Search Results (214)

To address the lack of a commercially available vaccine for goose circovirus (GoCV), we developed and evaluated a prokaryotically expressed subunit vaccine targeting the viral capsid (Cap) protein. A truncated Cap protein (GoCV-ΔCap) was expressed in Escherichia coli (E. coli) and formulated with aluminum hydroxide as a subunit vaccine (GoCVsubvac). Goslings were primed intramuscularly (i.m.) with high (75 µg) or low (15 µg) doses GoCVsubvac, followed by a boost 14 days later. At 14 days post-boost, goslings were challenged with GoCV and were administered a bivalent inactivated vaccine against Newcastle disease virus (NDV) and H9-subtype Avian influenza virus (AIV). Using our established gosling pathogenicity model, vaccine efficacy was evaluated via body weight, lesions, viral load, antibody titers, cytokine responses, and interference with NDV/AIV immunity. Results demonstrated that the GoCV-ΔCap vaccine, especially the high-dose formulation, provided effective immunoprotection. It elicited robust humoral and cellular immune responses, reduced lymphoid pathology, and decreased the viral detection rate in lymphoid tissues from 100% (5/5) in infected controls to 40% (2/5). Importantly, it alleviated GoCV-induced immunosuppression and preserved the immunogenicity of co-administered vaccines. This novel subunit vaccine is a promising candidate for controlling GoCV disease (GoCVD). Full article

During 2022–2024, a highly pathogenic avian influenza virus (HPAIV) H5N1 strain, designated A/Seagull/Hebei/qhd6/2024 (H5N1), was isolated from migratory birds in Beidaihe National Wetland Park, North China. Phylogenetic analyses revealed that its hemagglutinin (HA) gene belongs to the 2.3.4.4b clade, while the neuraminidase (NA) gene and internal genes clustered with strains originating from multiple continents, consistent with a transcontinental reassortment event. The virus also exhibited 90.1–98.1% nucleotide homology with human-derived H5N1 isolates. Molecular characterization identified key virulence-associated mutations, including the classic HPAIV HA cleavage site, HA-T160A (associated with enhanced human receptor-binding capacity), and NA-I117T (potentially linked to drug resistance). BALB/c mouse infection experiments confirmed systemic replication and high pathogenicity of strain qhd6, with a 50% lethal dose (LD50) of 0.95 log₁₀EID₅₀/mL. Antigenic analysis revealed good cross-reactivity with the widely used H5-Re14 vaccine strain. This study reports the identification, in Beidaihe National Wetland Park, of an HPAIV H5N1 strain whose genetic characteristics suggest intercontinental reassortment and indicate cross-species transmission risk. It clarifies the genetic characteristics and pathogenicity of this strain, providing an important theoretical and practical basis for precise surveillance, risk early warning, and comprehensive prevention and control of AIV at migratory bird stopover sites in North China. Full article

The global emergence of the avian influenza virus (AIV) H5N1 clade 2.3.4.4b since 2016 has caused substantial losses in wild bird and poultry populations, along with heightened risks of transmission to humans and other mammals. Vaccination of poultry has been a key strategy to curb the virus’s spread and mitigate its socioeconomic impact. This report describes an outbreak of high pathogenicity avian influenza virus (HPAIV) H5N1 clade 2.3.4.4b in a flock of 15,000 brown layer chickens (170 days old), all of which had received a four-dose vaccination regimen with H5N1/H5N8 commercial vaccines at 17, 50, 100, and 125 days of age. Despite this vaccination history, H5N1 infection was confirmed approximately seven weeks post-vaccination. H5N1 infection was confirmed by RT-qPCR, virus isolation, and full genome sequencing covering all eight gene segments, followed by phylogenetic and molecular analyses. Clinical signs included reduced feed intake, decreased egg production, and a cumulative mortality rate of 35% over 52 days. Hemagglutination inhibition (HI) testing with various H5 antigens revealed inconsistent antibody titers (geometric mean: 4.0 to 9.1 log2). Genetic analysis of the full-length HA and NA gene sequences further revealed strong similarity to contemporaneous H5N1 clade 2.3.4.4b strains circulating in Egypt, with multiple mutations in the HA head domain, particularly near immunogenic epitopes and receptor binding sites. These findings highlight the limitations of current vaccination strategies under conditions of antigenic mismatch and complex immunization schedules, emphasizing the need for improved vaccine matching and continuous molecular surveillance. To improve outbreak management in poultry, enhanced vaccination protocols, stringent biosecurity measures, and rigorous monitoring practices are critical. Full article

