#Safety, humoral and cellular immune responses to a pre-pandemic adjuvanted #influenza #H5N8 #vaccine

 

Abstract

Highly pathogenic avian influenza (HPAI) A(H5) viruses can be transmitted from infected birds to various mammalian species, including humans. Avian influenza viruses (AIVs), members of the Orthomyxoviridae family, possess segmented RNA genomes prone to reassortment, favoring the emergence of novel genetic traits that may alter transmissibility, pathogenicity, and antigenicity. Although no sustained human-to-human transmission has been reported, the potential adaptation of these viruses poses a significant pandemic threat. This study aimed to evaluate the non-clinical safety, toxicity, and humoral immune responses induced by an adjuvanted H5 influenza vaccine in rats and rabbits, to support future clinical safety trials in humans. Male and female Wistar rats and New Zealand rabbits were observed for 14, 28, and 90 days after receiving two intramuscular doses of the H5N8 vaccine (15 μg HA/dose) formulated with the IB160 oil-in-water emulsion adjuvant. No systemic comorbidities, central nervous system alterations, or relevant clinical signs were observed. Hematological parameters remained within normal ranges, with total and differential leukocyte counts showing only minor fluctuations (<1% of total leukocytes). Mild biochemical variations in urea and hepatic transaminase levels were not correlated with histopathological alterations. The vaccine elicited a robust humoral response soon after immunization, with all groups reaching protective HAI-antibody titers. Although antibody levels declined over time, particularly in males, they remained significantly above baseline, indicating durable immunological memory. Furthermore, the vaccine induced a specific cellular immune response, confirmed by IL-2 and TNF production by antigen-specific T lymphocytes in splenic cell cultures after the booster dose. In conclusion, the H5N8 vaccine with the IB160 adjuvant was well tolerated locally and systemically, without compromising vital organ function. The safety and immunogenicity findings are consistent with expectations for adjuvanted influenza vaccines, demonstrating strong and durable humoral and cellular immune responses.

Source: 

Link: https://www.sciencedirect.com/science/article/abs/pii/S0264410X26008583?via%3Dihub

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Sex differences in #vaccine-induced #neuraminidase cross-recognition impact #H5N1 #dissemination to the lower respiratory tract in mice

 

Abstract

H5N1 vaccines have been poorly immunogenic in humans, creating a challenge for vaccine development. Seasonal influenza vaccines offer some cross-protection against H5N1, but there has been no consideration of whether protection differs between the sexes. We investigated sex differences in antibody responses following receipt of either beta-propiolactone inactivated whole virus H1N1 or H5N1 (LAIV backbone) vaccines in C57BL/6 mice. Using systems serology assays, vaccination induced strong homologous and heterologous antibody responses, with females generating greater IgG titers than males against whole virus H1N1 and H5N1, which was primarily mediated by greater IgG responses to neuraminidase (NA) than hemagglutinin (HA) protein. Cross-reactive H5N1 IgG titers were greater among H1N1-vaccinated females, and primarily mediated by greater N1-specific IgG titers. IgG2b and IgG2c were the primary antibody isotypes generated in response to these vaccines, with females having greater IgG2b titers and enhanced binding to FcγRIV for avian and human NA than males following either homologous or heterologous vaccination. Antibody-dependent complement deposition was measured as an FcR-mediated non-neutralizing response against HA and NA and was more robust among H1N1 and H5N1 vaccinated females than their male counterparts in response to homologous HA only. Vaccinated females tended to have greater neutralizing antibody titers than males against the homologous vaccine strain, with limited cross-neutralizing antibodies detected in either sexes. Neuraminidase inhibition titers were greater in vaccinated females than males against the heterologous virus following H1N1 vaccination and against both the vaccine and heterologous viruses following H5N1 vaccination. When H1N1 and H5N1 vaccinated mice were challenged with a lethal dose of A/Texas/37/2024 H5N1, all H5N1 vaccinated mice were protected, regardless of sex. Among H1N1 vaccinated mice, while both sexes were protected against disease, H1N1 vaccinated females restricted virus to the upper respiratory tract and had lower pulmonary virus titers than males at 3 days post challenge. These findings highlight that sex differences in vaccine-induced NA-specific antibody responses are associated with differential respiratory dissemination of H5N1 and that sex should be considered in studies of vaccine-induced cross-reactive influenza immunity.

Competing Interest Statement

The authors have declared no competing interest.

