ETIDIOH - NEW

  Abstract Dipeptidyl peptidase-4 (DPP4) is an established receptor for Middle East respiratory syndrome-related coronaviruses (MERSr-CoVs), while recent studies have identified angiotensin-converting enzyme 2 (ACE2) usage in multiple merbecovirus clades . Yet, receptor usage of many genetically diverse bat MERSr-CoVs remains unclear . Here we show that broadly distributed HKU25 clade merbecoviruses use ACE2 , rather than DPP4, as their receptor. Cryo-electron microscopy revealed that HsItaly2011 and VsCoV-a7 strains engage ACE2 similarly to HKU5 but with remodelled interfaces and distinct orthologue selectivity, suggesting a shared evolutionary origin of ACE2 recognition . EjCoV-3, a close relative of the DPP4-using BtCoV422, showed broad multi-species ACE2 tropism and preadaptation to human ACE2 . Several ACE2 glycans and residues within or near the binding interface were identified as determinants of orthologue selectivity. These viruses remain sensitive to several broadly neutr...

  ABSTRACT Subtype H3N2 influenza A viruses (IAVs), which emerged in 1968 to cause a pandemic, have shown continual circulation and adaptation that has necessitated frequent updates of candidate vaccine viruses . Here, we sought to determine how genetic changes in the hemagglutinin (HA) and neuraminidase of 21 antigenically distinct H3N2 IAVs isolated from 1968 to 2019 correlate with mammalian fitness and adaptation. We found a surge of adaptation between 1997 and 2002 , resulting in the emergence of A/Fujian/411/2002 (H3N2) and poor vaccine efficacy , leading to an epidemic during the 2003–2004 season. This surge was characterized by a large reduction in binding to mammalian-type α2,6-linked sialic acids and increased infectivity and replication kinetics in humanized Madin-Darby canine kidney cells. HA glycosylation also increased most rapidly from 1968 to 2004 and then plateaued. Symptomatic infections were only evident in mice following inoculation with viruses isolated in the 1...

  ABSTRACT Systemic spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to extrapulmonary tissues has been observed following acute infections. Autopsy studies further indicate tissue-specific virus diversity, including in immune-privileged sites . Questions remain on the viral dynamics leading to the tissue tropism of SARS-CoV-2, including evolutionary trajectories and functional adaptations that could impact persistence and transmission . In this study, we characterized SARS-CoV-2 genomes from 27 distinct tissues collected from an autopsy case where the patient had a primary immune deficiency . We identified tissue-specific virus genotypes , in some instances coexisting within the same sites, with mutations primarily in the receptor-binding domain of the spike protein. Protein simulations and isolation of infectious virus indicate combinations of spike substitutions that would lead to increased protein stability and stronger binding of the virus to host cells. ...

  Abstract Avian influenza viruses (AIVs) are a global public health risk; human infection is typically associated with high mortality . While the relationship between several mammalian adaptive mutations and host factors have been described, it is unknown whether additional uncharacterised mutations lead to adaptation. Here, we combine phylogenetic analysis and complementary experimental methods to quantify the impact of novel mutations that emerge at the avian-mammal interface . We constructed phylogenetic trees of mammalian and avian influenza sequences for the polymerase (PA, PB1, PB2) and nucleoprotein (NP) segments and identified potential avian to mammal spillover events. We found >6500 mutations across the polymerase and NP, including known signatures of mammalian adaptation such as PB2 E627K and D701N which occurred independently in mammals 143 and 56 times respectively. We selected 95 mutations which were mostly undescribed and emerged independently multiple times in a...

