Left, control OCT cryosection of buy BMN 673 biofilm incubated without specific antiserum, but with anti-rabbit conjugated gold particles; no labeling with the gold particles occurred; Right, OCT cryosection of a biofilm incubated with rabbit antibodies to EPS, followed by anti-rabbit conjugated gold particles. The black dots are gold particles around the bacterial cells and in the residual biofilm matrix. CDK inhibitor Mannose is not present

in the H. somni LOS, but is the predominant component of the EPS. Therefore, a fluorescein isothionate-labeled, mannose-specific lectin (Morniga M [black mulberry]) was incubated with H. somni biofilms. This lectin bound to the matrix material between the cells of the biofilm of 2336 (Figure 9), indicating that the EPS was a major component of the H. somni biofilm. Analysis of the biofilm embedded in OCT resin with the sialic acid-reactive lectins (MAA [Maackia amurensis], WGA [Wheat Germ agglutinin], HHA

[Amaryllis], and SBA [soybean] further supported that Neu5Ac was also a component of the biofilm of 2336 (data not shown). SEM examination showed that the addition of Neu5Ac to chemically defined medium increased biofilm production by 2336, whereas biofilm formation by 129Pt was unchanged (Figure 10). Although the LOS of 2336 was sialylated when grown in the presence of Neu5Ac, there were no differences in LOS structure or sialylation levels AZD1080 in vivo when 2336 was grown as a biofilm, as planktonic cells, or on blood agar plates (additional file 1, Table S1). In the absence of supplemental Neu5Ac, of only LOS from 2336 grown on blood agar plates was sialylated, presumably due to the presence of Neu5Ac in the fresh blood. As previously reported [12], the LOS of 129Pt grown under any of the above conditions was not sialylated. Figure 9 H. somni biofilm labeled with Moringa M lectin. H. somni was grown as a biofilm on cover slips and stained with TO-PRO-3 to label the bacterial cells (top left), MNA (specific for α-mannose)-FITC to label mannose (top right), and were merged (bottom center) to demonstrate

the presence of mannose within the bacterial biofilm. Mannose is present in the H. somni EPS, but not in the LOS. Figure 10 SEM image of biofilm formation by H. somni 2336 and 129Pt. A1-A2, biofilm formation by 2336; B1- B2, enhanced biofilm formation by 2336 grown in the presence of Neu5Ac (50 μg/ml) in chemically defined medium; C1- C2, biofilm formation by 129Pt; D1- D2, biofilm formation by 129Pt grown in the presence of Neu5Ac in defined medium. There is no significant change in the density of the biofilm of 129Pt grown in the presence of Neu5Ac. Putative polysaccharide locus in H. somni 2336 To understand the genetic basis of EPS biosynthesis in H. somni, we sought to identify a locus of genes that could encode for enzymes involved in the synthesis and transport of a polysaccharide other than LOS.