Funder Information Declared

NIH/NIAID Johns Hopkins Center of Excellence for Influenza Research and Response, 75N93021C00045

Richard Eliasberg Family Foundation

Source: 

Link: https://www.biorxiv.org/content/10.64898/2026.05.26.728011v3

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Immunogen selection and prior #immunity shape #antibody breadth following immunisation with avian #H5 #hemagglutinin

 

Abstract

Avian influenza A viruses pose a persistent zoonotic threat to humans owing to their expanding host range and high case fatality rates. In particular, viruses from the 2.3.4.4b clade of the H5 subtype have now been detected in over 60 mammalian species, raising serious pandemic concerns. Understanding immune recognition of the H5 hemagglutinin (HA) is therefore critical for effective vaccine design and pandemic preparedness. To understand the breadth of cross-recognition induced by different H5 strains, we selected genetically diverse H5 human isolates from 2003-2023 and assessed neutralising antibody responses elicited by adjuvanted recombinant HA protein-based vaccines in C57BL/6 mice. Neutralisation activity of sera was determined against seven H5 HA variants using pseudotyped viruses and a PR8-reassortant virus in micro-neutralisation assays. Our results showed a wide variety of cross-strain neutralisation across H5 HA antigen variants. The conventional vaccine strain A/Indonesia/05/2005 displayed narrow activity against emerging clade 2.3.4.4b viruses, whereas ancestral variants exhibited cross-neutralisation profiles showing a diversity of breath but with limited potency. Polyvalent H5 HA formulations and nanoparticle-displayed H5 HA platforms substantially broadened cross-neutralisation against diverse H5 strains. To examine the impact of pre-existing immunity on H5 vaccine immunogenicity in mouse models, mice were primed with either seasonal influenza infection or quadrivalent influenza vaccine (QIV) prior to H5 HA immunisation. QIV pre-vaccination, but not prior influenza infection, enhanced subsequent neutralizing responses towards A/Fujian-Sanyuan/21099/2017 (clade 2.3.4.4b) H5. Collectively, our results demonstrate that immunogen selection and prior immunity shape antibody breadth following immunisation with avian A(H5) hemagglutinin.

Source: 

Link: https://www.biorxiv.org/content/10.64898/2026.09.01.748495v1

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Isolation and characterisation of #Nipah virus neutralising candidate therapeutic monoclonal #antibodies from an #mRNA-immunised pig

 

Abstract

Nipah virus (NiV) is a highly pathogenic zoonotic paramyxovirus with epidemic potential. Despite the threat NiV poses, no therapeutics are licensed to treat infection. Studies have shown that monoclonal antibodies (mAb) can protect animals against NiV and the related Hendra virus (HeV). The best studied mAb, m102.4, has been used to treat infected patients on a compassionate basis, and has entered clinical trials. However, there is a need to define additional mAbs with therapeutic potential, which could be combined with m102.4 to improve neutralising potency and breadth. Here, we isolated five high affinity mAbs from an mRNA immunised pig, which bound the G glycoprotein derived from NiV Malaysia strain (NiV-M), and one of which (mAb A2) also bound HeV G. Aligned with this, all mAbs neutralised NiV-M pseudovirus but only mAb A2 neutralised pseudovirus representing the NiV Bangladesh (NiV-B) strain. mAb A2 and the most potent NiV-M neutralising mAb, C1, showed minimal competition with each other and m102.4, suggesting recognition of non-overlapping epitopes. Single-particle cryogenic electron microscopy of the NiV-M G receptor binding domain complexed to A1 and C2 Fab fragments revealed distinct epitopes that did not overlap with the receptor-binding site, targeted by m102.4, suggesting action through steric impedance of receptor binding or interference downstream of receptor engagement. Inoculation of mAb A2 to hamsters did not provide complete protection against NiV-B challenge (60% survival), however, a split dose of mAb A2 and m102.4 provided the same protection as m102.4 alone (100% survival). Collectively, these data demonstrate the potential of the porcine model for isolation of therapeutic candidate mAbs, which contribute both to our understanding of the NiV G antigenic landscape, and the development of mAb combinations, that exert complementary mechanisms of neutralisation, for therapeutic intervention.

Competing Interest Statement

The authors have declared no competing interest.

Funder Information Declared

European Commission, https://ror.org/00k4n6c32, VetBioNet, EMJMD LIVE

Innovate UK, 971555

UK Research and Innovation, BBS/E/I/00007031, BBS/E/I/00007037, BBS/E/I/00007038, BBS/E/I/00007039, MR/S007555/1, BB/T008784/1

Wellcome Trust, 203141/Z/16/Z

Source: 

Link: https://www.biorxiv.org/content/10.64898/2026.08.28.745669v1?rss=1

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Impaired #Neuraminidase and Polymerase Activities Correspond With Limited Aerosol Infectivity of #B3.13 and #D1.1 #H5N1 Lineages in #Human Respiratory Cultures

 

