Characterization of surface saccharides in two Corynebacterium diphtheriae strains

Abstract

Two Corynebacterium diphtheriae strains were analyzed by assays employing a battery of highly purified fluorescent lectins. From 22 lectins tested only seven with affinity to receptor molecules containing N-acetylglucosamine (D-GlcNAc), Nacetylgalactosamine (D-GalNAc), galactose (D-Gal), mannose-like (D-Man-like) and sialic acid residues showed positive fluorescent labeling. A higher reactivity of Triticum vulgaris (WGA), which binds to sialic acid and/or L-D-GlcNAc-containing residues, and Bandeiraea simplicifolia II (BS-II), which recognizes K and L-D-GlcNAc units, was shown by the sucrosefermenting strain. Ricinus communis (RCA-I), which recognizes D-Gal units in addition to both Glycine max (SBA) and Artocarpus integrifolia (Jacaline) agglutinins that bind to D-GalNAc-containing residues, reacted preferentially with the sucrose-negative strain. Canavalia ensiformis (Con A), which recognizes D-Man-like receptors, reacted with both sucrosefermenting and non-sucrose-fermenting C. diphtheriae biotypes. However, higher interaction was observed with the nonsucrose-fermenting strain. Fluorescence of WGA binding was significantly decreased by neuraminidase treatment suggesting the presence of an exposed sialic acid moiety on C. diphtheriae surfaces. Binding assay using radiolabeled [IPS I]WGA essentially confirmed the lectin fluorescence studies. N-Acetylneuraminic acid moieties were detected in whole cell hydrolysates as assessed by thin-layer and gas-liquid chromatography. The data indicate differences on the cell surface saccharide ligands between the sucrose-fermenting and the non-sucrose-fermenting C. diphtheriae strains. z

Effects of Iron Limitation on Adherence and Cell Surface Carbohydrates of Corynebacterium diphtheriae Strains

“…We have observed an absolute reduction in the sialic acid residues and a significant increase in the amounts of D-GlcNAc, D-GalNAc, D-Gal, and D-Man residues in both strains. Sialic acid terminal constituents of cell moieties seemed to raise difficulties in C. diphtheriae adherence to HEp-2 cells, as previously observed with erythrocytes (11).…”

“…Sialic acid residues were expressed mainly on the surface of the nonhemagglutinating and highly glassadherent strain 241. It is known that sugar residues, particularly sialic acid residues, contribute to the expression of hydrophilic characteristics and adherence to glass by C. diphtheriae (11). Here we have demonstrated that iron has a regulatory role in the expression of surface carbohydrate moieties of C. diphtheriae strains.…”

“…The slides were then washed three times in PBS-5% BSA for 5 min each time, mounted in PBS-glycerol (50%), and observed under a fluorescence microscope (Universal Photomicroscope; Zeiss, Oberkochen, Germany). FITC-lectin binding to bacterial cells was inhibited by preincubation with the specific sugar hapten (0.1 M) (11).…”

“…The amount of iodinated lectin bound to the cells was determined with a gamma counter (Beckman Instruments, Inc., Palo Alto, Calif.). The specificity of binding was ascertained by performing parallel binding determinations in the presence of a 0.1 M concentration of the specific sugar inhibitor for each concentration of lectin used (11).…”

“…The hemagglutinating activity of nonfimbrial adhesion protein combination 67-72p is influenced by the concentrations of sugar residues expressed on bacterial surface (5). Sialic acid terminal moieties are expressed mainly on the surface of the nonhemagglutinating and highly glass-adherent sucrose-fermenting strain 241 (11). C. diphtheriae, which is generally considered to be an extracellular colonizer, also exhibits the ability to survive within cultured epithelial cells (9).…”