Deciphering transmission dynamics and spillover of avian influenza viruses from avian species to swine populations globally

Abstract

Genome sequences of eleven avian influenza virus (AIV) subtypes have been reported in swine populations from seven countries until August 2020. To unravel the transmission dynamics and spillover events of AIVs from avian reservoirs to swine, full-length hemagglutinin (HA) sequences of AIV subtypes (n = 11) reported from various avian species and swine were retrieved from the 'Influenza Research Database'. Phylogenetic analysis identified closely related avian and swine AIV sequences suggesting potential spillover events from multiple domestic and wild avian species, including chicken, duck, pigeon, goose, quail, and aquatic birds to swine. Furthermore, N-linked glycosylation analysis of these closely related AIV sequences supported the possibility of multiple spillover events of highly pathogenic H5N1 and low pathogenic H9N2 viruses from various avian species to swine. The principal coordinate analysis further validated these findings for H5N1 and H9N2 viruses; however, spillover events of the other nine AIV subtypes were limited. Interestingly, the presence of potential mammalian adaptation markers, particularly in some of the swine H5N1, H7N9, and H9N2 viruses, suggested that these viruses may have already adapted in swine. The occurrence and circulation of these AIVs in swine, especially the H5N1 and H9N2 viruses with numerous spillover events from the avian reservoirs to swine, pose a significant threat in terms of their reassortment with endemic swine viruses or circulating human influenza viruses within the swine which may facilitate the emergence of a novel influenza virus strain with pandemic potential.

Keywords: Avian influenza virus; Avian to swine spillover; IAV adaptation; Influenza pandemic; N-linked glycosylation; Phylogenetic analysis; Principal coordinate analysis; Virus evolution.

Fig. 1

A Neighbour-joining tree of full-length HA nucleotide sequences of avian and swine H5N1 (n = 3544) and H5N2 viruses (n = 1119) suggested multiple spillover events from chicken, ducks and wild birds to swine. The possibility of swine-to-swine transmission of H5N1 viruses in Indonesia was also observed. The cyan nodes represented H5N1 virus transmission among avian species, while the dark cyan nodes represented H5N2 virus transmission among avian species. The red nodes represented H5N1 and H5N2 virus transmission from avian species to swine. The cyan and red dots at the nodes of the subtree represented > 90% bootstrap support. B Seven different glycosylation patterns of swine and closely related avian H5N1 viruses suggested multiple avian to swine transmissions of H5N1 viruses. The identical glycosylation patterns of swine and closely related avian H5N1 viruses represented the possibility of spillover within these groups. Two different patterns of N-linked glycosylation suggested independent introductions of H5N2 viruses in Mexican and Korean swine. C The PCA plot further confirmed multiple spillover events of H5N1 and H5N2 viruses in swine populations. The red dots represented H5N1 and H5N2 viruses in swine, while the cyan and dark cyan dots represented H5N1 and H5N2 viruses, respectively, in avian species reported worldwide. D The 100% amino acid as well as nucleotide sequence identities for a few swine H5N1 viruses (highlighted within two blue boxes) along with identical glycosylation sites (highlighted with red boxes) suggested the possibility of swine-to-swine transmission of these swine H5N1 viruses in Indonesia

Fig. 2

A Neighbour-joining tree of full-length HA nucleotide sequences of 212 avian and one swine H7N2 viruses represented wild bird to swine transmission of the H7N2 virus in South Korea. Neighbour-joining tree of 748 full-length HA nucleotide sequences of swine and avian H7N9 viruses determined chicken to swine spillover of H7N9 viruses in China. A subtree highlighted the circulation of H7N9 viruses between chicken and swine in China. The blue and red dots at the nodes of the subtree represented > 90% bootstrap support. B The identical glycosylation pattern in HA proteins of H7N2 viruses obtained from multiple wild bird species and ducks suggested a broad circulation of H7N2 viruses in avian species in Southeast Asia with one spillover event to swine. The identical N-linked glycosylation patterns shared by swine and closely related avian (chicken) H7N9 viruses supported the findings of phylogenetics suggesting chicken to swine spillover of H7N9 virus in China. C PCA plot represented the global transmission of avian H7N2 viruses with one spillover event to swine and two avian to swine spillover events of H7N9 viruses. The cyan and dark cyan dots represented the circulation of H7N2 and H7N9 viruses, respectively, among avian reservoirs worldwide, while the red dots suggested the occurrence of swine H7N2 and H7N9 viruses