Abstract

The ongoing panzootic of clade 2.3.4.4b highly pathogenic avian influenza (HPAI) H5N1 viruses has reached a critical point, marked by unprecedented mammalian spillover and sustained outbreaks in U.S. dairy cattle. While these viruses remain highly lethal in traditional ferret models, human infections-primarily linked to the B3.13 and D1.1 lineages-have been notably mild, typically presenting as conjunctivitis with minimal respiratory involvement. In this study, we address this disconnect by evaluating the infectivity of recent H5N1 isolates using a physiologically relevant air-liquid interface (ALI) culture system that incorporates an aerosol settling chamber. We demonstrate that while direct liquid inoculation leads to efficient replication, aerosolized H5N1 strains exhibit a significant defect in their ability to infect human respiratory epithelium. In contrast, a prototypic H5N1 virus remains highly pathogenic and lethal in ferrets regardless of the inoculation route, showing systemic dissemination to the brain and other organs. Our findings identify two primary viral determinants driving this respiratory restriction: reduced neuraminidase (NA) enzymatic activity and impaired polymerase activity. Collectively, these results suggest that commonly used mammalian models may overstate current human pandemic risk. This work highlights the critical need for alternative risk-assessment platforms to identify the specific genetic shifts required for these viruses to overcome existing barriers to human adaptation.

Source: 

Link: https://www.biorxiv.org/content/10.64898/2026.08.18.745466v1

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Characterisation of a #human monoclonal #antibody targeting a conserved epitope at the base of the HA head of #influenza #H3N2 virus

 

Summary

Background

Numerous broadly reactive human monoclonal antibodies (mAbs) against the haemagglutinin (HA) of influenza A viruses have recognised conserved epitopes across HA subtypes or within subtypes. Most heterosubtypic mAbs target the HA stem, the receptor-binding site (RBS), or the trimeric interface. Although at least three H3-specific mAbs recognise epitopes outside these regions, the overall landscape of conserved H3-specific epitopes remains incompletely understood. This study aimed to identify and characterise conserved epitopes on H3-HA to inform the development of vaccines resilient to antigenic change.

Methods

We screened a panel of previously reported H3-HA-reactive human mAbs to identify mAbs recognising conserved epitopes. The candidate clone, 034-10040 4F02 (4F02), was evaluated for neutralising, haemagglutination inhibiting, and HA-mediated fusion-inhibitory, Fc receptor-mediated effector functions in vitro, and for protective efficacy in a lethal mouse challenge model. Cryo-electron microscopy was used to define the structural basis of 4F02 binding. Human sera were screened for antibodies targeting similar epitopes.

Findings

Clone 4F02 recognised the HA of human influenza A (H3N2) viruses that circulated across multiple decades. It neutralised multiple H3N2 viruses, exhibited weak haemagglutination inhibition, blocked HA-mediated fusion activity, activated Fc receptor-mediated signalling, and protected mice against lethal challenge. Cryo-electron microscopy revealed that 4F02 targets the base of the HA head at a head-stem interface spanning antigenic sites C, D, and E. Antibodies targeting similar epitopes were detected, albeit at a low level, in human sera.

Interpretation

Characterisation of the 4F02 epitope reveals a previously underappreciated site of vulnerability at the H3-HA head-stem interface. This finding expands our understanding of conserved epitopes and provides a target for the development of influenza vaccines resilient to antigenic change.

Funding

This work was supported by the Japan Agency for Medical Research and Development, JSPS KAKENHI, the National Institutes of Allergy and Infectious Diseases.

Source: 

Link: https://www.thelancet.com/journals/ebiom/article/PIIS2352-3964(26)00334-8/fulltext

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A mouse #monoclonal #antibody against #H7N9 #influenza virus cross-react with #human #platelets

 

Highlights

    • H7N9–98 mAb specifically binds human platelets and targets a ~60 kDa platelet protein.

    • H7N9 virus induces ITP via molecular mimicry with platelet antigens.

    • H7N9 virus-related ITP correlates with disease severity.

Abstract

Objective

To investigate the association and potential mechanisms between H7N9 influenza virus infection or vaccination and immune thrombocytopenia (ITP), providing foundational data for the prevention and treatment of related ITP.

Methods

Using laboratory-prepared anti-H7N9 influenza virus monoclonal antibodies (mAbs) (H7N9-98 and H7N9-120) as research subjects, the interactions between these antibodies and human platelets were analyzed through Western blotting (WB), immunohistochemistry (IHC), and immunofluorescence (IF) techniques.

Results

The mAb H7N9-98 exhibited specific binding to human platelets, showing positive results in both IHC and IF assays. Western blotting results demonstrated that this antibody could specifically recognize approximately 60 kDa human platelet proteins. The isotype control mAb H7N9-120 did not exhibit the aforementioned binding reactions, with all test results being negative.

Conclusion

These findings suggest that the specific antibodies induced by the H7N9 virus may mediate platelet damage through cross-reactivity with platelet autoantigens. This mechanism warrants further investigation to provide experimental evidence for the pathogenesis of secondary ITP associated with H7N9 infection.

