UNC-Chapel Hill Coronavirus Research // Carolina Digital Repository

Works (1049)

961. A Mouse-Adapted SARS-Coronavirus Causes Disease and Mortality in BALB/c Mice

Title Tesim:: A Mouse-Adapted SARS-Coronavirus Causes Disease and Mortality in BALB/c Mice
Creator:: Yount, Boyd, Heise, Mark, Sheahan, Tim, Baric, Ralph, Deming, Damon, Roberts, Anjeanette, Subbararo, Kanta, Cheng, Aaron, Genrich, Gillian L., Zaki, Sherif R., Herman, Brian D., Paddock, Christopher, and Vogel, Leatrice
Date of publication:: 2007
Abstract Tesim:: No single animal model for severe acute respiratory syndrome (SARS) reproduces all aspects of the human disease. Young inbred mice support SARS-coronavirus (SARS-CoV) replication in the respiratory tract and are available in sufficient numbers for statistical evaluation. They are relatively inexpensive and easily accessible, but their use in SARS research is limited because they do not develop illness following infection. Older (12- to 14-mo-old) BALB/c mice develop clinical illness and pneumonitis, but they can be hard to procure, and immune senescence complicates pathogenesis studies. We adapted the SARS-CoV (Urbani strain) by serial passage in the respiratory tract of young BALB/c mice. Fifteen passages resulted in a virus (MA15) that is lethal for mice following intranasal inoculation. Lethality is preceded by rapid and high titer viral replication in lungs, viremia, and dissemination of virus to extrapulmonary sites accompanied by lymphopenia, neutrophilia, and pathological changes in the lungs. Abundant viral antigen is extensively distributed in bronchial epithelial cells and alveolar pneumocytes, and necrotic cellular debris is present in airways and alveoli, with only mild and focal pneumonitis. These observations suggest that mice infected with MA15 die from an overwhelming viral infection with extensive, virally mediated destruction of pneumocytes and ciliated epithelial cells. The MA15 virus has six coding mutations associated with adaptation and increased virulence; when introduced into a recombinant SARS-CoV, these mutations result in a highly virulent and lethal virus (rMA15), duplicating the phenotype of the biologically derived MA15 virus. Intranasal inoculation with MA15 reproduces many aspects of disease seen in severe human cases of SARS. The availability of the MA15 virus will enhance the use of the mouse model for SARS because infection with MA15 causes morbidity, mortality, and pulmonary pathology. This virus will be of value as a stringent challenge in evaluation of the efficacy of vaccines and antivirals.
Resource type:: Article
Admin Note:: deposited as part of 1Science uploads
Affiliation Label Tesim:: Department of Microbiology and Immunology
Type:: http://purl.org/dc/dcmitype/Text
DOI:: https://doi.org/10.17615/5ehf-dw39
Identifier:: https://dx.doi.org/10.1371/journal.ppat.0030005
ISSN:: 1553-7366
Journal Issue:: 1
Journal Title:: PLoS Pathogens
Journal Volume:: 3
Keyword:: SARS Virus, RNA Virus Infections, Humans, Disease Models, Animal, Mice, Inbred BALB C, Molecular Sequence Data, Animals, Mice, and Severe Acute Respiratory Syndrome
Language Label:: English
ORCID:
Other Affiliation:: , National Institutes of Health, and Centers for Disease Control and Prevention
Page Start:: e5
Person:: Yount, Boyd, Heise, Mark, Sheahan, Tim, Baric, Ralph, Deming, Damon, Roberts, Anjeanette, Subbararo, Kanta, Cheng, Aaron, Genrich, Gillian L., Zaki, Sherif R., Herman, Brian D., Paddock, Christopher, and Vogel, Leatrice
Rights Statement Label:: In Copyright
Source:: 3n204358b