Fig. 3

A Neighbour-joining tree of 6323 full-length HA nucleotide sequences of avian and swine H9N2 viruses determined multiple spillover events from avian reservoirs to swine. The red nodes represented avian to swine transmission of H9N2 viruses in China and Hong Kong, while black nodes represented avian to swine spillover in South Korea. The cyan nodes represented the circulation of H9N2 viruses among avian species globally. The blue dots at the nodes in the subtree represented > 90% bootstrap support. B Four patterns of glycosylation sites in swine and avian H9N2 viruses suggested multiple introductions and evolutionary trajectories of H9N2 viruses in swine in Southeast Asia. C The PCA plot represented multiple spillover events of H9N2 viruses in swine

Fig. 4

A Neighbour-joining tree of 36 full-length HA nucleotide sequences of avian and swine H3N3 viruses determined the wild bird to swine transmission in Ontario. The blue dots at the nodes represented > 90% bootstrap support. B The loss of glycosylation at amino acid residue 499 (NGTY) appeared to have transmitted H3N3 virus to swine; however, it should be noted that the availability of a limited number of avian and swine H3N3 virus sequences restricted our ability to infer avian to swine spillover. C The PCA plot illustrated the occurrence of swine and avian H3N3 viruses globally

Fig. 5

A Neighbour-joining tree of avian and swine H4N1, H4N6, and H4N8 virus HA nucleotide sequences depicted duck to swine transmission of H4N1 and H4N8 viruses in China and mallard to swine transmission of H4N6 viruses in North America. The blue and red dots at the nodes in the subtree represented > 90% bootstrap support. B The identical N-linked glycosylation patterns of swine and duck H4N1 and H4N8 viruses supported the phylogenetic clustering. The glycosylation at amino acid position 14 (NSSQ) appeared crucial for duck to swine spillover of H4N1 virus. The identical glycosylation sites of swine and closely related mallard H4N6 viruses suggested mallard to swine transmission of H4N6 virus in North America. C The PCA plot represented the occurrence of swine and avian H4 subtypes which supported the phylogenetic analyses

Fig. 6

A Neighbour-joining tree of 421 full-length HA nucleotide sequences of swine and avian H6N6 viruses represented two separate events of avian to swine transmission in China highlighted with red nodes. The blue dots at the nodes in the subtree represented > 90% bootstrap support. B The identical glycosylation patterns of pigeon, duck, and swine H6N6 viruses in China suggested a broad circulation of H6N6 viruses in avian species and their sporadic spillover to swine. C The PCA plot confirmed the occurrence of two independent spillover events of H6N6 viruses in Chinese swine

Fig. 7

A Neighbour-joining tree of 91 full-length HA nucleotide sequences of H10N5 viruses reported from avian species and swine determined one spillover event from chicken to swine in China. The closely related chicken and swine H10N5 virus sequences were not contemporary suggesting a circulation of these viruses in avian species at a low frequency in the region. The blue dots at the nodes in the subtree represented > 90% bootstrap support. B The identical glycosylation pattern of chicken and duck H10N5 viruses suggested a common origin of circulating H10N5 viruses in China with one spillover event to swine. C The PCA plot represented the global transmission pattern of avian and swine H10N5 viruses

Fig. 8

A–G Schematic representation of potential mammalian adaptation markers (H3 numbering) [6] in mature HA proteins of swine IAVs. The analysis was conducted using the Sequence Variation (SNP) tool of the Influenza Research Database [65]. A key amino acid substitution for mammalian adaptation, Q226L, was observed in swine H4N6, H7N9 and some of the H9N2 viruses, while another potential mammalian adaptation marker, T160A, was present in several swine H5N1, H5N2, H7N2, H7N9 and H9N2 viruses. A potential swine adaptation marker, D225G [32], was present in all of the swine H9N2 viruses