In China, H9N2 avian influenza virus (AIV) has become widely prevalent in poultry, causing huge economic losses after secondary infection with other pathogens. Importantly, H9N2 AIV continuously infects humans, and its six internal genes frequently reassort with other influenza viruses to generate novel influenza viruses that infect humans, threatening public health. Inactivated whole-virus vaccines have been used to control H9N2 AIV in China for more than 20 years, and they can alleviate clinical symptoms after immunization, greatly reducing economic losses. However, H9N2 AIVs can still be isolated from immunized chickens and have recently become the main epidemic subtype. A more effective vaccine prevention strategy might be able to address the current situation. Herein, we analyze the current status and vaccination strategy against H9N2 AIV and summarize the progress in vaccine development to provide insight for better H9N2 prevention and control.

Influenza A virus (IAV) is an important zoonotic pathogen that can cause disease in animals such as poultry and pigs, and it can cause infection and even death in humans, posing a serious threat to public health. IAV is an enveloped virus that relies on host cell metabolic systems, especially lipid metabolism systems, to complete its life cycle in host cells. On the other side, host cells regulate their metabolic processes to prevent IAV replication and maintain their normal physiological functions. This review summarizes the roles of fatty acid, cholesterol, phospholipid and glycolipid metabolism in IAV infection, proposes future research challenges, and looks forward to the prospective application of lipid metabolism modification to limit IAV infection, which will provide new directions for the development of anti-influenza drugs.

The H9N2 subtype of avian influenza viruses (AIVs) is widely prevalent in poultry and wild birds globally, with occasional transmission to humans. In comparison to other H9N2 lineages, the BJ/94 lineage has raised more public health concerns; however, its evolutionary dynamics and transmission patterns remain poorly understood. In this study, we demonstrate that over three decades (1994–2023), BJ/94 lineage has undergone substantial expansion in its geographical distribution, interspecies transmission, and viral reassortment with other AIV subtypes, increasing associated public health risks. These changes were primarily driven by the emergence of a dominant genotype G57. In the first decade, G57 emerged in East China and rapidly adapted to chickens and spread across China. Since 2013, the G57 genotype has expanded beyond China into eight other countries and reassorted with various AIV subtypes to form new zoonotic reassortants. Chickens have played a key role in the generation and circulation of the G57 viruses, with ducks and other poultry species likely assuming an increasingly importantly role. Over the past decade, G57 has been more frequently detected in wild birds, mammals, and humans. Additionally, Vietnam has emerged as a new hotspot for the international spread of G57. Our results suggest that the BJ/94 lineage H9N2 virus may continue to overcome geographical and species barriers, with potentially more severe consequences.

The H9N2 avian influenza virus (AIV) is currently widespread worldwide, posing a severe threat to the poultry industry and public health. Reassortment is an important way for influenza viruses to adapt to a new host. In 2007, the PB2 gene of H9N2 AIV in China was reassorted, and the DK1-like lineage replaced the F/98-like lineage, forming a dominant genotype of G57. This genotype and its reassortants (such as H7N9, H10N8 and H5N6) showed higher mammalian adaptation, and caused increased human infections. However, the adaptive mechanisms of the DK1-like lineage PB2 gene remain unclear. Here, we confirmed that the PB2 lineage of the H9N2 AIV currently prevalent in China still belongs to the DK1-like lineage and, compared with the previously predominant F/98-like lineage, the DK1-like lineage PB2 gene significantly enhances H9N2 AIV to mammalian adaptation. Through transcriptomic analysis and qRT–PCR and western blot experiments, we identified a host factor, sphingosine kinase 1 (SphK1), that is closely related to viral replication. SphK1 inhibits the replication of DK1-like PB2 gene H9N2 AIV, but the ability of SphK1 protein to bind DK1-like PB2 protein is weaker than that of F/98-like PB2 protein, which may contribute to H9N2 AIV containing the DK1-like PB2 gene to escape the inhibitory effect of host factor SphK1 for efficient infection. This study broadens our understanding of the adaptive evolution of H9N2 AIV and highlights the necessity to pay close attention to the AIV that contains the adaptive PB2 protein in animals and humans.

The sustained circulation of H9N2 avian influenza viruses (AIVs) poses a significant threat for contributing to a new pandemic. Given the temporal and spatial uncertainty in the antigenicity of H9N2 AIVs, the immune protection efficiency of vaccines remains challenging. By developing an antigenicity prediction method for H9N2 AIVs, named PREDAC-H9, the global antigenic landscape of H9N2 AIVs was mapped. PREDAC-H9 utilizes the XGBoost model with 14 well-designed features. The XGBoost model was built and evaluated to predict the antigenic relationship between any two viruses with high values of 81.1 %, 81.4 %, 81.3 %, 81.1 %, and 89.4 % in accuracy, precision, recall, F1 value, and area under curve (AUC), respectively. Then the antigenic correlation network (ACnet) was constructed based on the predicted antigenic relationship for H9N2 AIVs from 1966 to 2022, and ten major antigenic clusters were identified. Of these, four novel clusters were generated in China in the past decade, demonstrating the unique complex situation there. To help tackle this situation, we applied PREDAC-H9 to calculate the cluster-transition determining sites and screen out virus strains with the high cross-protective spectrum, thus providing an in silico reference for vaccine recommendation. The proposed model will reduce the clinical monitoring workload and provide a useful tool for surveillance and control of H9N2 AIVs.

The interaction between migratory birds and domestic waterfowl facilitates viral co-infections, leading to viral reassortment and the emergence of novel viruses. In 2022, samples were collected from duck farms around Poyang Lake in Jiangxi Province, China, which is located within the East Asia–Australasia flyway. Three strains of H4N6 avian influenza virus (AIV) were isolated. Genetic and phylogenetic analyses showed that the isolated H4N6 avian influenza viruses (AIVs) belonged to new genotypes, G23 and G24. All isolated strains demonstrated dual receptor binding properties. Additionally, the isolated strains were able to replicate efficiently not only in avian cells but also in mammalian cells. Furthermore, the H4N6 AIV isolates could infect chickens, with viral replication detected in the lungs and extrapulmonary organs, and could transmit within chicken flocks through contact, with viral shedding detected only in oropharyngeal swabs from chickens in the contact group. Notably, the H4N6 AIV could infect mice without prior adaptation and replicate in the lungs with high viral titers, suggesting that it is a potential threat to humans. In conclusion, this study provides valuable insight into the characteristics of H4N6 strains currently circulating in China.