A recent publication in this Journal reported the emergence of an H5N6 strain originating from farm dogs in China. H5N6 subtype avian influenza strains in Clade 2.3.4.4 have circulated in poultry in China since 2013. In 2014, highly pathogenic avian influenza H5 GsGd (Clade 2.3.4.4) caused multiple outbreaks on several continents. Studies have shown that H5N6 AIV can infect not only poultry and wild birds, but also mammals such as pigs, cats, and even humans. China has adopted a vaccine immunization policy to prevent the spread and prevalence of influenza viruses among poultry. However, this strategy does not prevent the spread and evolution of influenza viruses in waterfowl, wild birds and mammals. Avian influenza viruses carried by wild birds can not only cause avian influenza outbreaks but also infect companion animals that go outdoors, as in the case of dogs in Guangxi. Therefore, monitoring avian influenza viruses carried by wild birds has become important.
In the course of avian influenza surveillance at the wild bird interface in the Pearl River Delta region, our laboratory isolated a wild-bird-derived H5N6 subtype AIV strain from wild bird fecal samples collected in Shenzhen in March 2023. This strain was named A/wild bird/Guangdong/SZ1/230322/ H5N6. We sequenced the whole genome of the virus and analyzed its genetic evolution. The HA phylogenetic tree showed that the isolate was located in Clade 2.3.4.4b, which is in Clade 2.3.4.4b with the strains currently prevalent in poultry hosts on the live bird market and the vaccine strains currently found in China (Re-14 and rHN5801). SZ1 belongs to the same branch as the H5N6 strain originating from farm dogs reported by Yao et al. in GISAID in 2023.
Genetic analyses showed that the cleavage site of the HA gene in A/Wildbird/Guangdong/SZ1/230322/H5N6 contained the contiguous basic amino acid REKRRKR↓GLF, which is consistent with the molecular profile of highly pathogenic strains. The SZ1 strain has mutations in the HA gene at the S137A and T192I loci, a double-site mutation that results in enhanced binding of the virus to the α-2,6-linked SAs (human-type receptors), which is characterized by enhanced binding to the mammalian receptor. 226Q and 228?G indicated that these strains have an affinity for the avian-type receptors. T33K, L89V, R477G, I495V, E627K, and 701D in the PB2 gene, L13P, D538, K578, and 614E in the PB1 gene, A150R and A184K in the NP gene, E133 and G631S in the PA gene, D97E in the NS gene, and N30D and T215A mutations in the M1 gene not only enhance receptor binding but also increase virulence in mammals. BLAST of the GenBank database showed that the sequences of the HA and NA fragments observed in this study showed the highest nucleotide sequence identity with A/Canine/China/GX30/2023 (H5N6) and A/duck/Sichuan/SC5698/2021 (H5N6), 98.59% and 98.62%, respectively. The sequences of the NS, PA, PB2, and PB1 fragments of the internal genes were similar to the canine-derived virus from Guangxi, China. The nucleotide similarity between the M segment sequence and the human source sequence from Guangdong, China, was 99.49% (Table S1). Fig. 2 shows a schematic diagram of the triple reassortment of human, canine, and avian gene fragments it carries.