ETIDIOH - NEW

  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 Comparably few lineages of influenza A virus (IAV) have evolved long-term sustained transmission in mammals . The reasons remain largely unknown , and the possibility of avian IAVs evolving sustained mammalian transmission is an ongoing concern . Here we measured the GC content and frequency of GC dinucleotides in 115,520 whole genomes of IAVs using bioinformatic analyses . We found that persistent mammalian lineages showed declining trends in GC-related content and could be reliably separated from IAVs circulating only in birds and those sporadically infecting mammals. Similarly, the earliest viruses of persistent mammalian lineages showed reduced GC-related content , suggesting that this trait might in part contribute to their eventual persistence . Recent highly pathogenic 2.3.4.4b H5 viruses that spread in mink, foxes and humans were also characterized by reduced GC-related content . While not sufficient, reduced GC-related content may be a necessary condition for su...

  Abstract Clade 2.2 H5N1 influenza viruses have caused an unusually high number of human infections , providing a unique opportunity to investigate early molecular steps associated with host adaptation . Although most work has focused on hemagglutinin (HA), the contribution of neuraminidase (NA) to these early adaptive events has remained unclear. By analyzing publicly available sequences from clade 2.2-infected patients , we identified 20 NA mutations and compared their phenotypes to 20 mutations acquired during diversification in primary human airway cells under drug-free conditions. Most patient-derived NA mutations resulted in modest reductions in sialidase activity , keeping activity within a functional range that supported improved replication in α2,6 sialylglycan (α2,6 Sia)-dominant environments , whereas excessive reduction impaired fitness . Notably, the phenotypes of culture-selected and patient-derived mutations were highly concordant , suggesting that these NA changes ...

  Abstract Highly pathogenic H5Ny influenza A viruses are causing unprecedented, season-independent outbreaks across avian and mammalian species , including dairy cattle , a novel reservoir. The sialoside-binding properties of influenza A hemagglutinin (HA) are strongly related to its ability to infect and transmit between hosts . Mucin-like O-glycans , omnipresent in respiratory tracts, have been understudied as viral receptors due to their complexity. To address this, we synthesized 25 O-linked glycans with diverse sialosides , including modifications by fucosides and sulfates . Our findings reveal that H5Ny 2.3.4.4b viruses bind core 3 sialyl-Lewisx and Sia-Gal-β3GalNAc, O-linked glycans not recognized by classical H5 or other avian viruses . By determining crystal structures, we resolved the structural features of four glycans in an H5 hemagglutinin (HA) from a 2016 2.3.4.4b virus . While these viruses do not bind human-type receptors , their broad receptor specificity enhances...

  Abstract Highly pathogenic avian influenza H5N1 viruses of clade 2.3.4.4b have caused widespread avian mortality and sporadic mammalian infections , raising concerns about their potential for efficient replication in the human population . Efficient replication in the human upper respiratory tract is considered a key barrier to transmission . Here, we demonstrate that an H5N1 virus isolated from bovine milk in Texas in 2024 (H5N1Tex/24) replicates as efficiently as the 2009 pandemic H1N1 virus (H1N1HH4/09) in well-differentiated human nasal epithelial cells . These cells express both avian- and human-type influenza receptors , indicating receptor adaptation is unnecessary for entry . H5N1Tex/24 replicates effectively at 33 degrees Celsius , reflecting nasal cavity temperature , whereas earlier avian H5N1 strains require 37 degrees Celsius, suggesting that H5N1Tex/24 has acquired another key adaptive feature to the human upper respiratory tract. H5N1Tex/24 remains sensitive to int...

  Abstract In 2024, an unprecedented outbreak of H5N1 high pathogenicity avian influenza was detected in dairy cattle in the USA resulting in spillbacks into poultry, wild birds and other mammals including humans . Here, we present molecular and virological evidence that the cattle B3.13 genotype H5N1 viruses rapidly accumulated adaptations in polymerase genes that enabled better replication in bovine cells and tissues , as well as cells of other mammals including humans . We find evidence of several mammalian adaptations in cattle including PB2 M631L , which is found in all cattle sequences, and PA K497R , which is found in the majority. Structurally, PB2 M631L maps to the polymerase-ANP32 interface , an essential host factor for viral genome replication. We show that this mutation adapts the polymerase to better interact with bovine ANP32 proteins , particularly ANP32A, and thereby enhances virus replication in bovine mammary systems and primary human airway cultures . We show th...