962. Analysis of Intraviral Protein-Protein Interactions of the SARS Coronavirus ORFeome

Title Tesim:: Analysis of Intraviral Protein-Protein Interactions of the SARS Coronavirus ORFeome
Creator:: Teepe, Carola, Simpson, Jeremy, Friedel, Caroline C., Pepperkok, Rainer, Roberts, Rhonda, Haas, Juergen, von Brunn, Albrecht, Baric, Ralph, and Zimmer, Ralf
Date of publication:: 2007
Abstract Tesim:: The severe acute respiratory syndrome coronavirus (SARS-CoV) genome is predicted to encode 14 functional open reading frames, leading to the expression of up to 30 structural and non-structural protein products. The functions of a large number of viral ORFs are poorly understood or unknown. In order to gain more insight into functions and modes of action and interaction of the different proteins, we cloned the viral ORFeome and performed a genome-wide analysis for intraviral protein interactions and for intracellular localization. 900 pairwise interactions were tested by yeast-two-hybrid matrix analysis, and more than 65 positive non-redundant interactions, including six self interactions, were identified. About 38% of interactions were subsequently confirmed by CoIP in mammalian cells. Nsp2, nsp8 and ORF9b showed a wide range of interactions with other viral proteins. Nsp8 interacts with replicase proteins nsp2, nsp5, nsp6, nsp7, nsp8, nsp9, nsp12, nsp13 and nsp14, indicating a crucial role as a major player within the replication complex machinery. It was shown by others that nsp8 is essential for viral replication in vitro, whereas nsp2 is not. We show that also accessory protein ORF9b does not play a pivotal role for viral replication, as it can be deleted from the virus displaying normal plaque sizes and growth characteristics in Vero cells. However, it can be expected to be important for the virus-host interplay and for pathogenicity, due to its large number of interactions, by enhancing the global stability of the SARS proteome network, or play some unrealized role in regulating protein-protein interactions. The interactions identified provide valuable material for future studies.
Resource type:: Article
Admin Note:: deposited as part of the 1Science upload
Affiliation Label Tesim:: University of North Carolina at Chapel Hill
Type:: http://purl.org/dc/dcmitype/Text
DOI:: https://doi.org/10.17615/bzbk-w739
Identifier:: https://dx.doi.org/10.1371/journal.pone.0000459
ISSN:: 1932-6203
Journal Issue:: 5
Journal Title:: PloS One
Journal Volume:: 2
Keyword:: Viral Proteins, Molecular Sequence Data, Fluorescent Antibody Technique, Genome, Amino Acid Sequence, Open Reading Frames, and SARS Virus
Language Label:: English
ORCID:
Other Affiliation:: Ludwig-Maximilians-Universitaet, European Molecular Biology Laboratory, and
Page Start:: e459
Person:: Teepe, Carola, Simpson, Jeremy, Friedel, Caroline C., Pepperkok, Rainer, Roberts, Rhonda, Haas, Juergen, von Brunn, Albrecht, Baric, Ralph, and Zimmer, Ralf
Rights Statement Label:: In Copyright
Source:: xk81jq99t

963. Evaluation of Serologic and Antigenic Relationships Between Middle Eastern Respiratory Syndrome Coronavirus and Other Coronaviruses to Develop Vaccine Platforms for the Rapid Response to Emerging Coronaviruses

Title Tesim:: Evaluation of Serologic and Antigenic Relationships Between Middle Eastern Respiratory Syndrome Coronavirus and Other Coronaviruses to Develop Vaccine Platforms for the Rapid Response to Emerging Coronaviruses
Creator:: Trevor D. Scobey, Rachel L. Graham, Sudhakar Agnihothram, Lisa E. Gralinski, Ralph S. Baric, Eric F. Donaldson, Vineet D. Menachery, Mark R. Denison, Maria Zambon, Boyd L. Yount Jr, and Robin Gopal
Date of publication:: 2013
Abstract Tesim:: Background. Middle East respiratory syndrome coronavirus (MERS-CoV) emerged in 2012, causing severe acute respiratory disease and pneumonia, with 44% mortality among 136 cases to date. Design of vaccines to limit the virus spread or diagnostic tests to track newly emerging strains requires knowledge of antigenic and serologic relationships between MERS-CoV and other CoVs.
Resource type:: Article
Type:: http://purl.org/dc/dcmitype/Text
DOI:: https://doi.org/10.17615/zdkg-xj11
ISSN:: 1537-6613 and 0022-1899
Journal Issue:: 7
Journal Title:: Journal of Infectious Diseases
Journal Volume:: 209
Keyword:: Antibodies, Viral, Middle Aged, MERS-CoV Vaccine Design, Humans, Synthetic Genomics, Chiroptera, Spike Glycoprotein, Coronavirus, Neutralization Tests, Animals, Serology, Mice, Inbred BALB C, Nucleocapsid Proteins, Antigens, Viral, Mice, Diagnostics , Coronaviridae, Blotting, Western, Enzyme-Linked Immunosorbent Assay, and Cross Reactions
Language Label:: English
ORCID:
Other Affiliation:: Department of Epidemiology and Department of Microbiology and Immunology, University of North Carolina, Chapel Hill, Departments of Pediatrics and Microbiology and Immunology, Vanderbilt University, Nashville, Tennessee, and Viral Zoonosis Unit, Public Health of England, London, United Kingdom
Page End:: 1006
Page Start:: 995
Person:: Trevor D. Scobey, Rachel L. Graham, Sudhakar Agnihothram, Lisa E. Gralinski, Ralph S. Baric, Eric F. Donaldson, Vineet D. Menachery, Mark R. Denison, Maria Zambon, Boyd L. Yount Jr, and Robin Gopal
Rights Statement Label:: In Copyright
Source:: 9p290f84v

