This study aimed to investigate whether bacterial ghosts (BGs), empty cell envelopes of a gram-negative bacterium, delivering envelope protein domain III (EDIII) of dengue virus (DENV) serotype 2 could induce protective immune responses against dengue infection. In this study, we constructed Salmonella Typhimurium BGs expressing and delivering EDIII (BG-EDIII) and evaluated these ghosts for their immunogenicity studies in C57BL/6 mice. Our results demonstrated that the mice vaccinated once orally with BG-EDIII followed by an intramuscular boosting with a recombinant EDIII protein elicited significantly higher humoral and cell-mediated immune responses compared to the BGs alone vaccinated group (p<0.001). Upon challenge with DENV2, significantly lower viral load and liver damage was observed in BG-EDIII vaccinated group than BGs alone control group (p<0.05). The outcomes of this study revealed the ability of BG- EDIII to stimulate immune response with no observable damage to the vital organs.

Salmonella Enteritidis (SE) infection, one of a major foodborne disease remains considerable public health concerns globally. In this study, we have constructed that the genetically inactivated SE ghost vaccine candidate, JOL2220 (Δlon ΔcpxR Δasd), which express the immunogenic hemagglutinin portion (HA1) of H1N1 virus on the Salmonella surface. The lysis gene cassette, holin-endolysin system originated from double DNA bacteriophage λ was cloned into the ghost vector plasmid pJHL464, which activated under the control of the convergent promoter system. Scanning electronic microscopic examination revealed that transmembrane tunnels of JOL2220 was formed following the 48 hr of the lysis mediated by the holin-endolysin system. In vitro expression of HA1 antigenic protein was also determined by Western blot analysis. The Chickens immunized with JOL2220 ghost via intramuscular and oral routes showed significant increase in IgG level against both SE and HA1 antigen. Further, we observed markedly enhancement of lymphocyte proliferation and T-cell differentiation in the primed PBMC of the immunized chicken, compared to those in the control group. Collectively, the results indicated that the SE ghosts expressing HA1 antigen may have immunostimulatory properties against Salmonella and influenza infection where the chicken is the predominant reservoir.

Shiga toxin 2e (Stx2e) has a pivotal role in the colonization and enterotoxicity of F18+Shiga toxin-producing Escherichia coli (STEC), which causes porcine edema disease (ED). In this study, a Stx2eA mutant, which has a Glu167Gln mutation in Stx2eA that inactivates N-glycosidase activity, was genetically engineered to evaluate its potential immunogenicity and protective efficacy. A significant increase in serum IgG1 (a Th2 indicator) was shown in mice immunized with the mutated Stx2eA. However, only 56% of the mice immunized with the toxoid (5 μg) survived following a challenge with a lethal dose 50 (LD50) of a virulent F18+STEC strain (JOL654), while mice immunized with Salmonella ghosts delivering selected antigens of F18+STEC showed an 86% survival rate. The results suggest that sole use of the mutated Stx2eA toxoid may not be an effective preventive strategy for the control of porcine ED.