16 research outputs found

    Mammary antigen-specific inflammatory response.

    No full text
    <p>A) Time-course of cell concentrations in milk (SCC) after infusion of 3 μg rEcOmpA through the teat canal of immunized and control cows. B) Time-course of cell concentrations in milk (SCC) after infusion of 9 μg rEcOmpA through the teat canal of immunized and control cows. Data are median values and interquartile (Q1; Q3).</p

    Assessment of rEcOmpA purity.

    No full text
    <p>5 μg of rEcOmpA were analysed by SDS-PAGE and Coomassie blue staining (A) and by western-blotting with a rabbit anti-ompA antibody (B). Bacterial pellets from the wt and Δrfb P4 strains were immunoblotted with rabbit antibodies to OmpA, showing that similar amounts of OmpA were produced by both strains (C).</p

    Humoral response to immunization with rEcOmpA.

    No full text
    <p>A) The response to immunization with rEcOmpA (on days 0 and 30) was assessed by ELISA using whole heat-killed smooth (P4) and rough (P4Δrfb) bacteria as coating antigen. Data are median values (12 immunized cows) and interquartile (Q1 and Q3). B) Reference curve of the pooled serum in ELISA with <i>E</i>. <i>coli</i> P4 and P4Δrfb as coating antigens and secondary antibody to bovine IgG(H+L) revealing IgG and IgM antibodies. C) Titration of natural antibodies detected before immunization, showing the correlation between titers of antibodies to <i>E</i>. <i>coli</i> P4 and P4Δrfb. D) Titration of antibodies to <i>E</i>. <i>coli</i> P4Δrfb in the IgM isotype and IgG1 and IgG2 subisotypes. As different reference sera, and different secondary antibodies were used for IgM and IgG antibodies, only titers variation can be compared, not their absolute values.</p

    Production of IL-17A and IFN-γ by blood cells stimulated with OmpA.

    No full text
    <p>A) Time-course production of IL-17A in the OmpA-specific whole blood assay. The blood of the 12 immunized cows was cultured in the presence of OmpA and the concentrations of IL-17A measured by ELISA. B) Time-course production of IFN-γ in the OmpA-specific whole blood assay. C) The effect of magnetic depletion of CD4+ cells from PBMC of two immunized cows on the production of IL-17A and IFN-γ. Means are expressed as the percentage of cytokine production by CD4-depeleted cells relative to the production by untreated PBMC (100%). Friedman test: p< 0.001 for IFN-g, followed by Dunn’s multiple comparison test versus D0, at D30 and D60 (ns at D45); IL -17A: Friedman test p < 0.0001, Dunn’s multiple comparison test: versus D0: D30 * D45 *** D60.</p

    Mass Spectrometry Analysis of the Extracellular Peptidome of <i>Lactococcus lactis</i>: Lines of Evidence for the Coexistence of Extracellular Protein Hydrolysis and Intracellular Peptide Excretion

    No full text
    We report here the use of a peptidomic approach to revisit the extracellular proteolysis of <i>Lactococcus lactis</i>. More than 1800 distinct peptides accumulate externally during growth of the plasmid-free protease-negative strain <i>L. lactis</i> IL1403 in a protein- and peptide-free medium. These peptides mainly originate from cell-surface- and cytoplasmic-located proteins, despite the fact that no cell lysis could be evidenced. Positioning each identified peptide on its parental protein sequence demonstrated the involvement of exo- and endopeptidase activities. The endopeptidases responsible for the release of surface and cytoplasmic peptides had distinct specificities. The membrane-anchored protease HtrA was responsible for the release of only a part of the surface peptides, and its preference for branched-chain amino acids in the N-terminal side of the cleaved bond was established in situ. Other yet uncharacterized surface proteases were also involved. Several lines of evidence suggest that surface and cytoplasmic peptides were produced by different routes, at least part of the latter being most likely excreted as peptides from the cells. The mechanism by which these cytoplasmic peptides are excreted remains an open question, as it is still the case for excreted cytoplasmic proteins

    CXCL8 secretion induced by stimulation of PS cells with live <i>E</i>. <i>coli</i> strains.

    No full text
    <p>PS cells were incubated for 3 hours with the indicated strains at a MOI of 1 in stimulation medium (A) or fresh milk (B). Experiments in stimulation medium were performed in the absence of rbCD14 (white bars) or in the presence of 0.5 μg/ml rbCD14 (black bars). Experiments in milk were done without the addition of rbCD14. Cells were then washed twice with HBSS and medium with gentamicin was added. Response was analysed by quantification of CXCL8 secretion by ELISA 24h after beginning of the experiment. Data presented are mean values and SD obtained from four (-CD14) or three (+ CD14) independent experiments with stimulations performed in duplicates. * indicates statistical significance (P < 0.05). p-values were calculated using exact permutation tests after global comparison using a Kruskal and Wallis test.</p

    Growth in milk of selected <i>E</i>. <i>coli</i> strains.

    No full text
    <p>Fresh whole milk was inoculated with 10<sup>4</sup> cfu/mL of strains MG1655 (▲), D6-117.29 (▽), B1171 (○) and P4 (◻). Inoculated milk was distributed in 1.5 mL microtubes (1.5 mL per tube) and bacteria from one tube were counted at each time point by 10-fold serial dilution and plating on TSA agar plates. Data presented are means of cfu/ml calculated from three independent experiments.</p
    corecore