Source: 

Link: https://www.sciencedirect.com/science/article/pii/S1567134826001346?via%3Dihub

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Trimester-dependent vertical #transmission of #H5N1 #influenza virus through #placental and mammary routes impairs offspring development

 

Abstract

Avian influenza H5N1 has pandemic potential and historically causes more severe disease in pregnant women than the general population. With increasing transmission of H5N1 detected among placental mammals, animal models are necessary for testing countermeasures, including during pregnancy. Pregnant outbred mice infected with a contemporary strain of bovine H5N1 during the second trimester equivalent causes in utero transmission, with infectious virus detected in the uterus, placenta, and fetus. Birth following third trimester infection results in offspring with decreased size, neurodevelopmental delays, and adolescent behavioral impairments, with infectious virus detected in the neonatal milk ring and lungs, as well as mammary tissues. H5N1 viral protein colocalizes with trophoblast cells in the placenta and epithelial cells in mammary tissue that spatially overlap with lectins for α2,3-linked SA. With the pandemic potential of H5N1, our vertical transmission model in placental mammals is essential for understanding viral spread and evaluating treatments during pregnancy.

Source: 

Link: https://www.nature.com/articles/s41467-026-76891-9

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#Nipah virus #Malaysia and #Bangladesh strain-induced #pathogenesis in mice lacking type I #interferon receptor signaling

 

Abstract

Nipah virus (NiV) is a zoonotic highly pathogenic Paramyxovirus inducing lethal outbreaks of encephalitis and Acute Respiratory Distress Syndrome (ARDS) with an average case-fatality rate of 75%. Two viral strains, NiV-Malaysia (NiV-Mal) and NiV-Bangladesh (NiV-Ban), associated to distinct route of transmission, symptoms and lethality have been described. Due to the permanent threat of these emerging infections and the lack of approved therapeutics, it is crucial to improve our understanding regarding NiV-associated pathogenesis. Mice represent a small and accessible animal model, provided with numerous biological tools for the functional assessment of different genes related to antiviral response. Here, we explore the susceptibility of mice deficient for type I interferon receptor (IFNAR KO) to inoculation with either NiV-Mal or NiV-Ban through intraperitoneal or intranasal routes. Our results complement observations showing that IFNAR KO mice are susceptible to NiV-Ban infection via intraperitoneal route, although to a lesser extent than NiV-Mal, and develop encephalitis and a pulmonary syndrome with viral dissemination to various organs. Additionally, intranasal administration of both viral strains exhibited a subclinical infection with viral replication in the brain and the lungs along to the production of neutralizing antibodies in some animals. These results suggest that IFNAR KO mice may represent a reliable model permitting comparative studies of the immunopathogenesis induced by both NiV-Mal and NiV-Ban infections.

Source: 

Link: https://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0013894

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A novel candidate #vaccine virus derived from #Japan's first #mammalian case of clade 2.3.4.4b #H5N1 highly pathogenic avian #influenza virus

 

Abstract

The development of candidate vaccine viruses (CVVs) for pre-pandemic preparedness requires attenuation of pathogenicity while maintaining immunogenicity. In this study, we developed and characterized NIID-002, a reassortant virus derived from A/Ezo red fox/Hokkaido/1/2022 (H5N1; clade 2.3.4.4b), to evaluate its suitability as a candidate vaccine. NIID-002 exhibited markedly reduced pathogenicity compared with its parental strain, while retaining broad antigenic reactivity and protein yield comparable to other clade 2.3.4.4b CVVs. In mammalian models, NIID-002 demonstrated strong attenuation, causing no lethal infection in mice and only minimal weight loss with limited viral replication in ferrets. Antisera raised against NIID-002 reacted broadly with recent wild-type H5N1 isolates, suggesting potential broad protection. Protein yield analysis confirmed a production efficiency comparable to that of other CVVs within the same clade, supporting its feasibility for large-scale vaccine manufacturing. Overall, NIID-002 fulfills the key requirements for the pandemic preparedness of CVV, combining reduced pathogenicity, broad antigenic reactivity, and adequate production efficiency. These findings highlight its potential as a candidate H5N1 vaccine and underscore the continued need for surveillance and refinement of influenza vaccine strategies to address evolving viral threats.

Source: 

Link: https://www.sciencedirect.com/science/article/abs/pii/S0264410X26008571?via%3Dihub

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Enhanced #Pathogenicity and Contact #Transmissibility of #Human-origin Avian #Influenza #H5N1 Clade 2.3.4.4b Genotype B3.13 Compared to D1.1 in #Ferrets

 