964. Receptor usage and cell entry of bat coronavirus HKU4 provide insight into bat-to-human transmission of MERS coronavirus

Title Tesim:: Receptor usage and cell entry of bat coronavirus HKU4 provide insight into bat-to-human transmission of MERS coronavirus
Creator:: Lili Wang, Jian Tang, Fang Li, Lanying Du, Cuiqing Ma, Ralph S. Baric, Shibo Jiang, Yang Yang, and Chang Liu
Date of publication:: 2014
Abstract Tesim:: A constant and long-term threat to human health is cross-species transmission of Middle East respiratory syndrome coronavirus (MERS-CoV) from bats to humans. However, this process is poorly understood. Examining the cross-species transmissibility of bat coronavirus HKU4, which is genetically related to MERS-CoV, can provide critical information about the likely causes of MERS-CoV infections in humans. Here we investigate the receptor usage and cell entry mechanism of HKU4 compared with MERS-CoV. Our results reveal that MERS-CoV has adapted to use human receptor and cellular proteases for efficient human cell entry, whereas HKU4 can potentially follow-up and also infect human cells. These findings are critical for evaluating emerging disease potentials of bat coronaviruses and for preventing and controlling their spread in humans.
Resource type:: Article
Type:: http://purl.org/dc/dcmitype/Text
DOI:: https://doi.org/10.17615/g09d-nd11
ISSN:: 1091-6490, 0744-2831, and 0027-8424
Journal Issue:: 34
Journal Title:: Proceedings of the National Academy of Sciences of the United States of America
Journal Volume:: 111
Keyword:: Humans, Dipeptidyl Peptidase 4, Disease Reservoirs, Host Specificity, Chiroptera, Spike Glycoprotein, Coronavirus, Coronavirus, Host-Pathogen Interactions, Virulence, Middle East, Respiratory Tract Infections, Animals, Receptors, Virus, and Coronavirus Infections
Language Label:: English
ORCID:
Other Affiliation:: Lindsley F. Kimball Research Institute, New York Blood Center, New York, NY, Department of Pharmacology, University of Minnesota Medical School, Minneapolis, MN, Department of Epidemiology, University of North Carolina, Chapel Hill, NC, and Lindsley F. Kimball Research Institute, New York Blood Center, New York, NY ; Key Laboratory of Medical Molecular Virology of Ministries of Education and Health, Shanghai Medical College and Institute of Medical Microbiology, Fudan University, Shangha
Page End:: 12521
Page Start:: 12516
Person:: Lili Wang, Jian Tang, Fang Li, Lanying Du, Cuiqing Ma, Ralph S. Baric, Shibo Jiang, Yang Yang, and Chang Liu
Publisher:: National Academy of Sciences of the United States of America
Rights Statement Label:: In Copyright
Source:: mw22vb04c

965. Pathogenic Influenza Viruses and Coronaviruses Utilize Similar and Contrasting Approaches To Control Interferon-Stimulated Gene Responses

