ETIDIOH - NEW

  Abstract Influenza virus mRNAs are stable and competent for nuclear export and translation because they receive a 5′ cap(1) structure in a process called cap snatching 1. During cap snatching, the viral RNA-dependent RNA polymerase (FluPol) binds to host RNA polymerase II (Pol II) and the emerging transcript2,3. The FluPol endonuclease then cleaves a capped RNA fragment that subsequently acts as a primer for the transcription of viral genes 4,5. Here we present the cryogenic electron microscopy structure of FluPol bound to a transcribing Pol II in complex with the elongation factor DSIF in the pre-cleavage state. The structure shows that FluPol directly interacts with both Pol II and DSIF , positioning the FluPol endonuclease domain near the RNA exit channel of Pol II . These interactions are important for the endonuclease activity of FluPol and FluPol activity in cells. A second structure , trapped after cap snatching, shows that the cleaved capped RNA rearranges within FluPol, ...

  Abstract Influenza A virus is a highly contagious respiratory pathogen, and its rapid and continuous adaptive mutations for immune escape have limited the efficacy of existing vaccines and antiviral drugs. Here, we report a multimechanism anti-influenza platform based on the conjugation of zanamivir (ZMV) with amantadine (Aman). Aman acts as a hydrophobic tag that promotes the degradation of neuraminidase and concurrently enhances the physicochemical properties of ZMV , leading to improved membrane permeability and a significantly prolonged half-life. Meanwhile, the ZMV moiety counteracts Aman-induced cytotoxic autophagy . The resulting conjugate, compound 7j , exhibits potent activity against a wide range of neuraminidase and M2 ion channel mutations . Notably, a single intravenous dose of 7j fully protected mice from a lethal H1N1 challenge . Our work demonstrates that the rational fusion of ZMV and Aman achieves synergistic multimechanistic antiviral activity with enhanced eff...

  Abstract Newly emerging influenza virus strains pose a constant threat as they encounter a population lacking neutralizing antibodies against the new strain . However, cross-reactive non-neutralizing antibodies (nnABs) may be present and assist in mitigating disease symptoms via various effector mechanisms, including antibody-dependent cellular cytotoxicity (ADCC). Although nnABs to influenza virus have received more attention lately , little information is available on their age-related prevalence , steady-state levels, functional properties , and changes in these parameters over time. Using longitudinal samples from adolescents, adults, and older adults , collected before and after the 2009 swine flu pandemic , we comprehensively characterized the specificity and functionality of nnAB responses against H1N1 pandemic 2009 (H1N1pdm09) virus . Remarkably, all participants exhibited cross-reactive antibodies to this virus before having encountered it through infection or vaccinatio...

  Abstract Background :  Genetic drift and host-associated adaptation in influenza A viruses threaten the long-term reliability of RT-qPCR-based diagnostics , particularly when nucleotide mismatches arise within primer and probe binding regions . Conventional assay evaluations often emphasize sequence conservation but rarely assess functional mismatch tolerance across divergent subtypes and hosts.  Methods :  We performed an in silico evaluation of a matrix (M) gene–targeted RT-qPCR assay by aligning primer and probe binding regions against 22 H1N1 isolates and representative H3N2 and H5N1 reference strains, including recent zoonotic isolates from avian and bovine hosts . Nucleotide mismatches were identified, quantified, and mapped relative to assay components and oligonucleotide termini. Mismatch burden was summarized by subtype and assay region.  Results :  H1N1 isolates exhibited complete conservation across primer and probe regions. In contrast, H3N2 a...

  ABSTRACT Swine influenza A virus (swIAV) is an important pathogen with regard to both the swine industry and public health . The pandemic A(H1N1) 2009 outbreak was caused by the swine-origin pandemic A(H1N1) 2009 [A(H1N1)pdm09] virus. Several reports have shown that several amino acid substitutions in the hemagglutinin (HA) antigenic sites can alter HA antigenicity. However, the impact of the amino acid deletion at position 155 on HA antigenicity remains unknown. In this study, we have isolated 11 samples of swIAVs from seven pig farms in Japan and found an amino acid deletion at position 155 of the HA region in one of the isolates of the H1N2 subtype . To examine the impact of this amino acid deletion on viral replication and HA antigenicity, we generated recombinant influenza A viruses possessing the H1 HA gene encoding either an artificial insertion or deletion of glycine at position 155. The growth kinetics of these recombinant viruses in two different cell lines demonstrated...

