8 research outputs found
Amylase activity analysis.
<p>Recombinant and nonrecombinant strains grew on the starch-containing LB plate before (A) and after (B) being stained by iodine. The integration of <i>cotC</i>::<i>ompC</i> gene fusion disrupts <i>amyE</i> gene and made the strain amylase deficient, transparent halo was produced around the <i>B</i>. <i>subtilis</i> 168, but in the recombinant <i>B</i>. <i>subtilis</i> SE2 clones, no transparent halo was produced.</p
Immunofluorescence microscopy analysis.
<p>Sporulation of <i>B</i>. <i>subtilis</i> strains was induced by the exhaustion method, and spores were collected in DSM after 48h. Spores were labeled with OmpC antiserum followed by Cy3-labeled goat anti-chicken IgG. Spores were visualized by phase-contrast (PC) and immunofluorescence (IF) microscopy.</p
Schematic representation of the construction of the recombinant spores.
<p>The <i>cotC</i>::<i>ompC</i> gene fusion and <i>cat</i> (chloramphenicol-resistance gene) gene carried by plasmid pDG364-<i>cotC</i>-<i>ompC</i> were integrated into the <i>amyE</i> gene locus of <i>B</i>. <i>subtilis</i> 168 chromosome by double cross-over recombination events. Arrows indicate direction of transcription.</p
PCR analysis with different primer pairs.
<p>Site-directed PCR analysis of <i>cotC</i>::<i>ompC</i> gene fusion integrated into the chromosome of <i>B</i>. <i>subtilis</i> 168. lane WT, <i>B</i>. <i>subtilis</i> 168; lane RT, recombinant <i>B</i>. <i>subtilis</i> SE2; primer pairs used in PCR are labeled above.</p
Serum IgG and intestinal mucosal SIgA antibodies responses.
<p>PI, pre-immune samples. Samples obtained from the mice immunized with recombinant spores (A and C) and wild-type spores (B and D); E, samples obtained from a naïve, untreated control group. ** indicates <i>p</i><0.01.</p
Protection of mice against challenge with <i>S</i>. Typhimurium strain SL1344.
<p>Protection of mice against challenge with <i>S</i>. Typhimurium strain SL1344.</p
DataSheet_1_Surface Display of porcine circovirus type 2 antigen protein cap on the spores of bacillus subtilis 168: An effective mucosal vaccine candidate.pdf
The oral mucosal vaccine has great potential in preventing a series of diseases caused by porcine circovirus type 2 (PCV2) infection. This study constructed a recombinant Bacillus subtilis RB with PCV2 Capsid protein (Cap) on its spore surface and cotB as a fusion partner. The immune properties of the recombinant strain were evaluated in a mouse model. IgA in intestinal contents and IgG in serum were detected by enzyme-linked immunosorbent assay (ELISA). The results demonstrated that recombinant spores could activate strong specific mucosal and humoral immune responses. In addition, spores showed good mucosal immune adjuvant function, promoting the proliferation of CD3+, CD4+ and CD8+ T cells and other immune cells. We also found that the relative expression of inflammatory cytokines such as IL-1β, IL-6, IL-10, TNF-α and IFN in the small intestinal mucosa was significantly up-regulated under the stimulation of recombinant bacteriophage. These effects are important for the balance of Th1/Th2-like responses. In summary, our results suggest that recombinant B. subtilis RB as a feed additive provides a new strategy for the development of novel and safe PCV2 mucosal subunit vaccines.</p