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1.
Indian J Med Microbiol ; 38(2): 210-212, 2020.
Article in English | MEDLINE | ID: covidwho-745217

ABSTRACT

Recent emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and subsequent containment procedures have impacted the world as never seen before. Therefore, there is considerable curiosity about the genome evolution related to the origin, transmission and vaccine impact of this virus. We have analysed genome sequences of SARS-CoV-2 isolated from Indian patients to gain an in-depth understanding of genomic evolution and transmission in India. Phylogenetic analysis and mutation profiling revealed major lineages being evolved by characteristic mutations. As the mutation frequency in spike protein is comparatively lesser, the candidate vaccines expected to have wide coverage worldwide including India.


Subject(s)
Antigens, Viral/genetics , Betacoronavirus/genetics , Coronavirus Infections/prevention & control , Genome, Viral , Pandemics/prevention & control , Pneumonia, Viral/prevention & control , Spike Glycoprotein, Coronavirus/genetics , Viral Vaccines/genetics , Animals , Antigens, Viral/immunology , Antigens, Viral/metabolism , Betacoronavirus/classification , Betacoronavirus/immunology , Betacoronavirus/pathogenicity , Chiroptera/virology , Coronavirus Infections/epidemiology , Coronavirus Infections/genetics , Coronavirus Infections/immunology , Coronavirus Infections/transmission , Disease Reservoirs/virology , Eutheria/virology , Evolution, Molecular , Humans , India/epidemiology , Mutation , Phylogeny , Pneumonia, Viral/epidemiology , Pneumonia, Viral/immunology , Pneumonia, Viral/transmission , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism , Viral Vaccines/biosynthesis , Viral Vaccines/immunology
2.
Emerg Microbes Infect ; 9(1): 1965-1973, 2020 Dec.
Article in English | MEDLINE | ID: covidwho-725731

ABSTRACT

Serology is a crucial part of the public health response to the ongoing SARS-CoV-2 pandemic. Here, we describe the development, validation and clinical evaluation of a protein micro-array as a quantitative multiplex immunoassay that can identify S and N-directed SARS-CoV-2 IgG antibodies with high specificity and sensitivity and distinguish them from all currently circulating human coronaviruses. The method specificity was 100% for SARS-CoV-2 S1 and 96% for N antigen based on extensive syndromic (n=230 cases) and population panel (n=94) testing that also confirmed the high prevalence of seasonal human coronaviruses. To assess its potential role for both SARS-CoV-2 patient diagnostics and population studies, we evaluated a large heterogeneous COVID-19 cohort (n=330) and found an overall sensitivity of 89% (≥ 21 days post onset symptoms (dps)), ranging from 86% to 96% depending on severity of disease. For a subset of these patients longitudinal samples were provided up to 56 dps. Mild cases showed absent or delayed, and lower SARS-CoV-2 antibody responses. Overall, we present the development and extensive clinical validation of a multiplex coronavirus serological assay for syndromic testing, to answer research questions regarding to antibody responses, to support SARS-CoV-2 diagnostics and to evaluate epidemiological developments efficiently and with high-throughput.


Subject(s)
Antibodies, Viral/blood , Betacoronavirus/immunology , Clinical Laboratory Techniques/methods , Coronavirus Infections/diagnosis , Nucleocapsid Proteins/blood , Pneumonia, Viral/diagnosis , Spike Glycoprotein, Coronavirus/blood , Aged , Antigens, Viral/blood , Antigens, Viral/immunology , Betacoronavirus/pathogenicity , Clinical Laboratory Techniques/standards , Coronavirus Infections/immunology , Coronavirus Infections/mortality , Coronavirus Infections/virology , Female , Humans , Longitudinal Studies , Male , Middle Aged , Middle East Respiratory Syndrome Coronavirus/immunology , Middle East Respiratory Syndrome Coronavirus/pathogenicity , Neutralization Tests , Nucleocapsid Proteins/immunology , Pandemics , Pneumonia, Viral/immunology , Pneumonia, Viral/mortality , Pneumonia, Viral/virology , Protein Array Analysis , SARS Virus/immunology , SARS Virus/pathogenicity , Sensitivity and Specificity , Severity of Illness Index , Spike Glycoprotein, Coronavirus/immunology
3.
Signal Transduct Target Ther ; 5(1): 156, 2020 08 14.
Article in English | MEDLINE | ID: covidwho-717099

