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Archives of Virology Jan 2021Genome sequences of members of a potential fourth rhinovirus (RV) species, provisionally denoted as rhinovirus A clade D, from patients with acute respiratory infection...
Genome sequences of members of a potential fourth rhinovirus (RV) species, provisionally denoted as rhinovirus A clade D, from patients with acute respiratory infection were determined. Bayesian coalescent analysis estimated that clade D emerged around the 1940s and diverged further around 2006-2007 into two distinctive sublineages (RV-A8-like and RV-A45-like) that harbored unique "clade-defining" substitutions. Similarity plots and bootscan mapping revealed a recombination breakpoint located in the 5'-UTR region of members of the RV-A8-like sublineage. Phylogenetic reconstruction revealed the distribution of clade D viruses in the Asia Pacific region and in Europe, underlining its worldwide distribution.
Topics: Amino Acid Sequence; Asia; Bayes Theorem; Enterovirus Infections; Genome, Viral; Genotype; Humans; Phylogeny; Picornaviridae Infections; Respiratory Tract Infections; Rhinovirus
PubMed: 33084935
DOI: 10.1007/s00705-020-04855-5 -
Proceedings of the National Academy of... Nov 2020Human rhinoviruses (RVs) are positive-strand RNA viruses that cause respiratory tract disease in children and adults. Here we show that the innate immune signaling...
Human rhinoviruses (RVs) are positive-strand RNA viruses that cause respiratory tract disease in children and adults. Here we show that the innate immune signaling protein STING is required for efficient replication of members of two distinct RV species, RV-A and RV-C. The host factor activity of STING was identified in a genome-wide RNA interference (RNAi) screen and confirmed in primary human small airway epithelial cells. Replication of RV-A serotypes was strictly dependent on STING, whereas RV-B serotypes were notably less dependent. Subgenomic RV-A and RV-C RNA replicons failed to amplify in the absence of STING, revealing it to be required for a step in RNA replication. STING was expressed on phosphatidylinositol 4-phosphate (PI4P)-enriched membranes and was enriched in RV-A16 compared with RV-B14 replication organelles isolated in isopycnic gradients. The host factor activity of STING was species-specific, as murine STING (mSTING) did not rescue RV-A16 replication in STING-deficient cells. This species specificity mapped primarily to the cytoplasmic, ligand-binding domain of STING. Mouse-adaptive mutations in the RV-A16 2C protein allowed for robust replication in cells expressing mSTING, suggesting a role for 2C in recruiting STING to RV-A replication organelles. Palmitoylation of STING was not required for RV-A16 replication, nor was the C-terminal tail of STING that mediates IRF3 signaling. Despite co-opting STING to promote its replication, interferon signaling in response to STING agonists remained intact in RV-A16 infected cells. These data demonstrate a surprising requirement for a key host mediator of innate immunity to DNA viruses in the life cycle of a small pathogenic RNA virus.
Topics: Carrier Proteins; Common Cold; Enterovirus; HeLa Cells; Host-Pathogen Interactions; Humans; Immunity, Innate; Interferon Regulatory Factor-3; Lipoylation; Membrane Proteins; Mutation; Protein Domains; Signal Transduction; Species Specificity; Viral Nonstructural Proteins; Virus Replication
PubMed: 33060297
DOI: 10.1073/pnas.2014940117 -
Indian Journal of Medical Microbiology 2019Human rhinovirus (HRV) and Enterovirus (ENV) are the major causes of childhood acute respiratory tract infections (ARTIs). This study sought to understand the...
INTRODUCTION
Human rhinovirus (HRV) and Enterovirus (ENV) are the major causes of childhood acute respiratory tract infections (ARTIs). This study sought to understand the distribution pattern of HRV subgroups, their seasonality and association with respiratory complications in patients at a tertiary care hospital.
RESULTS
Of the total 332 ARTI samples, 82 (24.7%) were positive for ENV/HRV. Twenty positive samples were processed further for phylogenetic analysis. Ten of the 20 samples were identified to be HRVs (70% HRV A and 30% HRV C) and nine were enteroviruses. HRV A clustered near three distinct HRV types (A12, A78 and A82). Four of the HRV strains (represented as SEQ 137 rhino, SEQ 282 rhino, SEQ 120 rhino and SEQ 82 rhino) had high sequence similarity. HRV C showed seasonality and was associated with disease severity.
CONCLUSION
The genotyping and phylogenetic analysis of the HRVs in the current study shows its circulatory pattern, association with risk factors and evolutionary dynamics.