Abstract

Since its emergence in 2020, multiple genotypes of the H5N1 clade 2.3.4.4b have been identified, with B3.13 and D1.1 emerging in the USA as two major and concerning genotypes. However, their relative pathogenicity and transmissibility in mammals have not been fully elucidated. We compared the pathogenicity and transmissibility of the first two human H5N1 clade 2.3.4.4b cases caused by B3.13 in Texas (A/Texas/37/2024; HPhTX B3.13) and D1.1 in Louisiana (A/Louisiana/12/2024; HPhLA D1.1) in a ferret model of infection and transmission. HPhTX B3.13 infection resulted in more severe clinical disease and enhanced viral shedding, with evidence of increased transmission relative to HPhLA D1.1. Histopathological analysis revealed more extensive lung pathology in animals infected with HPhTX B3.13, consistent with increased viral loads and inflammatory responses. Importantly, both genotypes showed no significant differences in reactivity to ferret sera raised against candidate vaccine virus (CVV) strains, receptor binding properties, or neuraminidase (NA) activity and thermostability. Whole-genome sequencing revealed no adaptive mutations in HPhTX B3.13 following infection or transmission. In contrast, HPhLA D1.1 showed rapid acquisition of the mammalian-adaptive mutation E627K in infected ferrets and both E627K and Q194K in the only fatal contact animal. Both mutations were associated with enhanced polymerase activity and computational analyses suggested that they enhance interactions with the mammalian host factors ANP32A and B. Our findings indicate that B3.13 is already well adapted for mammalian infection and transmission whereas D1.1 retains evolutionary potential through the rapid acquisition of adaptive mutations, highlighting important genotype-specific differences relevant to zoonotic risk assessment and pandemic preparedness.

Competing Interest Statement

The A.G.-S. laboratory has received research support from Avimex, Dynavax, Pharmamar, and Accurius, outside of the reported work within the last three years. A.G.-S. has consulting agreements for the following companies involving cash and/or stock within the last three years: Castlevax, Amovir, Vivaldi Biosciences, Contrafect, Avimex, Pagoda, Accurius, Applied Biological Laboratories, Pharmamar, CureLab Oncology, CureLab Veterinary, Virofend, Prosetta and A.A.C.T., outside of the reported work. A.G.-S. has been an invited speaker in meeting events within the last three years organized by Seqirus, Novavax and Hipra. A.G.-S. is inventor on patents and patent applications on the use of antivirals and vaccines for the treatment and prevention of virus infections and cancer, owned by the Icahn School of Medicine at Mount Sinai, New York, outside of the reported work. The Icahn School of Medicine at Mount Sinai has licensed some of these inventions to Medimmune, Avimex, Leinco Technologies, Castlevax, Virofend, Kerafast, Cell Signaling, EMD Millipore, Genentech, Paratus and Nura Bio, and as a result receives financial compensation. Subject to Mount Sinai receiving such financial consideration, AG-S will receive a portion of that consideration pursuant to the terms of the Mount Sinai Intellectual Property Policy. All other authors declare no commercial or financial conflict of interest.

Funder Information Declared

NIH/NIAID, 75N93021C00014

Horizon Europe Program, KAPPA-FLU no. 101084171

Source: 

Link: https://www.biorxiv.org/content/10.64898/2026.08.10.744032v1

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#Lassa fever and Argentine hemorrhagic fever #treatment in guinea pigs using broad-spectrum cap-dependent #endonuclease #inhibitors

 

ABSTRACT

The class Bunyaviricetes encompasses several highly pathogenic viruses that cause lethal hemorrhagic fevers. Due to their limited prevention and treatment options and high pathogenicity, these viruses require handling in biosafety level-4 facilities. Within the bunyaviruses, arenaviruses are particularly notable for their pathogenicity and ability to cause severe hemorrhagic disease in humans. The cap-dependent endonuclease (CEN) is a unique and crucial enzyme involved in the replication cycle of these viruses. As humans do not possess a similar enzyme, CEN represents an ideal target for antiviral drug development with reduced risk of side effects. Recently, we identified a promising CEN inhibitor (CENi) demonstrating potent inhibition of virus replication. In this manuscript, we demonstrate the successful therapeutic efficacy of CENis against Lassa fever and Argentine hemorrhagic fever virus infections in guinea pig models of lethal hemorrhagic fever. In addition, we identified several CENis with antiviral activity against other highly pathogenic arenaviruses. These findings further support the potential of CENis as therapeutic agents for arenavirus infections that cause severe and often lethal hemorrhagic fever. Collectively, our results suggest that CENis are promising candidates for pan-arenavirus therapy and may also have broader utility against other CEN-containing viruses for which no approved antiviral treatments currently exist.

