Generation and evaluation of the trivalent inactivated reassortant vaccine using human, avian, and swine influenza A viruses.

Reassortant technology was used to obtain three interspecific reassortant influenza viruses using three influenza viruses of A/Puerto Rico/8/34(H1N1), A/swine/Hebei/1/2005(H3N2) and A/chicken/Guangdong/126/2002(H9N2). The high-growth reassortant strains were H9/PR8, H3/H9N2 and H1/H9N2 that contained hemagglutinin (HA) and neuraminidase (NA) genes from the inactivated parental viruses and the other 6 internal genes from the live parental viruses. The trivalent formalin-inactivated vaccine, containing H1, H3 and H9 subtype antigens from human, swine and avian influenza viruses respectively, was prepared using these reassortant viruses. Animal studies showed that the vaccine was safe and immunogenic. Two-dosing regimen of the influenza vaccine induced high titers of hemagglutination inhibiting (HI) antibodies and influenza-specific IgG antibodies without antigenic cross-interference. It protected 100% chickens from challenge of A/chicken/Guangdong/126/2002 virus and protected 100% mice against challenges with different combinations of the three infective parental viruses. These results indicated that the trivalent vaccine could offer multi-protection against multi-influenza viruses synchronously. This kind of multivalent inactivated reassortant influenza vaccine maybe enlightens the pandemic influenza preparedness as the emergency measure.

[Expression of recombinant Vp6 gene of porcine rotavirus A with non-antibiotic Lactobacillus vector in Escherichia coli].

The antigenic determinants of Vp6 gene of porcine rotavirus A was amplified from infected MA 104 cell by the reverse transcription-polymerase chain reaction (RT-PCR), the product of which was a 1194bp cDNA segment. Using T-A cloning technique, the PCR product was cloned into pGEM-T vector. Cloning plasmid pGEM-T-Vp6 and the prokaryotic shuttle expression vector pW425t between E. coli and Lactobacillus, were digested by SacI and KpnI double enzymes, respectively. The purified Vp6 gene was subcloned into the expression vector pW425t. Thus, the recombinant pW425t-Vp6 was constructed, which then was transformed into the competence thyA gene-mutant E. coli X13. Treated lysates of bacterium were loaded directly onto SDS-PAGE, on which approximately 44.88 kD fusion protein was observed. The protein was further analyzed using Western blot, which indicated that the protein was reactive with the antibody of rotavirus A. The results lay foundation for further studies on the Lactobacillus subunit vaccine and DNA vaccine of Vp6 gene for prevention and control of porcine rotavirus.