Topics: Adolescent; Enterovirus; Enterovirus Infections; Female; Humans; India; Male; Nasopharynx; Phylogeny; Prospective Studies; RNA, Viral; Respiratory Tract Infections
PubMed: 32436882
DOI: 10.4103/ijmm.IJMM_20_23 -
Clinical Chemistry Jul 2020More than 2 months separated the initial description of SARS-CoV-2 and discovery of its widespread dissemination in the United States. Despite this lengthy interval,...
BACKGROUND
More than 2 months separated the initial description of SARS-CoV-2 and discovery of its widespread dissemination in the United States. Despite this lengthy interval, implementation of specific quantitative reverse transcription (qRT)-PCR-based SARS-CoV-2 tests in the US has been slow, and testing is still not widely available. Metagenomic sequencing offers the promise of unbiased detection of emerging pathogens, without requiring prior knowledge of the identity of the responsible agent or its genomic sequence.
METHODS
To evaluate metagenomic approaches in the context of the current SARS-CoV-2 epidemic, laboratory-confirmed positive and negative samples from Seattle, WA were evaluated by metagenomic sequencing, with comparison to a 2019 reference genomic database created before the emergence of SARS-CoV-2.
RESULTS
Within 36 h our results showed clear identification of a novel human Betacoronavirus, closely related to known Betacoronaviruses of bats, in laboratory-proven cases of SARS-CoV-2. A subset of samples also showed superinfection or colonization with human parainfluenza virus 3 or Moraxella species, highlighting the need to test directly for SARS-CoV-2 as opposed to ruling out an infection using a viral respiratory panel. Samples negative for SARS-CoV-2 by RT-PCR were also negative by metagenomic analysis, and positive for Rhinovirus A and C. Unlike targeted SARS-CoV-2 qRT-PCR testing, metagenomic analysis of these SARS-CoV-2 negative samples identified candidate etiological agents for the patients' respiratory symptoms.
CONCLUSION
Taken together, these results demonstrate the value of metagenomic analysis in the monitoring and response to this and future viral pandemics.
Topics: Betacoronavirus; COVID-19; Coronavirus Infections; Enterovirus; Humans; Metagenomics; Nasopharynx; Pandemics; Phylogeny; Pneumonia, Viral; RNA, Viral; Real-Time Polymerase Chain Reaction; SARS-CoV-2; Sequence Analysis, RNA; Superinfection
PubMed: 32379863
DOI: 10.1093/clinchem/hvaa106 -
American Journal of Respiratory and... Jul 2020
Topics: Asthma; Child; Enterovirus; Humans; Lymphocytes; Picornaviridae Infections; Rhinovirus
PubMed: 32240597
DOI: 10.1164/rccm.202003-0634ED -
American Journal of Respiratory and... Jul 2020Individuals with asthma have heightened antibody responses to rhinoviruses (RVs), although those specific for RV-C are lower than responses specific for RV-A,... (Comparative Study)
Comparative Study
Individuals with asthma have heightened antibody responses to rhinoviruses (RVs), although those specific for RV-C are lower than responses specific for RV-A, suggesting poor immunity to this species. To ascertain and compare T-cell memory responses induced by RV-A and RV-C in children with and without asthma. Peripheral blood mononuclear cells from 17 children with asthma and 19 control subjects without asthma were stimulated with peptide formulations to induce representative species-specific responses to RV-A and RV-C. Molecular profiling (RNA sequencing) was used to identify enriched pathways and upstream regulators. Responses to RV-A showed higher expression of IFNG and STAT1 compared with RV-C, and significant expression of CXCL9, 10, and 11 was not found for RV-C. There was no reciprocal increase of T-helper cell type 2 (Th2) cytokine genes or the Th2 chemokine genes CCL11, CCL17, and CCL22. RV-C induced higher expression of CCL24 (eotaxin-2) than RV-A in the responses of children with and without asthma. Upstream regulator analysis showed both RV-A and, although to a lesser extent, RV-C induced predominant Th1 and inflammatory cytokine expression. The responses of children with asthma compared with those without asthma were lower for both RV-A and RV-C while retaining the pattern of gene expression and upstream regulators characteristic of each species. All groups showed activation of the IL-17A pathway. RV-C induced memory cells with a lower IFN-γ-type response than RV-A without T-helper cell type 2 (Th2) upregulation. Children with asthma had lower recall responses than those without asthma while largely retaining the same gene activation profile for each species. RV-A and RV-C, therefore, induce qualitatively different T-cell responses.
