H5N8 and H5N1 high pathogenicity avian influenza viruses (HPAIVs) caused outbreaks in poultry farms in Japan from November 2021 to May 2022. Hemagglutinin genes of these viruses belong to clade 2.3.4.4B and can be divided phylogenetically into the following groups: 20A, 20E, and 21E. In this study, we compared the infectivity and transmissibility of HPAIVs from three groups of chickens. Representative strains from 20A, 20E, and 21E groups are A/chicken/Akita/7C/2021(H5N8)(Akita7C), A/chicken/Kagoshima/21A6T/2021(H5N1)(Kagoshima6T), and A/chicken/Iwate/21A7T/2022(H5N1)(Iwate7T), respectively. Fifty percent lethal dose of Akita7C in chickens (10^3.83 fifty percent egg infectious dose (EID50)) was up to seven times lower than those of Kagoshima6T and Iwate7T (10^4.50 and 10^4.68 EID_50, respectively). Mean death times for Akita7C- and Kagoshima6T-infected chickens (3.45 and 3.30 days, respectively) were at least a day longer than that of Iwate7T (2.20 days). Viral titers of the trachea and cloaca of Iwate7T-infected chicken were the highest detected. The transmission rate of the Akita7C strain (100%) was markedly higher than those of the two strains (<50%). These data suggest that the infectivity and transmissibility of the Akita7C strain (H5N8) in chickens are higher than those of H5N1 viruses, providing fundamental information needed for formulating effective prevention and control strategies for HPAI outbreaks.

Genetically related highly pathogenic avian influenza viruses (HPAIVs) of H5N6 subtype caused outbreaks simultaneously in East Asia and Europe—geographically distinct regions—during winter 2017–2018. This situation prompted us to consider whether the application of phylogeographic analysis to a particular gene segment of AIVs could provide clues for understanding how AIV had been disseminated across the continent. Here, the N6 NA genes of influenza viruses isolated across the world were subjected to phylogeographic analysis to illustrate the inter- and intracontinental dissemination of AIVs. Those isolated in East Asia during winter and in Mongolia/Siberia during summer were comingled within particular clades of the phylogeographic tree. For AIVs in one clade, their dissemination in eastern Eurasia extended from Yakutia, Russia, in the north to East Asia in the south. AIVs in western Asia, Europe, and Mongolia were also comingled within other clades, indicating that Mongolia/Siberia plays an important role in the dissemination of AIVs across the Eurasian continent. Mongolia/Siberia may therefore have played a role in the simultaneous outbreaks of H5N6 HPAIVs in Europe and East Asia during the winter of 2017–2018. In addition to the long-distance intracontinental disseminations described above, intercontinental disseminations of AIVs between Eurasia and Africa and between Eurasia and North America were also observed. Integrating these results and known migration flyways suggested that the migration of wild birds and the overlap of flyways, such as that observed in Mongolia/Siberia and along the Alaskan Peninsula, contributed to the long-distance intra- and intercontinental dissemination of AIVs. These findings highlight the importance of understanding the movement of migratory birds and the dynamics of AIVs in breeding areas—especially where several migration flyways overlap—in forecasting outbreaks caused by HPAIVs.

In Japan during the 2016–2017 winter season, clade 2.3.4.4 highly pathogenic avian influenza viruses (HPAIVs) of the H5N6 subtype caused 12 outbreaks in chicken and Muscovy duck farms. These viruses have been circulating in Vietnam and China since 2014. In this study, we evaluated the susceptibility of chicken, Pekin duck, and Muscovy duck to H5N6 HPAIVs that originated in Japan, Vietnam, and China. The H5N6 HPAIVs examined in this study were highly lethal to chickens compared with their pathogenicity in Pekin duck and Muscovy duck. One of five chickens infected with A/Muscovy duck/Aomori/1‐3T/2016 (MusDk/Aomori) survived despite viral shedding, although all of the chickens infected with the other viruses died. The 50% chicken lethal dose differed among the Japanese strains that shared the same gene constellation, indicating that gene constellation was not a major determinant of pathogenicity in chicken. MusDk/Aomori, A/chicken/Niigata/1‐1T/2016 (Ck/Niigata), and A/duck/Hyogo/1/2016 (Dk/Hyogo) infected all Muscovy ducks inoculated; Ck/Niigata killed 50% of the ducks it infected whereas the other two did not kill any ducks. A/chicken/Japan/AnimalQuarantine‐HE144/2016 (HE144) isolated from chicken meat that originated in China was highly pathogenic to Pekin duck: all of the ducks died within 3.75 days of inoculation. This study shows that the pathogenicity of the clade 2.3.4.4 H5N6 HPAIVs differs not only between hosts but also within the same host species.

