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2.
Cell ; 2020.
Article | WHO COVID | ID: covidwho-756809

ABSTRACT

Summary Initially, children were thought to be spared from disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) However, a month into the epidemic, a novel multisystem inflammatory syndrome in children (MIS-C) emerged Herein, we report on the immune profiles of nine MIS-C cases All MIS-C patients had evidence of prior SARS-CoV-2 exposure, mounting an antibody response with intact neutralization capability Cytokine profiling identified elevated signatures of inflammation (IL-18 and IL-6), lymphocytic and myeloid chemotaxis and activation (CCL3, CCL4, and CDCP1) and mucosal immune dysregulation (IL-17A, CCL20, CCL28) Immunophenotyping of peripheral blood revealed reductions of non-classical monocytes, and subsets of NK- and T- lymphocytes, suggesting extravasation to affected tissues Finally, we profiled the auto-antigen reactivity of MIS-C plasma, which revealed both known disease-associated autoantibodies (anti-La) and novel candidates that recognize endothelial, gastrointestinal and immune-cell antigens All patients were treated with anti-IL6R antibody and/or IVIG, which led to rapid disease resolution

3.
Nat Commun ; 11(1): 4378, 2020 09 01.
Article in English | MEDLINE | ID: covidwho-740036

ABSTRACT

Children are strikingly underrepresented in COVID-19 case counts. In the United States, children represent 22% of the population but only 1.7% of confirmed SARS-CoV-2 cases as of April 2, 2020. One possibility is that symptom-based viral testing is less likely to identify infected children, since they often experience milder disease than adults. Here, to better assess the frequency of pediatric SARS-CoV-2 infection, we serologically screen 1,775 residual samples from Seattle Children's Hospital collected from 1,076 children seeking medical care during March and April of 2020. Only one child was seropositive in March, but seven were seropositive in April for a period seroprevalence of ≈1%. Most seropositive children (6/8) were not suspected of having had COVID-19. The sera of seropositive children have neutralizing activity, including one that neutralized at a dilution > 1:18,000. Therefore, an increasing number of children seeking medical care were infected by SARS-CoV-2 during the early Seattle outbreak despite few positive viral tests.

4.
J Infect Dis ; 2020 Aug 29.
Article in English | MEDLINE | ID: covidwho-733374

ABSTRACT

Over three million infections with the severe acute respiratory syndrome corona virus 2 (SARS-CoV-2) were confirmed globally by May, 2020. While PCR-based assays are used for diagnosis, high through-put, rapid serologic methods are needed. A Luminex binding assay was developed and used to assess simultaneously the presence of COVID-19-specific antibodies in human serum and plasma. Clear differentiation was achieved between specimens from infected and uninfected subjects, and a wide range of serum/plasma antibody levels were delineated in infected subjects. All 25 specimens from 18 COVID-19 patients were positive in the assays with both, the trimeric spike and the receptor-binding domain (RBD) proteins. None of the 13 specimens from uninfected subjects displayed antibodies to either antigen. There was a highly statistically significant difference between the antibody levels COVID-19-infected and -uninfected specimens (p<0.0001). This high through-put antibody assay is accurate, requires only 2.5 hours, and uses 5 nanograms of antigen per test.

6.
Immunity ; 2020 Jul 30.
Article in English | MEDLINE | ID: covidwho-710374

ABSTRACT

SARS-CoV-2 infection has emerged as a serious global pandemic. Because of the high transmissibility of the virus and the high rate of morbidity and mortality associated with COVID-19, developing effective and safe vaccines is a top research priority. Here, we provide a detailed evaluation of the immunogenicity of lipid nanoparticle-encapsulated, nucleoside-modified mRNA (mRNA-LNP) vaccines encoding the full-length SARS-CoV-2 spike protein or the spike receptor binding domain in mice. We demonstrate that a single dose of these vaccines induces strong type 1 CD4+ and CD8+ T cell responses, as well as long-lived plasma and memory B cell responses. Additionally, we detect robust and sustained neutralizing antibody responses and the antibodies elicited by nucleoside-modified mRNA vaccines do not show antibody-dependent enhancement of infection in vitro. Our findings suggest that the nucleoside-modified mRNA-LNP vaccine platform can induce robust immune responses and is a promising candidate to combat COVID-19.