Topics: Adolescent; Asthma; Cells, Cultured; Child; Child, Preschool; Enterovirus; Female; Gene Expression Regulation, Viral; Healthy Volunteers; Humans; Lymphocytes; Male; Picornaviridae Infections; Th2 Cells
PubMed: 32142615
DOI: 10.1164/rccm.201908-1670OC -
Clinical and Translational Allergy 2019Rhinovirus A and C infections are important contributors to asthma induction and exacerbations. No data exist on the interaction of local immune responses in rhinovirus...
BACKGROUND
Rhinovirus A and C infections are important contributors to asthma induction and exacerbations. No data exist on the interaction of local immune responses in rhinovirus infection. Therefore, we aimed to determine the tonsillar immune responses according to rhinovirus A, B and C infections.
METHODS
We collected tonsillar samples, nasopharyngeal aspirates and peripheral blood from 42 rhinovirus positive tonsillectomy patients. Fifteen respiratory viruses or their types were investigated from nasopharynx and tonsil tissue, and rhinovirus species were typed. The expression of 10 cytokines and 4 transcription factors (IFN-α, IFN-β, IFN-γ, IL-10, IL-13, IL-17, IL-28, IL-29, IL-37, TGF-β, FOXP3, GATA3, RORC2 and Tbet) were studied from tonsil tissue by quantitative PCR. A standard questionnaire of respiratory symptoms and health was filled by the patient or his/her guardian. The patients were divided into three groups by the determination of rhinovirus species.
RESULTS
Overall, 16 patients had rhinovirus A, 12 rhinovirus B and 14 rhinovirus C infection. In rhinovirus B positive group there were significantly less men ( = 0.0072), less operated in spring ( = 0.0096) and more operated in fall ( = 0.030) than in rhinovirus A or C groups. Rhinovirus A positive patients had more respiratory symptoms ( = 0.0074) and particularly rhinitis ( = 0.036) on the operation day. There were no significant differences between the groups in virus codetection. In adjusted analysis, rhinovirus C infections were associated with increased IFN-α ( = 0.045) and decreased RORC2 expression ( = 0.025).
CONCLUSIONS
Rhinovirus species associated differently with clinical characteristics and tonsillar cytokine responses.
PubMed: 31827765
DOI: 10.1186/s13601-019-0302-7 -
Science Translational Medicine Oct 2019Human enterovirus A71 (HEVA71) causes hand, foot, and mouth disease (HFMD) in young children and is considered a major neurotropic pathogen but lacks effective...
Human enterovirus A71 (HEVA71) causes hand, foot, and mouth disease (HFMD) in young children and is considered a major neurotropic pathogen but lacks effective antivirals. To identify potential therapeutic agents against HFMD, we screened a 502-compound flavonoid library for compounds targeting the HEVA71 internal ribosome entry site (IRES) that facilitates translation of the HEVA71 genome and is vital for the production of HEVA71 viral particles. We validated hits using cell viability and viral plaque assays and found that prunin was the most potent inhibitor of HEVA71. Downstream assays affirmed that prunin disrupted viral protein and RNA synthesis and acted as a narrow-spectrum antiviral against enteroviruses A and B, but not enterovirus C, rhinovirus A, herpes simplex 1, or chikungunya virus. Continuous HEVA71 passaging with prunin yielded HEVA71-resistant mutants with five mutations that mapped to the viral IRES. Knockdown studies showed that the mutations allowed HEVA71 to overcome treatment-induced suppression by differentially regulating recruitment of the IRES trans-acting factors Sam68 and hnRNPK without affecting the hnRNPA1-IRES interaction required for IRES translation. Furthermore, prunin effectively reduced HEVA71-associated clinical symptoms and mortality in HEVA71-infected BALB/c mice and suppressed hepatitis C virus at higher concentrations, suggesting a similar mechanism of prunin-mediated IRES inhibition for both viruses. These studies establish prunin as a candidate for further development as a HEVA71 therapeutic agent.
Topics: Animals; Anti-Bacterial Agents; Cell Death; DNA-Binding Proteins; Drug Evaluation, Preclinical; Drug Resistance, Viral; Enterovirus A, Human; Enterovirus Infections; Flavonoids; Genes, Reporter; Hepacivirus; Heterogeneous Nuclear Ribonucleoprotein A1; Humans; Internal Ribosome Entry Sites; Luciferases; Mice, Inbred BALB C; Mutation; Phlorhizin; Reproducibility of Results; Virus Replication
PubMed: 31666401
DOI: 10.1126/scitranslmed.aar5759 -
American Journal of Respiratory Cell... Mar 2020Rhinovirus (RV) exposure evokes exacerbations of asthma that markedly impact morbidity and mortality worldwide. The mechanisms by which RV induces airway...