Source: 

Link: https://journals.asm.org/doi/10.1128/mbio.00880-26

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Innate Immune Responses Induced by #H9N2 #Influenza A Virus and #Klebsiella pneumoniae Co-Infection

 

Abstract

Klebsiella pneumoniae infection following H9N2 Influenza A virus (IAV) infection causes severe pneumonia. But the underlying pathogenic mechanisms of H9N2 IAV and K. pneumoniae co-infection are complex and need to be further explored. In this study, the lung transcriptomes of mice with H9N2 IAV and K. pneumoniae co-infection were characterized by transcriptomic profiling. As a result, GO enrichment analysis revealed that the differential genes were primarily involved in the activation of immune responses, cellular components of membranes and extracellular spaces, and defense responses against pathogen infections. According to KEGG enrichment, the differentially expressed genes (DEGs) were concentrated in TLR signaling pathways, RLR signaling pathways, TNF signaling pathways and NLRP3 signaling pathways. Furthermore, in vitro cell models were established to investigate the innate immune responses induced by H9N2 IAV and K. pneumoniae CPS co-stimulation. K. pneumoniae CPS stimulation influenced the cytokine profiles of mink lung epithelial cells infected with H9N2 IAV, worsened cell viability, and aggravated apoptosis, indirectly inhibiting H9N2 IAV replication. The findings demonstrated that K. pneumoniae superinfection modulated the innate immune responses induced by H9N2 IAV infection, contributing to its pathogenesis.

Source: 

Link: https://www.mdpi.com/1999-4915/18/8/900

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Identification and characterization of #PB2 #mutations associated with #mammalian #adaptation of highly pathogenic #H5N1 avian #influenza viruses

 

Abstract

The highly pathogenic avian influenza virus (HPAIV) subtype H5N1 has been continuously circulating among wild bird populations and domestic poultry. It’s ongoing circulation has led to outbreaks in poultry and U.S. dairy cattle populations, as well as sporadic severe infections in individuals engaged in poultry and dairy farming. These occurrences have raised concerns about the potential evolution of this virus into a pandemic strain. To elucidate the molecular determinants facilitating H5N1 cross-species adaptation and to evaluate its implications for public health, we conducted serials of sequence analysis and specific-site mutations on the viral polymerase subunit PB2 to determine its effect on polymerase activity and viral infectivity. The results showed that three mutations in the PB2 protein (E362G, D441N and M631L) were presented cooperative effects associated with enhanced viral replication in mammalian cells. Compared to the original isolated strain of the 2.3.4.4b clade, A/chicken/NL/FAV-0033/2021, these three mutations were predominantly identified in isolates obtained from cattle and other mammalian hosts between 2021 and 2024. The M631L mutation, identified as the primary determinant of increased polymerase activity in mammalian cells, significantly enhanced the binding affinity of PB2 to ANP32A. The mutation E362G and D441N did not increased polymerase activity and viral replication significantly but enhanced binding affinity of PB2 to ANP32A. The combined mutations with E362G, D441N and M631L resulted in a significantly increased polymerase activity and viral replication in H5N1 virus, and significantly elevated viral loads and aggravated pulmonary pathology in lungs of mice with H5N1 infection. These findings indicate that the PB2-M631L mutation constitutes a crucial molecular marker for the adaptation of H5N1 to mammalian hosts, whereas the E362G and D441N mutations likely function as supportive modulatory factors that optimize this host-adaptation process.

Source: 

Link: https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2026.1867604/full

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#Risk #assessment of avian #influenza #H5N5 virus from the first #human case using the #ferret model

 

ABSTRACT

The incursion of Eurasian-origin genotype A6 A(H5N5) virus into North America expanded the genetic diversity among North American highly pathogenic avian influenza viruses and heightened concern about zoonotic risk. Following a fatal human infection with the A(H5N5) virus A/Washington/2148/2025, viral replication was assessed in polarized human bronchial epithelial cells, and pathogenicity, transmissibility in direct contact and respiratory droplet models, and airborne virus shedding were evaluated in ferrets to inform pandemic risk assessment. A(H5N5) displayed robust replication in Calu-3 cells at 33°C and 37°C, showing kinetics and peak titers comparable to those of contemporary genotype B3.13 and D1.1 A(H5N1) viruses. In ferrets, A(H5N5) replicated efficiently in the respiratory tract, disseminated to extrapulmonary tissues, and caused fatal disease in all inoculated animals. Airborne transmission was not observed, and infrequent, low-level detection of virus in air samples paralleled that of A(H5) viruses that are not transmissible via air in ferrets. In a direct contact model, limited transmission was detected within 4 days of exposure, with evidence of lower respiratory tract replication in contact animals. These findings indicate that the A(H5N5) virus has the capacity for robust replication in an airway epithelial cell line and can cause severe systemic infection and mortality in ferrets but has not acquired adaptations for airborne spread in mammals. Collectively, these results underscore heterogeneity among clade 2.3.4.4b A(H5Nx) viruses in North America and the need for genotype-by-genotype evaluation of newly emerged viruses to understand public health risk.