ABSTRACT

The global Coronavirus disease 2019 (COVID-19) pandemic caused by SARS-CoV-2 has affected more than eight million people. There is an urgent need to investigate how the adaptive immunity is established in COVID-19 patients. In this study, we profiled adaptive immune cells of PBMCs from recovered COVID-19 patients with varying disease severity using single-cell RNA and TCR/BCR V(D)J sequencing. The sequencing data revealed SARS-CoV-2-specific shuffling of adaptive immune repertories and COVID-19-induced remodeling of peripheral lymphocytes. Characterization of variations in the peripheral T and B cells from the COVID-19 patients revealed a positive correlation of humoral immune response and T-cell immune memory with disease severity. Sequencing and functional data revealed SARS-CoV-2-specific T-cell immune memory in the convalescent COVID-19 patients. Furthermore, we also identified novel antigens that are responsive in the convalescent patients. Altogether, our study reveals adaptive immune repertories underlying pathogenesis and recovery in severe versus mild COVID-19 patients, providing valuable information for potential vaccine and therapeutic development against SARS-CoV-2 infection.


Subject(s)
B-Lymphocytes/immunology , Betacoronavirus/pathogenicity , Coronavirus Infections/immunology , Immunity, Cellular , Immunity, Humoral , Pneumonia, Viral/immunology , T-Lymphocytes/immunology , Antigens, Viral/genetics , Antigens, Viral/immunology , B-Lymphocytes/classification , B-Lymphocytes/virology , Betacoronavirus/immunology , Case-Control Studies , China , Convalescence , Coronavirus Infections/genetics , Coronavirus Infections/pathology , Coronavirus Infections/virology , Disease Progression , Gene Expression , High-Throughput Nucleotide Sequencing , Host-Pathogen Interactions/immunology , Humans , Immunologic Memory , Pandemics , Pneumonia, Viral/genetics , Pneumonia, Viral/pathology , Pneumonia, Viral/virology , Receptors, Antigen, B-Cell/classification , Receptors, Antigen, B-Cell/genetics , Receptors, Antigen, B-Cell/immunology , Receptors, Antigen, T-Cell/classification , Receptors, Antigen, T-Cell/genetics , Receptors, Antigen, T-Cell/immunology , Severity of Illness Index , Single-Cell Analysis , T-Lymphocytes/classification , T-Lymphocytes/virology
4.
BMC Res Notes ; 13(1): 372, 2020 Aug 06.
Article in English | MEDLINE | ID: covidwho-696522

ABSTRACT

OBJECTIVE: COVID19 has caused a global and ongoing pandemic. The need for population seroconversion data is apparent to monitor and respond to the pandemic. Using a lateral flow assay (LFA) testing platform, the seropositivity in 63 New York Blood Center (NYBC) Convelescent Plasma (CP) donor samples were evaluated for the presence of COVID19 specific IgG and IgM. RESULTS: CP donors showed diverse antibody result. Convalescent donor plasma contains SARS-CoV-2 specific antibodies. Weak antibody bands may identify low titer CP donors. LFA tests can identify antibody positive individuals that have recovered from COVID19. Confirming suspected cases using antibody detection could help inform the patient and the community as to the relative risk to future exposure and a better understanding of disease exposure.


Subject(s)
Antibodies, Viral/blood , Antigens, Viral/immunology , Betacoronavirus/immunology , Blood Donors , Clinical Laboratory Techniques/methods , Convalescence , Coronavirus Infections/diagnosis , Immunoassay/methods , Immunoglobulin G/blood , Immunoglobulin M/blood , Nucleocapsid Proteins/immunology , Pandemics , Pneumonia, Viral/diagnosis , Point-of-Care Testing , Spike Glycoprotein, Coronavirus/immunology , Antibody Specificity , Coronavirus Infections/therapy , Gold Colloid , Humans , Immunization, Passive , Plasma , Protein Domains , Recombinant Proteins/immunology , Reproducibility of Results , Sensitivity and Specificity , Seroconversion
5.
Drug Des Devel Ther ; 14: 2607-2611, 2020.
Article in English | MEDLINE | ID: covidwho-683522

ABSTRACT

In March 2020, the WHO declared the COVID-19 disease as a pandemic disease. There have been studies on the COVID-19 to find a certain treatment, but yet, there is no certain cure. In this article, we present a possible way to treat severe cases of COVID-19. Based on the previous studies, there are similarities between the spike antigens of SARS-CoV and SARS-CoV-2 viruses. It is expected that these similarities (structural and affinity to the receptor of ACE2) can lead to the same pathophysiological activity of the virus by the use of ACE2 and FcγRII (the antibody-dependent enhancement mechanism). Therefore, we propose a way of washing out (by plasmapheresis) the possible antibodies against the spike protein of the virus out of patients' plasma to stop the antibody-dependent enhancement (ADE)-mediated infection of the immune system cells at the first phase of the treatment and simultaneous use of the anti-ACE2 with anti-FcγRII monoclonal antibodies at the second phase. We propose these procedures for the patients that have no significant response for typical anti-viral, ARDS and conservative therapies, and the disease persists or progresses despite sufficient therapies.