  Abstract The polymerase acidic (PA) protein is a subunit of the trimeric influenza A virus (IAV) RNA-dependent RNA polymerase and the target of the anti-influenza drug baloxavir marboxil (BXM). As with other direct-acting antivirals , treatment with BXM can lead to selection of viruses carrying resistance mutations . If these mutations have negligible fitness costs, resistant viruses can spread widely and render existing treatments obsolete . Multiple BXM resistance mutations in the nuclease domain of PA have been identified, with I38T and I38M amino acid substitutions occurring frequently. These mutations have minimal to no effects on viral polymerase activity , virus replication , or transmission . However, for reasons that are not well understood, viruses with BXM resistance substitutions have not been able to compete with parental wild-type strains . The IAV genome segment encoding PA also encodes the host shutoff nuclease PA-X, which shares the endonuclease domain with PA bu...

Abstract Key to the success of influenza A virus as a pathogen are its numerous tactics of immune evasion. To suppress anti-viral cellular and organismal responses , influenza A virus encodes several immunomodulatory proteins , including the endoribonuclease PA-X . PA-X decreases inflammation and immune responses in in vivo infections by limiting host gene expression . PA-X is conserved in 99% of all influenza A viral strains , pointing to its importance as a crucial immunomodulator . However, it is not yet known how PA-X activity alters the antiviral response in the human airway or how it benefits the virus. To define how influenza A virus uses this protein to evade immune responses , we characterized the impacts of PA-X on the host response to infection in the infected and bystander cells of the airway epithelium using a 3D ex vivo model. We discovered that PA-X exerts a dual action on immune responses , dampening aspects of both the innate and adaptive immune systems . Consistent wi...

  Abstract Influenza A virus (IAV) is a zoonotic pathogen with a broad host range, posing an ongoing threat to global public health . As the core subunit of the IAV polymerase , polymerase basic protein 1 (PB1) is essential for viral replication and transcription , and its mutations are key drivers of viral evolution . This review evaluates the impact of PB1 mutations on IAV evolution, with a focus on polymerase activity, host adaptation, transmissibility, and virulence. Additionally, it discusses the implications of these mutations for vaccine development. The review aims to provide insights that can inform influenza surveillance, identify novel antiviral targets, and guide vaccine design. Source:  Link:  https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2026.1768665/full ____

  Abstract Influenza-associated encephalitis/encephalopathy (IAE) is a severe neurological complication characterized by central nervous system dysfunction and structural damage following influenza virus infection . Predominantly affecting infants and young children , IAE exhibits its highest incidence in those under five years of age . Key clinical manifestations of IAE include acute seizures, sudden high fever, and impaired consciousness , frequently progressing to coma . Neuroimaging , particularly magnetic resonance imaging (MRI), often reveals multifocal brain lesions involving multiple brain regions, including the cerebellum, brainstem, and corpus callosum . The prognosis of IAE is poor , with a mortality rate reaching 30%. Current diagnosis relies heavily on clinical presentation and characteristic neuroimaging findings, as the precise pathogenesis of IAE remains elusive . While various research models, including cell lines, brain organoids, and animal models , have been dev...

  Abstract Selection of advantageous mutations drives the emergence of dominant variants during seasonal influenza epidemics . However, within-host detection of such variants remains rare , limiting our understanding of how selection operates at the scale of individual hosts. In this study, we used a controlled human infection model to examine the within-host evolutionary dynamics in thirteen participants intranasally infected with a seasonal H3N2 influenza A virus . Although this clinical trial is ongoing , our work represents a pre-planned, interim, exploratory analysis. Results in this system were contrasted with those observed in a ferret model of infection. The inoculum, used in both humans and ferrets, carried standing diversity that enabled evaluation of variant trajectories during infection. Although the dynamics were variable among participants, in humans , the minor variants in the PA and NP gene segments tended to increase in frequency as infection progressed. Variant dy...

