ETIDIOH - NEW

  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...

  Abstract Currently, no immunomodulatory agents have been conclusively shown to benefit severe influenza . The World Health Organization conditionally advises against the use of systemic corticosteroids, macrolides, plasma therapy, mechanistic target of rapamycin inhibitors , and nonsteroidal anti-inflammatory drugs for such patients. High-dose systemic corticosteroids may increase mortality and morbidity in severe influenza ; the potential of low-dose corticosteroids merits further study given survival benefits in patients with severe coronavirus disease 2019 ( COVID-19 ). Passive immunotherapy using convalescent plasma or intravenous immunoglobulin (IVIG) from healthy donors has not proven effective , suggesting that future research should focus on hyperimmune plasma or IVIG from recent infections . An open-label randomized controlled trial (RCT) found that a triple combination of oseltamivir, clarithromycin, and naproxen improved outcomes in severe influenza. One RCT has indica...

  Significance Since pigs serve as intermediate hosts between humans and the natural reservoir of influenza viruses in wild birds, they play a key role in the emergence of influenza strains with pandemic potential , as demonstrated by the 2009 pandemic. Therefore, influenza pandemic preparedness will benefit from the development of vaccines that broadly protect pigs against diverse influenza A strains. However, progress is limited by our poor molecular understanding of porcine antibody responses to influenza virus. This study isolates and characterizes a panel of broadly neutralizing influenza antibodies from pigs . Our findings not only have significant implications for the development of broadly protective influenza vaccines for pigs, but also reveal the molecular differences in the antibody responses between pigs and humans. Abstract Antibody responses to the influenza virus hemagglutinin (HA) stem, a major target for broadly protective vaccine development , have been extensivel...

  Abstract Influenza A virus (IAV) infection during pregnancy is associated with stillbirth and preterm birth , possibly by disrupting placental and fetal immunity . To investigate this, pregnant pigtail macaques were inoculated with IAV [A/California/07/2009 (H1N1)] and examined at necropsy 5 days post-infection (N=11) versus uninfected controls (N=16). Stillbirth occurred in 18% of infected pregnancies but not in controls. While vertical transmission was not observed, low levels of viral RNA were detected in two placentas . Maternal IAV infection was associated with increased placental IL-1β and IFN-β levels and an upregulated type I interferon and integrated stress transcriptional response. Fetuses exposed to IAV had greater frequencies of innate immune cells in lymph nodes and CD4+ T cells in lungs . These results suggest that placental and fetal immune environments undergo immune activation independent of the severity of maternal lung infection. Influenza vaccination during pr...

  Abstract Increase in the occurrence of human H5N1 spillover infections resulting from dissemination of highly pathogenic avian influenza (HPAI) virus into bird and mammal populations raises concerns about HPAI adapting to become human transmissible . Studies identified hemagglutinin (HA) acid stability and receptor preference as essential traits that shape host tropism . Mutations that increase HA stability and affinity for α-2,6-linked sialic acids have been shown to confer airborne transmissibility in a ferret model , however mechanisms of activation of H5 subtype HA have not been probed and the effect of adaptive mutations on HA function has been largely inferred from static structures. Here, we use hydrogen/deuterium-exchange mass spectrometry to dissect activation dynamics for two ancestral HPAI H5 HA, their matched HA with adaptive mutations, and a contemporary H5 HA. By measuring dynamics, we identify variation in active site flexibility among the HA and demonstrate that a...

  Abstract Influenza virus cross-subtype antibodies targeting the hemagglutinin (HA) head are rare . Here, we found that a large proportion of monoclonal antibodies (mAbs) isolated from individuals immunized with the 2021-22 seasonal influenza vaccine bound to an epitope on the HA head of both the H1N1 vaccine strain and H3N2 strains from the mid-1990s. These H1/H3 cross-reactive antibodies were also found in polyclonal sera , but only in samples from individuals born in the 1990s . Ferrets sequentially exposed to an H3N2 virus from the 1990s and a contemporary seasonal influenza vaccine produced the same type of H1/H3 cross-reactive antibodies. We found evidence that H1N1 viruses are currently evolving within the human population to abrogate the binding of these antibodies . Together, our study demonstrates how prior influenza virus exposures can influence the specificity of antibodies elicited by entirely different influenza virus subtypes, and how viruses evolve to escape these ...

  Abstract The polymerase acidic (PA) protein is a subunit of the trimeric influenza A virus (IAV) RNAdependent 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 Influenza A viruses (IAV) remain a persistent One Health threat, and whole-genome sequencing from wastewater offers a promising surveillance tool . However, IAV is at low abundance in wastewater , making it difficult to sequence. We benchmarked four targeted enrichment methods suited for whole-genome sequencing including custom and off-the-shelf amplicon and probe-based methods. Our custom HA tiled-amplicon panel was sensitive, fast, and cost-effective, making it suitable for monitoring low-abundance seasonal variants of known subtypes . However, its reliance on conserved and intact primer-binding sites limited primer design to fewer subtypes. A previously published universal amplicon method targeted all IAV subtypes , but it performed poorly in wastewater due to its reliance on intact genome segments. Probe-capture methods were resilient to RNA degradation and mismatches , potentially enabling broader surveillance and detection of emerging strains. However, probes were ...

  Abstract Influenza A viruses remain a global health threat, yet no universal antibody therapy exists . Clinical programs have centered on neutralizing mAbs , only to be thwarted by strain specificity and rapid viral escape . We instead engineered three non-neutralizing IgG2a mAbs that target distinct, overlapping epitopes within the conserved N terminus of the M2 ectodomain (M2e). Combined at low dose, this “triple M2e-mAb” confers robust prophylactic and therapeutic protection in mice challenged with diverse human and zoonotic IAV strains, including highly pathogenic variants. Therapeutic efficacy depends on Fc-mediated effector activity via FcγRI, FcγRIII, and FcγRIV, rather than in vitro neutralization. Serial passaging in triple M2e-mAb–treated immunocompetent and immunodeficient hosts failed to generate viral escape mutants. Our findings redefine the influenza-specific antibody therapeutic design and support Fc-optimized, non-neutralizing M2e-mAbs as a broadly effective, mut...