7.
Non-conventional | WHO COVID | ID: covidwho-665205

ABSTRACT

Severe acute respiratory syndrome CoV-2 (SARS-CoV-2) is currently causing a worldwide pandemic with high morbidity and mortality Development of animal models that recapitulate important aspects of coronavirus disease 2019 (COVID-19) is critical for the evaluation of vaccines and antivirals, and understanding disease pathogenesis SARS-CoV-2 has been shown to use the same entry receptor as SARS-CoV-1, human angiotensin-converting enzyme 2 (hACE2)(1-3) Due to amino acid differences between murine and hACE2, inbred mouse strains fail to support high titer viral replication of SARS-CoV-2 virus Therefore, a number of transgenic and knock-in mouse models, as well as viral vector-mediated hACE2 delivery systems have been developed Here we compared the K18-hACE2 transgenic model to adenovirus-mediated delivery of hACE2 to the mouse lung We show that K18-hACE2 mice replicate virus to high titers in both the lung and brain leading to lethality In contrast, adenovirus-mediated delivery results in viral replication to lower titers limited to the lung, and no clinical signs of infection with a challenge dose of 10 (4) plaque forming units The K18-hACE2 model provides a stringent model for testing the ability of vaccines and antivirals to protect against disease, whereas the adenovirus delivery system has the flexibility to be used across multiple genetic backgrounds and modified mouse strains

8.
Non-conventional | WHO COVID | ID: covidwho-663795

ABSTRACT

Initially, the global outbreak of COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spared children from severe disease However, after the initial wave of infections, clusters of a novel hyperinflammatory disease have been reported in regions with ongoing SARS-CoV-2 epidemics While the characteristic clinical features are becoming clear, the pathophysiology remains unknown Herein, we report on the immune profiles of eight Multisystem Inflammatory Syndrome in Children (MIS-C) cases We document that all MIS-C patients had evidence of prior SARS-CoV-2 exposure, mounting an antibody response with normal isotype-switching and neutralization capability We further profiled the secreted immune response by high-dimensional cytokine assays, which identified elevated signatures of inflammation (IL-18 and IL-6), lymphocytic and myeloid chemotaxis and activation (CCL3, CCL4, and CDCP1) and mucosal immune dysregulation (IL-17A, CCL20, CCL28) Mass cytometry immunophenotyping of peripheral blood revealed reductions of mDC1 and non-classical monocytes, as well as both NK- and T- lymphocytes, suggesting extravasation to affected tissues Markers of activated myeloid function were also evident, including upregulation of ICAM1 and FcR1 in neutrophil and non-classical monocytes, well-documented markers in autoinflammation and autoimmunity that indicate enhanced antigen presentation and Fc-mediated responses Finally, to assess the role for autoimmunity secondary to infection, we profiled the auto-antigen reactivity of MIS-C plasma, which revealed both known disease-associated autoantibodies (anti-La) and novel candidates that recognize endothelial, gastrointestinal and immune-cell antigens All patients were treated with anti- IL6R antibody or IVIG, which led to rapid disease resolution tracking with normalization of inflammatory markers

9.
J Immunol ; 205(4): 915-922, 2020 08 15.
Article in English | MEDLINE | ID: covidwho-616100

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for millions of infections and hundreds of thousands of deaths globally. There are no widely available licensed therapeutics against SARS-CoV-2, highlighting an urgent need for effective interventions. The virus enters host cells through binding of a receptor-binding domain within its trimeric spike glycoprotein to human angiotensin-converting enzyme 2. In this article, we describe the generation and characterization of a panel of murine mAbs directed against the receptor-binding domain. One mAb, 2B04, neutralized wild-type SARS-CoV-2 in vitro with remarkable potency (half-maximal inhibitory concentration of <2 ng/ml). In a murine model of SARS-CoV-2 infection, 2B04 protected challenged animals from weight loss, reduced lung viral load, and blocked systemic dissemination. Thus, 2B04 is a promising candidate for an effective antiviral that can be used to prevent SARS-CoV-2 infection.