Subject(s)
Antibodies, Monoclonal/administration & dosage , Coronavirus Infections/therapy , Plasmapheresis/methods , Pneumonia, Viral/therapy , Antibodies, Monoclonal/immunology , Antibodies, Viral/immunology , Antigens, Viral/immunology , Coronavirus Infections/immunology , Coronavirus Infections/virology , Humans , Pandemics , Peptidyl-Dipeptidase A/immunology , Pneumonia, Viral/immunology , Pneumonia, Viral/virology , Receptors, IgG/immunology , Severity of Illness Index
6.
Curr Opin Rheumatol ; 32(5): 458-461, 2020 09.
Article in English | MEDLINE | ID: covidwho-654334

ABSTRACT

BACKGROUND: Shortly after its emergence in December 2019, the coronavirus disease 2019 (COVID-19) was declared as a pandemic by the World Health Organization. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is the seventh member of the Coronaviridae family of viruses that causes disease in humans. THE PROBLEM: Despite the established role of molecular diagnostics, COVID-19 serodiagnosis remains a poorly discovered and enigmatic area. Although there are numerous commercial serological products available globally, there is a severe paucity of high-quality peer-reviewed literature on their true performance characteristics. That being said, publications including in-house developed serological tests started to shed light on the kinetics of the humoral response. SUMMARY: In spite of intense focus of assessing the performance characteristics of the commercially-available kits, the main issue remains rather invisible, that is, lack of solid science behind COVID-19 serology its clinical usefulness thereof. This short review summarizes the key points as to why COVID-19 is not jest ready to fly. PURPOSE OF REVIEW: Despite having been mentioned as a testing option, COVID-19 serology has significant shortcomings that needs discussing. This short review is meant to shed light on one of those aspects.


Subject(s)
Antibody-Dependent Enhancement , Antigens, Viral/immunology , Betacoronavirus/immunology , Coronavirus Infections/immunology , Pneumonia, Viral/immunology , Humans , Pandemics , Serologic Tests
7.
Cell ; 182(5): 1284-1294.e9, 2020 09 03.
Article in English | MEDLINE | ID: covidwho-652603

ABSTRACT

The spike protein of SARS-CoV-2 has been undergoing mutations and is highly glycosylated. It is critically important to investigate the biological significance of these mutations. Here, we investigated 80 variants and 26 glycosylation site modifications for the infectivity and reactivity to a panel of neutralizing antibodies and sera from convalescent patients. D614G, along with several variants containing both D614G and another amino acid change, were significantly more infectious. Most variants with amino acid change at receptor binding domain were less infectious, but variants including A475V, L452R, V483A, and F490L became resistant to some neutralizing antibodies. Moreover, the majority of glycosylation deletions were less infectious, whereas deletion of both N331 and N343 glycosylation drastically reduced infectivity, revealing the importance of glycosylation for viral infectivity. Interestingly, N234Q was markedly resistant to neutralizing antibodies, whereas N165Q became more sensitive. These findings could be of value in the development of vaccine and therapeutic antibodies.


Subject(s)
Antigens, Viral/genetics , Betacoronavirus/pathogenicity , Mutation , Spike Glycoprotein, Coronavirus/genetics , A549 Cells , Animals , Antigens, Viral/immunology , Betacoronavirus/genetics , Betacoronavirus/immunology , Binding Sites , Cattle , Chlorocebus aethiops , Cricetinae , Dogs , Glycosylation , HEK293 Cells , HeLa Cells , Humans , Macaca mulatta , Madin Darby Canine Kidney Cells , Mice , RAW 264.7 Cells , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism , Swine , Vero Cells , Virulence/genetics
8.
Vaccine ; 38(35): 5734-5739, 2020 07 31.
Article in English | MEDLINE | ID: covidwho-640754

ABSTRACT

Several protein vaccine candidates are among the COVID-19 vaccines in development. The Brighton Collaboration Viral Vector Vaccines Safety Working Group (V3SWG) has prepared a standardized template to describe the key considerations for the benefit-risk assessment of protein vaccines. This will help key stakeholders to assess potential safety issues and understand the benefit-risk of such a vaccine platform. The structured and standardized assessment provided by the template would also help contribute to improved public acceptance and communication of licensed protein vaccines.


