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Avian influenza (AI) is an extremely contagious viral disease of domestic and wild birds that can spread rapidly among bird populations, inducing serious economic losses in the poultry industry. During the winter season 2021-2022, we isolated seventeen highly pathogenic avian influenza (HPAI) H5N8 viruses from outbreaks involving ducks in Egypt, occurring in both backyard and farm settings. The aim of this study was to pinpoint genetic key substitutions (KSs) that could heighten the risk of a human pandemic by influencing the virus's virulence, replication ability, host specificity, susceptibility to drugs, or transmissibility. To understand their evolution, origin, and potential risks for a human pandemic, whole-genome sequencing and phylogenetic analysis were conducted. Our analysis identified numerous distinctive mutations in the Egyptian H5N8 viruses, suggesting potential enhancements in virulence, resistance to antiviral drugs, and facilitation of transmission in mammals. In this study, at least five genotypes within one genome constellation of H5N8 viruses were identified, raising concerns about the potential emergence of novel viruses with altered characteristics through reassortment between different genotypes and distinct groups. These findings underscore the role of ducks in the virus's evolutionary process and emphasize the urgent need for enhanced biosecurity measures in domestic duck farms to mitigate pandemic risk.

We isolated highly pathogenic avian influenza (HPAI) H5N5 and H5N1 viruses from crows in Hokkaido, Japan, during winter 2023-24. They shared genetic similarity with HPAI H5N5 viruses from northern Europe but differed from those in Asia. Continuous monitoring and rapid information sharing between countries are needed to prevent HPAI virus transmission.

In the 2021/22 winter, one H5N1 and nine H5N8 high pathogenicity avian influenza viruses (HPAIVs) of clade 2.3.3.4b were isolated from the water in crane roosts on the Izumi plain, Japan. Additionally, we isolated low pathogenicity avian influenza viruses (LPAIVs) of five subtypes: H1N1, H4N2, H4N6, H7N7, and H10N4. H5N8 HPAIVs belonging to the G2a group were isolated throughout winter, whereas H5N1 HPAIV belonging to the G2b group were isolated only in early winter. These findings suggest co-circulation of both G2a and G2b HPAIVs in early winter. Although two H7N7 LPAIVs were isolated from cranes' roost water collected on the same day, the gene constellations of the two isolates were clearly different, indicating the contemporary invasion of at least two different genotypes of H7N7 LPAIVs in the Izumi plain. This study underscores the importance of monitoring both HPAIVs and LPAIVs to understand avian influenza virus ecology in migratory waterfowl populations., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

During the 2020/21 winter season, 29 and 10 H5N8 high pathogenicity avian influenza viruses (HPAIVs) were isolated from environmental water and wild birds, respectively, in Kagoshima prefecture, Japan. Furthermore, seven subtypes of low pathogenicity avian influenza viruses (LPAIVs) were also isolated; H1N1, H2N9, H3N2, H3N6, H3N8, H4N6, and H6N6 subtypes. While the H5 hemagglutinin (HA) genes of the G1 cluster were isolated throughout the winter season, those of the G2 cluster were also detected in late winter, suggesting that H5 HPAIVs possessing H5 HA genes from the two different clusters were individually introduced into Kagoshima prefecture. Intriguingly, genetic constellations revealed that the H5N8 HPAIVs could be classified into six genotypes, including four previously reported genotypes (E1, E2, E3, and E7), and two new genotypes (tentatively named E8 and E9). The PB1 and PA gene segments of genotypes E8 and E9 shared high similarity with those of LPAIVs, whereas the remaining gene segments were close to those of genotype E1. Furthermore, LPAIVs whose PA gene segment was close to that of genotype E9 were isolated from the environmental water. Overall, we revealed that various HPAIV genotypes circulated in Kagoshima prefecture during the 2020/21 winter season. This study highlights the importance of monitoring both HPAIV and LPAIV to better understand AIV ecology in migratory waterfowl populations.

Environmental water-targeted surveillance of migratory aquatic birds at overwintering sites is potentially one of the most effective approaches for understanding the ecology of avian influenza viruses (AIVs). In this study, we improved the method for AIV isolation from environmental water samples by making a minor modification to our previously reported process. We experimentally demonstrated that the AIV recovery efficiency of the modified method was 10-100-fold higher than that of the original method. This improved isolation method allowed us to isolate a considerably larger number of AIV isolates from environmental water samples collected at an overwintering site for tens of thousands of migratory aquatic birds in Japan during the 2018/2019 winter season, compared with those during previous winter seasons. Genetic and phylogenetic analyses revealed that AIVs of the same subtypes with multiple genetic constellations were circulating in a single overwintering site during a single winter season. These findings indicate that our improved isolation method contributes to enhance environmental water-targeted surveillance and to a better understanding of AIV ecology in migratory aquatic bird populations by monitoring ongoing AIV circulation., (© 2022 Wiley-VCH GmbH.)

Genetic analyses of highly pathogenic avian influenza H5 subtype viruses isolated from the Izumi Plain, Japan, revealed cocirculation of 2 genetic groups of clade 2.3.4.4b viruses among migratory waterfowl. Our findings demonstrate that both continuous surveillance and timely information sharing of avian influenza viruses are valuable for rapid risk assessment.