Highly pathogenic avian influenza A (H7N9) remains a threat to poultry health and poses a zoonotic risk, highlighting the need for vaccine antigens capable of inducing both systemic and mucosal immunity. In this study, we evaluated X33CLS-H7, a clarified cell-lysate supernatant derived from glycoengineered Pichia pastoris expressing H7 hemagglutinin, in BALB/c mice following intramuscular(i.m.) injection, nebulized inhalation, or intranasal instillation. H7 expression and hemagglutination activity were confirmed by Western blotting and hemagglutination assay, respectively. Serum HA7-specific IgG and IgA responses, hemagglutination inhibition(HI) activity, H7N9 pseudovirus neutralization, bronchoalveolar lavage fluid (BALF) antibodies, and safety readouts were assessed. After two i.m. immunizations, X33CLS-H7 induced the strongest systemic antibody responses, with an HI geometric mean titer of 1:1622 95% CI, 1:1108–1:2348 and a mean log10 NT₅₀ of 4.62. Respiratory immunization also elicited antibody responses. After four doses, high-dose nebulized delivery produced the strongest responses among the respiratory delivery regimens, with serum IgG and IgA titers of 1.02 × 10⁵ and 2.24 × 10³, respectively, an endpoint HI GMT r of 1:457 95% CI, 1:211–1:971, and a mean log10 NT₅₀ of 3.77 compared with 2.02 in saline controls. High-dose nebulized delivery also generated detectable HA7-specific IgG and IgA responses in bronchoalveolar lavage fluid. No overt local or systemic toxicity signals were observed under the tested conditions. These findings indicate that X33CLS-H7 retains HA7-associated antigenicity and can induce systemic and respiratory mucosal antibody responses, supporting its further evaluation as a simplified and scalable H7N9 vaccine antigen candidate. Full article

Highly pathogenic avian influenza (HPAI) H5N1 viruses of the goose/Guangdong (Gs/GD) lineage have driven a global panzootic since 2020, with clade 2.3.4.4b establishing sustained transmission in wild birds. In South America, early outbreaks were largely associated with the North American-derived B3.2 genotype, which showed limited diversification after its introduction. Here, we report the genomic characterization of eight H5N1 viruses detected in Uruguay during the reemergence of avian influenza in February–March 2026. Complete genomes were obtained from wild birds exhibiting neurological signs, predominantly Coscoroba coscoroba. All viruses belong to clade 2.3.4.4b but exhibit a reassortant genomic constellation distinct from B3.2. The HA, NA, and MP segments retain the Eurasian backbone, whereas internal genes derive from both South American and North American low-pathogenicity avian influenza lineages. PB2 variation distinguishes two closely related viral groups differing in PB2 origin, whereas the remaining genomic segments retain a shared background. Sequence variation in the neuraminidase gene reduced the sensitivity of a widely used N1-specific RT-qPCR assay, highlighting limitations of existing diagnostic tools during viral evolution. These findings confirm the presence of reassortant H5N1 viruses in Uruguay and, together with recent reports from Argentina and Brazil, support an emerging pattern of genomic diversification in southern South America. Full article

The H5N1 subtype of highly pathogenic avian influenza (HPAI) poses a major zoonotic threat due to its high fatality rate and capacity for cross species transmission. In early 2025, Argentina detected a novel triple reassortant A(H5N1) virus in Chaco Province, combining Eurasian, North American, and South American lineage segments. Genomic analyses of subsequent outbreaks in Buenos Aires and Entre Ríos confirmed persistence of this reassortant and additional HA substitutions (T204K, P251S) potentially linked to increased mammalian receptor affinity. Although PB2 sequences lacked canonical mammalian-adaptive markers (E627K, Q591K, D701N), all contained I292M, a mutation associated with human adaptation. Phylogenetic analyses revealed distinct genotypes and increasing divergence. These findings indicate ongoing viral evolution and adaptation within Argentina, emphasizing the urgent need for sustained genomic surveillance, timely data sharing, and integrated One Health strategies to mitigate zoonotic and socioeconomic risks associated with H5N1 spread in South America. Full article