Subject(s)
Antibodies, Monoclonal/therapeutic use , Antibodies, Neutralizing/therapeutic use , Antibodies, Viral/therapeutic use , Betacoronavirus/drug effects , Coronavirus Infections/drug therapy , Coronavirus Infections/immunology , Pneumonia, Viral/drug therapy , Pneumonia, Viral/immunology , Animals , Antibodies, Monoclonal/immunology , Antibodies, Monoclonal/pharmacology , Antibodies, Neutralizing/immunology , Antibodies, Neutralizing/pharmacology , Antibodies, Viral/immunology , Antibodies, Viral/pharmacology , Chlorocebus aethiops , Coronavirus Infections/virology , Disease Models, Animal , Epitope Mapping , Female , HEK293 Cells , Humans , Immunodominant Epitopes/immunology , Mice , Mice, Inbred C57BL , Pandemics , Peptidyl-Dipeptidase A/genetics , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/virology , Protein Interaction Domains and Motifs/genetics , Protein Interaction Domains and Motifs/immunology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism , Transfection , Vero Cells
10.
Curr Protoc Microbiol ; 58(1): e108, 2020 09.
Article in English | MEDLINE | ID: covidwho-614201

ABSTRACT

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in the city of Wuhan, Hubei Province, China, in late 2019. Since then, the virus has spread globally and caused a pandemic. Assays that can measure the antiviral activity of antibodies or antiviral compounds are needed for SARS-CoV-2 vaccine and drug development. Here, we describe in detail a microneutralization assay, which can be used to assess in a quantitative manner if antibodies or drugs can block entry and/or replication of SARS-CoV-2 in vitro. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Microneutralization assay to test inhibition of virus by antibodies (purified antibodies or serum/plasma) Basic Protocol 2: Screening of anti-SARS-CoV-2 compounds in vitro Support Protocol: SARS-CoV-2 propagation.


Subject(s)
Antibodies, Viral/immunology , Betacoronavirus/immunology , Drug Evaluation, Preclinical/methods , Neutralization Tests/methods , Animals , Chlorocebus aethiops , Coronavirus Infections/immunology , Coronavirus Infections/virology , Humans , Mice , Pandemics , Pneumonia, Viral/immunology , Pneumonia, Viral/virology , Vero Cells , Virus Internalization/drug effects , Virus Replication/drug effects
11.
Proc Natl Acad Sci U S A ; 117(28): 16587-16595, 2020 07 14.
Article in English | MEDLINE | ID: covidwho-611003

ABSTRACT

At the end of 2019, a novel coronavirus (severe acute respiratory syndrome coronavirus 2; SARS-CoV-2) was detected in Wuhan, China, that spread rapidly around the world, with severe consequences for human health and the global economy. Here, we assessed the replicative ability and pathogenesis of SARS-CoV-2 isolates in Syrian hamsters. SARS-CoV-2 isolates replicated efficiently in the lungs of hamsters, causing severe pathological lung lesions following intranasal infection. In addition, microcomputed tomographic imaging revealed severe lung injury that shared characteristics with SARS-CoV-2-infected human lung, including severe, bilateral, peripherally distributed, multilobular ground glass opacity, and regions of lung consolidation. SARS-CoV-2-infected hamsters mounted neutralizing antibody responses and were protected against subsequent rechallenge with SARS-CoV-2. Moreover, passive transfer of convalescent serum to naïve hamsters efficiently suppressed the replication of the virus in the lungs even when the serum was administrated 2 d postinfection of the serum-treated hamsters. Collectively, these findings demonstrate that this Syrian hamster model will be useful for understanding SARS-CoV-2 pathogenesis and testing vaccines and antiviral drugs.