Subject(s)
Viral Vaccines/adverse effects , Viral Vaccines/immunology , Antigens, Viral/administration & dosage , Antigens, Viral/adverse effects , Antigens, Viral/immunology , Coronavirus Infections/immunology , Coronavirus Infections/prevention & control , Humans , Patient Safety , Risk Assessment , Vaccines, Synthetic/administration & dosage , Vaccines, Synthetic/adverse effects , Vaccines, Synthetic/immunology , Viral Proteins/administration & dosage , Viral Proteins/adverse effects , Viral Proteins/immunology , Viral Vaccines/administration & dosage
9.
J Clin Virol ; 129: 104544, 2020 08.
Article in English | MEDLINE | ID: covidwho-634673

ABSTRACT

The emergence of the severe acute respiratory syndrome coronavirus-2 (SARS CoV-2) has been followed by the rapid development of antibody tests. To assess the utility of the tests for clinical use and seroepidemiologic studies, we examined the sensitivity of commercial antibody tests from Roche, Abbott, Novatec, Virotech Siemens, Euroimmun, and Mediagnost in a prospective diagnostic study. The tests were evaluated with 73 sera from SARS CoV-2 RNA positive individuals with mild to moderate disease or asymptomatic infection. Sera were obtained at 2-3 weeks (N = 25) or > 4 weeks (N = 48) after symptom onset and viral RNA test. The overall sensitivity of the tests ranged from 64.4-93.2%. The most sensitive assays recognized 95.8-100 % of the sera obtained after 4 weeks or later. Sera drawn at 2-3 weeks were recognized with lower sensitivity indicating that the optimal time point for serologic testing is later than 3 weeks after onset of the disease. Nucleoprotein- and glycoproteinbased assays had similar sensitivity indicating that tests with both antigens are suitable for serological diagnostics. Breakdown of the test results showed that nucleoprotein- and glycoprotein-based tests of comparable sensitivity reacted with different sets of sera. The observation indicates that a combination of nucleoprotein- and glycoprotein-based tests would increase the percentage of positive results.


Subject(s)
Antibodies, Viral/blood , Antigens, Viral/immunology , Betacoronavirus/isolation & purification , Clinical Laboratory Techniques/methods , Coronavirus Infections/diagnosis , Pneumonia, Viral/diagnosis , Serologic Tests/methods , Viral Structural Proteins/immunology , Betacoronavirus/immunology , Glycoproteins/immunology , Humans , Nucleoproteins/immunology , Pandemics , Prospective Studies , Sensitivity and Specificity , Time Factors
10.
Emerg Microbes Infect ; 9(1): 1712-1721, 2020 Dec.
Article in English | MEDLINE | ID: covidwho-632216

ABSTRACT

Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) has resulted in a pandemic and is continuing to spread rapidly around the globe. No effective vaccine is currently available to prevent COVID-19, and intense efforts are being invested worldwide into vaccine development. In this context, all technology platforms must overcome several challenges resulting from the use of an incompletely characterized new virus. These include finding the right conditions for virus amplification for the development of vaccines based on inactivated or attenuated whole viral particles. Here, we describe a shotgun tandem mass spectrometry workflow, the data produced can be used to guide optimization of the conditions for viral amplification. In parallel, we analysed the changes occurring in the host cell proteome following SARS-CoV-2 infection to glean information on the biological processes modulated by the virus that could be further explored as potential drug targets to deal with the pandemic.


Subject(s)
Antigens, Viral/biosynthesis , Betacoronavirus/immunology , Proteomics/methods , Viral Vaccines/immunology , Virion/immunology , Animals , Antigens, Viral/immunology , Chlorocebus aethiops , Tandem Mass Spectrometry , Vero Cells
11.
J Infect ; 81(3): e31-e32, 2020 Sep.
Article in English | MEDLINE | ID: covidwho-625901

ABSTRACT

We followed-up a mild COVID-19 patient for 91 days and serially monitored his serum antibodies to four SARS-CoV-2 related antigens (NP, RBD, S1 and ECD) and neutralization activities. Our data revealed a profile of serial antibody responses during the progress and a quick decline of neutralization activities after discharge.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Betacoronavirus/immunology , Coronavirus Infections/immunology , Pneumonia, Viral/immunology , Adult , Antigens, Viral/immunology , Enzyme-Linked Immunosorbent Assay , Humans , Longitudinal Studies , Male , Neutralization Tests , Pandemics , RNA, Viral , Retrospective Studies
12.
Science ; 369(6504): 650-655, 2020 08 07.
Article in English | MEDLINE | ID: covidwho-610891