Wastewater surveillance emerged as a critical public health tool during the COVID-19 pandemic, enabling early detection of community-level pathogen circulation independent of clinical testing. Its ability to capture signals from both symptomatic and asymptomatic individuals highlighted the importance of optimizing sampling methodologies to improve sensitivity and reliability. A key question is whether the several-fold increase in SARS-CoV-2 detectability observed when using passive tampon swab sampling compared with paired grab samples also applies to other respiratory viruses, including influenza A (including its avian influenza H5N1 subtype), influenza B, and respiratory syncytial virus (RSV). We collected 24 h passive swab samples with same-day grab samples from Detroit sewersheds, concentrated and purified nucleic acids, and using RT-ddPCR, quantified respiratory syncytial virus, SARS-CoV-2, influenza A, influenza B, and H5N1 influenza A viruses using markers RSV, SC2, InfA, InfB, and H5, respectively. Samples testing positive for H5 (marker for H5N1 influenza A) were further analyzed by targeted PCR and amplicon sequencing. Across three sites, median 24 h swab:grab ratios of virus copies were 7.0 for RSV, 9.2 for SC2, 9.9 for InfA, and 3.6 for InfB. A 239 bp hemagglutinin sequence from a sample with a strong H5 signal (795 copies/10 mL) had 100% identity to avian influenza viruses from Canada geese. Twenty-four-hour swab sampling greatly improves viral detectability across diverse targets and enabled the first confirmed detection of H5 in Detroit wastewater. Combined with magnetic bead purification, the overall sensitivity gain over conventional PEG-NaCl-Qiagen methods is approximately 36-fold, enabling earlier warning of community pathogens than grab samples. By integrating 24 hour passive swab sampling with high-efficiency nucleic acid purification, we expand the sensitivity of wastewater surveillance to enable detection and confirmation of low-abundance pathogens like avian influenza (H5). Full article

H5 subtype avian influenza virus (AIV) can infect both chickens and ducks, leading to substantial economic losses. Nevertheless, certain strains cause silent infections in ducks. In this study, a goose-origin clade 2.3.4.4h H5N6 AIV was isolated, which caused high mortality in mixed-gender white leghorn chickens but no deaths in mixed-gender mallard ducks. After independent serial in vitro passage in duck embryo fibroblasts (DEFs) and in vivo passage in specific-pathogen-free (SPF) ducks, the DEF-passage 10 (P10) virus induced markedly higher mortality rates and viral loads in SPF ducks compared to the DEF-P1 virus and the original parental virus prior to passage. Similarly, the in vivo-passaged P3 and P4 viruses exhibited significantly higher mortality rates than the P1 virus in SPF ducks, with 100% mortality and markedly increased viral titers in the organs. A whole-genome SNP analysis identified seven high-frequency mutations in the M1, NA and NS1 proteins. The NS1-F161L substitution virus exhibited significantly increased mortality rates, viral loads in multiple tissues, and a robustly induced innate immune response in ducks. Furthermore, dynamic evolutionary variations in the NS1 protein among global H5 avian influenza viruses revealed that the NS1-F161L substitution became dominant in clade 2.3.4.4b viruses in 2021 and subsequent years. Collectively, our findings demonstrate that host-driven adaptation can rapidly increase the pathogenicity of H5N6 AIVs in ducks and identify NS1-F161L as a critical virulence marker. These results offer novel insights relevant to the molecular surveillance, virulence prediction, and risk assessment of circulating H5 AIVs in waterfowl. Full article

Highly pathogenic avian influenza (HPAI) H5N1 clade 2.3.4.4b continues to circulate globally across wild birds, poultry, and an expanding range of mammalian hosts, highlighting the need for antiviral strategies that address the animal–environment–human interface. The influenza A polymerase acidic (PA) endonuclease, a key enzyme in viral transcription, represents a conserved antiviral target across host species. In this study, we present a computational prioritization framework integrating homology modeling, molecular docking, molecular dynamics simulations, and physicochemical filtering to identify candidate PA endonuclease inhibitors relevant to a One Health context. Homology models of contemporary H5N1 clade 2.3.4.4b PA sequences were constructed based on the crystallographic template 6FS8 and used for cross-host docking against a targeted ligand library. Docking analysis identified baloxavir, a reference inhibitor, and entecavir, a nucleoside analog, as compounds of interest, with entecavir demonstrating favorable binding behavior, particularly in the poultry-associated model. Molecular dynamics simulations of the poultry PA–entecavir complex indicated stable interaction over 170 ns, supported by low structural deviation and favorable binding free energy (ΔG ≈ −85 kJ/mol). Physicochemical profiling suggested that entecavir possesses properties such as high polarity and predicted aqueous solubility, which were considered within the translational filtering step of this computational workflow. However, these properties do not establish antiviral efficacy or practical suitability for field use. The study provides a structured framework for integrating cross-host structural analysis with basic translational considerations, supporting the identification of candidate compounds for further biochemical and virological evaluation within the context of H5N1 control. Full article

