Research Note: The immune enhancement ability of inulin on ptfA gene DNA vaccine of avian Pasteurella multocida.

To evaluate the ability of inulin to enhance the immune response of a ptfA gene DNA vaccine for avian Pasteurella multocida, inulin was added as an adjuvant to the ptfA-DNA vaccine, obtaining an inulin-adjuvant DNA vaccine. The DNA vaccine was administered to chickens; a fimbria protein vaccine and an attenuated live vaccine were used as positive controls. The levels of the serum antibody and concentrations of interferon-γ (IFN-γ), interleukin-2 (IL-2), and interleukin-4 (IL-4) were determined, and a lymphocyte proliferation assay was performed. After being challenged with virulent P. multocida, the protective efficacy was evaluated. The results showed that the serum antibodies induced by the ptfA-DNA vaccine were not enhanced by inulin. The stimulation index values and the concentrations of IL-2 and IFN-γ in chickens vaccinated with inulin-adjuvant DNA vaccine were significantly higher than those in chickens vaccinated with the DNA vaccine, those with the fimbria protein vaccine, and the chickens gavaged with inulin. The concentrations of IL-4 in the inulin-adjuvant DNA vaccine group and the fimbria protein vaccine group were higher than those in the DNA vaccine group and the inulin-gavage group. The protective efficacy rates of the attenuated live vaccine group, the fimbria protein vaccine group, the DNA vaccine group, the inulin-adjuvant DNA vaccine group, and the inulin-gavage group were 90, 70, 55, 65, and 55%, respectively.

Immune efficacy of DNA vaccines based on oprL and oprF genes of Pseudomonas aeruginosa in chickens.

Pseudomonas aeruginosa (P. aeruginosa) is a zoonotic pathogen that can infect a variety of animals, including poultry. However, as there is no commercial vaccine available it is imperative that new and effective vaccines are developed. In this study, 2 monovalent DNA vaccines (pOPRL and pOPRF), one divalent combination DNA vaccine (pOPRL+pOPRF) and one fusion DNA vaccine (pOPRLF) were constructed based on the oprL and oprF genes of P. aeruginosa. These vaccines were administered to chickens, an outer membrane protein vaccine (OMP vaccine) and inactivated vaccine used as positive controls. The serum antibody, interferon-γ (IFN-γ), interleukin-2 (IL-2) and interleukin-4 (IL-4) concentrations were determined and lymphocyte proliferation assays were performed. After challenging with virulent P. aeruginosa, protective efficacy was evaluated. Following vaccination, serum antibodies, stimulation index (SI) values, concentrations of IL-2 and IFN-γ in chickens vaccinated with the bivalent combination DNA vaccine and fusion DNA vaccine were found to be significantly higher than in those chickens vaccinated with the 2 monovalent DNA vaccines. Moreover, the immune indexes in the bivalent combination DNA vaccine group were higher than those in the fusion DNA vaccine group. However, the concentrations of IL-4 in the 4 DNA vaccine groups were of no significant difference. The protective efficacy rate provided by pOPRL, pOPRF, pOPRLF, pOPRL+pOPRF, inactivated vaccine and OMP vaccine were 53.3%, 40%, 66.7%, 80%, 93.3%, and 80%, respectively. The results indicate that DNA vaccines constructed with the oprL and oprF genes of P. aeruginosa, particularly the divalent combination DNA vaccine, represent better potential vaccines. This study has laid a foundation for the design and application of future DNA vaccines of P. aeruginosa.

Construction of a ptfA chitosan nanoparticle DNA vaccine against Pasteurella multocida and the immune response in chickens.

The aim of this study was to evaluate the effect of chitosanon the immune response induced by a DNA vaccine based on the ptfA gene of avian Pasteurella multocida. Naked DNA vaccine was packed with chitosanmolecules, resulting in a chitosannanoparticle DNA vaccine. The encapsulation efficiency, shape, size and resistance to DNA degradation were determined. The vaccine was administered to chickens and serum antibody, interferon-γ (IFN-γ), interleukin-2 (IL-2) and interleukin-4 (IL-4) concentrations were determined and lymphocyte proliferation assays were performed. After challenge with virulent avian P. multocida, protective efficacy was evaluated. The encapsulation efficiency of the chitosan nanoparticle DNA vaccine was 95.3%. The particle size was approximately 200nm and close to spherical in shape and it could effectively resist degradation by DNases. Following vaccination, serum antibodies, stimulation index (SI) value and concentrations of IFN-γ and IL-2 in chickens vaccinated with the chitosan nanoparticle DNA vaccine were significantly higher than those that were vaccinated with the naked DNA vaccine (P-values are 0.026, 0.045, 0.039 and 0.024, respectively). However, the concentrations of IL-4 in the two DNA vaccines group were no significant difference (P=0.157). The protective efficacy rate provided by naked DNA vaccine, chitosan nanoparticle DNA vaccine and the attenuated live vaccine were 56%, 68% and 88%, respectively. The results indicated that chitosan was able to enhance the immune response to a naked DNA vaccine based on the ptfA gene of P. multocida.

Evaluation of immunogenicity and protective efficacy of recombinant ptfA of avian Pasteurella multocida.

Avian Pasteurella multocida is the causative agent of fowl cholera, a disease much affecting the poultry industry. In order to study the efficacy of the recombinant subunit vaccine constructed with ptfA gene of avian P. multocida, the ptfA gene fragment amplified by PCR from avian P. multocida was cloned into the prokaryotic expression vector pET32a and the recombinant plasmid pET32a-ptfA was obtained. The pET32a-ptfA was expressed in Escherichiacoli BL21(DE3) and the target protein rPtfA was purified. The purified protein was then mixed with Freund's adjuvant and the recombinant subunit vaccine was obtained. Three groups of chickens labeled as rPtfA, attenuated live vaccine and PBS were vaccinated with the recombinant subunit vaccine, attenuated live vaccine and PBS, respectively. Serum antibodies, peripheral blood lymphocyte proliferation (PBLP) and interferon-γ (IFN-γ) level secreted by peripheral blood lymphocyte were tested. The immunized chickens were finally challenged with virulent avian P. multocida and the protection rate was counted. Indirect ELISA showed the levels of antibodies in rPtfA and attenuated vaccine groups were most significantly higher than the other groups (P<0.01), and the former was slightly lower than the latter. Peripheral blood lymphocyte proliferation experiments and IFN-γ experiments indicated that SI value and the levels of IFN-γ induced by ConA in the two vaccine groups were significantly higher than those of the PBS groups (P<0.01), and that the attenuated vaccine group was higher than the rPtfA group. The protection rates of rPtfA and attenuated live vaccines were 45% and 75%, respectively. The results indicated that the PtfA recombinant subunit vaccine was capable of improving the immunity level and inducing a protective effect for the vaccinated chickens, but it was barely satisfactory.