure and replacement of these devices. The presence of both bacterial and yeast strains has been suggested to be crucial for the development of voice prosthetic biofilms. Polymicrobial biofilms that include and are the leading cause of voice prosthesis failure.

Cell surface shaving of Candida albicans biofilms, hyphae, and yeast form cells.

We used a brief trypsin treatment followed by peptide separation and identification using nano-LC followed by off-line MS/MS to identify the surface proteins on live Candida albicans organisms growing in biofilms and planktonic yeast cells and hyphae. One hundred thirty-one proteins were present in at least two of the three replicates of one condition and distributed in various combinations of the three growth conditions. Both previously reported and new surface proteins were identified and these were distributed between covalently attached proteins and noncovalently attached proteins of the cell wall.

Proteomic analysis of cytoplasmic and surface proteins from yeast cells, hyphae, and biofilms of Candida albicans.

Candida albicans is a human commensal and opportunistic pathogen that participates in biofilm formation on host surfaces and on medical devices. We used DIGE analysis to assess the cytoplasmic and non-covalently attached cell-surface proteins in biofilm formed on polymethylmethacrylate and planktonic yeast cells and hyphae. Of the 1490 proteins spots from cytoplasmic and 580 protein spots from the surface extracts analyzed, 265 and 108 were differentially abundant respectively (>or=1.

Candida albicans cell wall proteins.

The Candida albicans cell wall maintains the structural integrity of the organism in addition to providing a physical contact interface with the environment. The major components of the cell wall are fibrillar polysaccharides and proteins. The proteins of the cell wall are the focus of this review.

Evidence for the presence of complex carbohydrates in Candida albicans cell wall glycoproteins.

In order to test the hypothesis that cell wall glycoproteins of Candida albicans contained non-mannan oligosaccharides, the sugar composition of cell wall extracts and fractions of cell wall extracts was examined by means of fluorophore-assisted carbohydrate electrophoresis (FACE). In addition to the expected mannose, glucose, and N-acetyl-glucosamine, this analysis showed the presence of galactose, N-acetyl-galactosamine, fucose, and sialic acid and two unknown sugars. These sugars are also associated with complex oligosaccharides of mammalian glycoproteins.

Farnesol-mediated inhibition of Candida albicans yeast growth and rescue by a diacylglycerol analogue.

During Candida albicans yeast cell growth to early stationary phase, metabolites accumulate in the medium, including the quorum-sensing molecule farnesol. We found that besides germ tube inhibition, 40 microM farnesol also inhibited C. albicans yeast growth under yeast growth permissive conditions.

Defining Candida albicans stationary phase by cellular and DNA replication, gene expression and regulation.

Stationary phase Candida albicans yeast cells harbour properties of better adherence, virulence and elevated drug resistance. C. albicans stationary phase is not well characterized in vitro either physiologically or molecularly.

Role for cell density in antifungal drug resistance in Candida albicans biofilms.

Biofilms of Candida albicans are less susceptible to many antifungal drugs than are planktonic yeast cells. We investigated the contribution of cell density to biofilm phenotypic resistance. Planktonic yeast cells in RPMI 1640 were susceptible to azole-class drugs, amphotericin B, and caspofungin at 1 x 10(3) cells/ml (standard conditions) using the XTT [2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide sodium salt] assay.

Analysis of RNA species of various sizes from stationary-phase planktonic yeast cells of Candida albicans.

We initiated a comparison of Candida albicans stationary-phase gene expression with other growth states. The widely used hot acid phenol method for RNA extraction did not extract rRNA from late stationary-phase cells. The RNA from growing yeast cells, hyphae and biofilm was biased towards small-sized RNA.

Candida albicans SNO1 and SNZ1 expressed in stationary-phase planktonic yeast cells and base of biofilm.

The Candida albicans homologues of the most studied Saccharomyces cerevisiae stationary-phase genes, SNO1 and SNZ1, were used to test the hypothesis that, within a biofilm, some cells reach stationary phase within continuously fed, as well as static, C. albicans biofilms grown on dental acrylic. The authors first studied the expression patterns of these two genes in planktonic growth conditions.

Non-glucan attached proteins of Candida albicans biofilm formed on various surfaces.

Non-glucan attached proteins of the cell surface and extracellular matrix of Candida albicans biofilms formed on two catheter surfaces and denture acrylic were examined. The SDS-PAGE protein profiles of these proteins compared with that obtained from planktonic yeast cells and germ tubes were generally similar. This observation suggested that this class of biofilm surface proteins is not composed of a unique set of extracellular proteins or that one or a few proteins dominate the non-glucan attached proteins of biofilm.

Non-conventional protein secretion in yeast.

Many proteins are transported to the cell surface of Saccharomyces cerevisiae and Candida albicans to be either integrated into the cell-wall structure or exported to the external medium. Secretion of many of these proteins through the classical endoplasmic reticulum-Golgi pathway is driven by a canonical N-terminal signal peptide. However, several surface proteins lacking this motif can also access the cell surface and remain loosely bound to the wall.

