Evolution of enzootic H9N2 influenza virus in China and its role in the genesis and development of other zoonotic viruses

Abstract

The H9N2 influenza virus has been enzootic for two decades in land-based poultry in multiple Asian countries, causing sporadic infections in mammals and economic loss. With the lineages maintained majorly by chickens, the ecology and evolutionary behavior of the H9N2 viruses have been shaped by the farming practice and poultry trade in China. Recently, novel human-infecting viruses including H5N6, H7N9 and H10N8 subtypes emerged all through reassortment, using the backbone of the predominant H9N2 virus. Therefore, it is critical to define the dynamics of the H9N2 virus in chickens and its role in the genesis and development of novel zoonotic viruses. Through our systematic surveillance at live poultry markets in six provinces in southern China from 2006-2014, the epidemiology and evolutionary pattern of the most prevalent H9N2 viruses in chicken have been addressed. Additional data from recent outbreaks of the H7N9 influenza virus were utilized for the characterization of the viruses and factors that driving the emergence and dissemination of the novel reassortants in current influenza ecosystem in China. Phylogenetic analysis of the full genomes of 337 chicken H9N2 viruses generated in this study and the public available sequences revealed an unexpected reduction in the genetic heterogeneity accompanied with the increase in prevalence of the Zhejiang/HJ-like H9N2 virus. Genotype replacement of the H9N2 viruses and antigenic drift of the predominant Zhejiang/HJ-like H9 were detected. The novel antigenic H9N2 viruses rapidly expanded their geographic distribution. A decrease in the relative genetic diversity of the Zhejiang/HJ-like H9 may be a reflection of the selection bottleneck associated with the clonal expansion of the antigenically drifted variant. From 2013 to 2014, several novel subtypes of influenza virus harboring the Zhejiang/HJ-like internal gene cassette emerged in chickens. Different from other novel subtypes, the H7N9 virus disseminated quickly in chickens in China, led to hundreds of infections in human. By analyzing the 438 H7N9 genomes and 263 related viruses sampled during the second wave of the outbreak, the H7N9 virus was found diverged into three distinct clades based on their surface genes and became establishment in the field. Frequent reassortment between H7N9 and contemporary H9N2 viruses have caused the fast generation of genetic variants. The establishment of the novel H7N9 subtype in chickens was likely associated with the advantage in high genetic compatibility between the antigenically escaped surface genes and the highly efficient internal gene cassette of the H9N2 virus, the absence of immune pressure and the frequent transportation of chickens. These findings demonstrated the significant role of enzootic chicken viruses in facilitating the emergence and development of novel zoonotic viruses in China, as exemplified by the H9N2 and H7N9 subtypes. The association of poultry transportation with avian influenza virus evolution and disease spreading highlighted the additional strategy for outbreak control and viral elimination under current influenza ecosystem.