Highly pathogenic avian influenza virus (HPAIV) H5N1 has been detected in dairy cattle in the United States, with high viral loads observed in milk from infected animals. This raises public health concerns regarding potential transmission through exposure to raw milk. The sale of raw milk via vending machines represents a well-established distribution model in many European countries, including Switzerland. Although a notice must be posted on these milk vending machines stating that it is raw milk, together with appropriate processing instructions (heating to over 70 °C required, storage below 5 °C, consumption within 3 days), these notices are sometimes missing, and consumers often do not follow these guidelines. Over a four-month period, spanning from June 2025 to September 2025, 124 raw milk samples were collected from vending machines across Switzerland. Samples were screened for influenza A using reverse-transcription quantitative PCR (RT-qPCR). No samples tested positive for influenza A virus. The data from this study demonstrate the feasibility of implementing a sampling and detection system for HPAIV H5N1 in direct-to consumer raw milk samples and highlight the currently very low risk of HPAIV in raw milk samples sold via vending machines in Switzerland. Full article

The ongoing panzootic of the highly pathogenic avian influenza (HPAI) H5N1 virus, dominated by clade 2.3.4.4b, constitutes a significant global threat to wildlife, animal health, and public health. Once characterized by sporadic outbreaks, H5N1 has evolved into a sustained, year-round infection with an expanded host range that now includes numerous mammalian species. Its high pathogenicity is primarily driven by the acquisition of a polybasic haemagglutinin cleavage site, enabling systemic viral spread, alongside emerging endothelial and neurotropic properties that contribute to severe disease and high mortality in mammals. Although zoonotic transmission remains limited, H5N1 continues to accumulate mutations associated with mammalian adaptation, particularly within the haemagglutinin and polymerase complex. Notably, recent outbreaks in U.S. dairy cattle highlight the emergence of novel mammalian reservoirs with increased human exposure risk. Concurrently, vaccination strategies are advancing beyond traditional adjuvanted inactivated vaccines toward next-generation platforms, including mRNA and virus-like particle vaccines, designed for rapid deployment and broader immune protection. However, ongoing viral evolution, constrained vaccine availability, and gaps in coordinated surveillance underscore the urgent need for an integrated One Health approach to reduce panzootic risk. Full article

Background/Objectives: Biosecurity and stamping out are key control measures against H5 high pathogenicity avian influenza (HPAI) outbreaks. Vaccination in poultry is an additional tool to reduce disease risk and facilitate timely containment. This study aimed to establish a candidate vaccine strain against H5 HPAI in Asia and validate its protective efficacy. Methods: Based on genetic and antigenic analyses, a representative HPAI virus, A/duck/Vietnam/HU16-DD3/2023 (H5N1), collected in northern Vietnam, was selected to generate a candidate vaccine strain, rgPR8/VN23HA∆KRRK-NA (rgPR8/VN23; H5N1), using reverse genetics, followed by formulation of an inactivated oil-adjuvanted vaccine. Vaccine efficacy was evaluated by measuring humoral antibody responses after intramuscular vaccination and by assessing mortality and virus recovery following intranasal challenge with a clade 2.3.4.4b virus, A/Ezo red fox/Hokkaido/1/2022 (H5N1). Results were compared with those obtained using an antigenically homologous vaccine to the challenge strain and a Japanese stockpiled vaccine. Results: All vaccinated juvenile chickens developed sufficient immunity to survive the challenge at 21 days post-vaccination. The rgPR8/VN23 (H5N1) and homologous vaccines markedly reduced virus recovery, suggesting near-sterile protection, whereas low-titer viruses were transiently detected in chickens vaccinated with the stockpiled vaccine. The rgPR8/VN23 (H5N1) vaccine conferred clinical protection in juvenile chickens as early as 8 days post-vaccination. A single dose of the rgPR8/VN23 (H5N1) vaccine provided incomplete protection in laying hens, whereas a double-volume regimen improved protective efficacy. Conclusions: The rgPR8/VN23 (H5N1) vaccine conferred strong immunity to juvenile chickens; however, a refined vaccination strategy may be required to achieve complete protection in laying hens. Full article