  Abstract The H7N9 influenza viruses, which are capable of causing severe respiratory syndrome in humans , were first discovered to infect humans in 2013 and continue to pose a persistent public health threat . Quail has been proposed as a potential intermediate host that may facilitate the emergence of novel reassorted influenza A viruses with the capacity to infect humans across species barriers; however, information on the biological characterization of quail H7N9 remains limited. In this study, we isolated and identified an avian H7N9 influenza virus from quails , designated as A/quail/Hebei/CH06-07/2018 (H7N9) and abbreviated as CH06-07, in Hebei, China . Phylogenetic analyses revealed that both the HA gene and the NA gene of CH06-07 were clustered in the Eurasian lineage . Furthermore, CH06-07 exhibited binding affinity for both α2,3-linked and α2,6-linked sialic acid receptors and demonstrated high pathogenicity in both quails and mice . Notably, transmission studies reveal...

  Abstract SARS-like betacoronaviruses (sarbecoviruses) endemic in bats pose a significant zoonotic threat to humans . Genetic pathways associated with spillover of bat sarbecoviruses into humans are incompletely understood. We previously showed that the wild-type spike of the rhinolophid bat coronavirus SHC014-CoV has poor entry activity and uncovered two distinct genetic pathways outside the receptor-binding domain (RBD) that increased spike opening, ACE2 binding , and cell entry. Herein, we show that the widely studied bat sarbecovirus WIV1-CoV is likely a cell culture-adapted variant whose progenitor bears a spike resembling that of Rs3367-CoV, which was sequenced from the same population of rhinolophid bats as SHC014-CoV. Our findings suggest that the acquisition of a single amino-acid substitution in the ‘630-loop’ of the S1 subunit was the key spike adaptation event during the successful isolation of WIV1-CoV, and that it enhances spike opening, virus-receptor recognition, a...

  Abstract Biodiversity loss is one of the greatest challenges facing Earth today. The most direct information on species losses comes from recent extinctions . However, our understanding of these recent, human-related extinctions is incomplete across life, especially their causes and their rates and patterns among clades, across habitats and over time. Furthermore, prominent studies have extrapolated from these extinctions to suggest a current mass extinction event . Such extrapolations assume that recent extinctions predict current extinction risk and are homogeneous among groups, over time and among environments. Here, we analyse rates and patterns of recent extinctions (last 500 years). Surprisingly, past extinctions did not strongly predict current risk among groups. Extinctions varied strongly among groups , and were most frequent among molluscs and some tetrapods , and relatively rare in plants and arthropods . Extinction rates have increased over the last five centuries , b...

  ABSTRACT SARS-CoV-2 is under strong evolutionary selection to acquire mutations in its spike protein that reduce neutralization by human polyclonal antibodies. Here, we use pseudovirus-based deep mutational scanning to measure how mutations to the spike from the recent KP.3.1.1 SARS-CoV-2 strain affect cell entry, binding to the ACE2 receptor, RBD up/down motion, and neutralization by human sera and clinically relevant antibodies. The spike mutations that most affect serum antibody neutralization sometimes differ between sera collected before versus after recent vaccination or infection, indicating that these exposures shift the neutralization immunodominance hierarchy. The sites where mutations cause the greatest reduction in neutralization by post-vaccination or infection sera include receptor-binding domain (RBD) sites 475, 478, and 487, all of which have mutated in recent SARS-CoV-2 variants . Multiple mutations outside the RBD affect sera neutralization as strongly as any RB...

  Abstract The antigenic relationship between Eurasian avian-like H1N1 swine influenza viruses (EA H1N1) and human pandemic 2009 H1N1 viruses (2009/H1N1) remains a critical question for influenza surveillance and vaccine efficacy . This study systematically investigated the antigenic differences between strains A/swine/Tianjin/312/2016 (TJ312, EA H1N1) and A/Guangdong-Maonan/SWL1536/2019 (GD1536, 2009/H1N1). Cross-hemagglutination inhibition (HI) assays revealed a significant antigenic disparity, with a 16-fold reduction in heterologous versus homologous HI titers . Comparative sequence analysis identified 22 amino acid differences across the five major antigenic sites (Sa, Sb, Ca1, Ca2, and Cb) of the HA1 subunit. Using reverse genetics , a panel of mutant viruses was generated. This study revealed that a single histidine (H)-to-asparagine (N) substitution at residue 128 (H3 numbering) in the Sa antigenic site acts as a primary determinant of antigenic variation , sufficient to ca...

