A Replicating Modified Vaccinia Tiantan Strain Expressing an Avian-Derived Influenza H5N1 Hemagglutinin Induce Broadly Neutralizing Antibodies and Cross-Clade Protective Immunity in Mice

Xiao, Haixia; Liu, Li; Zhu, Qingyu; Tan, Zhiwu; Yu, Wenbo; Tang, Xian; Zhan, Dawei; Du, Yanhua; Wang, Haibo; Liu, Di; Li, Zhixin; Yuen, Kwok-Yung; Ho, David D.; Gao, George F.; Chen, Zhiwei

HA XJ /WSN stands for the recombinant influenza A virus with its HA gene derived from the human strain A/Xingjiang/1/06 (H5N1) and the other seven genes from A/WSN/33 (H1N1). This reverse genetics system was kindly provided by Yoshiro Kawaoka [25] . Two wild-type H5N1 influenza viruses, A/BhG/QH/1/05 and A/Vietnam/1194/2004 (A/VN/1194/04), were used for challenge in this study. A/BhG/QH/1/05 and A/VN/1194/04 were cultivated in 10-day-old embryonated SPF chicken eggs (Beijing Merial Vital Laboratory Animal Technology Co., Ltd., Beijing, China), while A/HA XJ /WSN was propagated on Madin-Darby canine kidney (MDCK) cells. MDCK cells and Vero cells were cultured in Dulbecco's modified Eagle's minimum essential medium (DMEM) supplemented with 10% fetal calf serum (FCS) at 37uC in an atmosphere of 5% CO 2 . Allantoic fluids and cell cultures were harvested and stored at 270uC. The fifty percent mouse lethal dose (MLD 50 ) of each strain was determined in 6week-old female BALB/c mice and the fifty percent tissue culture infectious dose (TCID 50 ) was titrated in MDCK cells. All of these experiments were conducted in a bio-safety level-3 laboratory. A group of diverse H5N1 pseudoviruses were prepared according to recently described methods [10] .