Highly pathogenic influenza A viruses of the H5 subtype continue to diversify worldwide through mutation and genetic reassortment, generating novel variants with unpredictable consequences under the One Health approach. Between 2024 and 2025, five outbreaks of avian influenza A viruses were detected in backyard poultry across Michoacán, Estado de México, and Ciudad de México. We conducted molecular and genetic characterization of five highly pathogenic H5N2 viruses isolated from these events. All cases tested positive for influenza A virus and the H5 hemagglutinin, exhibiting high pathogenicity with intravenous pathogenicity index values ranging from 2.88 to 3.0. Whole-genome sequencing revealed novel reassortants containing hemagglutinin from Eurasian H5N1 clade 2.3.4.4b and neuraminidase from the endemic Mexican H5N2 lineage. The viral genome of the isolate from Michoacán contained six segments derived from Eurasian H5N1 viruses introduced into North America in 2021–2022, while nucleoprotein and neuraminidase originated from Mexican H5N2 viruses. In contrast, viruses from Estado de México and Ciudad de México contained five H5N1-derived segments and incorporated polymerase basic protein 1, nucleoprotein, and neuraminidase from low-pathogenic H5N2 viruses circulating in 2024. Phylogenetic analyses confirmed the emergence of a distinct H5N2 Mexican sublineage, providing evidence of active viral reassortment and local evolutionary processes in Mexico. Full article

Highly pathogenic avian influenza A/H5N1 remains a persistent threat to public health and poultry production. H5N1 antigens are typically poorly immunogenic and require effective adjuvants for antigen dose-sparing. Here, we evaluated chitosan microparticles (CSMs) and nanoparticles (CSNs) as polymeric nano-adjuvants for an H5N1 influenza vaccine, focusing on the roles of antigen dose and particle size. A purified hemagglutinin antigen was adsorbed onto chitosan particles at doses ranging from 0.15 to 5.0 µg. Both CSNs and CSMs showed consistently high loading efficiency (97–99%). BALB/c mice were immunized intramuscularly in a prime–boost schedule. Chitosan nanoparticles significantly enhanced IgG and hemagglutination inhibition (HI) titers at low antigen doses compared with aluminum hydroxide and antigen-only controls (p < 0.05). Immune responses reached saturation at a 1.5 µg dose of antigen for chitosan nanoparticles and 3.0 µg for chitosan microparticles. IgG subtype analysis suggested a balanced IgG1/IgG2a profile. Collectively, these findings support chitosan-based polymeric nanoparticles as promising adjuvants enabling dose-sparing H5N1 vaccination. Full article

Avian influenza, a disease caused by avian influenza virus (AIV), mainly infects birds but can also infect mammals, which poses a serious threat to public health. Therefore, thorough understanding of its pathogenic mechanism and the identification of antiviral targets are essential for the prevention, control, and treatment of AIV. The polymerase acidic protein (PA) is a core component of the viral RNA-dependent RNA polymerase complex and plays a central role in viral transcription through its cap-snatching activity during early infection. We employed a multi-omics approach combining transcriptome analysis with PA interaction proteomics to characterize host responses during AIV infection and explore the PA–host interaction network. Transcriptomics revealed a polarized host response marked by activated translation-related processes, mitochondrial energy metabolism, and innate immune signaling, alongside broad suppression of nuclear transcriptional regulation and cell cycle pathways. Immunoprecipitation–mass spectrometry identified host proteins associated with PA that were enriched in RNA metabolism, ribosome biogenesis, and protein homeostasis. Integrative analysis of transcriptomic and interactome data, along with protein–protein interaction network analysis, prioritized a subset of high-confidence PA-interacting host factors. Among these, ribosomal protein RPS27A was validated to interact with PA and to support viral replication during early infection in this study. Full article