Recombinant Fowlpox Virus Highly Expressing HA From Subtype H9N2 Of Avian Influenza Virus And Its Protective Efficacy Against Homologous Challenge In Chickens

H9 subtype avian influenza virus (AIV) is widly distributed and has resulted in considerable economic losses to poultry industry in China in recent years. In order to protect poultry against this midly pathogenic subtype of AIV, inactivated vaccines in oil emulsion have been developed and proved to be effective in reducing severity of disease and spread of AIV in field situation. However, the use of inactived vaccines was restricted because of the disadvantages they inherited, including higer cost and interference of routine surveillance by serological test. In order to develop new genetic engineering vaccines which are safe, convenient for use and with high protective efficacy, synthetic promoter,Ps , which is 3.5 times stronger than the P7.5 promoter of vaccina virus was used for the construction of recombinant FPV highly expressing HA from subtype H9N2 of Avian influenza virus. The recombinant FPV (rFPV-Ps-HA) was evaluated for its protective efficacy in vaccination trial with spacific-pathogen-free (SPF) chickens, in comparision with recombinant FPV (rFPV-P7.5-HA) and inactived vaccine in oil emulsion. 1. Construction of recombinant FPV highly expressing HA from avain inflenza virusAIV HA gene of F strain in pUCl8HA was removed and inserted into SK to construct plasmid SKHA. Then, the HA gene was inserted into the downstream of Ps to form transfer vector pFGl 18HA. Recombinant FPV was derived by FuGENE?-mediated transfection with transfervector on chicken embryo fibroblast (CEF) monolayer cultures which were infected by wild type FPV Chinese vaccine strain 282E4 3-4 hours earlier. Recombinant FPV was selected and purified by blue plaques expressing β -galactosidase. The expression of foreign gene in rFPV was confirmed by PCR and IFA. 2. Vaccination trails for evaluating protective efficacy of rFPV against homologousATV challengeTo examine the protective efficacy of rFPV-Ps-HA,3 groups of 1-day-old SPF chickens were vaccinated with rFPV-Ps-HA, rFPV-P7.5-HA and inactived vaccine in oil emulsion by subcutaneous route at the neck at the dosage of 105 PFU. All groups were challenged with 107 ELD50 ATV (H9N2 F strain) 16 days postvaccination. Cloacal swabs were taken from all chickens 5 days postchallenged for isolating challenged AIV. It is better for rFPV-Ps-HA to inhabit virus shedding than rFPV-P7.5-HA. Cloacal and tracheal swabs were taken from all chickens 2 5, 7 9 and 11 days postchalleng for isolating challenged AIV. We found that all the vaccinated groups inhaibited virus shedding; The rate of virus shedding was greater for cloaca specimens than for tracheal specimens.