Subject(s)
Coronavirus Infections/virology , Disease Models, Animal , Lung/pathology , Pneumonia, Viral/virology , Animals , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , Betacoronavirus/pathogenicity , Betacoronavirus/physiology , Cell Line , Chlorocebus aethiops , Coronavirus Infections/pathology , Coronavirus Infections/therapy , Cricetinae , Humans , Immunization, Passive , Lung/diagnostic imaging , Lung/virology , Mesocricetus , Pandemics , Pneumonia, Viral/pathology , Ribonucleoproteins/chemistry , Vero Cells , Viral Proteins/chemistry , Virus Replication
12.
Lancet Infect Dis ; 2020 Jun 15.
Article in English | MEDLINE | ID: covidwho-607554
13.
Lancet Infect Dis ; 2020 Jun 15.
Article in English | MEDLINE | ID: covidwho-598576
14.
Science ; 369(6501): 297-301, 2020 07 17.
Article in English | MEDLINE | ID: covidwho-418857

ABSTRACT

New York City (NYC) has emerged as one of the epicenters of the current severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. To identify the early transmission events underlying the rapid spread of the virus in the NYC metropolitan area, we sequenced the virus that causes coronavirus disease 2019 (COVID-19) in patients seeking care at the Mount Sinai Health System. Phylogenetic analysis of 84 distinct SARS-CoV-2 genomes indicates multiple, independent, but isolated introductions mainly from Europe and other parts of the United States. Moreover, we found evidence for community transmission of SARS-CoV-2 as suggested by clusters of related viruses found in patients living in different neighborhoods of the city.


Subject(s)
Betacoronavirus/genetics , Coronavirus Infections/epidemiology , Coronavirus Infections/transmission , Genome, Viral , Pneumonia, Viral/epidemiology , Pneumonia, Viral/transmission , Adult , Aged , Aged, 80 and over , Coronavirus Infections/mortality , Epidemiological Monitoring , Female , Geography, Medical , Humans , Male , Middle Aged , New York City/epidemiology , Pandemics , Phylogeny , Pneumonia, Viral/mortality , Residence Characteristics , Young Adult
17.
Cell ; 181(7): 1489-1501.e15, 2020 06 25.
Article in English | MEDLINE | ID: covidwho-260045

ABSTRACT

Understanding adaptive immunity to SARS-CoV-2 is important for vaccine development, interpreting coronavirus disease 2019 (COVID-19) pathogenesis, and calibration of pandemic control measures. Using HLA class I and II predicted peptide "megapools," circulating SARS-CoV-2-specific CD8+ and CD4+ T cells were identified in ∼70% and 100% of COVID-19 convalescent patients, respectively. CD4+ T cell responses to spike, the main target of most vaccine efforts, were robust and correlated with the magnitude of the anti-SARS-CoV-2 IgG and IgA titers. The M, spike, and N proteins each accounted for 11%-27% of the total CD4+ response, with additional responses commonly targeting nsp3, nsp4, ORF3a, and ORF8, among others. For CD8+ T cells, spike and M were recognized, with at least eight SARS-CoV-2 ORFs targeted. Importantly, we detected SARS-CoV-2-reactive CD4+ T cells in ∼40%-60% of unexposed individuals, suggesting cross-reactive T cell recognition between circulating "common cold" coronaviruses and SARS-CoV-2.


Subject(s)
Betacoronavirus/physiology , Coronavirus Infections/immunology , Epitopes, T-Lymphocyte , Pneumonia, Viral/immunology , Betacoronavirus/genetics , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Convalescence , Coronavirus Infections/blood , Coronavirus Infections/metabolism , Coronavirus Infections/prevention & control , Coronavirus Infections/virology , Cross Reactions , Humans , Leukocytes, Mononuclear/immunology , Pandemics , Pneumonia, Viral/blood , Pneumonia, Viral/metabolism , Pneumonia, Viral/virology , Spike Glycoprotein, Coronavirus/metabolism , Viral Proteins/metabolism , Viral Vaccines/immunology
18.
Nat Med ; 26(7): 1033-1036, 2020 07.
Article in English | MEDLINE | ID: covidwho-244499

ABSTRACT

Here, we describe a serological enzyme-linked immunosorbent assay for the screening and identification of human SARS-CoV-2 seroconverters. This assay does not require the handling of infectious virus, can be adjusted to detect different antibody types in serum and plasma and is amenable to scaling. Serological assays are of critical importance to help define previous exposure to SARS-CoV-2 in populations, identify highly reactive human donors for convalescent plasma therapy and investigate correlates of protection.