On 5 November 2020, a confirmed outbreak due to an H5N8 highly pathogenic avian influenza virus (HPAIV) occurred at an egg-hen farm in Kagawa prefecture (western Japan). This virus, A/chicken/Kagawa/11C/2020 (Kagawa11C2020), was the first HPAI poultry isolate in Japan in 2020 and had multiple basic amino acids—a motif conferring high pathogenicity to chickens—at the hemagglutinin cleavage site. Mortality of chickens was 100% through intravenous inoculation tests performed according to World Organization for Animal Health criteria. Phylogenetic analysis showed that the hemagglutinin of Kagawa11C2020 belongs to clade 2.3.4.4B of the H5 Goose/Guangdong lineage and clusters with H5N8 HPAIVs isolated from wild bird feces collected in Hokkaido (Japan) and Korea in October 2020. These H5N8 HPAIVs are closely related to H5N8 HPAIVs isolated in European countries during the winter of 2019–2020. Intranasal inoculation of chickens with 10⁶ fifty-percent egg infectious doses of Kagawa11C2020 revealed that the 50% chicken lethal dose was 10^4.63 and the mean time to death was 134.4 h. All infected chickens demonstrated viral shedding beginning on 2 dpi—before clinical signs were observed. These results suggest that affected chickens could transmit Kagawa11C2020 to surrounding chickens in the absence of clinical signs for several days before they died.

The flyways of many different wild waterfowl pass through the Caspian Sea region. The western coast of the middle Caspian Sea is an area with many wetlands, where wintering grounds with large concentrations of birds are located. It is known that wild waterfowl are a natural reservoir of the influenza A virus. In the mid-2000s, in the north of this region, the mass deaths of swans, gulls, and pelicans from high pathogenicity avian influenza virus (HPAIV) were noted. At present, there is still little known about the presence of avian influenza virus (AIVs) and different avian paramyxoviruses (APMVs) in the region’s waterfowl bird populations. Here, we report the results of monitoring these viruses in the wild waterfowl of the western coast of the middle Caspian Sea from 2017 to 2020. Samples from 1438 individuals of 26 bird species of 7 orders were collected, from which 21 strains of AIV were isolated, amounting to a 1.46% isolation rate of the total number of samples analyzed (none of these birds exhibited external signs of disease). The following subtypes were determined and whole-genome nucleotide sequences of the isolated strains were obtained: H1N1 (n = 2), H3N8 (n = 8), H4N6 (n = 2), H7N3 (n = 2), H8N4 (n = 1), H10N5 (n = 1), and H12N5 (n = 1). No high pathogenicity influenza virus H5 subtype was detected. Phylogenetic analysis of AIV genomes did not reveal any specific pattern for viruses in the Caspian Sea region, showing that all segments belong to the Eurasian clades of classic avian-like influenza viruses. We also did not find the amino acid substitutions in the polymerase complex (PA, PB1, and PB2) that are critical for the increase in virulence or adaptation to mammals. In total, 23 hemagglutinating viruses not related to influenza A virus were also isolated, of which 15 belonged to avian paramyxoviruses. We were able to sequence 12 avian paramyxoviruses of three species, as follows: Newcastle disease virus (n = 4); Avian paramyxovirus 4 (n = 5); and Avian paramyxovirus 6 (n = 3). In the Russian Federation, the Newcastle disease virus of the VII.1.1 sub-genotype was first isolated from a wild bird (common pheasant) in the Caspian Sea region. The five avian paramyxovirus 4 isolates obtained belonged to the common clade in Genotype I, whereas phylogenetic analysis of three isolates of Avian paramyxovirus 6 showed that two isolates, isolated in 2017, belonged to Genotype I and that an isolate identified in 2020 belonged to Genotype II. The continued regular monitoring of AIVs and APMVs, the obtaining of data on the biological properties of isolated strains, and the accumulation of information on virus host species will allow for the adequate planning of epidemiological measures, suggest the most likely routes of spread of the virus, and assist in the prediction of the introduction of the viruses in the western coastal region of the middle Caspian Sea.