Influenza H7N9 Virus Hemagglutinin with T169A Mutation Possesses Enhanced Thermostability and Provides Effective Immune Protection against Lethal H7N9 Virus Challenge in Chickens

Abstract

H7N9 avian influenza virus (AIV) has caused huge losses in the poultry industry and impacted human public health security, and still poses a potential threat. Currently, immune prevention and control of avian influenza relies on traditional inactivated vaccines; however, they have some limitations and genetically engineered avian influenza subunit vaccines may be potential candidate vaccines. In this study, a T169A mutation in the HA protein derived from H7N9 AIV A/Chicken/Guangdong/16876 (H7N9-16876) was generated using the baculovirus expression system (BVES). The results showed that the mutant (HAm) had significantly increased thermostability compared with the wild-type HA protein (HA-WT). Importantly, immunizing chickens with HAm combined with ISA 71VG elicited higher cross-reactive hemagglutination inhibition (HI) antibody responses and cytokine (IFN-γ and IL-4) secretion. After a lethal challenge with heterologous H7N9 AIV, the vaccine conferred chickens with 100% (10/10) clinical protection and effectively inhibited viral shedding, with 90% (9/10) of the chickens showing no virus shedding. The thermostability of HAm may represent an advantage in practical vaccine manufacture and application. In general, the HAm generated in this study represents a promising subunit vaccine candidate for the prevention and control of H7N9 avian influenza.

MDCK-Adaptive Mutation of A169S Changes Glycosylation Pattern of Hemagglutinin and Enhances MDCK-Based H7N9 Vaccine Virus Production without Loss of Antigenicity and Immunogenicity

“…The signature oxonium ions of Hex 2 NAc (m/z 366.14), and Hex 3 NAc (m/z 528. 19), and the intact peptide carrying GlcNAc (m/z 2056.97, 1+) were observed in collisioninduced fragments of the glycan portion. A number of fragment peaks were observed at m/z 2584.16 (1+), m/z 2422.11 (1+), m/z 1028.98 (2+), and m/z 926.95 (2+), which reflected a sequential loss of 162 Da (Hex) and 203 Da (HexNac) from this triply charged peptide ion (Figure 2c).…”

“…The removal of glycan from the N149 of the H7N1 HA protein is able to restrict virus replication in MDCK cells [ 18 ]. Chen et al demonstrated that the deletion of N149 glycan can improve the thermal stability and immunogenicity of recombinant HA protein (A/Anhui/1/2013 H7N9) [ 19 ]. Additionally, Chen et al presented that a combination of the glycosylated N149 and other residue substitutions in H7N9 HA improves virus growth in eggs [ 20 ].…”