ABSTRACT

Developing therapeutics against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) could be guided by the distribution of epitopes, not only on the receptor binding domain (RBD) of the Spike (S) protein but also across the full Spike (S) protein. We isolated and characterized monoclonal antibodies (mAbs) from 10 convalescent COVID-19 patients. Three mAbs showed neutralizing activities against authentic SARS-CoV-2. One mAb, named 4A8, exhibits high neutralization potency against both authentic and pseudotyped SARS-CoV-2 but does not bind the RBD. We defined the epitope of 4A8 as the N-terminal domain (NTD) of the S protein by determining with cryo-eletron microscopy its structure in complex with the S protein to an overall resolution of 3.1 angstroms and local resolution of 3.3 angstroms for the 4A8-NTD interface. This points to the NTD as a promising target for therapeutic mAbs against COVID-19.


Subject(s)
Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Betacoronavirus/immunology , Coronavirus Infections/immunology , Pneumonia, Viral/immunology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Adult , Animals , Antibodies, Monoclonal/blood , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/metabolism , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/chemistry , Antibodies, Neutralizing/metabolism , Antibodies, Viral/blood , Antibodies, Viral/chemistry , Antibodies, Viral/metabolism , Antibody Affinity , Antibody Specificity , Antigens, Viral/immunology , B-Lymphocytes/immunology , Chlorocebus aethiops , Coronavirus Infections/therapy , Cryoelectron Microscopy , Enzyme-Linked Immunosorbent Assay , Genes, Immunoglobulin Heavy Chain , Humans , Immunologic Memory , Middle Aged , Mutation , Nucleocapsid Proteins/immunology , Pandemics , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/therapy , Protein Domains , Protein Interaction Domains and Motifs/immunology , Receptors, Virus/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Vero Cells , Young Adult
13.
J Clin Virol ; 129: 104521, 2020 08.
Article in English | MEDLINE | ID: covidwho-610625

ABSTRACT

BACKGROUND: Knowledge of the COVID-19 epidemic extent and the level of herd immunity is urgently needed to help manage this pandemic. METHODS: We used a panel of 167 samples (77 pre-epidemic and 90 COVID-19 seroconverters) and SARS-CoV1, SARS-CoV2 and MERS-CoV Spike and/or Nucleopcapsid (NC) proteins to develop a high throughput multiplex screening assay to detect IgG antibodies in human plasma. Assay performances were determined by ROC curves analysis. A subset of the COVID-19+ samples (n = 36) were also tested by a commercial NC-based ELISA test and the results compared with those of the novel assay. RESULTS: On samples collected ≥14 days after symptoms onset, the accuracy of the assay is 100 % (95 % CI: 100-100) for the Spike antigen and 99.9 % (95 % CI:99.7-100) for NC. By logistic regression, we estimated that 50 % of the patients have seroconverted at 5.7 ± 1.6; 5.7 ± 1.8 and 7.9 ± 1.0 days after symptoms onset against Spike, NC or both antigens, respectively and all have seroconverted two weeks after symptoms onset. IgG titration in a subset of samples showed that early phase samples present lower IgG titers than those from later phase. IgG to SARS-CoV2 NC cross-reacted at 100 % with SARS-CoV1 NC. Twenty-nine of the 36 (80.5 %) samples tested were positive by the commercial ELISA while 31/36 (86.1 %) were positive by the novel assay. CONCLUSIONS: Our assay is highly sensitive and specific for the detection of IgG antibodies to SARS-CoV2 proteins, suitable for high throughput epidemiological surveys. The novel assay is more sensitive than a commercial ELISA.


Subject(s)
Antibodies, Viral/blood , Betacoronavirus/immunology , Immunoglobulin G/blood , Middle East Respiratory Syndrome Coronavirus/immunology , SARS Virus/immunology , Serologic Tests/methods , Adult , Aged , Aged, 80 and over , Antigens, Viral/immunology , Coronavirus Infections/diagnosis , Coronavirus Infections/immunology , Coronavirus Infections/virology , Female , Humans , Male , Middle Aged , Nucleocapsid Proteins/immunology , Pandemics , Pneumonia, Viral/diagnosis , Pneumonia, Viral/immunology , Pneumonia, Viral/virology , ROC Curve , Sensitivity and Specificity , Severe Acute Respiratory Syndrome/diagnosis , Severe Acute Respiratory Syndrome/immunology , Severe Acute Respiratory Syndrome/virology , Spike Glycoprotein, Coronavirus/immunology , Time Factors
14.
Science ; 369(6504): 643-650, 2020 08 07.
Article in English | MEDLINE | ID: covidwho-599037