Rhinovirus (RV) exposure evokes exacerbations of asthma that markedly impact morbidity and mortality worldwide. The mechanisms by which RV induces airway hyperresponsiveness (AHR) or by which specific RV serotypes differentially evoke AHR remain unknown. We posit that RV infection evokes AHR and inflammatory mediator release, which correlate with degrees of RV infection. Furthermore, we posit that rhinovirus C-induced AHR requires paracrine or autocrine mediator release from epithelium that modulates agonist-induced calcium mobilization in human airway smooth muscle. In these studies, we used an model to measure bronchoconstriction and mediator release from infected airways in human precision cut lung slices to understand how RV exposure alters airway constriction. We found that rhinovirus C15 (RV-C15) infection augmented carbachol-induced airway narrowing and significantly increased release of IP-10 (IFN-γ-induced protein 10) and MIP-1β (macrophage inflammatory protein-1β) but not IL-6. RV-C15 infection of human airway epithelial cells augmented agonist-induced intracellular calcium flux and phosphorylation of myosin light chain in co-cultured human airway smooth muscle to carbachol, but not after histamine stimulation. Our data suggest that RV-C15-induced structural cell inflammatory responses are associated with viral load but that inflammatory responses and alterations in agonist-mediated constriction of human small airways are uncoupled from viral load of the tissue.
Topics: Asthma; Calcium Signaling; Carbachol; Cells, Cultured; Chemokine CXCL10; Enterovirus; Enterovirus Infections; Histamine; Humans; Inflammation Mediators; Muscle Contraction; Muscle, Smooth; Myocytes, Smooth Muscle; Myosin Light Chains; Phosphorylation; Protein Processing, Post-Translational; RNA, Viral; Respiratory Hypersensitivity; Viral Load
PubMed: 31533004
DOI: 10.1165/rcmb.2019-0004OC -
PloS One 2019Antibodies are essential to functional immunity, yet the epitopes targeted by antibody repertoires remain largely uncharacterized. To aid in characterization, we...
Antibodies are essential to functional immunity, yet the epitopes targeted by antibody repertoires remain largely uncharacterized. To aid in characterization, we developed a generalizable strategy to predict antibody-binding epitopes within individual proteins and entire proteomes. Specifically, we selected antibody-binding peptides for 273 distinct sera out of a random library and identified the peptides using next-generation sequencing. To predict antibody-binding epitopes and the antigens from which these epitopes were derived, we tiled the sequences of candidate antigens into short overlapping subsequences of length k (k-mers). We used the enrichment over background of these k-mers in the antibody-binding peptide dataset to predict antibody-binding epitopes. As a positive control, we used this approach, termed K-mer Tiling of Protein Epitopes (K-TOPE), to predict epitopes targeted by monoclonal and polyclonal antibodies of well-characterized specificity, accurately recovering their known epitopes. K-TOPE characterized a commonly targeted antigen from Rhinovirus A, predicting four epitopes recognized by antibodies present in 87% of sera (n = 250). An analysis of 2,908 proteins from 400 viral taxa that infect humans predicted seven enterovirus epitopes and five Epstein-Barr virus epitopes recognized by >30% of specimens. Analysis of Staphylococcus and Streptococcus proteomes similarly predicted 22 epitopes recognized by >30% of specimens. Twelve of these common viral and bacterial epitopes agreed with previously mapped epitopes with p-values < 0.05. Additionally, we predicted 30 HSV2-specific epitopes that were 100% specific against HSV1 in novel and previously reported antigens. Experimentally validating these candidate epitopes could help identify diagnostic biomarkers, vaccine components, and therapeutic targets. The K-TOPE approach thus provides a powerful new tool to elucidate the organisms, antigens, and epitopes targeted by human antibody repertoires.
Topics: Adolescent; Adult; Aged; Algorithms; Antibodies; Antigens, Bacterial; Antigens, Viral; Child; Enterovirus; Epitopes; Humans; Middle Aged; Proteome; Proteomics; Sequence Analysis, Protein; Staphylococcus; Streptococcus
PubMed: 31490930
DOI: 10.1371/journal.pone.0217668