IMPORTANCE

The emergence of Eurasian-origin genotype A6 highly pathogenic avian influenza A(H5N5) virus in North America has increased viral diversity and raised concerns about zoonotic and pandemic risk. In this study, we evaluated the replication kinetics, pathogenesis, and transmission of A/Washington/2148/2025 A(H5N5) virus, which was isolated from the first reported human infection with this influenza virus subtype, using polarized human bronchial epithelial cells and the ferret model. The A(H5N5) virus replicated efficiently in vitro at temperatures representative of the upper and lower respiratory tracts and caused fatal systemic disease in inoculated ferrets. Limited transmission was observed during 4 days of direct contact. Airborne virus detection was infrequent and did not result in airborne transmission. These findings show that A(H5N5) virus can replicate robustly in mammalian cells and cause severe disease but lacks adaptations supporting efficient airborne spread, informing assessment of the pandemic risk posed by genotype A6 influenza viruses.

Source: 

Link: https://journals.asm.org/doi/10.1128/jvi.00856-26

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Protective effect of #H5N8 stockpiled #vaccine against a virus genetically identical to a #human isolate of #bovine #H5N1 #influenza virus

 

Summary

Background

Since early 2024, highly pathogenic avian influenza A(H5N1) viruses of clade 2.3.4.4b have caused extensive outbreaks in dairy cattle in the United States, with spillover into mammalian species, including humans. A bovine-derived A(H5N1) virus isolated from a human case retains high pathogenicity and transmissibility in mammalian models, highlighting its pandemic potential. Stockpiled pre-pandemic influenza vaccines are intended to provide early protection before strain-matched vaccines are available; however, their protective efficacy against bovine A(H5N1) viruses has not been directly evaluated in vivo.

Methods

In this study, we assessed the protective efficacy of an AS03-adjuvanted A/Astrakhan/3212/2020 (H5N8) clade 2.3.4.4b-based influenza vaccine stockpiled in Japan using mouse and ferret models. Vaccinated and unvaccinated animals were challenged with a virus genetically identical to a human isolate of bovine A(H5N1) virus. Neutralising antibody responses, viral replication in organs, and survival were evaluated.

Findings

Vaccination with the AS03-adjuvanted A(H5N8)-based stockpiled vaccine induced robust neutralising antibody responses in both animal models, significantly suppressed viral replication, and conferred complete protection against lethal challenge. In contrast, all unvaccinated mice and ferrets succumbed to infection. These findings demonstrate that the AS03-adjuvanted A(H5N8)-based stockpiled vaccine provides strong cross-protective efficacy against bovine A(H5N1) viruses.

Interpretation

An AS03-adjuvanted A(H5N8)-based vaccine stockpiled in Japan could serve as an immediate countermeasure against bovine A(H5N1) viruses during the early phase of a pandemic.

Funding

This work was supported by grants from the Japan Program for Infectious Diseases Research and Infrastructure (JP20wm0125002) and the Japan Initiative for World-leading Vaccine Research and Development Centers (JP223fa627001) from the Japan Agency for Medical Research and Development.

Source: 

Link: https://www.thelancet.com/journals/ebiom/article/PIIS2352-3964(26)00314-2/fulltext

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A One-Shot Multivalent Live-Attenuated Candidate #Influenza #Vaccine against Divergent #Zoonotic #H5N1 Clades

 

Abstract

The continued emergence of genetically diverse high pathogenicity avian influenza (HPAI) H5N1 viruses with zoonotic potential highlights the urgent need for developing vaccines capable of providing broad protection against multiple circulating clades. Here, we developed a one-shot, multivalent, live-attenuated influenza vaccine (LAIV) based on the temperature-sensitive (ts), cold-adapted (ca), and attenuated (att) influenza A/Ann Arbor/6/1960 master donor virus (MDV) that incorporates the hemagglutinin (HA) and neuraminidase (NA) glycoproteins from representative clades 2.3.4.4b (A/Louisiana/12/2024), 2.3.2.1a (A/Victoria/149/2024), and 2.3.2.1e (A/Cambodia/2302009/2023) H5N1 viruses. A single intranasal (IN) immunization of C57BL/6 mice with the multivalent LAIV elicited robust humoral immune responses, with immune sera exhibiting broad cross-reactivity against antigens from all three H5N1 clades included in the vaccine. Following homologous viral challenge, vaccinated C57BL/6 mice were completely protected from disease, demonstrating the immunogenicity and protective efficacy of the multivalent LAIV. By simultaneously targeting antigenically distinct H5N1 lineages with pandemic potential, this strategy expands antigenic coverage within a single LAIV to confirm pan-H5N1 protection. Together, these findings support the development and implementation of this multivalent LAIV as a broadly protective pan-H5N1 LAIV for pandemic preparedness.