ABSTRACT

The rapid spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has had a large impact on global health, travel, and economy. Therefore, preventative and therapeutic measures are urgently needed. Here, we isolated monoclonal antibodies from three convalescent coronavirus disease 2019 (COVID-19) patients using a SARS-CoV-2 stabilized prefusion spike protein. These antibodies had low levels of somatic hypermutation and showed a strong enrichment in VH1-69, VH3-30-3, and VH1-24 gene usage. A subset of the antibodies was able to potently inhibit authentic SARS-CoV-2 infection at a concentration as low as 0.007 micrograms per milliliter. Competition and electron microscopy studies illustrate that the SARS-CoV-2 spike protein contains multiple distinct antigenic sites, including several receptor-binding domain (RBD) epitopes as well as non-RBD epitopes. In addition to providing guidance for vaccine design, the antibodies described here are promising candidates for COVID-19 treatment and prevention.


Subject(s)
Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Betacoronavirus/immunology , Coronavirus Infections/immunology , Pneumonia, Viral/immunology , Spike Glycoprotein, Coronavirus/immunology , Adult , Aged , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , Antibody Affinity , Antigens, Viral/immunology , B-Lymphocyte Subsets/immunology , Broadly Neutralizing Antibodies/immunology , Cell Line, Tumor , Coronavirus Infections/prevention & control , Coronavirus Infections/therapy , Epitopes/immunology , Female , Humans , Immunologic Memory , Immunophenotyping , Male , Middle Aged , Pandemics/prevention & control , Pneumonia, Viral/prevention & control , Pneumonia, Viral/therapy , Protein Domains , Protein Interaction Domains and Motifs/immunology , Receptors, Virus/metabolism , Spike Glycoprotein, Coronavirus/chemistry
15.
Clin Microbiol Infect ; 26(8): 1082-1087, 2020 Aug.
Article in English | MEDLINE | ID: covidwho-594311

ABSTRACT

OBJECTIVES: To evaluate the diagnostic performance of seven rapid IgG/IgM tests and the Euroimmun IgA/IgG ELISA for antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in COVID-19 patients. METHODS: Specificity was evaluated in 103 samples collected before January 2020. Sensitivity and time to seropositivity was evaluated in 167 samples from 94 patients with COVID-19 confirmed with RT-PCR on nasopharyngeal swab. RESULTS: Specificity (confidence interval) of lateral flow assays (LFAs) was ≥91.3% (84.0-95.5) for IgM, ≥90.3% (82.9-94.8) for IgG, and ≥85.4% (77.2-91.1) for the combination IgM OR IgG. Specificity of the ELISA was 96.1% (90.1-98.8) for IgG and only 73.8% (64.5-81.4) for IgA. Sensitivity 14-25 days after the onset of symptoms was between ≥92.1% (78.5-98.0) and 100% (95.7-100) for IgG LFA compared to 89.5% (75.3-96.4) for IgG ELISA. Positivity of IgM OR IgG for LFA resulted in a decrease in specificity compared to IgG alone without a gain in diagnostic performance, except for VivaDiag. The results for IgM varied significantly between the LFAs with an average overall agreement of only 70% compared to 89% for IgG. The average dynamic trend to seropositivity for IgM was not shorter than for IgG. At the time of hospital admission the sensitivity of LFA was <60%. CONCLUSIONS: Sensitivity for the detection of IgG antibodies 14-25 days after the onset of symptoms was ≥92.1% for all seven LFAs compared to 89.5% for the IgG ELISA. The results for IgM varied significantly, and including IgM antibodies in addition to IgG for the interpretation of LFAs did not improve the diagnostic performance.


Subject(s)
Antibodies, Viral/analysis , Antigens, Viral/immunology , Betacoronavirus/immunology , Coronavirus Infections/diagnosis , Pneumonia, Viral/diagnosis , Adult , Aged , Aged, 80 and over , Coronavirus Infections/immunology , Diagnostic Tests, Routine , Female , Humans , Immunoglobulin A/metabolism , Immunoglobulin G/metabolism , Immunoglobulin M/metabolism , Male , Middle Aged , Pandemics , Pneumonia, Viral/immunology , Sensitivity and Specificity , Time Factors , Young Adult
16.
Vaccine ; 38(32): 5071-5075, 2020 07 06.
Article in English | MEDLINE | ID: covidwho-592568

