Waterfowl are considered to be the natural hosts of avian influenza virus. In 2017, two reassortant highly pathogenic H5N6 avian influenza viruses of clade 2.3.4.4, subclade II, were identified in wild birds in eastern China. Genome sequencing and phylogenetic and antigenicity analysis showed that the viruses originated from multiple reassortments. To evaluate their pathogenicity in mammals, 15 BALB/c mice were infected with these viruses, and survival and weight loss were monitored for 14 days. Infection was associated with moderate pathogenicity in the mice, and the viruses could replicate in the lungs without prior adaptation. Thus, the existence of these viruses poses a continuous threat to both birds and humans.