  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 In 2024, a human infection with clade 2.3.4.4b highly pathogenic avian influenza A(H5N1) virus was identified in the United States in an individual with no known exposure . Genetic analysis revealed two hemagglutinin (HA) substitutions, P136S and A156T, which may alter viral antigenicity . Virus isolation was unsuccessful, preventing timely serologic analysis. To overcome this limitation, we generated recombinant viruses by reverse genetics and characterized the effects of the substitutions on antigenicity, receptor binding , and replicative fitness. The A156T substitution introduced a potential N-linked glycosylation site , resulting in altered antigenicity and reduced replication in primary human nasal epithelial cells and ferrets . Importantly, the A(H5N1) candidate vaccine virus (CVV) IDCDC-RG80A, which possesses HA-T156, remained antigenically effective against viruses with and without these substitutions. These findings highlight the importance of sequencing, rever...

  {Excerpt} Highly pathogenic avian influenza caused by H5N1 viruses emerged in East Asia in the late 1990s and spread to Europe, Africa, and the Middle East , circulating in wild and domestic birds and occasionally spilling over into mammals and humans . These viruses are classified into genetic lineages called clades based on differences in the gene encoding hemagglutinin , a key surface protein involved in cell entry. In 2021, one such lineage, clade 2.3.4.4b , crossed from Europe to Canada . It spread rapidly, reaching the southern tip of South America in less than 2 years and causing massive mortality in seabirds and marine mammals , including South American sea lions (Otaria flavescens) and southern elephant seals (Mirounga leonina) (1). However, human mortality has been markedly lower in the present H5N1 outbreak than in past ones. If the virus evolves the capacity to transmit from human to human, understanding this reduction will be essential for mounting an effective respo...

  ABSTRACT H10 subtype avian influenza viruses primarily circulate among wild waterfowl but can occasionally infect mammals , including humans , and recent sporadic human cases have raised significant public health concerns . In this study, we sequenced and analysed 59 H10 subtype viruses isolated from wild birds in Taiwan . Results showed that all isolates were genetically distinct from human and other mammalian H10 subtype isolates. Taiwanese isolates exhibited high genetic diversity and could be categorized into 34 distinct genotypes , with each genotype circulating only in a single migratory season and not recurring during subsequent seasons. Additional analyses revealed that certain gene pools frequently circulate in the Pacific Rim , with evidence of North American lineage genes establishing long-term populations in Eurasia and vice versa. Although no characteristics indicative of mammalian adaptation was found in the Taiwanese isolates, temporal changes in the haemagglutinin...

  Abstract Human influenza viruses rapidly acquire mutations in their hemagglutinin (HA) protein that erode neutralization by antibodies from prior exposures. Here, we use a sequencing-based assay to measure neutralization titers for 78 recent H3N2 HA strains against a large set of children and adult sera , measuring ~10,000 total titers . There is substantial person-to-person heterogeneity in the titers against different viral strains, both within and across age cohorts. The growth rates of H3N2 strains in the human population in 2023 are highly correlated with the fraction of sera with low titers against each strain. Notably, strain growth rates are less correlated with neutralization titers against pools of human sera, demonstrating the importance of population heterogeneity in shaping viral evolution . Overall, these results suggest that high-throughput neutralization measurements of human sera against many different viral strains can help explain the evolution of human influen...

