Development of American-Lineage Influenza H5N2 Reassortant Vaccine Viruses for Pandemic Preparedness

Abstract

Novel low-pathogenic avian influenza (LPAI) H5N2 viruses hit poultry farms in Taiwan in 2003, and evolved into highly pathogenic avian influenza (HPAI) viruses in 2010. These viruses are reassortant viruses containing HA and NA genes from American-lineage H5N2 and six internal genes from local H6N1 viruses. According to a serological survey, the Taiwan H5N2 viruses can cause asymptomatic infections in poultry workers. Therefore, a development of influenza H5N2 vaccines is desirable for pandemic preparation. In this study, we employed reverse genetics to generate a vaccine virus having HA and NA genes from A/Chicken/CY/A2628/2012 (E7, LPAI) and six internal genes from a Vero cell-adapted high-growth H5N1 vaccine virus (Vero-15). The reassortant H5N2 vaccine virus, E7-V15, presented high-growth efficiency in Vero cells (512 HAU, 107.6 TCID50/mL), and passed all tests for qualification of candidate vaccine viruses. In ferret immunization, two doses of inactivated whole virus antigens (3 μg of HA protein) adjuvanted with alum could induce robust antibody response (HI titre 113.14). In conclusion, we have established reverse genetics to generate a qualified reassortant H5N2 vaccine virus for further development.

Development and Evaluation of Vero Cell-Derived Master Donor Viruses for Influenza Pandemic Preparedness

“…Several egg-derived CVVs, NIBRG-14 (A/Vietnam/1194/2004 (H5N1) × PR8) and NIBRG-268 (A/Anhui/1/2013 (first H7N9 wave) × PR8) were provided by National Institute for Biological Standards and Control (NIBSC), Herts, UK, and IDCDC-RG6 (A/Anhui/1/2005 (H5N1) × PR8), IDCDC-RG30 (A/Hubei/1/2010 (H5N1) × PR8), IDCDC-RG32A (A/Shanghai/2/2013 (1st H7N9 wave) × PR8) were provided by the Centers for Disease Control and Prevention (CDC), Atlanta, GA, USA. Wild-type virus, A/CK/YL/0502/2012 (American-lineage H5N2 virus, R3) was provided by Animal Health Research Institute, Taiwan [ 7 ].…”

“…Our laboratory previously adapted an egg-derived high-growth H5N1 clade 1 CVV in Vero cells (Vero-15) [ 6 ] ( Figure 1 , strategy one) and applied Vero-15 as an MDV to generate high-growth reassortant viruses for avian influenza viruses (AIVs). Our previous data shows that Vero-15 could generate HGRs of H5N1 clade 2 viruses but not American-lineage H5N2 viruses [ 7 ]. Therefore, it is desirable to develop other MDVs to establish the Vero cell-based production platform for multiple AIVs.…”

Synthesis of Asymmetric N-Glycans as Common Core Substrates for Structural Diversification through Selective Enzymatic Glycosylation

“…With a large library of well-defined asymmetric N - glycan isomers in hand, we studied the receptor binding specificity of seven strains of highly pathogenic avian origin influenza viruses of trimeric form (A/Chicken/Taiwan/0502/2012; H5N2). We prepared a glycan array according to the reported protocol , on the N -hydroxysuccinimide (NHS) activated glass slides to understand context based avidity during HA-mediated infection.…”