The influenza A virus (IAV) has been a major cause of several pandemics, underscoring the importance of elucidating its transmission dynamics. This review investigates potential intermediate hosts in the cross-species transmission of IAV to humans, focusing on the factors that facilitate zoonotic events. We evaluate the roles of various animal hosts, including pigs, galliformes, companion animals, minks, marine mammals, and other animals, in the spread of IAV to humans.

H9N2 avian influenza A viruses (AIVs) cause economic losses in the poultry industry and provide internal genomic segments for the evolution of H5N1 and H7N9 AIVs into more detrimental strains for poultry and humans. In addition to the endemic Y439/Korea-lineage H9N2 viruses, the Y280-lineage spread to Korea since 2020. Conventional recombinant H9N2 vaccine strains, which bear mammalian pathogenic internal genomes of the PR8 strain, are pathogenic in BALB/c mice. To reduce the mammalian pathogenicity of the vaccine strains, the PR8 PB2 was replaced with the non-pathogenic and highly productive PB2 of the H9N2 vaccine strain 01310CE20. However, the 01310CE20 PB2 did not coordinate well with the hemagglutinin (HA) and neuraminidase (NA) of the Korean Y280-lineage strain, resulting in a 10-fold lower virus titer compared to the PR8 PB2. To increase the virus titer, the 01310CE20 PB2 was mutated (I66M-I109V-I133V) to enhance the polymerase trimer integrity with PB1 and PA, which restored the decreased virus titer without causing mouse pathogenicity. The reverse mutation (L226Q) of HA, which was believed to decrease mammalian pathogenicity by reducing mammalian receptor affinity, was verified to increase mouse pathogenicity and change antigenicity. The monovalent Y280-lineage oil emulsion vaccine produced high antibody titers for homologous antigens but undetectable titers for heterologous (Y439/Korea-lineage) antigens. However, this defect was corrected by the bivalent vaccine. Therefore, the balance of polymerase and HA/NA activities can be achieved by fine-tuning PB2 activity, and a bivalent vaccine may be more effective in controlling concurrent H9N2 viruses with different antigenicities.

Viruses have evolved sophisticated mechanisms to manipulate host cell processes and utilize intracellular organelles to facilitate their replication. These complex interactions between viruses and cellular organelles allow them to hijack the cellular machinery and impair homeostasis. Moreover, viral infection alters the cell membrane’s structure and composition and induces vesicle formation to facilitate intracellular trafficking of viral components. However, the research focus has predominantly been on the immune response elicited by viruses, often overlooking the significant alterations that viruses induce in cellular organelles. Gaining a deeper understanding of these virus-induced cellular changes is crucial for elucidating the full life cycle of viruses and developing potent antiviral therapies. Exploring virus-induced cellular changes could substantially improve our understanding of viral infection mechanisms.

The 2009 influenza A H1N1 pandemic (pdm09) originated from the influenza A virus of swine (IAV‐S) through multiple reassortment events with avian and human IAVs. The pdm09 reportedly reintroduced the virus to pigs, contributing to the evolution and diversity of IAV‐S through frequent reassortment and drifts. Surveillance and whole‐genome sequencing of IAV‐S from conventional pig farms in Korea during 2021–2022 revealed that the genetic diversity of H1 and H3 IAV‐S was continuously enriched after human‐to‐swine spillover of pdm09 viruses with long‐term maintenance, persistence, and reassortment of virus lineages. Evidence of additional human‐to‐swine spillover of viruses that are different from the 2009 virus but close to that of the recent H1N1pdm09 human vaccine was identified in this study. The identification of swine‐adapted pdm09 viruses, which have accumulated amino acid mutations with potentially altered antigenicity and a unique potential N‐glycosylation site within the haemagglutinin (HA) gene, suggests the distinctive evolution of spillover pdm09 viruses in swine. The genetic constellation of the recently emerging Eurasian avian‐like swine lineage and the preexisting classical swine lineage H1 viruses in Korea has been expanded through reassortment with cocirculating pdm09 viruses and/or H3N2 IAV‐S harboring the pdm09 M gene (H3N2pM). Collectively, after the major shift of Korean IAV‐S from the classical swine lineage to the pdm09 lineage in 2009, the frequent spillover of pdm09 viruses and the circulation of IAV‐S harboring pdm09 gene segments led to the continuous diversification of IAV‐S through antigenic drift and shift, raising concerns about the potential reintroduction of these viruses to humans.

Influenza A virus (IAV) is a major global health threat, causing seasonal epidemics and occasional pandemics. Particularly, Influenza A viruses from avian species pose significant zoonotic threats, with PB2 adaptation serving as a critical first step in cross-species transmission. A comprehensive risk assessment framework based on PB2 sequences is necessary, which should encompass detailed analyses of specific residues and mutations while maintaining sufficient generality for application to non-PB2 segments. In this study, we developed two complementary approaches: a regression-based model for accurately distinguishing among risk groups, and a SHAP-based risk assessment model for more meaningful risk analyses. For the regression-based risk models, we compared various methodologies, including tree ensemble methods, conventional regression models, and deep learning architectures. The optimized regression model, combined with SHAP value analysis, identified and ranked individual residues contributing to zoonotic potential. The SHAP-based risk model enabled intra-class analyses within the zoonotic risk assessment framework and quantified risk yields from specific mutations. Experimental analyses demonstrated that the Random Forest regression model outperformed other models in most cases, and we validated the target value settings for risk regression through ablation studies. Our SHAP-based analysis identified key residues (271A, 627K, 591R, 588A, 292I, 684S, 684A, 81M, 199S, and 368Q) and mutations (T271A, Q368R/K, E627K, Q591R, A588T/I/V, and I292V/T) critical for zoonotic risk assessment. Using the SHAP-based risk assessment model, we found that influenza A viruses from Phasianidae showed elevated zoonotic risk scores compared to those from other avian species. Additionally, mutations I292V/T, Q368R, A588T/I, V598A/I/T, and E/V627K were identified as significant mutations in the Phasianidae. These PB2-focused quantitative methods provide a robust and generalizable framework for both rapid screening of avians’ zoonotic potential and analytical quantification of risks associated with specific residues or mutations.

Lee CY. J Vet Sci. 2023 Oct;24:e78. https://doi.org/10.4142/jvs.23105