Elsevier

The Lancet

Volume 381, Issue 9881, 1–7 June 2013, Pages 1926-1932
The Lancet

Articles
Origin and diversity of novel avian influenza A H7N9 viruses causing human infection: phylogenetic, structural, and coalescent analyses

https://doi.org/10.1016/S0140-6736(13)60938-1Get rights and content

Summary

Background

On March 30, 2013, a novel avian influenza A H7N9 virus that infects human beings was identified. This virus had been detected in six provinces and municipal cities in China as of April 18, 2013. We correlated genomic sequences from avian influenza viruses with ecological information and did phylogenetic and coalescent analyses to extrapolate the potential origins of the virus and possible routes of reassortment events.

Methods

We downloaded H7N9 virus genome sequences from the Global Initiative on Sharing Avian Influenza Data (GISAID) database and public sequences used from the Influenza Virus Resource. We constructed phylogenetic trees and did 1000 bootstrap replicates for each tree. Two rounds of phylogenetic analyses were done. We used at least 100 closely related sequences for each gene to infer the overall topology, removed suspicious sequences from the trees, and focused on the closest clades to the novel H7N9 viruses. We compared our tree topologies with those from a bayesian evolutionary analysis by sampling trees (BEAST) analysis. We used the bayesian Markov chain Monte Carlo method to jointly estimate phylogenies, divergence times, and other evolutionary parameters for all eight gene fragments. We used sequence alignment and homology-modelling methods to study specific mutations regarding phenotypes, specifically addressing the human receptor binding properties.

Findings

The novel avian influenza A H7N9 virus originated from multiple reassortment events. The HA gene might have originated from avian influenza viruses of duck origin, and the NA gene might have transferred from migratory birds infected with avian influenza viruses along the east Asian flyway. The six internal genes of this virus probably originated from two different groups of H9N2 avian influenza viruses, which were isolated from chickens. Detailed analyses also showed that ducks and chickens probably acted as the intermediate hosts leading to the emergence of this virulent H7N9 virus. Genotypic and potential phenotypic differences imply that the isolates causing this outbreak form two separate subclades.

Interpretation

The novel avian influenza A H7N9 virus might have evolved from at least four origins. Diversity among isolates implies that the H7N9 virus has evolved into at least two different lineages. Unknown intermediate hosts involved might be implicated, extensive global surveillance is needed, and domestic-poultry-to-person transmission should be closely watched in the future.

Funding

China Ministry of Science and Technology Project 973, National Natural Science Foundation of China, China Health and Family Planning Commission, Chinese Academy of Sciences.

Introduction

On March 30, 2013, a novel avian influenza A H7N9 virus causing human infections was identified in China.1 As of April 18, 2013, the virus had spread to six provinces and municipal cities—ie, Shanghai, Anhui, Jiangsu, Zhejiang, Beijing, and Henan. To our knowledge, this outbreak represents the first time that the H7N9 subtype has infected people and caused fatal cases (as of April 18, 2013, 87 people have been infected and 17 have died). In modern times, H7 subtype avian influenza viruses, including H7N1, H7N2, H7N3 and H7N7, have caused more than 100 human infections, including a fatal case in the Netherlands.2, 3, 4 In wild birds, H7 and N9 avian influenza viruses have evolved to American, Oceanian, and Eurasian lineages.5, 6 Preliminary analyses1 have shown that the H7N9 viruses causing the 2013 outbreak in China are novel reassortants, the HA gene originated from avian influenza viruses circulating in ducks in Zhejiang Province, the NA gene is related to avian influenza viruses isolated from wild birds, and the internal genes probably orinigated from an earlier H9N2 lineage. Kageyama and colleagues7 have postulated that the internal genes originated from poultry viruses. In this study, we integrated data from phylogenetic analyses, coalescent analyses, and host ecology to infer the potential origins and genetic diversity of the novel avian influenza A H7N9 viruses.

Section snippets

Sequence alignment

We downloaded the H7N9 virus genome sequences from the Global Initiative on Sharing Avian Influenza Data (GISAID) database—specifically, A/Anhui/1/2013 (Anhui/1), EPI439503∼EPI439510; A/Shanghai/1/2013 (Shanghai/1), EPI439486∼EPI439491, EPI439493, EPI439494; A/Shanghai/2/2013 (Shanghai/2), EPI439495∼EPI439502; and A/Hangzhou/1/2013 (Hangzhou/1), EPI440095∼EPI440097. We downloaded all the public sequences used from the Influenza Virus Resource.8 CLC Main Workbench (CLC Bio, Aarhus, Denmark) was

Results

The phylogeny of the H7 gene sequences available showed three main independent lineages—ie, American, Oceanian, and Eurasian lineages. The novel H7N9 virus fell within the Eurasian lineage, and was genetically close to sequences isolated from ducks in Zhejiang Province in 2011 (Figure 1, Figure 2). The H7 phylogenetic tree also showed that varied H7 viruses were circulating in wild ducks along the east Asian flyway, which covers eastern China, South Korea, and Japan. Genetic exchange between

Discussion

Tracing the origin of the novel H7N9 virus is of vital importance for formulation of effective prevention and surveillance policies. We propose at least four possible gene-segment origins are proposed for the novel H7N9 virus (panel).

The estimated time to most recent common ancestor of the novel H7N9 strain was during the wintering period for wild birds. Thus, corresponding H7 strains might have been circulating in poultry (most likely in ducks) for at least 1 year.

An earlier H11N9 strain of

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These authors contributed equally to this work

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