Competing Interest Statement

The A.G.-S. laboratory has received research support from Avimex, Dynavax, Pharmamar, and Accurius, outside of the reported work within the last three years. A.G.-S. has consulting agreements for the following companies involving cash and/or stock within the last three years: Castlevax, Amovir, Vivaldi Biosciences, Contrafect, Avimex, Pagoda, Accurius, Applied Biological Laboratories, Pharmamar, CureLab Oncology, CureLab Veterinary, Virofend, Prosetta and A.A.C.T., outside of the reported work. A.G.-S. has been an invited speaker in meeting events within the last three years organized by Seqirus, Novavax and Hipra. A.G.-S. is inventor on patents and patent applications on the use of antivirals and vaccines for the treatment and prevention of virus infections and cancer, owned by the Icahn School of Medicine at Mount Sinai, New York, outside of the reported work. The Icahn School of Medicine at Mount Sinai has licensed some of these inventions to Medimmune, Avimex, Leinco Technologies, Castlevax, Virofend, Kerafast, Cell Signaling, EMD Millipore, Genentech, Paratus and Nura Bio, and as a result receives financial compensation. Subject to Mount Sinai receiving such financial consideration, AG-S will receive a portion of that consideration pursuant to the terms of the Mount Sinai Intellectual Property Policy. All other authors declare no commercial or financial conflict of interest.

Source: 

Link: https://www.biorxiv.org/content/10.64898/2026.08.02.742385v1

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The #Environmental Polycyclic Aromatic Hydrocarbon (PAH) #Benzopyrene (BP) Alters #SARS-CoV-2 #Pathogenesis in a Mouse #Model of Disease

 

Abstract

Since emerging in late 2019, SARS-CoV-2 has caused over 7 million deaths globally and remains a public health concern. Understanding SARS-CoV-2 pathogenesis is vital, especially as factors like environmental exposures are still poorly understood. Polycyclic aromatic hydrocarbons (PAHs), like benzo[a]pyrene (BP), found in pollutants like cigarette smoke, diesel exhaust, and charcoal-broiled steaks, are known to injure the lungs. We aimed to evaluate if BP exacerbates SARS-CoV-2 pathogenesis in a mouse model of disease. One day following intranasal administration of BP (20 mg/kg) or vehicle control, we infected male and female K18-hACE2 mice with ancestral SARS-CoV-2 and assessed lung viral load, weight change, clinical scores, immune cell recruitment, and survival in the presence and absence of BP exposure. We found that BP-exposed mice had decreased survival compared to mock-exposed mice. Additionally, BP did not alter innate or adaptive immune cell populations in the lungs of SARS-CoV-2-infected mice. These findings suggest that PAH exposure exacerbates severe COVID-19 outcomes by unknown mechanisms, highlighting the need to further explore environmental impacts on SARS-CoV-2 infection.

Source: 

Link: https://www.mdpi.com/1999-4915/18/8/823

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A lethal #human #H5N5 #influenza virus isolate exhibits low #pandemic #risk traits

 

Abstract

In fall of 2025, a fatal infection of highly pathogenic avian influenza (HPAI) virus H5N5 occurred. To define the risk of this emerging virus to humans, we performed a comprehensive analysis based on our established triage. Serological analysis revealed that humans across all birth years had no detectable neutralizing antibodies to this H5N5 isolate. Further characterization revealed a lack of phenotypic signatures associated with epidemiologically successful influenza viruses in humans, including reduced replication in human airway cells and an avian-like pH of inactivation. Additionally, assessment of H5N5 in ferrets revealed a lack of direct contact transmission and moderate disease severity. H5N5 infection in ferrets with prior immunity against the 2009 H1N1 pandemic strain resulted in fewer clinical signs and reduced viral shedding. Together our data suggest that the current H5N5 HPAI lineage poses a low pandemic risk.

Source: 

Link: https://www.biorxiv.org/content/10.64898/2026.07.20.739507v1

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Elicitation of #stem-directed #antibodies in rhesus #macaques by a conventional #hemagglutinin immunogen

 

Abstract

Because they can bind many strains of influenza, antibodies targeting the hemagglutinin (HA) stem have been attractive targets for vaccine development. Many monoclonal antibodies (mAbs) directed at the HA stem have been isolated from humans, and these mAbs have mediated broad protection in animal models. We describe here HA stem-directed mAbs isolated from rhesus macaques immunized with an "ordinary" H1 HA trimer. All immunized rhesus macaques developed high serum titers with broad reactivity to diverse H1N1 and H5N1 viruses, and 7 isolated mAbs strongly blocked canonical stem antibody CR6261 binding to H1. MAb DH726.1 robustly protected mice from lethal challenge with H1N1 and H5N1 viruses, and cryo-EM showed the binding footprint overlapped that of some human mAbs. These findings suggest that vaccination with the standard, trimeric HA immunogens may be sufficient to elicit stem antibodies at titers adequate to protect against zoonotic H5N1 influenza.

Competing Interest Statement

The authors have declared no competing interest.

Funder Information Declared

NIH NIAID, Division of Microbiology and Infectious Diseases, P01-AI089618

NIH NIAID Division of AIDS, Center for HIV/AIDS Vaccine Immunology, U19-AI067854

Source: 

Link: https://www.biorxiv.org/content/10.64898/2026.07.16.738984v1

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