Anaerobic growth of Candida albicans does not support biofilm formation under similar conditions used for aerobic biofilm.

C. albicans is an opportunistic fungus causing life-threatening systemic infections particularly in immunocompromised individuals. The organism is a commensal in humans and grows either aerobically, e.

Evidence for the presence of pir-like proteins in Candida albicans.

Pir proteins are unique proteins with internal repeat sequences that are reported to be present in the cell wall of Saccharomyces cerevisiae. They are covalently attached to the cell wall and can be released by mild alkali treatment. In this study the biotinylated cell wall preparations from Candida albicans and S.

Cell surface proteins of Candida albicans: preparation of extracts and improved detection of proteins.

We have reexamined the detection of the components in a beta-mercaptoethanol and ammonium carbonate buffer extract of surface proteins of Candida albicans and the effects of postextraction manipulation of the extract on recovery of extract components. Following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), preferential staining of some moieties was observed when bands detected by a commercial silver staining method or a Coomassie Brilliant Blue (CBB) staining method were compared. Additional protein bands that were either not detected or poorly detected by a single method alone were readily observed by a combined silver-CBB staining method.

Oral colonization by Candida albicans.

Candida albicans is a commensal yeast normally present in small numbers in the oral flora of a large proportion of humans. Colonization of the oral cavity by C. albicans involves the acquisition and maintenance of a stable yeast population.

Immunodetection of CD45 epitopes on the surface of Candida albicans cells in culture and infected human tissues.

Candida albicans is a leading cause of disseminated fungal infection in immunocompromised patients. Candida-host cell interactions are mediated at the cell surface. Since blood-group I epitopes have been detected on the surface of C albicans cells, we investigated whether CD45, the molecule that carries the I antigen on human lymphocytes, is present on the C albicans cell surface, in culture and in human tissue specimens of human candidiasis.

Interactions between Candida albicans and the human extracellular matrix component tenascin-C.

Tenascins are large multimeric proteins that contain repeated structural motifs that include epidermal growth factor (EGF)-like repeats, fibronectin type III repeats and a globular fibrinogen-like domain, and are involved in tissue and organ morphogenesis, as well as in adhesion and migration of cells. C. albicans germ-tubes, but not blastospores, were able to bind to soluble human tenascin-C as revealed by an indirect immunofluorescence assay.

Variations in mRNA transcript levels of cell wall-associated genes of Saccharomyces cerevisiae following spheroplasting.

mRNA transcript levels of 38 genes from Saccharomyces cerevisiae were investigated during attempted spheroplast regeneration. Many of the genes selected are involved in cell wall biosynthesis. Spheroplasts did not regenerate into osmotically competent cells during the experiment.

Enolase is present in the cell wall of Saccharomyces cerevisiae.

Non-covalently attached or soluble cell wall proteins of Saccharomyces cerevisiae were extracted using a high pH/2-mercaptoethanol procedure and were separated for peptide sequencing using 2D-PAGE. A partial N-terminal sequence of a major protein spot was obtained and showed high identity with enolase gene products. Western blotting with an anti-enolase antibody confirmed that enolase was present in the cell wall extract.

Cell wall and secreted proteins of Candida albicans: identification, function, and expression.

The cell wall is essential to nearly every aspect of the biology and pathogenicity of Candida albicans. Although it was initially considered an almost inert cellular structure that protected the protoplast against osmotic offense, more recent studies have demonstrated that it is a dynamic organelle. The major components of the cell wall are glucan and chitin, which are associated with structural rigidity, and mannoproteins.

Serologic response to cell wall mannoproteins and proteins of Candida albicans.

The cell wall of Candida albicans not only is the structure in which many biological functions essential for the fungal cells reside but also is a significant source of candidal antigens. The major cell wall components that elicit a response from the host immune system are proteins and glycoproteins, the latter being predominantly mannoproteins. Both the carbohydrate and protein moieties are able to trigger immune responses.

3-phosphoglycerate kinase: a glycolytic enzyme protein present in the cell wall of Candida albicans.

We have used a polyclonal antiserum to cell wall proteins of Candida albicans to isolate several clones from a cDNA lambda gt11 expression library. Affinity-purified antibody prepared to the fusion protein of one clone identified a 40 kDa moiety present in cell wall extracts from both morphologies of the organism. Indirect immunofluorescence demonstrated expression of this moiety at the C.

Ubiquitin-like epitopes associated with Candida albicans cell surface receptors.

We have recently reported the cloning of a Candida albicans polyubiquitin gene and the presence of ubiquitin in the cell wall of this fungus. The polyubiquitin cDNA clone was isolated because of its reactivity with antibodies generated against the candidal 37-kDa laminin-binding protein. In the present study, we have further investigated the relationship between ubiquitin and cell wall components displaying receptor-like activities, including the 37-kDa laminin receptor, the 58-kDa fibrinogen-binding mannoprotein, and the candidal C3d receptor.