Genetic Characterization of Continually Evolving Highly Pathogenic H5N6 Influenza Viruses in China, 2012-2016

H5N6 is a highly pathogenic avian influenza (HPAI) and a zoonotic disease that causes recurring endemics in East Asia. At least 155 H5N6 outbreaks, including 15 human infections, have been reported in China. These repeated outbreaks have increased concern that the H5N6 virus may cross over to humans and cause a pandemic. In February, 2016, peafowls in a breeding farm exhibited a highly contagious disease. Post-mortem examinations, including RT-PCR, and virus isolation, confirmed that the highly pathogenic H5N6 influenza virus was the causative agent, and the strain was named A/Pavo Cristatus/Jiangxi/JA1/2016. In animal experiments, it exhibited high pathogenicity in chickens and an estimated median lethal dose in mice of ~10^4.3 TCID₅₀. A phylogenetic analysis showed that JA1/2016 was clustered in H5 clade 2.3.4.4. FG594-like H5N6 virus from Guangdong Province was the probable predecessor of JA1/2016, and the estimated divergence time was June 2014. Furthermore, we found that H5N6 influenza viruses can be classified into the two following groups: Group 1 and Group 2. Group 2 influenza viruses have not been detected since the end of 2014, whereas Group 1 influenza viruses have continually evolved and reassorted with the "gene pool" circulating in south China, resulting in the rise of novel subtypes of this influenza virus. An increase in the number of its identified hosts, the expanding range of its distribution, and the continual evolution of H5N6 AIVs enhance the risk that an H5N6 virus may spread to other continents and cause a pandemic.