Adjuvant Effect Of LBP On Influenza Inactivated Vaccine And Preliminary Study On Construction And Immunogenicity Of Multi-epitope Influenza DNA Vaccines

The paper consisits of three parts:(1) brief introduction of influenza virus, influenza vaccine and adjuvants; (2) adjuvant effect of Lycium barbarum polysaccharide on influenza inactivated whole-virion vaccine; (3) construction and immunogenicity of multi-epitope influenza DNA vaccines.The first part serves as an overview which outlines the basic characteristics of influenza virus, research progress on influenza vaccines and adjuvants.In the second part, adjuvant effect of Lycium barbarum polysaccharide on inactivated whole-virion influenza vaccine was reported. Mice were intraperitoneally immunized with the whole-virion inactivated influenza virus (A/Vietnam/1194/2004 (H5N1)) vaccine formulated with different dosages of Lycium barbarum polysaccharide. Mice injected with aluminium hydroxide-adjuvanted vaccine and vaccine alone were used as controls. Three weeks after the immunization, ELISA method was used to detect the virus specific IgG, IgG 1 and IgG 2a. HI method was used for determination of neutralizing antibody. Challenge test was also performed at the time by intranasal administration with a lethal dose (10×LD50) of A/Chicken/Henan/12/2004(H5N1) virus. The protective effect after immunization were evaluated by detection of body weight changes, lung virus titers and survival rates. Results showed that, when compared with the vaccine alone, LBP-adjuvanted vaccine could significantly enhance the antibody levels and protective abilities of mice against lethal dose virus challenge. The adjuvant effect of LBP were comparable to that of alum adjuvant. LBP may be used as a potential adjuvant of inactivated influenza virus vaccine.In the third part, multiple dominant epitopes in antigens of H1, H3, H5 and H9 subtypes were selected based on the bioinformatics. Two fusion genes expressing multiple epitopes were designed and synthesized, and were then cloned into expression plasmid pCAGGSP7 to construct two recombinants pCAGGSP7/MP and pCAGGSP7/ME, respectively. The immunogenicity of the multi-epitope DNA vaccines was preliminarily analysed in BALB/c mice. Mice were immunized three times at a 2-week interval. Sera of immunized mice were collected weekly to analyze with ELISA detection. Two weeks after the third immunization, the splenic lymphocytes were separated and analyzed for detecting theSubgroups, and mice were challenged with lethal dose of H1, H3, H5 and H9 subtype viruses, respectively. The results showed that pCAGGSP7/MP and pCAGGSP7/ME immunized mice acquired satisfactory humoral and cellular immune responses to several serotypes (H1/H3/H5/H9) of influenza A virus; and pCAGGSP7/MP and pCAGGSP7/ME afforded partial protection in mice against lethal challenge with different subtype viruses. It suggested that DNA vaccines based on epitope design may be a useful tool to fight against different subtypes of influenza infection.