ABSTRACT

SARS-CoV-2 is the cause of the worldwide outbreak of COVID-19 that has been characterized as a pandemic by the WHO. Since the first report of COVID-19 on December 31, 2019, 179,111 cases were confirmed in 160 countries/regions with 7426 deaths as of March 17, 2020. However, there have been no vaccines approved in the world to date. In this study, we analyzed the biological characteristics of the SARS-CoV-2 Spike protein, Pro330-Leu650 (SARS-CoV-2-SPL), using biostatistical methods. SARS-CoV-2-SPL possesses a receptor-binding region (RBD) and important B (Ser438-Gln506, Thr553-Glu583, Gly404-Aps427, Thr345-Ala352, and Lys529-Lys535) and T (9 CD4 and 11 CD8 T cell antigenic determinants) cell epitopes. High homology in this region between SARS-CoV-2 and SARS-CoV amounted to 87.7%, after taking the biological similarity of the amino acids into account and eliminating the receptor-binding motif (RBM). The overall topology indicated that the complete structure of SARS-CoV-2-SPL was with RBM as the head, and RBD as the trunk and the tail region. SARS-CoV-2-SPL was found to have the potential to elicit effective B and T cell responses. Our findings may provide meaningful guidance for SARS-CoV-2 vaccine design.


Subject(s)
Betacoronavirus/chemistry , Drug Design , Models, Immunological , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Viral Vaccines/chemistry , Viral Vaccines/immunology , Amino Acid Sequence , Antigens, Viral/chemistry , Antigens, Viral/immunology , Betacoronavirus/immunology , Coronavirus Infections/epidemiology , Coronavirus Infections/immunology , Coronavirus Infections/prevention & control , Coronavirus Infections/virology , Epitopes, B-Lymphocyte/chemistry , Epitopes, B-Lymphocyte/immunology , Epitopes, T-Lymphocyte/chemistry , Epitopes, T-Lymphocyte/immunology , Humans , Models, Molecular , Pandemics/prevention & control , Pneumonia, Viral/epidemiology , Pneumonia, Viral/immunology , Pneumonia, Viral/prevention & control , Pneumonia, Viral/virology , Sequence Alignment , Vaccines, Subunit/chemistry , Vaccines, Subunit/immunology
17.
Viruses ; 12(6)2020 06 10.
Article in English | MEDLINE | ID: covidwho-592405

ABSTRACT

The ongoing Coronavirus Disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) signals an urgent need for an expansion in treatment options. In this study, we investigated the anti-SARS-CoV-2 activities of 22 antiviral agents with known broad-spectrum antiviral activities against coronaviruses and/or other viruses. They were first evaluated in our primary screening in VeroE6 cells and then the most potent anti-SARS-CoV-2 antiviral agents were further evaluated using viral antigen expression, viral load reduction, and plaque reduction assays. In addition to remdesivir, lopinavir, and chloroquine, our primary screening additionally identified types I and II recombinant interferons, 25-hydroxycholesterol, and AM580 as the most potent anti-SARS-CoV-2 agents among the 22 antiviral agents. Betaferon (interferon-ß1b) exhibited the most potent anti-SARS-CoV-2 activity in viral antigen expression, viral load reduction, and plaque reduction assays among the recombinant interferons. The lipogenesis modulators 25-hydroxycholesterol and AM580 exhibited EC50 at low micromolar levels and selectivity indices of >10.0. Combinational use of these host-based antiviral agents with virus-based antivirals to target different processes of the SARS-CoV-2 replication cycle should be evaluated in animal models and/or clinical trials.


Subject(s)
Antiviral Agents/pharmacology , Betacoronavirus/drug effects , Coronavirus Infections/drug therapy , Pneumonia, Viral/drug therapy , Animals , Antigens, Viral/immunology , Betacoronavirus/immunology , Betacoronavirus/metabolism , Chlorocebus aethiops , Coronavirus Infections/virology , Humans , Interferons/metabolism , Lipogenesis/drug effects , Pandemics , Pneumonia, Viral/virology , Signal Transduction/drug effects , Vero Cells , Viral Load/drug effects , Viral Plaque Assay , Virus Replication/drug effects
18.
Sci Transl Med ; 12(550)2020 07 01.
Article in English | MEDLINE | ID: covidwho-591374