Title Tesim:: Pathogenic Influenza Viruses and Coronaviruses Utilize Similar and Contrasting Approaches To Control Interferon-Stimulated Gene Responses
Creator:: Katze, Michael G. , Proll, Sean , Smith, Richard D. , Webb-Robertson, Bobbie-Jo , Menachery, Vineet D. , Fan, Shufang , Baric, Ralph S. , Chang, Jean , Li, Chengjun , Green, Richard , Eisfeld, Amie J. , Josset, Laurence , Shukla, Anil K. , Kawaoka, Yoshihiro , Matzke, Melissa M. , Neumann, Gabriele , Waters, Katrina M. , Tilton, Susan C. , Gralinski, Lisa E. , Schäfer, Alexandra , Weiss, Jeffrey , Metz, Thomas O. , Sims, Amy C., Long, Casey , Williams, Christopher M. , and Schepmoes, Athena A.
Date of publication:: 2014
Abstract Tesim:: ABSTRACT The broad range and diversity of interferon-stimulated genes (ISGs) function to induce an antiviral state within the host, impeding viral pathogenesis. While successful respiratory viruses overcome individual ISG effectors, analysis of the global ISG response and subsequent viral antagonism has yet to be examined. Employing models of the human airway, transcriptomics and proteomics datasets were used to compare ISG response patterns following highly pathogenic H5N1 avian influenza (HPAI) A virus, 2009 pandemic H1N1, severe acute respiratory syndrome coronavirus (SARS-CoV), and Middle East respiratory syndrome CoV (MERS-CoV) infection. The results illustrated distinct approaches utilized by each virus to antagonize the global ISG response. In addition, the data revealed that highly virulent HPAI virus and MERS-CoV induce repressive histone modifications, which downregulate expression of ISG subsets. Notably, influenza A virus NS1 appears to play a central role in this histone-mediated downregulation in highly pathogenic influenza strains. Together, the work demonstrates the existence of unique and common viral strategies for controlling the global ISG response and provides a novel avenue for viral antagonism via altered histone modifications.IMPORTANCEThis work combines systems biology and experimental validation to identify and confirm strategies used by viruses to control the immune response. Using a novel screening approach, specific comparison between highly pathogenic influenza viruses and coronaviruses revealed similarities and differences in strategies to control the interferon and innate immune response. These findings were subsequently confirmed and explored, revealing both a common pathway of antagonism via type I interferon (IFN) delay as well as a novel avenue for control by altered histone modification. Together, the data highlight how comparative systems biology analysis can be combined with experimental validation to derive novel insights into viral pathogenesis.
Resource type:: Article
Affiliation Label Tesim:: Department of Epidemiology
Type:: http://purl.org/dc/dcmitype/Text
DOI:: https://doi.org/10.17615/nabc-b354
ISSN:: 2161-2129 and 2150-7511
Journal Issue:: 3
Journal Title:: MBio
Journal Volume:: 5
Keyword:: Middle East Respiratory Syndrome Coronavirus, Influenza, Human, Transcription Factors, Models, Biological, Host-Pathogen Interactions, Proteomics, Influenza A virus, Cluster Analysis, Gene Expression Profiling, Humans, Influenza A Virus, H5N1 Subtype, Gene Expression Regulation, Animals , SARS Virus, Histones, Coronavirus, Viral Nonstructural Proteins, Coronavirus Infections, Interferons, Protein Binding, Cell Line, and Interferon Type I
Language Label:: English
ORCID:
Other Affiliation:: University of Washington, Pacific Northwest National Laboratory, , and University of Wisconsin—Madison
Page End:: 11
Page Start:: 1
Person:: Katze, Michael G. , Proll, Sean , Smith, Richard D. , Webb-Robertson, Bobbie-Jo , Menachery, Vineet D. , Fan, Shufang , Baric, Ralph S. , Chang, Jean , Li, Chengjun , Green, Richard , Eisfeld, Amie J. , Josset, Laurence , Shukla, Anil K. , Kawaoka, Yoshihiro , Matzke, Melissa M. , Neumann, Gabriele , Waters, Katrina M. , Tilton, Susan C. , Gralinski, Lisa E. , Schäfer, Alexandra , Weiss, Jeffrey , Metz, Thomas O. , Sims, Amy C., Long, Casey , Williams, Christopher M. , and Schepmoes, Athena A.
Publisher:: American Society for Microbiology
Rights Statement Label:: In Copyright
Source:: q811kq365