The circulation of highly pathogenic avian influenza viruses (HPAIVs) of various subtypes (e.g., H5N1, H5N6, H5N8, and H7N9) in poultry remains a global concern for animal and public health. Migratory waterfowls play important roles in the transmission of these viruses across countries. To monitor virus spread by wild birds, active surveillance for avian influenza in migratory waterfowl was conducted in Mongolia from 2015 to 2019. In total, 5000 fecal samples were collected from lakesides in central Mongolia, and 167 influenza A viruses were isolated. Two H5N3, four H7N3, and two H7N7 viruses were characterized in this study. The amino acid sequence at hemagglutinin (HA) cleavage site of those isolates suggested low pathogenicity in chickens. Phylogenetic analysis revealed that all H5 and H7 viruses were closely related to recent H5 and H7 low pathogenic avian influenza viruses (LPAIVs) isolated from wild birds in Asia and Europe. Antigenicity of H7Nx was similar to those of typical non-pathogenic avian influenza viruses (AIVs). While HPAIVs or A/Anhui/1/2013 (H7N9)-related LPAIVs were not detected in migratory waterfowl in Mongolia, sporadic introductions of AIVs including H5 and H7 viruses into Mongolia through the wild bird migration were identified. Thus, continued monitoring of H5 and H7 AIVs in both domestic and wild birds is needed for the early detection of HPAIVs spread into the country.

During the 2016-2017 winter season, we isolated 33 highly pathogenic avian influenza viruses (HPAIVs) of H5N6 subtype and three low pathogenic avian influenza viruses (LPAIVs) from debilitated or dead wild birds, duck faeces, and environmental water samples collected in the Izumi plain, an overwintering site for migratory birds in Japan. Genetic analyses of the H5N6 HPAIV isolates revealed previously unreported phylogenetic variations in the PB2, PB1, PA, and NS gene segments and allowed us to propose two novel genotypes for the contemporary H5N6 HPAIVs. In addition, analysis of the four gene segments identified close phylogenetic relationships between our three LPAIV isolates and the contemporary H5N6 HPAIV isolates. Our results implied the co-circulation and co-evolution of HPAIVs and LPAIVs within the same wild bird populations, thereby highlighting the importance of avian influenza surveillance targeting not only for HPAIVs but also for LPAIVs., (© 2018 Blackwell Verlag GmbH.)

The Izumi plain in Kagoshima Prefecture, Japan, is an overwintering site of endangered cranes (hooded cranes and white-naped cranes) and of many other migratory birds (including wild ducks) that are considered carriers of avian influenza viruses (AIVs). To assess the risks of a highly pathogenic avian influenza outbreak in the crane populations, we tested various environmental samples for AIVs in this area. In the 2014-2015 winter season, we isolated one AIV of the H6N2 subtype from the cranes' roost water and two AIVs of the H11N9 subtype from a crane fecal sample and a cloacal swab of a dead spot-billed duck. Genetic analysis of these AIV isolates indicated that our H6N2 isolate is genetically close to AIVs isolated from wild birds in Southeast Asian countries, except that the PB1 and NS genes belong to the North American virus lineage. All genes of the two H11N9 isolates are related to AIVs belonging to the Eurasian virus lineage. Notably, in our phylogenetic trees, H11 HA and N9 NA genes showing high sequence similarity to the corresponding genes of isolates from wild birds in South Africa and Spain, respectively, did not cluster in the major groups with recent wild-bird isolates from East Asia. These results suggest that AIVs with viral gene segments derived from various locations and bird species have been brought to the Izumi plain. These findings imply a possible association of dynamic movements of wild birds with AIV evolution.

An influenza A virus of H4N6 subtype was isolated from the Izumi plain, Japan, in 2013. Genetic analyses revealed that two viral genes (M and NS gene segments) of this isolate were genetically distinct from those of the H4N6 virus isolated from the same place in 2012. Furthermore, three viral genes (PB2, PB1 and M gene segments) of this isolate share high similarity with those of the North American isolates of 2014. These results suggest a high frequency of genetic reassortment of avian influenza viruses in Asian waterfowl and intercontinental movements of avian influenza viruses via migratory waterfowl., (© 2017 The Societies and John Wiley & Sons Australia, Ltd.)

The Izumi plain in Kagoshima prefecture, Japan, is an overwintering site of more than 10,000 cranes. The wet paddy areas are artificially created to provide roosting sites for the cranes every winter. Since wild ducks, known to be a natural reservoir of influenza A viruses, also overwinter in this area, the cranes' roost water likely serves as a source of influenza A virus infection. To assess this potential risk, we collected 126 water samples from the cranes' roost in the 2012/2013 winter season for virus isolation. We isolated six influenza viruses of three subtypes (H3N8, H4N6, and H4N8) from the water samples collected in the months of November and December. Genetic analysis of our isolates indicated that these viruses were genetically similar to the low-pathogenic avian influenza viruses circulating among Eurasian waterfowl. These findings suggest the possibility of the cranes becoming infected with the avian influenza viruses that are present in their roost water.