| A poultry avian influenza of Avian Influenza is caused by type A influenza virus infection and disease syndrome, the prevalence of poultry can be expressed as respiratory diseases,Sub-clinical symptoms and no obvious symptoms of the virus invisible infection, such as a series of clinical symptoms. Avian influenza viruses can be divided into highly pathogenic highly pathogenic avian influenza virus (HPAIV), a low pathogenic avian influenza virus (LPAIV) and non-pathogenic avian influenza virus (NPAIV), depending on the pathogenicity of these three classes.China’s most widely popular H5N1AIV belongs to HPAIV, with infection more object types, a wide host range, variability, and lethal, and other characteristics, and H5N1subtype AIV has been reported for human infection. H5N1AIV subtype caused a serious threat to China’s animal husbandry industry and social and public health and safety. It is by for the necessary and urgent for H5N1subtype of the HPAIV the prevention and control.The vaccine is the primary means of prevention and control of the H5N1subtype of AIV inactivated vaccine by virtue of the immunogenicity and high security features, is widely used. But insufficient to interfere with the clinical and quarantine and epidemiological investigation of inactivated vaccine for avian flu prevention and detection of certain difficulties. Therefore, new vaccines against the H5N1subtype of AIV research is particularly important. Known as the3rd generation of new vaccines, DNA vaccines can simultaneously stimulate the body to generate cellular and humoral immune response and immune duration of the length; easy to build, low cost, a variety of advantages, it has broad prospects.In this study, the characteristics of the high variability of the H5N1subtype of AIV through5000obtained from the NCBI influenza database H5AIV HA protein sequence comparison, analysis of a synonymous HA protein, corresponds to thethe nucleotide sequence of the chicken body Preference of codon optimization, synthetic-Cons,-HA5gene was cloned into the eukaryotic expression vector pCAGGS construct a recombinant expression plasmid pCACons-HA5. And SPF chickens were immunized effectiveness assessment pCACons-HA5. Provide the necessary experimental data for further use of research and clinical applications of pCACons-HA5.For assessment of immune efficacy of pCACons-HA5is the H5N1subtype AIV against different antigens branched immune protection and immune duration of the HI antibody levels of different immunization dose and immune to verify a comprehensive assessment. With pCACons-HA5,15μg,30μg dose of immune SPF chickens after immunization chicken a good mental state, without any clinical and pathological changes in SPF chicken has a good security. The protection force for GS/GD/1/96(CladeO) strains immune protective efficacy trials indicate that two doses of15μg pCACons-HA5SPF chickens can produce the highest value of the average HI antibody levels up to4.33Log2arising as100%.30μg inoculated SPF chickens can produce the highest value of the average HI antibody levels up to4.80Log2protection force to100%. Strain of immune protection against DK/FJ/31/2007(Clade2.3.4a) the effectiveness of the tests showed the the15μg pCACons-HA5inoculated SPF chickens can produce the maximum value of the average HI antibody levels up to2.65Log2, the protection force of100%.30μg inoculated SPF chickens can produce the highest value of the average HI antibody levels up to2.78Log2protection force to100%. Strains of immune protection against DK/GD/S1322/2010(Clade2.3.2c) the effectiveness of the test a15μg pCACons-HA5inoculated SPF chickens can produce the highest value of the average HI antibody levels up0.5Log2, protect the force generated by50%.30μg inoculated SPF chickens can produce the highest value of the average HI antibody levels up to0.83Log2protection force generated by70%. Effectiveness test for immune protection of DK/HuB/S1513/2010(Clade2.3.2b) strains showed that15μg pCACons-HA5SPF chickens after inoculation can produce the maximum value of the average HI antibody levels up to3.23Log2protection force generated by70%. The30μg inoculated SPF chickens can produce the highest value of the average HI antibody levels up to4.30Log2, the protection force of100percent immune protection for CK/SD/A-10/2011(Clade7.2) strain effect test showed that15μg pCACons-HA5.SPF chickens after inoculation can produce the maximum value of the average HI antibody levels up to4.80Log2protection force generated by60%. The30μg inoculated SPF chickens can produce the highest value of the average HI antibody levels up to4.70Log2protection force for the60%. To the15μg pCACons-HA5electrical transduction immune SPF chickens, respectively, with these five kinds of different antigens branch GS/GD/1/96、DK/FJ/31/2007、DK/GD/S1322/2010DK/HuB/S1513/2010、CK/SD/A-10/2011inactivated antigen determination of HI antibody levels average, the highest value of the results of the HI antibody levels were6.1log25.5log2,2.3log25.3log2,5.4log2. The fatal attack protection force and DK/GD/S1322/2010strains up to80%. By immune duration of the test results show that the SPF chickens vaccinated with15μg dose4months after immunization continued observation for five kinds of GS/GD/1/96、DK/FJ/31/2007、 DK/GD/S1322/2010、DK/HuB/S1513/2010、CK/SD/A-10/2011inactivated antigens of different antigens branched average HI antibody were2.2log21.33log2,1.1log21.27log2,3.0log2. The average HI antibody produced by a30ug dose inoculation of SPF chickens were3.0log22.04log2,1.3log22.0log2,3.41og2.Constructed in this study pCACons-HA5vaccination15μg DK/FJ/31/2007(Clade2.3.4a), and GS/GD/1/96(CladeO) to produce a comprehensive protection, when inoculated with30μg of DK/HuB/S1513/2010(Clade2.3.2b), the DK/FJ/31/2007(Clade2.3.4a) and GS/GD/1/96(CladeO) to produce a comprehensive protection, DK/GD/S1322/2010(Clade2.3.2c) and CK/SD/A-10/2011(Clade7.2) has a protective capacity. By electroporation the transduction pCACons-HA5vaccination may be to some extent, improve the immune efficacy of recombinant plasmids in vivo. The results showed that the recombinant plasmid pCACons-HA5the use of anti-H5subtype avian influenza virus DNA vaccines and further reconstruction and significance. |
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