966. A Mouse Model for Betacoronavirus Subgroup 2c Using a Bat Coronavirus Strain HKU5 Variant

Title Tesim:: A Mouse Model for Betacoronavirus Subgroup 2c Using a Bat Coronavirus Strain HKU5 Variant
Creator:: Rachel L. Graham, Mark Heise, Eric F. Donaldson, Maria Zambon, Heather L. Osswald, Ralph S. Baric, Robin Gopal, Trevor D. Scobey, Michelle M. Becker, Arun K. Ghosh, Andrew Mesecar, Jeremy Huynh, Lisa E. Gralinski, Alan Whitmore, Boyd L. Yount, Jr., Vineet D. Menachery, Sudhakar Agnihothram, Sakshi Tomar, and Mark R. Denison
Date of publication:: 2014
Abstract Tesim:: ABSTRACT Cross-species transmission of zoonotic coronaviruses (CoVs) can result in pandemic disease outbreaks. Middle East respiratory syndrome CoV (MERS-CoV), identified in 2012, has caused 182 cases to date, with ~43% mortality, and no small animal model has been reported. MERS-CoV and Pipistrellus bat coronavirus (BtCoV) strain HKU5 of Betacoronavirus (β-CoV) subgroup 2c share >65% identity at the amino acid level in several regions, including nonstructural protein 5 (nsp5) and the nucleocapsid (N) protein, which are significant drug and vaccine targets. BtCoV HKU5 has been described in silico but has not been shown to replicate in culture, thus hampering drug and vaccine studies against subgroup 2c β-CoVs. We report the synthetic reconstruction and testing of BtCoV HKU5 containing the severe acute respiratory syndrome (SARS)-CoV spike (S) glycoprotein ectodomain (BtCoV HKU5-SE). This virus replicates efficiently in cell culture and in young and aged mice, where the virus targets airway and alveolar epithelial cells. Unlike some subgroup 2b SARS-CoV vaccines that elicit a strong eosinophilia following challenge, we demonstrate that BtCoV HKU5 and MERS-CoV N-expressing Venezuelan equine encephalitis virus replicon particle (VRP) vaccines do not cause extensive eosinophilia following BtCoV HKU5-SE challenge. Passage of BtCoV HKU5-SE in young mice resulted in enhanced virulence, causing 20% weight loss, diffuse alveolar damage, and hyaline membrane formation in aged mice. Passaged virus was characterized by mutations in the nsp13, nsp14, open reading frame 5 (ORF5) and M genes. Finally, we identified an inhibitor active against the nsp5 proteases of subgroup 2c β-CoVs. Synthetic-genome platforms capable of reconstituting emerging zoonotic viral pathogens or their phylogenetic relatives provide new strategies for identifying broad-based therapeutics, evaluating vaccine outcomes, and studying viral pathogenesis.IMPORTANCEThe 2012 outbreak of MERS-CoV raises the specter of another global epidemic, similar to the 2003 SARS-CoV epidemic. MERS-CoV is related to BtCoV HKU5 in target regions that are essential for drug and vaccine testing. Because no small animal model exists to evaluate MERS-CoV pathogenesis or to test vaccines, we constructed a recombinant BtCoV HKU5 that expressed a region of the SARS-CoV spike (S) glycoprotein, thereby allowing the recombinant virus to grow in cell culture and in mice. We show that this recombinant virus targets airway epithelial cells and causes disease in aged mice. We use this platform to (i) identify a broad-spectrum antiviral that can potentially inhibit viruses closely related to MERS-CoV, (ii) demonstrate the absence of increased eosinophilic immune pathology for MERS-CoV N protein-based vaccines, and (iii) mouse adapt this virus to identify viral genetic determinants of cross-species transmission and virulence. This study holds significance as a strategy to control newly emerging viruses.
Resource type:: Article
Type:: http://purl.org/dc/dcmitype/Text
DOI:: https://doi.org/10.17615/dwgs-ep23
ISSN:: 2161-2129 and 2150-7511
Journal Issue:: 2
Journal Title:: MBio
Journal Volume:: 5
Keyword:: Eosinophilia, Coronavirus Infections, Animals, Mice, Disease Models, Animal, Drug Carriers, Vaccines, Synthetic, Genetic Vectors, Respiratory System, Mice, Inbred BALB C, Encephalitis Virus, Venezuelan Equine, Viral Vaccines, Chiroptera, and Coronavirus
Language Label:: English
ORCID:
Other Affiliation:: Departments of Epidemiology and Microbiology and Immunology, University of North Carolina, Chapel Hill, North Carolina, USA, Viral Zoonosis Unit, Public Health of England, London, United Kingdom, Departments of Chemistry and Medicinal Chemistry, Purdue University, West Lafayette, Indiana, USA, Departments of Pediatrics and Pathology and Microbiology and Immunology, Vanderbilt University, Nashville, Tennessee, USA, and Department of Chemistry, Purdue University, West Lafayette, Indiana, USA
Page End:: 12
Page Start:: 1
Person:: Rachel L. Graham, Mark Heise, Eric F. Donaldson, Maria Zambon, Heather L. Osswald, Ralph S. Baric, Robin Gopal, Trevor D. Scobey, Michelle M. Becker, Arun K. Ghosh, Andrew Mesecar, Jeremy Huynh, Lisa E. Gralinski, Alan Whitmore, Boyd L. Yount, Jr., Vineet D. Menachery, Sudhakar Agnihothram, Sakshi Tomar, and Mark R. Denison
Publisher:: American Society for Microbiology
Rights Statement Label:: In Copyright
Source:: zw12z965n

