O
not annotated - annotated - LINNAEUS only
20951743
Development and validation of a lateral flow immunoassay using colloidal gold for the identification of serotype-specific foot-and-mouth disease virus O, A and Asia 1.
A lateral flow immunoassay (LFI) was developed to identify and diagnose foot-and-mouth disease virus (FMDV) serotypes Ofoot-and-mouth disease virus (FMDV) serotypesfoot-and-mouth disease virus (FMDV) serotypes O, A and Asia 1. Antibodies obtained from rabbits and guinea pigs immunized with cell-culture-adapted virus strains (O/CHA/99, A/GS/LX/66, Asia 1/CHN/05) and suckling-mouse adapted virus strains (O/AV99(L), A/AV88(L), Asia 1/YNBS/58) were used as capture antibodies. The diagnostic kit included three immunochromatographic strips of types O, A and Asia 1, and the type-specific results were confirmed by color on the test lines of the three strips. The LFI was evaluated using epithelial and vesicular samples (n=396) prepared from current and historical field samples (provide by the National Foot-and-Mouth Disease Reference Laboratory of China at Lanzhou Veterinary Research Institute). Negative samples (n=95) were collected from healthy animals. The diagnostic sensitivity of the LFI for FMDV serotypes OFMDV serotypesFMDV serotypes O, A and Asia 1 was 88.3% compared to 89.7% obtained by the reference method of indirect-sandwich ELISA. The sensitivity of the LFI for FMDV type Asia 1 was higher at 92.1% compared to 90.5% for the ELISA. The specificity of the LFI was 97.1% compared with 97.4%.
21057010
Biochemical characterization of UDP-Gal:GlcNAc-pyrophosphate-lipid Beta-1,4-Galactosyltransferase WfeD, a new enzyme from Shigella boydii type 14 that catalyzes the second step in O-antigen repeating-unit synthesis.
The O antigen is the outer part of the lipopolysaccharide (LPS) in the outer membrane of Gram-negative bacteria and contains many repeats of an oligosaccharide unit. It contributes to antigenic variability and is essential to the full function and virulence of bacteria. Shigella is a Gram-negative human pathogen that causes diarrhea in humans. The O antigen of Shigella boydii type 14 consists of repeating oligosaccharide units with the structure [->6-d-Galpalpha1->4-d-GlcpABeta1->6-d-GalpBeta1->4-d-GalpBeta1->4-d-GlcpNAcBeta1->]n. The wfeD gene in the O-antigen gene cluster of Shigella boydii type 14 was proposed to encode a galactosyltransferase (GalT) involved in O-antigen synthesis. We confirmed here that the wfeD gene product is a Beta4-GalT that synthesizes the GalBeta1-4GlcNAcalpha-R linkage. WfeD was expressed in Escherichia coli, and the activity was characterized by using UDP-[^3H]Gal as the donor substrate as well as the synthetic acceptor substrate GlcNAcalpha-pyrophosphate-(CH2)11-O-phenyl. The enzyme product was analyzed by liquid chromatography-mass spectrometry (LC-MS), high-performance liquid chromatography (HPLC), nuclear magnetic resonance (NMR), and galactosidase digestion. The enzyme was shown to be specific for the UDP-Gal donor substrate and required pyrophosphate in the acceptor substrate. Divalent metal ions such as Mn^2(+), Ni^2(+), and, surprisingly, also Pb^2(+) enhanced the enzyme activity. Mutational analysis showed that the Glu101 residue within a DxD motif is essential for activity, possibly by forming the catalytic nucleophile. The Lys211 residue was also shown to be required for activity and may be involved in the binding of the negatively charged acceptor substrate. Our study revealed that the Beta4-GalT WfeD is a novel enzyme that has virtually no sequence similarity to mammalian Beta4-GalT, although it catalyzes a similar reaction.
21148734
Relating the physical properties of Pseudomonas aeruginosa lipopolysaccharides to virulence by atomic force microscopy.
Lipopolysaccharides (LPS) are an important class of macromolecules that are components of the outer membrane of Gram-negative bacteria such as Pseudomonas aeruginosa. P. aeruginosa contains two different sugar chains, the homopolymer common antigen (A band) and the heteropolymer O antigen (B band), which impart serospecificity. The characteristics of LPS are generally assessed after isolation rather than in the context of whole bacteria. Here we used atomic force microscopy (AFM) to probe the physical properties of the LPS of P. aeruginosa strain PA103 (serogroup O11) in situ. This strain contains a mixture of long and very long polymers of O antigen, regulated by two different genes. For this analysis, we studied the wild-type strain and four mutants, DeltaWzz1 (producing only very long LPS), DeltaWzz2 (producing only long LPS), DDeltaM (with both the wzz1 and wzz2 genes deleted), and Wzy::GM (producing an LPS core oligosaccharide plus one unit of O antigen). Forces of adhesion between the LPS on these strains and the silicon nitride AFM tip were measured, and the Alexander and de Gennes model of steric repulsion between a flat surface and a polymer brush was used to calculate the LPS layer thickness (which we refer to as length), compressibility, and spacing between the individual molecules. LPS chains were longest for the wild-type strain and DeltaWzz1, at 170.6 and 212.4 nm, respectively, and these values were not statistically significantly different from one another. Wzy::GM and DDeltaM have reduced LPS lengths, at 34.6 and 37.7 nm, respectively. Adhesion forces were not correlated with LPS length, but a relationship between adhesion force and bacterial pathogenicity was found in a mouse acute pneumonia model of infection. The adhesion forces with the AFM probe were lower for strains with LPS mutations, suggesting that the wild-type strain is optimized for maximal adhesion. Our research contributes to further understanding of the role of LPS in the adhesion and virulence of P. aeruginosa.