  Abstract Hemagglutinins (HA) from different influenza A virus subtypes share as little as ~40% amino acid identity , yet their protein structure and cell entry function are highly conserved . Here we examine the extent that sequence constraints on HA differ across three subtypes . To do this, we first use pseudovirus deep mutational scanning to measure how all amino-acid mutations to an H7 HA affect its cell entry function. We then compare these new measurements to previously described measurements of how all mutations to H3 and H5 HAs affect cell entry function . We find that ~50% of HA sites display substantially diverged preferences for different amino acids across the HA subtypes. The sites with the most divergent amino-acid preferences tend to be buried and have biochemically distinct wildtype amino acids in the different HA subtypes. We provide an example of how rewiring the interactions among contacting residues has dramatically shifted which amino acids are tolerated at s...

  ABSTRACT Avian influenza virus cross-species infection in humans poses a major threat to global public health . Species-specific differences between avian ANP32A and mammalian ANP32 proteins create a natural barrier against viral cross-species infection by directly impairing the functional interaction between the avian-origin viral RNA polymerase and mammalian ANP32 proteins , thereby restricting viral genome replication . The key to overcoming this barrier lies in the adaptation of viral RNA polymerase to host ANP32 family proteins . This mini-review summarizes the mechanisms and variations in influenza virus adaptation to ANP32 proteins across different species. Influenza viruses adapt to species-specific ANP32 proteins through various mutations and display distinct preferences for specific ANP32 family members within the same host. Additionally, alternative splicing variants of ANP32A within a single species further modulate viral RNA polymerase adaptability. Despite this dive...

  ABSTRACT Multiple genes are involved in the pathogenicity of influenza A virus . Our previous study reported two naturally occurring amino acid mutations in the polymerase acidic (PA) protein as crucial determinants of the virulence of Eurasian avian-like H1N1 (EA H1N1) influenza viruses. PA-X, an accessory protein encoded by the PA gene , is thought to play a role in viral pathogenicity and regulation of host immune response , but its specific function remains unclear. In this study, we found that two genetically similar EA H1N1 influenza viruses , A/swine/Liaoning/FX38/2017 (FX38) and A/swine/Liaoning/SY72/2018 (SY72), induced significantly different suppression levels of host protein synthesis . The difference in host shutoff activity induced by PA-X protein was the key factor affecting the inhibition of host gene expression . Loss of PA-X expression significantly reduced its host shutoff activity , thereby enhancing host antiviral immune response . PA-X deficiency had no appa...

  ABSTRACT The 2009 pandemic H1N1 (pH1N1) influenza A virus (IAV) is a reassortant virus with two polymerase components, PA and PB2, originating from avian IAV . Avian IAV polymerase does not function efficiently in mammalian cells without host-adaptive mutations . The mechanism by which pH1N1 replicates in human hosts is not fully elucidated , as pH1N1 does not contain the host-adaptive PB2 E627K mutation required for species-specific interaction with ANP32 , which facilitates replicase (polymerase oligomer) formation. Our previous research revealed that mutations in PA played a key role in mammalian host adaptation of pH1N1. These mutations were found in two separate domains of PA, the C-terminal (CTD) and N-terminal domains (NTD). We reported that the NTD mutations increase the expression of NP through enhanced association of GRSF1 with the mRNA transcripts. However, the role of CTD mutations, which are located at the interface of the polymerase oligomers , has not been elucidat...

  Highlights •  Epidemiological parameters differ by pathogen and by setting. •  Unsupervised machine learning classifies pathogens into distinct epidemiological archetypes. •  Pathogens can be allocated into defined groups outlining plausible parameter ranges across epidemiologically similar pathogens. Abstract Introduction In the light of the COVID-19 pandemic many countries are trying to widen their pandemic planning from its traditional focus on influenza . However, it is impossible to draw up detailed plans for every pathogen with epidemic potential. We set out to try to simplify this process by reviewing the epidemiology of a range of pathogens with pandemic potential and seeing whether they fall into groups with shared epidemiological traits. Methods We reviewed the epidemiological characteristics of 19 different pathogens with pandemic potential (those on the WHO priority list of pathogens, different strains of influenza and Mpox). We extracted data on key pa...