967. Mouse Dipeptidyl Peptidase 4 Is Not a Functional Receptor for Middle East Respiratory Syndrome Coronavirus Infection

Title Tesim:: Mouse Dipeptidyl Peptidase 4 Is Not a Functional Receptor for Middle East Respiratory Syndrome Coronavirus Infection
Creator:: Sudhakar S. Agnihothram, Mark T. Heise, Trevor Scobey, Adam S. Cockrell, Nicole R. Curnes, Ralph S. Baric, Boyd L. Yount, and Kayla M. Peck
Date of publication:: 2014
Abstract Tesim:: Human dipeptidyl peptidase 4 (hDPP4) was recently identified as the receptor for Middle East respiratory syndrome coronavirus (MERS-CoV) infection, suggesting that other mammalian DPP4 orthologs may also support infection. We demonstrate that mouse DPP4 cannot support MERS-CoV infection. However, employing mouse DPP4 as a scaffold, we identified two critical amino acids (A288L and T330R) that regulate species specificity in the mouse. This knowledge can support the rational design of a mouse-adapted MERS-CoV for rapid assessment of therapeutics.
Resource type:: Article
Type:: http://purl.org/dc/dcmitype/Text
DOI:: https://doi.org/10.17615/v26a-qn52
ISSN:: 1070-6321, 1098-5514, and 0022-538X
Journal Issue:: 9
Journal Title:: Journal of Virology
Journal Volume:: 88
Keyword:: Species Specificity, Molecular Sequence Data, DNA Mutational Analysis, Animals, Dipeptidyl Peptidase 4, Humans, Receptors, Virus, Virus Attachment, Mice, Amino Acid Sequence , Coronavirus Infections, Protein Conformation, Coronavirus, Host Specificity, and Models, Molecular
Language Label:: English
ORCID:
Other Affiliation:: Department of Epidemiology, University of North Carolina—Chapel Hill, Chapel Hill, North Carolina, USA, Departments of Genetics, Microbiology and Immunology, University of North Carolina—Chapel Hill, Chapel Hill, North Carolina, USA, Department of Epidemiology, University of North Carolina, Chapel Hill, North Carolina, USA, Department of Genetics, University of North Carolina—Chapel Hill, Chapel Hill, North Carolina, USA, Departments of Epidemiology, Microbiology and Immunology, University of North Carolina—Chapel Hill, Chapel Hill, North Carolina, USA, Department of Epidemiology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA ; Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA, and Department of Biology, University of North Carolina—Chapel Hill, Chapel Hill, North Carolina, USA
Page End:: 5199
Page Start:: 5195
Person:: Sudhakar S. Agnihothram, Mark T. Heise, Trevor Scobey, Adam S. Cockrell, Nicole R. Curnes, Ralph S. Baric, Boyd L. Yount, and Kayla M. Peck
Publisher:: American Society for Microbiology
Rights Statement Label:: In Copyright
Source:: 0g354k74z

968. Effects of Human Anti-Spike Protein Receptor Binding Domain Antibodies on Severe Acute Respiratory Syndrome Coronavirus Neutralization Escape and Fitness

