Intranasal vaccination elicits secretory IgA (SIgA) antibodies in the airways, which is required for cross-protection against influenza. To enhance the breadth of immunity induced by a killed swine influenza virus antigen (KAg) or conserved T cell and B cell peptides, we adsorbed the antigens together with the TLR3 agonist poly(I:C) electrostatically onto cationic alpha-D-glucan nanoparticles (Nano-11) resulting in Nano-11-KAg-poly(I:C) and Nano-11-peptides-poly(I:C) vaccines. In vitro, increased TNF-α and IL-1ß cytokine mRNA expression was observed in Nano-11-KAg-poly(I:C)-treated porcine monocyte-derived dendritic cells. Nano-11-KAg-poly(I:C), but not Nano-11-peptides-poly(I:C), delivered intranasally in pigs induced high levels of cross-reactive virus-specific SIgA antibodies secretion in the nasal passage and lungs compared to a multivalent commercial influenza virus vaccine administered intramuscularly. The commercial and Nano-11-KAg-poly(I:C) vaccinations increased the frequency of IFNγ secreting T cells. The poly(I:C) adjuvanted Nano-11-based vaccines increased various cytokine mRNA expressions in lymph nodes compared to the commercial vaccine. In addition, Nano-11-KAg-poly(I:C) vaccine elicited high levels of virus neutralizing antibodies in bronchoalveolar lavage fluid. Microscopic lung lesions and challenge virus load were partially reduced in poly(I:C) adjuvanted Nano-11 and commercial influenza vaccinates. In conclusion, compared to our earlier study with Nano-11-KAg vaccine, addition of poly(I:C) to the formulation improved cross-protective antibody and cytokine response.

Salmonella enterica serovar Enteritidis (S. Enteritidis, SE) infection in broilers causes a huge economic loss and public health risk. We previously demonstrated that orally delivered chitosan based (CS) Salmonella subunit nanoparticle (NP) vaccine containing immunogenic outer membrane proteins (OMP) and flagellin (FLA) of SE [CS-NP(OMP+FLA)] induces immune response in broilers. The objective of this study was to evaluate the dose-and age-dependent response and efficacy of CS-NP(OMP+FLA) vaccine in broilers. Three-day old birds were vaccinated and boosted once or twice. Additional groups were vaccinated at three weeks with no booster or boosted once a week later. Each dose of CS-NP vaccine had either 10 or 50 µg of OMP+FLA antigens. Our data revealed that two doses of vaccine were required to induce substantial immune response. Birds received 2 doses of CS-NP(OMP+FLA) vaccine at 3 days and 3 weeks of age with 10 µg antigens, and birds inoculated twice at 3 and 4 weeks of age with 50 µg antigens had lowest challenged bacterial load in the cecal contents with over 0.5 log 10 reduction. In CS-NP(OMP+FLA) vaccinated birds, antigen-specific splenocyte proliferation, mucosal and systemic antibody response and the frequency of IFNγ-producing T cells were increased compared to control groups. At the molecular level, in the cecal tonsils of CS-NP(OMP+FLA) immunized birds, mRNA levels of toll-like receptor (TLR) 2 and TLR 4, and cytokines IL-4 and IL-10 were upregulated. The CS-NP(OMP+FLA) vaccine given orally has the potential to induce a protective immune response against SE infection in broilers.

Parenteral administration of killed/inactivated swine influenza A virus (SwIAV) vaccine in weaned piglets provides variable levels of immunity due to the presence of preexisting virus specific maternal derived antibodies (MDA). To overcome the effect of MDA on SwIAV vaccine in piglets, we developed an intranasal deliverable killed SwIAV antigen (KAg) encapsulated chitosan nanoparticles called chitosan-based NPs encapsulating KAg (CS NPs-KAg) vaccine. Further, to target the candidate vaccine to dendritic cells and macrophages which express mannose receptor, we conjugated mannose to chitosan (mCS) and formulated KAg encapsulated mCS nanoparticles called mannosylated chitosan-based NPs encapsulating KAg (mCS NPs-KAg) vaccine. In MDA-positive piglets, prime-boost intranasal inoculation of mCS NPs-KAg vaccine elicited enhanced homologous (H1N2-OH10), heterologous (H1N1-OH7), and heterosubtypic (H3N2-OH4) influenza virus-specific secretory IgA (sIgA) antibody response in nasal passage compared to CS NPs-KAg vaccinates. In vaccinated upon challenged with a heterologous SwIAV H1N1, both mCS NPs-KAg and CS NPs-KAg vaccinates augmented H1N2-OH10, H1N1-OH7, and H3N2-OH4 virus-specific sIgA antibody responses in nasal swab, lung lysate, and bronchoalveolar lavage (BAL) fluid; and IgG antibody levels in lung lysate and BAL fluid samples. Whereas, the multivalent commercial inactivated SwIAV vaccine delivered intramuscularly increased serum IgG antibody response. In mCS NPs-KAg and CS NPs-KAg vaccinates increased H1N2-OH10 but not H1N1-OH7 and H3N2-OH4-specific serum hemagglutination inhibition titers were observed. Additionally, mCS NPs-KAg vaccine increased specific recall lymphocyte proliferation and cytokines IL-4, IL-10, and IFNγ gene expression compared to CS NPs-KAg and commercial SwIAV vaccinates in tracheobronchial lymph nodes. Consistent with the immune response both mCS NPs-KAg and CS NPs-KAg vaccinates cleared the challenge H1N1-OH7 virus load in upper and lower respiratory tract more efficiently when compared to commercial vaccine. The virus clearance was associated with reduced gross lung lesions. Overall, mCS NP-KAg vaccine intranasal immunization in MDA-positive pigs induced a robust cross-reactive immunity and offered protection against influenza virus.

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Controlling Salmonella enterica serovar Enteritidis (SE) infection in broilers is a huge challenge. In this study, our objective was to improve the efficacy of a chitosan nanoparticle (CS)-based Salmonella subunit vaccine for SE, containing immunogenic outer membrane proteins (OMP) and flagellin (FLA), called the CS(OMP+FLA) vaccine, by surface conjugating it with mannose to target dendritic cells, and comparing the immune responses and efficacy with a commercial live Salmonella vaccine in broilers. The CS(OMP+FLA)-based vaccines were administered orally at age 3 days and as a booster dose after three weeks, and the broilers were challenged with SE at 5 weeks of age. Birds were sacrificed 10 days post-challenge and it was observed that CS(OMP+FLA) vaccine surface conjugated with both mannose and FLA produced the greatest SE reduction, by over 1 log10 colony forming unit per gram of the cecal content, which was comparable to a commercial live vaccine. Immunologically, specific mucosal antibody responses were enhanced by FLA-surface-coated CS(OMP+FLA) vaccine, and mannose-bound CS(OMP+FLA) improved the cellular immune response. In addition, increased mRNA expression of Toll-like receptors and cytokine was observed in CS(OMP+FLA)-based-vaccinated birds. The commercial live vaccine failed to induce any such substantial immune response, except that they had a slightly improved T helper cell frequency. Our data suggest that FLA-coated and mannose-modified CS(OMP+FLA) vaccine induced robust innate and adaptive cell-mediated immune responses and substantially reduced the Salmonella load in the intestines of broilers.