Surface engineered polyanhydride-based oral Salmonella subunit nanovaccine for poultry

Sankar Renu,1,2,* Ashley D Markazi,3,* Santosh Dhakal,1,2 Yashavanth S Lakshmanappa,1,2 Suren R Gourapura,1,2 Revathi Shanmugasundaram,3 Sujata Senapati,4 Balaji Narasimhan,4 Ramesh K Selvaraj,5 Gourapura J Renukaradhya1,2 1Food Animal Health Research Program, Ohio Agricultural Research and Development Center, The Ohio State University, Wooster, OH 44691, USA; 2Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210, USA; 3Department of Animal Sciences, Ohio Agricultural Research and Development Center, The Ohio State University, Wooster, OH 44691, USA; 4Department of Chemical and Biological Engineering, Iowa State University, Ames, IA 50011, USA; 5Department of Poultry Science, University of Georgia, Athens, GA 30602, USA *These authors contributed equally to this work Purpose: Salmonellosis is a severe economic threat in poultry and a public health concern. Currently available vaccines are ineffective, and thus, developing effective oral Salmonella vaccine is warranted. Especially, a potent oral vaccine such as the mucoadhesive polyanhydride nanoparticle (PNP) protects the vaccine cargo and delivers to intestinal immune sites to elicit robust mucosal immunity and mitigate Salmonella colonization and shedding.Materials and methods: We designed a Salmonella subunit vaccine using PNP containing immunogenic Salmonella outer membrane proteins (OMPs) and flagellar (F) protein-entrapped and surface F-protein-coated PNPs (OMPs-F-PNPs) using a solvent displacement method. Using high-throughput techniques, we characterized the OMPs-F-PNPs physicochemical properties and analyzed its efficacy in layer birds vaccinated orally.Results: The candidate vaccine was resistant in acidic microenvironment and had ideal physicochemical properties for oral delivery in terms of particle size, charge, morphology, biocompatibility, and pH stability. In vitro, in vivo, and ex vivo studies showed that F-protein surface-anchored nanoparticles were better targeted to chicken immune cells in peripheral blood and splenocytes and intestinal Peyer’s patch sites. In layer chickens inoculated orally with OMPs-F-PNPs, substantially higher OMPs-specific IgG response and secretion of Th1 cytokine IFN-γ in the serum, enhanced CD8+/CD4+ cell ratio in spleen, and increased OMPs-specific lymphocyte proliferation were observed. OMPs-F-PNPs vaccination also upregulated the expression of toll-like receptor (TLR)-2 and -4, TGF-β, and IL-4 cytokines’ genes in chicken cecal tonsils (lymphoid tissues). Importantly, OMPs-F-PNPs vaccine cleared Salmonella cecal colonization in 33% of vaccinated birds.Conclusion: This pilot in vivo study demonstrated the targeted delivery of OMPs-F-PNPs to ileum mucosal immune sites of chickens and induced specific immune response to mitigate Salmonella colonization in intestines. Keywords: Salmonella antigens, polyanhydride nanoparticles, oral delivery, ileum, chickens&nbsp

Not Available

Not AvailableWe have recently demonstrated the effectiveness of an influenza A virus (IAV) subunit vaccine based onbiodegradable polyanhydride nanoparticles delivery in mice. In the present study, we evaluated the effi-cacy of 200 nm polyanhydride nanoparticles encapsulating inactivated swine influenza A virus (SwIAV)as a vaccine to induce protective immunity against a heterologous IAV challenge in pigs. Nursery pigswere vaccinated intranasally twice with inactivated SwIAV H1N2 (KAg) or polyanhydride nanoparticle-encapsulated KAg (KAg nanovaccine), and efficacy was evaluated against a heterologous zoonotic virulentSwIAV H1N1 challenge. Pigs were monitored for fever daily. Local and systemic antibody responses,antigen-specific proliferation of peripheral blood mononuclear cells, gross and microscopic lung lesions,and virus load in the respiratory tract were compared among the groups of animals. Our pre-challengeresults indicated that KAg nanovaccine induced virus-specific lymphocyte proliferation and increasedthe frequency of CD4+CD8aa+T helper and CD8+cytotoxic T cells in peripheral blood mononuclear cells.KAg nanovaccine-immunized pigs were protected from fever following SwIAV challenge. In addition, pigsimmunized with the KAg nanovaccine presented with lower viral antigens in lung sections and had 6 to8-fold reduction in nasal shedding of SwIAV four days post-challenge compared to control animals.Immunologically, increased IFN-csecreting T lymphocyte populations against both the vaccine and chal-lenge viruses were detected in KAg nanovaccine-immunized pigs compared to the animals immunizedwith KAg alone. However, in the KAg nanovaccine-immunized pigs, hemagglutination inhibition, IgGand IgA antibody responses, and virus neutralization titers were comparable to that in the animals immu-nized with KAg alone. Overall, our data indicated that intranasal delivery of polyanhydride-based SwIAVnanovaccine augmented antigen-specific cellular immune response in pigs, with promise to induce cross-protective immunityNot Availabl

