Multiple amino acid substitutions involved in the adaption of three avian-origin H7N9 influenza viruses in mice

Jianru Qin,Lang Gong,Xiaoting Shen,Chunyi Xue,Ouyang Peng,Yongchang Cao

doi:10.1186/s12985-018-1109-1

Abstract

BackgroundAvian influenza A H7N9 virus has caused five outbreak waves of human infections in China since 2013 and posed a dual challenge to public health and poultry industry. The number of reported H7N9 virus human cases confirmed by laboratory has surpassed that of H5N1 virus. However, the mechanism for how H7N9 influenza virus overcomes host range barrier has not been clearly understood.MethodsTo generate mouse-adapted H7N9 influenza viruses, we passaged three avian-origin H7N9 viruses in mice by lung-to-lung passages independently. Then, the characteristics between the parental and mouse-adapted H7N9 viruses was compared in the following aspects, including virulence in mice, tropism of different tissues, replication in MDCK cells and molecular mutations.ResultsAfter ten passages in mice, MLD50 of the H7N9 viruses reduced >750-3,160,000 folds, and virus titers in MDCK cells increased 10-200 folds at 48 hours post-inoculation. Moreover, the mouse-adapted H7N9 viruses showed more expanded tissue tropism and more serious lung pathological lesions in mice. Further analysis of the amino acids changes revealed 10 amino acid substitutions located in PB2 (E627K), PB1 (W215R and D638G), PA (T97I), HA (H3 numbering: R220G, L226S, G279R and G493R) and NA (P3Q and R134I) proteins. Moreover, PB2 E627K substitution was shared by the three mouse-adapted viruses (two viruses belong to YRD lineage and one virus belongs to PRD lineage), and PA T97A substitution was shared by two mouse-adapted viruses (belong to YRD lineage).ConclusionsOur result indicated that the virulence in mice and virus titer in MDCK cells of H7N9 viruses significantly increased after adapted in mouse model. PB2 E627K and PA T97A substitutions are vital in mouse adaption and should be monitored during epidemiological study of H7N9 virus.

Highlights

Avian influenza A H7N9 virus has caused five outbreak waves of human infections in China since 2013 and posed a dual challenge to public health and poultry industry
H7N9-53 and H7N9-MCX belong to Yangtza River Delta (YRD) lineage and H7N9-ZSM belongs to Pearl River Delta (PRD) lineage
Adaption of H7N9 influenza viruses to mice Firstly, the pathogenicity of parental H7N9-53, H7N9-MCX and H7N9-ZSM were evaluated in mice, and the three H7N9 viruses were unlethal to mice even at a high dose of 108.0 50% egg infectious dose (EID50) (Table 1)

Summary

Introduction

Avian influenza A H7N9 virus has caused five outbreak waves of human infections in China since 2013 and posed a dual challenge to public health and poultry industry. The number of reported H7N9 virus human cases confirmed by laboratory has surpassed that of H5N1 virus. A novel reassortant avian influenza H7N9 virus was first detected in February 2013 and since it has posed an unprecedented threat to both public health and poultry industry [1, 2]. The number of laboratory confirmed human infections with H7N9 virus has surpassed that of H5N1 virus [3]. A lot of previous work has convincingly proved that influenza virus could increase virulence in mice through lung-to-lung passage and many critical virulence-related sites were discovered by this method [18,19,20]

Methods

Results

Conclusion