  Abstract Identifying animal species that are susceptible to the plethora of existing and emerging viruses is critical for predicting and containing disease outbreaks . Current efforts to assess viral tropism largely rely on experimental infection models , but such experiments are logistically and ethically infeasible for many wildlife species. To tackle this challenge, we developed a panel of airway organoids from ten taxonomically diverse wildlife and livestock species and evaluated their susceptibility to influenza viruses of mammalian (pH1N1) and avian (H5N1) origin . Our analyses revealed large species-specific differences in infection rate and cytopathogenicity that aligned with known in vivo data and field observations. Furthermore, we demonstrated that this organoid panel can serve as a powerful tool to elucidate receptor-binding mechanisms, viral dynamics, and early host adaptation in poorly characterized animal species. In summary, this work provides a robust and ethical...

  Abstract The development of T-cell-based influenza vaccines relies on eliciting broad CD8+ T-cell immunity, wherein T stem cell-like memory (TSCM) cells serve as the ultimate long-lived reservoir for immune memory, thereby unlocking the potential for durable protection against viral drift and shift. However, the specific immunological cues that drive the robust expansion and functional preservation of this self-renewing, multipotent subset remain unknown. Here, utilizing multi-omic systems immunology in a pediatric cohort immunized with live attenuated influenza vaccine , we identified the determinants governing the expansion of influenza virus-reactive TSCM cells . We show that a pre-existing state of innate antiviral readiness , defined by a plasmacytoid dendritic cell-associated type I interferon signature , is the requisite condition for a robust TSCM expansion. Mechanistically, this baseline innate state enhances antigen priming and enforces a qualitative divergence in T-cel...

  Abstract Influenza remains a significant global public health concern. Live-attenuated influenza vaccines (LAIVs) are recognized as effective interventions for influenza prevention. Currently, two types of LAIVs are licensed for human use: one developed through cold-adapted viral gene mutation and the other through the deletion of the viral NS1 gene . However, the similarities and differences in these two LAIVs’ efficacy, safety, and immune responses have not been thoroughly studied. This study constructed a gene-deficient live-attenuated vaccine strain, CA4-dNS1, and a gene locus-mutated attenuated vaccine strain, CA4-cold , to compare their in vivo and in vitro replication capacity , broad-spectrum protective efficacy , safety, and immunogenicity . The results showed that both LAIVs provide comparable broad-spectrum protection against lethal H1N1 and H5N1 influenza challenges in mice and induce similar humoral and mucosal immune responses . Notably, the CA4-cold vaccine strain ...

  Abstract Maintaining tissue function while eliminating infected cells is fundamental, and inflammatory damage plays a major contribution to lethality after lung infection . We tested 50 immunomodulatory regimes to determine their ability to protect mice from lethal infection . Only neutrophil depletion soon after infection prevented death from influenza. This result suggests that the infected host passed an early tipping point after which limiting innate damage alone could not rescue lung function. We investigated treatments that could have efficacy when administered later in infection. We found that partial limitation of viral spread together with enhancement of epithelial repair, by interferon blockade or limiting CD8+ T cell–mediated killing of epithelial cells , reduced lethality . This finding highlights the importance of rebalancing repair and damage processes in the survival of pulmonary infections. Source:  Link:  https://www.science.org/doi/10.1126/science.adr4...

  Abstract Influenza virus is transmitted via respiratory expulsions, but detection of infectious virus in such expulsions has been challenging . Here, we describe quantification and genotyping of infectious virus in respiratory particles using a Modular Influenza Sampling Tunnel (MIST). The particles deposit on cell monolayers , enabling culture, quantification, and sequencing of viruses. Concomitantly, water-sensitive paper and fine particle samplers yield respiratory particle counts over a broad size range. Using the MIST, we captured infectious virus from humans experimentally infected with influenza virus on multiple days post-inoculation . The recovered respiratory particles varied in quantity over three orders of magnitude and contained viral genetic variation that was also detected in samples from infected individuals. Expulsion of infectious virus was associated with infectious viral load in saliva and nasopharyngeal swabs and with clinical symptoms . These data reveal the...