Title Tesim:: Effects of Human Anti-Spike Protein Receptor Binding Domain Antibodies on Severe Acute Respiratory Syndrome Coronavirus Neutralization Escape and Fitness
Creator:: Robert C. Liddington, Quan Karen Zhu, Meagan Deming, Barry Rockx, Jianhua Sui, Wayne A. Marascoa, and Ralph S. Baric
Date of publication:: 2014
Abstract Tesim:: The receptor binding domain (RBD) of the spike (S) glycoprotein of severe acute respiratory syndrome coronavirus (SARS-CoV) is a major target of protective immunity in vivo. Although a large number of neutralizing antibodies (nAbs) have been developed, it remains unclear if a single RBD-targeting nAb or two in combination can prevent neutralization escape and, if not, attenuate viral virulence in vivo. In this study, we used a large panel of human nAbs against an epitope that overlaps the interface between the RBD and its receptor, angiotensin-converting enzyme 2 (ACE2), to assess their cross-neutralization activities against a panel of human and zoonotic SARS-CoVs and neutralization escape mutants. We also investigated the neutralization escape profiles of these nAbs and evaluated their effects on receptor binding and virus fitness in vitro and in mice. We found that some nAbs had great potency and breadth in neutralizing multiple viral strains, including neutralization escape viruses derived from other nAbs; however, no single nAb or combination of two blocked neutralization escape. Interestingly, in mice the neutralization escape mutant viruses showed either attenuation (Urbani background) or increased virulence (GD03 background) consistent with the different binding affinities between their RBDs and the mouse ACE2. We conclude that using either single nAbs or dual nAb combinations to target a SARS-CoV RBD epitope that shows plasticity may have limitations for preventing neutralization escape during in vivo immunotherapy. However, RBD-directed nAbs may be useful for providing broad neutralization and prevention of escape variants when combined with other nAbs that target a second conserved epitope with less plasticity and more structural constraint.
Resource type:: Article
Type:: http://purl.org/dc/dcmitype/Text
DOI:: https://doi.org/10.17615/3qjm-qq07
ISSN:: 1070-6321, 1098-5514, and 0022-538X
Journal Issue:: 23
Journal Title:: Journal of Virology
Journal Volume:: 88
Keyword:: Severe Acute Respiratory Syndrome, Virulence, In Vitro Techniques, Immunity, Virus Attachment, Carrier Proteins, Homo sapiens, Coronavirus Infections, Receptors, Angiotensin, Mortality, ACE2, Infection, Antibodies, Neutralizing, Ace2 , Immunotherapy, Glycoproteins, Epitopes, Enzymes, Mus musculus, and Antibodies
Language Label:: English
ORCID:
Other Affiliation:: Infectious and Inflammatory Disease Center, Stanford-Burnham Medical Research Institute, La Jolla, California, USA, Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute; Department of Medicine, Harvard Medical School, Boston Massachusetts, USA, and Departments of Epidemiology and Microbiology and Immunology, University of North Carolina, Chapel Hill, North Carolina, USA
Page End:: 13780
Page Start:: 13769
Person:: Robert C. Liddington, Quan Karen Zhu, Meagan Deming, Barry Rockx, Jianhua Sui, Wayne A. Marascoa, and Ralph S. Baric
Publisher:: American Society for Microbiology
Rights Statement Label:: In Copyright
Source:: pr76f789v

969. Coronavirus Replicase-Reporter Fusions Provide Quantitative Analysis of Replication and Replication Complex Formation

Title Tesim:: Coronavirus Replicase-Reporter Fusions Provide Quantitative Analysis of Replication and Replication Complex Formation
Creator:: Xiaotao Lu, Megan Culler Freeman, Christopher T. Peek, Rachel L. Graham, and Mark R. Denison
Date of publication:: 2014
Abstract Tesim:: The replication of coronaviruses occurs in association with multiple virus-induced membrane structures that evolve during the course of infection; however, the dynamics of this process remain poorly understood. Previous studies of coronavirus replication complex organization and protein interactions have utilized protein overexpression studies and immunofluorescence of fixed cells. Additionally, live-imaging studies of coronavirus replicase proteins have used fluorescent reporter molecules fused to replicase proteins, but expressed from nonnative locations, mostly late-transcribed subgenomic mRNAs, in the presence or absence of the native protein. Thus, the timing and targeting of native replicase proteins expressed in real time from native locations in the genome remain unknown. In this study, we tested whether reporter molecules could be expressed from the replicase polyprotein of murine hepatitis virus as fusions with nonstructural protein 2 or 3 and whether such reporters could define the targeting and activity of replicase proteins during infection. We demonstrate that the fusion of green fluorescent protein and firefly luciferase with either nonstructural protein 2 or 3 is tolerated and that these reporter-replicase fusions can be used to quantitate replication complex formation and virus replication. The results show that the replicase gene has flexibility to accommodate a foreign gene addition and can be used directly to study replicase complex formation and evolution during infection as well as to provide highly sensitive and specific markers for protein translation and genome replication.
Resource type:: Article
Type:: http://purl.org/dc/dcmitype/Text
DOI:: https://doi.org/10.17615/gwrw-ms81
ISSN:: 0022-538X, 1098-5514, and 1070-6321
Journal Issue:: 10
Journal Title:: Journal of Virology
Journal Volume:: 88
Keyword:: Proteins, Cells, Coronavirus Infections, Ambp, Mus musculus, RNA, Fluorescent Antibody Technique, Polyproteins, Membranes, Genome, Green Fluorescent Proteins, Genes, Hepatitis, Luciferases, Firefly, Severe Acute Respiratory Syndrome, Virus Replication, Infection, and Protein Biosynthesis
Language Label:: English
ORCID:
Other Affiliation:: Departments of Pediatrics and Elizabeth B. Lamb Center for Pediatric Research, Vanderbilt University Medical Center, Nashville, Tennessee, USA, Departments of Pathology, Microbiology, and Immunology, and Elizabeth B. Lamb Center for Pediatric Research, Vanderbilt University Medical Center, Nashville, Tennessee, USA, Department of Epidemiology, University of North Carolina, Chapel Hill, North Carolina, USA, and Departments of Pediatrics and Pathology, Microbiology, and Immunology, and Elizabeth B. Lamb Center for Pediatric Research, Vanderbilt University Medical Center, Nashville, Tennessee, USA
Page End:: 5327
Page Start:: 5319
Person:: Xiaotao Lu, Megan Culler Freeman, Christopher T. Peek, Rachel L. Graham, and Mark R. Denison
Publisher:: American Society for Microbiology
Rights Statement Label:: In Copyright
Source:: 0z7092114

