Data are presented as the mean SEM of 6 to 13 birds per group

Data are presented as the mean SEM of 6 to 13 birds per group. 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 load in the intestines of broilers. Enteritidis, chitosan nanoparticle, mannose modification, antibody response, innate immunity, cell-mediated immunity 1. Introduction serovar Enteritidis (SE) is a Gram-negative bacterium that causes the majority of foodborne illness associated with broilers and is responsible for major economic losses to the U.S poultry industry [1]. Approximately 9% of all-trans-4-Oxoretinoic acid samples from poultry production are positive for [2]. contamination accounted for the greatest number of FDA-regulated food recalls during 2003 through 2011 [3]. Through an effective vaccination approach, Salmonellosis in humans can be substantially decreased by reducing colonization in poultry. Unfortunately, there are limited commercially available vaccines for use in broilers, and none of them provides protective immunity until slaughter. Commercial live vaccines are unsafe as the Rabbit polyclonal to PAX2 live vaccine strains (attenuated by all-trans-4-Oxoretinoic acid natural selection or genetic engineering) are potentially released into the environment and contaminate the human food chain [4]. Currently, due to a lack of effective vaccines and safety reasons, less than 1% of broilers receive a live spray vaccine once [5], and FDA regulations prohibit its use within 21 days of slaughter. Consumption of poultry meat contaminated with is an important cause of infections in humans. Therefore, there is a pressing demand for development of novel control methods that protect broilers from the day of hatch until slaughter against infection. Our previous all-trans-4-Oxoretinoic acid vaccine trial in broilers inoculated orally with chitosan nanoparticles (CS) entrapped with SE outer membrane proteins (OMP) and flagellin (FLA) and surface-coated with FLA, called the CS(OMP+FLA)-F vaccine, was shown to reduce the challenge SE load by 0.7 log10 CFU/g in the cecal content [5]. This outcome was associated with the secretion of increased antigen-specific mucosal and all-trans-4-Oxoretinoic acid systemic antibodies, splenocytes proliferation, and the frequency of IFN-producing T-cell responses. A similar study in layer chickens with CS(OMP+FLA)-F vaccine delivered orally targeted intestinal immune sites and induced mucosal antibody and cell-mediated immune responses, resulting in reduced challenge SE load [6]. Additionally, CS(OMP+FLA)-F-vaccine-treated chicken immune cells showed enhancement of various Toll-like receptors (TLRs) and Th1 and Th2 cytokine gene expression [6]. Mannose-ligand-binding C-type mannose receptor is mainly expressed in the dendritic cells (DCs) and macrophages [7]. In an earlier study, mannose-ligand-modified CS carrying vaccine cargo administered orally was found to target and deliver the loaded antigen to gut DCs in mice [8]. Protein-antigen-encapsulated mannosylated chitosan microspheres delivered orally were shown to bind with mannose receptors on macrophages and induce mucosal antibody responses in mice [9]. all-trans-4-Oxoretinoic acid Mannose-conjugated nanoparticles further improves its adjuvant effect [10], resulting in heightened immunity in the intestines of mice [11]. Therefore, in our present study, to improve the efficacy of the CS(OMP+FLA)-F vaccine, we conjugated mannose with or without FLA on the surface, CS(OMP+FLA)-F&M, and CS(OMP+FLA)-M formulations. These vaccine candidates were administered orally to broiler birds and evaluated for induced immune responses and efficacy compared to an orally delivered commercial live vaccine (Poulvac? ST). The Poulvac? ST is a genetically modified typhimurium (ST) strain, modified by deleting the aroA gene, and provides cross-protection against Kentucky, Enteritidis, Heidelberg and Hadar in birds [12]. 2. Material and Methods 2.1. Experimental Animals, Bacteria, and Vaccines Formulation Day-old Cornish Cross breed broilers were purchased from a commercial hatchery (Ashland, OH, USA). Birds were confirmed for 30 min centrifugation, suspended in milli-Q-water and used for vaccination. The CS (OMP+FLA)-F and CS (OMP+FLA)-M vaccines were prepared similarly but without mannose or FLA. In each dose of vaccines, an equal amount (5 g each) of OMP and FLA were entrapped. 2.2. Experimental Design On the day of hatch, 65 0.05. 2.8. Ethics Statement In accordance with the recommendations of Public Health Service Policy, the United States Department of Agriculture Regulations, the National Research Councils Guide for the Care and Use of Laboratory Animals, and the Federation of Animal Science Societies Guide.