NS gene of influenza virus A/swine/IA/15/30 increases the replication rate of A/Puerto Rico/8/34 in MDCK and Vero cells

Abstract

The reverse genetics (rg) technique has become an alternative approach to produce influenza virus vaccine strains in mammalian cells. The most frequently used genetic backbone for rg–derived viruses is from A/Puerto Rico/8/34 (H1N1) (PR8) virus which is currently approved for vaccine production. Previously, we reported that reassortant viruses with 6 internal genes from A/swine/IA/15/1930 (H1N1) (IA30) and HA and NA from A/chicken/Thailand/KU14/2004 (H5N1) replicated at higher titers than the corresponding PR8 reassortant. In this study, we aim to examine the IA30 genes that can enhance PR8 virus replication. Growth kinetics of PR8 viruses encoding PB2, PB1, PA, NP or NS of IA30 was compared to that of the wild type PR8 (PR8/WT) virus. The replication rate of PR8/IA30NS was significantly higher than the PR8/WT virus in both MDCK and Vero cells. Plaque analysis was also performed in MDCK cells infected with the PR8 viruses encoding IA30 NS or PB1 (PR8/IA30NS or PR8/IA30PB1). It produced significant larger plaque size in the monolayer of MDCK cells, suggesting that PR8/IA30NS could spread to bystander cells more efficiently than PR8/IA30PB1. Our results reveal the function of IA30 NS as a replication enhancer. We suggest the use of IA30 NS in combination with 5 internal genes of PR8 plus HA and NA of the circulating strain in a 5:1:2 reverse genetics system to produce high growth reassortants.