  Abstract We detected H5N1 high pathogenicity avian influenza in captive Greater Rhea (Rhea americana). Viral genetic analysis revealed the mammalian associated PB2-E627K mutation , indicating selection of mammalian-relevant mutations in ratites . Pathologic investigation of available tissues demonstrated severe multifocal necrotising inflammation , and a strong vasculotropism. Competing Interest Statement The authors have declared no competing interest. Source: BioRxIV,  https://www.biorxiv.org/content/10.1101/2025.09.08.674895v1 ____

  Abstract The incursion of high pathogenicity avian influenza A virus (IAV) into US dairy cows is unprecedented in the era of molecular diagnosis and pathogen sequencing. This raises questions over the likelihood of further outbreaks and whether dairy cattle could be a mixing vessel for novel strains of IAV. Using a panel of BSL2-safe reassortant viruses representing clade 2.3.4.4b H5 epizootic lineages circulating since 2020, we found that a cow B3.13 isolate displayed enhanced replication in cow mammary gland cells , along with increased viral polymerase activity and stronger interferon antagonism in cow cells compared to an earlier EA-2020-C genotype virus. However, multiple avian and mammalian IAV strains , including other clade 2.3.4.4b high pathogenicity genotypes, were replication competent in bovine cells, particularly those of the mammary gland , suggesting that there is a diverse circulating IAV pool with the potential to infect cows. Moreover, we show that cow mammary c...

  Abstract Monkeypox virus (MPXV) experienced an unprecedented global outbreak in 2022, characterized by a significant departure from historical patterns: a rapid spread of the epidemic to more than 110 non-traditional endemic countries , with more than 90,000 confirmed cases; a fundamental shift in the mode of transmission , with human-to-human transmission (especially among men who have sex with men (MSM)) becoming the dominant route (95.2%); and genetic sequencing revealing a key adaptive mutation in a novel evolutionary branch ( Clade IIb ) that triggered the outbreak. These features highlight the significant evolution of MPXV in terms of host adaptation, transmission efficiency, and immune escape ability . The aim of this paper is to provide insights into the viral adaptive evolutionary mechanisms driving this global outbreak, with a particular focus on the role of immune escape (e.g., novel mechanisms of M2 proteins targeting the T cell co-stimulatory pathway) in enhancing vi...

  ABSTRACT Alterations in the PB2-627 domain of avian influenza virus (AIV) can potentially increase the risk of cross-host species infections in humans and mammals . Recently, there has been a rise in human cases of AIV infections without the presence of the known mammalian determinant PB2-E627K . Here, we identified a variant, PB2-627V , which has evolved in poultry and has contributed to the increase in human AIV infections . By screening global PB2 sequences , we discovered a new independent cluster of PB2-627V that emerged in the 2010s , prevalent in avian, mammalian, and human AIV isolates , including those of H9N2, H7N9, H3N8, 2.3.4.4b H5N1 , and other subtypes. We functionally assessed its host adaptation , fitness , and transmissibility across three subtypes of AIVs (H9N2, H7N9, and H3N8) in different host models . PB2-627V combines the viral properties of avian-like PB2-627E and human-like PB2-627K , facilitating AIVs to efficiently infect and replicate in chickens and mi...

Abstract In early 2024, an unprecedented outbreak of H5N1 high pathogenicity avian influenza was detected in dairy cattle in the USA . As of mid-2025 the epidemic is ongoing, 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 mammalian species including humans and pigs . We find evidence of several mammalian adaptations gained early in the evolution of these viruses 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 ...

  Abstract SARS-CoV and MERS-CoV are two coronaviruses that have received significant attention due to their high pathogenicity and mortality rates in human populations . In this study, we compared their evolutionary dynamics to provide a One Health perspective on their differences in terms of the results of disease control. The phylogenetic network of SARS-CoVs showed that human isolates gathered into a “super-spreader” cluster and were distinct from civet isolates . In contrast, dromedary camel- and human-isolated MERS-CoVs were clustered together. Thus, most clades of MERS-CoV can infect humans , and MERS-CoVs seem to more easily spill over the animal-to-human interface. Additionally, the civet can be easily controlled , while the intermediate host (dromedary camels) of MERS-CoV is an important livestock species , so it is impossible to eliminate all animals. This further leads to difficulties in disease control in MERS. Although MERS-CoVs are endemic to dromedary camels in both...