  Highlights •  Human influenza A virus N2 neuraminidases were analyzed by antigenic cartography •  N2 antigenic evolution was discordant with antigenic evolution of H3 hemagglutinin •  Important epitopes were identified at the NA active site but also elsewhere in NA •  Changes in charge, volume, and hydropathy of epitope residues caused antigenic drift Summary Human influenza A viruses escape antibody-mediated immunity through changes in the hemagglutinin (HA) and neuraminidase (NA) glycoproteins . HA antigenic evolution has been studied extensively, with more recent interest in NA due to its importance in influenza vaccine efficacy. Here, the antigenic properties of the NA of more than 300 A( H3N2 ) and A( H2N2 ) viruses isolated since 1957 were quantified with a NA inhibition enzyme-linked lectin assay and visualized using antigenic cartography , with follow-up molecular studies using recombinant viruses. The antigenic evolution of N2 NA was more gradual than...

  Abstract Influenza B viruses cause substantial respiratory disease and seasonal outbreaks. Despite decades of circulation in humans , only the B/Victoria lineage persisted after the COVID-19 pandemic. Continual evolution has generated hemagglutinin deletion variants at residues 162-164 that drive successive epidemics , yet their functional consequences remain poorly understood. Using integrated phylodynamics and reverse genetics , we show that Clade V1A.1 viruses carrying a two-amino acid deletion exhibit enhanced replication and increased virulence compared with ancestral viruses lacking deletions. The recently prevailing Clade V1A.3 , which harbors a three-amino acid deletion together with the K136E substitution, has completely displaced V1A.1 and causes more severe disease in mice . Both clades bound efficiently to alpha 2-3 and 2-6 sialylated glycans and exhibited broad tolerance to acidic pH and elevated temperatures . These findings reveal that specific combinations of HA d...

  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 Since its emergence in 2012, Middle East respiratory syndrome coronavirus (MERS-CoV) has posed a significant threat to human health . Recently, novel MERS-like coronaviruses with the potential for cross-species transmission have been identified. In this study, we focused on two newly isolated bat strains with putative health concern: BatCoV/Ii/GD/2014-422 (2014-422) and BtTp-BetaCoV/GX2012 (GX2012). We determined the cryo-EM structures of the spike glycoprotein trimer in the closed state for these two viruses. These structures display a more compact conformation compared to MERS-CoV spike . Biochemical characterization demonstrates that the spike receptor-binding domains (RBDs) of 2014-422 and GX2012 can bind to human dipeptidyl peptidase 4 (hDPP4). To investigate the structural determinants of pseudovirus infection, we solved the cryo-EM structures of 2014-422 RBD-hDPP4 and GX2012 RBD-hDPP4 complexes. The binding mode of the complex is conserved, but the angle of the RB...

  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 Enterovirus D68 (EV-D68), a neurotropic respiratory pathogen, poses a considerable clinical threat through its link to pediatric acute flaccid myelitis (AFM) and severe respiratory illness . The possibility of recurrent epidemics , evidenced since the 2014 outbreak, remains a major concern . Genomic determinants of virulence are central to this threat. Sequence variations that affect host–receptor interactions , immune evasion, and replication efficiency serve as critical modifiers of pathogenicity. This article systematically reviews the evidence for specific genomic sites that enhance EV-D68 virulence , focusing on three critical regions: the VP1 receptor-binding site , the 2Apro/TRAF3 cleavage site, and the 3Cpro immunoregulatory region . Mutations in the VP1 receptor-binding site can alter affinity for host receptors such as sialic acid, heparan sulfate, and MFSD6 , thereby shaping viral entry and tissue tropism . Alterations in the 2Apro/TRAF3 cleavage site may impair pro...

  Abstract The global spread of highly pathogenic avian influenza A(H5N1) clade 2.3.4.4b viruses has recently extended to include diverse mammalian species , raising new concerns about pandemic risk . In 2023, this clade was first detected in Russian marine mammals during a mass mortality event among northern fur seals in the Far East . Genetic analyses revealed the causative viruses to belong to genotype A3 of European origin , which is known to have circulated in wild birds across the Far East since 2022. Notably, these isolates harbor the mammalian-adaptive substitutions PB2-K482R and NP-N319K—mutations previously linked to enhanced virulence in non-H5 avian influenza viruses , but whose impact on A(H5N1) clade 2.3.4.4b viruses remained to be characterized. The heightened virulence of A3 genotype viruses is confirmed by data obtained via a mouse model . However, despite these adaptive changes, ferret transmission models showed no evidence of airborne transmission of the fur seal...

