Development And Protective Efficacy Of A Universal Vaccine Against H1Subtype Influenza Viruses

Swine influenza is an acute respiratory disease caused by swine influenza virus which belongs tothe family of the Orthomyxoviridae.It often shows high fever, dullness, loss of appetite, laboredabdominal breathing,coughing and caused huge economic losses in pig industry worldwide. H1N1influenza a virus respectively brought global pandemic in1918and2009. The pandemic of1918caused as many as21million people deaded. In2009Influenza a (H1N1) virus which characterized byrapid transmission and large scale epidemics out break, had caused huge impacts on the health ofpeople. Currently, vaccination is the most effective strategy for prophylaxis and control of influenza,but because required continuous manufacturing update, it makes the delay of vaccine investigation andwest resources.So a safe and effective universal vaccine is urgently needed.Genetic vaccination using plasmid DNA represents an exciting means of induce protectivecellular and humoral immune responses and DNA vaccines expressed HA gene had been shudied bysome groups.However, developments of DNA vaccines were limited because of low transfectionefficiency and protein expression level. In this study we first investigate the immune potency of DNAvaccines by optimizing the codon of HA gene then focus on the immunogenicity and protective efficacyof a universal DNA vaccine which expressed consensus HA sequence of all the H1subtype influenzaviruses.On the other hand we also investigate the protective efficacy of M2e peptide vaccine challengedby different subtypes influenza viruses.1.Protective efficacy of DNA vaccine encoding codon-optimized HA gene of H1subtype seineinfluenza virus.The immunogenicity of DNA vaccine directly related to antigen gene expression andimmunogenicity of antigen expression In order to enhance the immunogenicity of DNA vaccine,the HAgene of H1subtype swine influenza virus A/Swine/Guangdong/1/01(H1N1) and recombinant rPan09(the HA and NA genes come from A/California/04/09, the remaining six genes come from PR8)influenza virus were optimized according to the mammalian bias codon usages,then inserted into thepCAGGS vector named pCA-opti-G11and pCA-opti-r09,respectively. The wild-type HA genes werealso constructed and named pCA-G11and pCA-r09. The result of the immune and protection efficacyshowed that the optimized codon of DNA vaccine can significantly enhance the level of humoralimmunity and cellular immunity response and completely suppress viral replication in lungs.2. Protective efficacy of universal vaccine of H1subtype influenza virus.Haemagglutinin is themajor surface glycoproteins of influenza virus, and it is also one of the most important protectiveantigen which can induce neutralizing antibody. In this study, we selected all of the HA gene sequencesof H1subtype classical swine influenza virus and influenza A virus from the GenBank, a consensus HAwas found and optimized according to the swine bias codon usages（optimized consensus, opti-CS）, wealso selected another two HA genes and optimized(one is from A/Swine/Guangdong/1/01strain and theother one is from rPan09strain)then the three optiHAs were inserted into pCAGGS vector,andconstructed three recombinant plasmids named pCA-opti-CS,pCA-opti-G11and pCA-opti-r09 respectively. Then immune BALB/c mouse and challenged by different influenza viruses.The resultshowed, pCA-opti-CS DNA vaccine could induce special humoral immunity and cellular immunityagainst different influenza viruses, reduce the pathological changes from different influenza viruschallenge in lung. It indicated that this vaccine can protect mouse against from heterogeny influenzaviruses.3. Matrix protein2(M2) is a membrane protein of influenza A virus which the N-terminalextracellular-doman is conserved among influenza A viruses, and is considered as a target for designinguniversal influenza vaccines. In this study, we designed a tetra-brabched multiple antigenic peptide(MAP) based vaccine, each peptide contains four copies of M2e and one copy of T helper (Th) cellepitope named M2e-MAP+Freund. The result of immunization and challenge showed that M2e-MAPcould induce strong M2e-specific IgG antibody response and protect mouse against challenge ofheterologous influenza viruses.In conclusion: codon optimization can strongly improve the protective efficacy of DNA vaccine,DNA vaccine expressing consensus HA sequence of H1subtype influenza virus could induce effectiveimmune response and protect mouse against influenza of different subtypes, indicated that plasmidcoding optimizated consensus HA sequence of H1subtype influenza is a putative candidate of universalvaccine investigation. M2e peptide is also a candidate of swine influenza virus vaccine. Therefore, thisstudy set up a foundation for developing a universal vaccine against influenza viruses.