Not Available

Not AvailableSwine influenza virus (SwIV) causes considerable economic loss to pig industry, and some SwIV are zoonotic. This study was conducted to evaluate the cross-protective efficacy of PLGA (poly lactic-co-glycolic acid) nanoparticles (NPs) encapsulated SwIV vaccine in pigs. Killed SwIV H1N2 (δ lineage) antigens (KAg) were encapsulated in PLGA NPs of 200–300 nm (PLGA-KAg NPs), and influenza antibody-free pigs were prime-boost vaccinated intranasally as mist and challenged using a heterologous, virulent and zoonotic SwIV H1N1 (γ lineage). PLGA-KAg NPs induced maturation of pig macrophages and dendritic cells in vitro. In vaccinated pigs, PLGA-KAg NPs induced antigen specific lymphocyte proliferation and enhanced the frequency of T-helper/memory cells and cytotoxic T cells in peripheral blood mononuclear cells (PBMCs). In virus challenged pigs, the PLGA-KAg NPs vaccine rescued virus induced clinical fever, reduced the gross lung pathology, reduced the virus load in the lung sections with complete clearance of the virus from the lungs of most of the pigs; but the nasal virus shedding was not reduced. Immunologically, at post-challenge day 6 in a recall response in PBMCs of PLGA KAg NPs vaccinated pigs, a significant increase in IFN-γ secreting T cells against both vaccine and challenge viruses were detected. However, humoral immune response in those pigs was not augmented. In conclusion, intranasal delivery of PLGA NPs based SwIV induced cross-protective response through specific cell-mediated response. Future studies are aimed at boosting the mucosal antibody response.Not Availabl

Not Available

Not AvailablePorcine reproductive and respiratory syndrome (PRRS) is a leading cause of economic burden to the pork industry worldwide. The routinely used modified live PRRS virus vaccine (PRRS-MLV) induces clinical protection, but it has safety concerns. Therefore, in an attempt to develop a safe and protective inactivated PRRSV vaccine, we generated PRRS-virus-like-particles (PRRS-VLPs) containing the viral surface proteins GP5-GP4-GP3-GP2a-M or GP5-M using a novel baculovirus expression system. Our in vitro results indicated that the desired PRRSV proteins were incorporated in both the VLPs preparations based on their reactivity in immunogold electron microscopy and ELISA. To boost their immunogenicity in pigs, we entrapped the PRRS-VLPs in PLGA nanoparticles and coadministered them intranasally with a potent adjuvant. We then evaluated their efficacy in pigs against a viral challenge using a virulent heterologous field isolate. Our results indicated that PRRS-VLPs induced an anamnestic immune response, since we observed boosted IgG and IFN-γ production in vaccinated and virus-challenged animals, but not during the pre-challenge period. Importantly, a two-log reduction in the lung viral load was detected in PRRS-VLP-vaccinated animals. In conclusion, we generated PRRS-VLPs containing up to five viral surface proteins and demonstrated their immunogenicity in pigs, but further studies are required to improve its immunogenicity and efficacy as a vaccine candidate.Not Availabl

Not Available

Not AvailablePorcine epidemic diarrhea virus (PEDV) is an economically devastating enteric disease in the swine industry. The virus infects pigs of all ages, but it cause severe clinical disease in neonatal suckling pigs with up to 100% mortality. Currently, available vaccines are not completely effective and feedback methods utilizing PEDV infected material has variable success in preventing reinfection. Comprehensive information on the levels and duration of effector/memory IgA and IgG antibody secreting B cell response in the intestines and lymphoid organs of PEDV-infected sows, and their association with specific antibody levels in clinical samples such as plasma, oral fluid, and feces is important. Therefore, our goal in this study was to quantify PEDV specific IgA and IgG B cell responses in sows at approximately 1 and 6 months post-infection in commercial swine herds, including parity one and higher sows. Our data indicated that evaluation of both PEDV specific IgA and IgG antibody levels in the plasma and oral fluid (but not feces) samples is beneficial in disease diagnosis. PEDV specific B cell response in the intestine and spleen of infected sows decline by 6 months, and this associates with specific antibody levels in the plasma and oral fluid samples; but the virus neutralization titers in plasma remains high beyond 6 months post-infection. In conclusion, in sows infected with PEDV the presence of effector/memory B cell response and strong virus neutralization titers in plasma up to 6 months post-infection, suggests their potential to protect sows from reinfection and provide maternal immunity to neonates, but challenge studies are required to confirm such responses.Not Availabl