  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...

  Abstract In a new study, Zhao et al. (2025) obtain 959 whole genome sequences of H1N1 subtype swine influenza virus (SIV) isolated from China . Their analysis of the sequences , isolated between 1977 and 2020, reveals how H1N1 lineages have co-evolved and contributed to instances of zoonotic transmission within the region. This study’s findings characterize the long-term evolutionary effects of frequent viral reassortment in SIV and highlight its potential to drive future pandemics. Source:  Link:  https://academic.oup.com/evolut/advance-article/doi/10.1093/evolut/qpaf262/8400336 ____

  Abstract Highly pathogenic avian influenza H5N1 poses an increasing public health risk, particularly following its spillover into dairy cows and associated human infections in the U.S. since March 2024. Here, we systematically identified critical PB2 mutations emerged during avian-to-cattle transmission and subsequent adaptation in cattle , notably PB2 M631L, which conferred pathogenicity in mice comparable to the well-characterized PB2 E627K mutation . Retrospective analysis reveals that PB2 631L also circulated in avian and human H5N1 strains during the 2013–2014 outbreaks in Cambodia and Vietnam . Additional adaptive mutations include established markers ( E627K, Q591R, D701N ), and novel variants ( I647V, G685R, K736R ). These mutations enhance polymerase activity by improving the utilization of both bovine and human ANP32A proteins , thereby increasing viral fitness and pathogenicity in mammals . The convergence of these adaptations highlights the elevated zoonotic risk of c...

  Abstract Highly pathogenic avian influenza (HPAI) H5N8 virus, first identified in late 2016 in Egypt, continues to circulate and has replaced the previously dominant HPAI H5N1 virus of clade 2.2.1. In this study, HPAI H5N8 was detected on 23 commercial poultry farms in Egypt . Complete genome sequences of three isolates collected in 2021 were obtained using next-generation sequencing (NGS) and subjected to genetic characterization. Phylogenetic analysis showed these isolates to belong to clade 2.3.4.4b, comprising two genotypes: EA-2021-Q and EA-2020-A . Molecular analysis of the haemagglutinin (HA) protein revealed the presence of T156A and V538A substitutions in the duck isolate and an N183S substitution in the chicken isolate . Several additional nonsynonymous mutations were identified, including 147I and 504V in the PB2 protein , 127V, 672L, and 550L in the PA protein , 64F and 69P in the M2 protein , and 42S in the NS1 protein . Comparative analysis of HA antigenic sites bet...

  Summary Background Highly pathogenic avian influenza H5N1 viruses of the A/Goose/Guangdong/1/1996 lineage pose a global threat to wildlife, domestic animals, and humans . Cross-species transmission events to mammals, including humans , in the past 4 years highlight this threat. For influenza A viruses, crucial determinants of cross-species and intraspecies transmission to and among mammals include attachment to and replication in respiratory airway epithelial cells . Although these determinants have been studied for H5N1 viruses in the past, limited studies for clade 2.3.4.4b viruses exist. Therefore, the aim of this study was to determine the ability of recent clade 2.3.4.4b H5N1 viruses to attach to human respiratory tissues, to replicate in human airway epithelial cells and the associated immune response. Methods In this in-vitro study, we investigated three H5N1 clade 2.3.4.4b viruses (H5N1Gull2022, H5N1Polecat2022, and H5N1Bovine2024) in comparison with previously studied 2....

  Abstract The MERS-COV virus is a zoonotic coronavirus that emerged in 2012 in KSA and caused viral illness with a case fatality rate up to 35 %. Over a decade later, the virus is still evolving and circulating . The aim of this review is to discuss the current epidemiology of MERS-COV both in humans and animals, during and post the COVID-19 pandemic. We have found that MERS-COV is still evolving in camels with new lineages being detected in Saudi Arabia . Although the number of human cases has decreased , there is a gradual resurgence in the number of cases. Furthermore, many cases are being reported without exposure to camels and/or raw products, nor contact with known human cases . This necessitates global efforts in the surveillance of asymptomatic carriers in the community, role of unknown animal reservoirs in the virus spread if any, as well as extensive genomic surveillance of the virus. This is in order to unveil and assess the genetic changes that the virus is undergoing ...