Development of a subtype H5 vaccine for pandemic avian influenza

ALEJANDRA CAPOZZO, MANUEL MARTÍNEZ, WIM SCHIELEN,

Buenos Aires

Congreso; III International Clinical Virology Symposium and Advances in Vaccines.; 2010

CEMIC y Pan American Society for Clinical Virology

Background and aims. Influenza viruses have been isolated from a variety of animals. Birds constitute the reservoir of the 16 subtypes of influenza virus A, among them the highly pathogenic avian H5N1. Since 1996, a zoonotic threat of avian influenza (AI) has been identified. The disease is now widespread among poultry and migratory birds in many countries in Southeast Asia. Although person-to-person spread of current H5N1 strains is unlikely, the virus is a potential source of a future influenza pandemic. Moreover, infections with H5N1 strains of AIV have resulted in death in 60% of the cases reported. Infection of poultry with highly pathogenic AI virus is devastating in terms of flock morbidity and mortality. Several studies have shown that the viral hemagglutinin protein (HA) is the principal target of immune response and that response is protective against lethal challenge with live virus. Recombinant HA vaccine may be an attractive alternative to the current inactivated vaccine. Our vaccine approach targets the HA1 external domain of HA, and the recombinant protein is expressed in the baculovirus/insect cell system. Methods. Viral RNA was extracted from inactivated virus (A/Viet Nam 1203/2004, provided by CDC, USA) using Trizol LS reagent (Life Technologies). cDNA synthesis and PCR were performed using One Step RT-PCR Kit (Qiagen), with specific primers targeting the HA1 region. The HA1 fragment was cloned in the baculovirus (Bv) transfer vector pVL1393 (BD Biosciences). Recombinant Bv (rBv) were obtained by co-transfection of Sf9 cells with pVL1393-HA1 and the linearized Bv genome (BaculoGold). Results. rBvs expressing the HA1 region of HA were obtained. Viral particles were cloned by limiting dilution and further amplified in Sf9 cells through a limited number of passages. The stocks were titrated and subjected to a first selection based on protein expression by SDS-PAGE and western blot, and further selection by maximum titers and amount of protein production. Conclusions. The recombinant H5/HA1 protein obtained in insect cells was properly expressed. The optimization of the production and purification processes of the antigen is in course. To count with a vaccine for potential pandemic H5H1 influenza in Argentina is critical for public health.