ABSTRACT

Multiple vaccine candidates against SARS-CoV-2 based on viral spike protein are under development. However, there is limited information on the quality of antibody responses generated with these vaccine modalities. To better understand antibody responses induced by spike protein-based vaccines, we performed a qualitative study by immunizing rabbits with various SARS-CoV-2 spike protein antigens: S ectodomain (S1+S2; amino acids 16 to 1213), which lacks the cytoplasmic and transmembrane domains (CT-TM), the S1 domain (amino acids 16 to 685), the receptor binding domain (RBD) (amino acids 319 to 541), and the S2 domain (amino acids 686 to 1213, lacking the RBD, as control). Resulting antibody quality and function were analyzed by enzyme-linked immunosorbent assay (ELISA), RBD competition assay, surface plasmon resonance (SPR) against different spike proteins in native conformation, and neutralization assays. All three antigens (S1+S2 ectodomain, S1 domain, and RBD), but not S2, generated strong neutralizing antibodies against SARS-CoV-2. Vaccination-induced antibody repertoire was analyzed by SARS-CoV-2 spike genome fragment phage display libraries (SARS-CoV-2 GFPDL), which identified immunodominant epitopes in the S1, S1-RBD, and S2 domains. Furthermore, these analyses demonstrated that the RBD immunogen elicited a higher antibody titer with five-fold higher affinity antibodies to native spike antigens compared with other spike antigens, and antibody affinity correlated strongly with neutralization titers. These findings may help guide rational vaccine design and facilitate development and evaluation of effective therapeutics and vaccines against COVID-19 disease.


Subject(s)
Antibodies, Viral/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Neutralizing/immunology , Antibody Formation/immunology , Antigens, Viral/immunology , Epitopes/immunology , Female , Immunization , Neutralization Tests , Rabbits
19.
F1000Res ; 9: 285, 2020.
Article in English | MEDLINE | ID: covidwho-619112

ABSTRACT

SARS-CoV-2 is the coronavirus agent of the COVID-19 pandemic causing high mortalities. In contrast, the widely spread human coronaviruses OC43, HKU1, 229E, and NL63 tend to cause only mild symptoms. The present study shows, by in silico analysis, that these common human viruses are expected to induce immune memory against SARS-CoV-2 by sharing protein fragments (antigen epitopes) for presentation to the immune system by MHC class I. A list of such epitopes is provided. The number of these epitopes and the prevalence of the common coronaviruses suggest that a large part of the world population has some degree of specific immunity against SARS-CoV-2 already, even without having been infected by that virus. For inducing protection, booster vaccinations enhancing existing immunity are less demanding than primary vaccinations against new antigens. Therefore, for the discussion on vaccination strategies against COVID-19, the available immune memory against related viruses should be part of the consideration.


Subject(s)
Antigens, Viral/immunology , Coronavirus Infections/immunology , Immunologic Memory , Pneumonia, Viral/immunology , Betacoronavirus , Coronavirus/classification , Epitopes/immunology , Humans , Pandemics
20.
Clin Chem ; 66(8): 1030-1046, 2020 08 01.
Article in English | MEDLINE | ID: covidwho-326949

ABSTRACT

BACKGROUND: The ongoing outbreak of the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has posed a challenge for worldwide public health. A reliable laboratory assay is essential both to confirm suspected patients and to exclude patients infected with other respiratory viruses, thereby facilitating the control of global outbreak scenarios. CONTENT: In this review, we focus on the genomic, transmission, and clinical characteristics of SARS-CoV-2, and comprehensively summarize the principles and related details of assays for SARS-CoV-2. We also explore the quality assurance measures for these assays. SUMMARY: SARS-CoV-2 has some unique gene sequences and specific transmission and clinical features that can inform the conduct of molecular and serological assays in many aspects, including the design of primers, the selection of specimens, and testing strategies at different disease stages. Appropriate quality assurance measures for molecular and serological assays are needed to maintain testing proficiency. Because serological assays have the potential to identify later stages of the infection and to confirm highly suspected cases with negative molecular assay results, a combination of these two assays is needed to achieve a reliable capacity to detect SARS-CoV-2.


Subject(s)
Betacoronavirus/genetics , Coronavirus Infections/diagnosis , Genome, Viral , Pneumonia, Viral/diagnosis , Antibodies, Viral/blood , Antigens, Viral/genetics , Antigens, Viral/immunology , Betacoronavirus/classification , Betacoronavirus/isolation & purification , Coronavirus Infections/transmission , Coronavirus Infections/virology , Humans , Pandemics , Phylogeny , Pneumonia, Viral/transmission , Pneumonia, Viral/virology , Quality Control , RNA, Viral/metabolism , RNA, Viral/standards , Reverse Transcriptase Polymerase Chain Reaction/methods , Reverse Transcriptase Polymerase Chain Reaction/standards
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