Subject(s)
Betacoronavirus/immunology , Clinical Laboratory Techniques/methods , Coronavirus Infections/diagnosis , Coronavirus Infections/immunology , Pneumonia, Viral/diagnosis , Pneumonia, Viral/immunology , Seroconversion , Adult , Betacoronavirus/pathogenicity , Case-Control Studies , Coronavirus Infections/blood , Coronavirus Infections/therapy , Coronavirus Infections/virology , Electrophoresis, Polyacrylamide Gel , Enzyme-Linked Immunosorbent Assay , Humans , Immunization, Passive , Longitudinal Studies , Middle Aged , Neutralization Tests , Pandemics , Pneumonia, Viral/therapy , Pneumonia, Viral/virology , Young Adult
19.
Curr Protoc Microbiol ; 57(1): e100, 2020 06.
Article in English | MEDLINE | ID: covidwho-72322

ABSTRACT

In late 2019, cases of atypical pneumonia were detected in China. The etiological agent was quickly identified as a betacoronavirus (named SARS-CoV-2), which has since caused a pandemic. Several methods allowing for the specific detection of viral nucleic acids have been established, but these only allow detection of the virus during a short period of time, generally during acute infection. Serological assays are urgently needed to conduct serosurveys, to understand the antibody responses mounted in response to the virus, and to identify individuals who are potentially immune to re-infection. Here we describe a detailed protocol for expression of antigens derived from the spike protein of SARS-CoV-2 that can serve as a substrate for immunological assays, as well as a two-stage serological enzyme-linked immunosorbent assay (ELISA). These assays can be used for research studies and for testing in clinical laboratories. © 2020 The Authors. Basic Protocol 1: Mammalian cell transfection and protein purification Basic Protocol 2: A two-stage ELISA for high-throughput screening of human serum samples for antibodies binding to the spike protein of SARS-CoV-2.


Subject(s)
Antibodies, Viral/blood , Antigens, Viral/biosynthesis , Antigens, Viral/isolation & purification , Clinical Laboratory Techniques/methods , Coronavirus Infections/diagnosis , Pneumonia, Viral/diagnosis , Spike Glycoprotein, Coronavirus/biosynthesis , Spike Glycoprotein, Coronavirus/isolation & purification , Betacoronavirus/immunology , Coronavirus Infections/blood , Coronavirus Infections/immunology , HEK293 Cells , Humans , Pandemics , Pneumonia, Viral/blood , Pneumonia, Viral/immunology , Recombinant Proteins/immunology , Recombinant Proteins/isolation & purification , Seroconversion , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology
20.
Immunity ; 52(4): 583-589, 2020 04 14.
Article in English | MEDLINE | ID: covidwho-20439

ABSTRACT

SARS-CoV-2, the causal agent of COVID-19, first emerged in late 2019 in China. It has since infected more than 870,000 individuals and caused more than 43,000 deaths globally. Here, we discuss therapeutic and prophylactic interventions for SARS-CoV-2 with a focus on vaccine development and its challenges. Vaccines are being rapidly developed but will likely come too late to affect the first wave of a potential pandemic. Nevertheless, critical lessons can be learned for the development of vaccines against rapidly emerging viruses. Importantly, SARS-CoV-2 vaccines will be essential to reducing morbidity and mortality if the virus establishes itself in the population.


Subject(s)
Betacoronavirus/pathogenicity , Coronavirus Infections/prevention & control , Drug Development , Pandemics/prevention & control , Pneumonia, Viral/prevention & control , Viral Vaccines , Animals , Betacoronavirus/immunology , China , Clinical Trials as Topic , Coronavirus/pathogenicity , Coronavirus Infections/epidemiology , Disease Models, Animal , Humans , Pneumonia, Viral/epidemiology , Time Factors , Viral Vaccines/therapeutic use
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