970. Synthetic Reconstruction of Zoonotic and Early Human Severe Acute Respiratory Syndrome Coronavirus Isolates That Produce Fatal Disease in Aged Mice

Title Tesim:: Synthetic Reconstruction of Zoonotic and Early Human Severe Acute Respiratory Syndrome Coronavirus Isolates That Produce Fatal Disease in Aged Mice
Creator:: Cameron, Mark, Donaldson, Eric, Rockx, Barry, Pickles, Raymond, Harkema, Jack, Sims, Amy, Sheahan, Timothy, Kelvin, David, Heise, Mark, and Baric, Ralph
Date of publication:: 2007
Abstract Tesim:: The severe acute respiratory syndrome (SARS) epidemic was characterized by high mortality rates in the elderly. The molecular mechanisms that govern enhanced susceptibility of elderly populations are not known, and robust animal models are needed that recapitulate the increased pathogenic phenotype noted with increasing age. Using synthetic biology and reverse genetics, we describe the construction of a panel of isogenic SARS coronavirus (SARS-CoV) strains bearing variant spike glycoproteins that are representative of zoonotic strains found in palm civets and raccoon dogs, as well as isolates spanning the early, middle, and late phases of the SARS-CoV epidemic. The recombinant viruses replicated efficiently in cell culture and demonstrated variable sensitivities to neutralization with antibodies. The human but not the zoonotic variants replicated efficiently in human airway epithelial cultures, supporting earlier hypotheses that zoonotic isolates are less pathogenic in humans but can evolve into highly pathogenic strains. All viruses replicated efficiently, but none produced clinical disease or death in young animals. In contrast, severe clinical disease, diffuse alveolar damage, hyaline membrane formation, alveolitis, and death were noted in 12-month-old mice inoculated with the palm civet HC/SZ/61/03 strain or early-human-phase GZ02 variants but not with related middle- and late-phase epidemic or raccoon dog strains. This panel of SARS-CoV recombinants bearing zoonotic and human epidemic spike glycoproteins will provide heterologous challenge models for testing vaccine efficacy against zoonotic reintroductions as well as provide the appropriate model system for elucidating the complex virus-host interactions that contribute to more-severe and fatal SARS-CoV disease and acute respiratory distress in the elderly.
Resource type:: Article
Admin Note:: deposited as part of the 1Science upload
Affiliation Label Tesim:: Department of Microbiology and Immunology and Department of Epidemiology
Type:: http://purl.org/dc/dcmitype/Text
DOI:: https://doi.org/10.17615/1tjz-xj23
Identifier:: https://dx.doi.org/10.1128/JVI.00505-07
ISSN:: 0022-538X
Journal Issue:: 14
Journal Title:: Journal of Virology
Journal Volume:: 81
Keyword:: Human, Isolate, Rodentia, Acute, Animal, Mouse, Death, Coronavirus, Virology, Virus, Mammalia, and Vertebrata
Language Label:: English
ORCID:
Other Affiliation:: University of Toronto, , and Michigan State University
Page End:: 7423
Page Start:: 7410
Person:: Cameron, Mark, Donaldson, Eric, Rockx, Barry, Pickles, Raymond, Harkema, Jack, Sims, Amy, Sheahan, Timothy, Kelvin, David, Heise, Mark, and Baric, Ralph
Rights Statement Label:: In Copyright
Source:: x633f5302

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