Development of a Mucosal Immune-Enhancing Oral Vaccine Against Porcine Epidemic Diarrhea Virus Using Lactobacillus paracasei
Description
We attempted to enhance the specific immune ability of the mucosa, Lactobacillus paracasei served as the host strain to express the fusion of the protective antigen S1 of PEDV, the M cell-targeting peptide Co1, the dendritic cell-targeting peptides (6aa), and the mucosal immune adjuvant LTB, thereby obtaining a candidate strain pPG-Co1-6aa-S1-LTB/27-2. Pregnant mice that received oral immunization with recombinant microorganisms demonstrated a notable increase in serum IgG antibodies that are specific to anti-PEDV, in addition to intestinal mucus and fecal SIgA antibodies, both of which exhibited in vitro neutralizing activity. Additionally, pregnant mice had considerably higher serum levels of IFN-γ, IL-2, IL-4, and IL-10. Notably, the level of anti-PEDV specific SIgA antibodies of the newborn mice delivered by the pregnant mice was also significantly elevated. The aforementioned findings show that the recombinant bacteria pPG-Co1-6aa-S1-LTB/27-2 can produce maternal antibodies and strongly stimulate humoral, cellular, and mucosal immune responses in pregnant mice. Figure 1. Illustrative diagram showing the creation of recombinant plasmids. Figure 2. Immunization protocol and sampling schedule. Figure 3. Identification and protein expression analysis of recombinant L.paracasei 27-2. A. PCR identification of recombinant plasmids. M: Trans 2k plus DNA marker. B. Western blot analysis was used to confirm the fusion protein's expression. M: Pre-stained protein marker. C. Analysis of binding activity of recombinant lactobacillus expressing LTB protein. Figure 4. Proteins from different species were presented on the surface of Lactobacillus paracasei. Figure 5. Biological characterization analysis. A. Growth curve determination. B. The protein stability of the recombinant strains was assessed through Western blot analysis. Figure 6. Anti-PEDV-specific IgG antibody levels in the serum, anti-PEDV-specific SIgA antibody levels in the intestinal mucus and fecal are listed in A, B, and C, respectively. Figure 7. Measurement of neutralizing activity in intestinal serum IgG and mucus SIgA antibody levels in vaccinated pregnant mice. Figure 8. Measure the blood cytokine levels in pregnant mice that have received recombinant strains and PBS vaccinations. The mean ± SEM of three separate experiments is used to illustrate the results. Figure 9. Quantities of anti-PEDV specific SIgA antibodies in the intestinal mucus of neonatal mice.