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1.
Cell Rep ; 37(13): 110169, 2021 12 28.
Article in English | MEDLINE | ID: covidwho-1616407

ABSTRACT

The importance of pre-existing immune responses to seasonal endemic coronaviruses (HCoVs) for the susceptibility to SARS-CoV-2 infection and the course of COVID-19 is the subject of an ongoing scientific debate. Recent studies postulate that immune responses to previous HCoV infections can either have a slightly protective or no effect on SARS-CoV-2 pathogenesis and, consequently, be neglected for COVID-19 risk stratification. Challenging this notion, we provide evidence that pre-existing, anti-nucleocapsid antibodies against endemic α-coronaviruses and S2 domain-specific anti-spike antibodies against ß-coronavirus HCoV-OC43 are elevated in patients with COVID-19 compared to pre-pandemic donors. This finding is particularly pronounced in males and in critically ill patients. Longitudinal evaluation reveals that antibody cross-reactivity or polyclonal stimulation by SARS-CoV-2 infection are unlikely to be confounders. Thus, specific pre-existing immunity to seasonal coronaviruses may increase susceptibility to SARS-CoV-2 and predispose individuals to an adverse COVID-19 outcome, guiding risk management and supporting the development of universal coronavirus vaccines.


Subject(s)
COVID-19/immunology , Coronavirus/immunology , SARS-CoV-2/immunology , Adult , Antibodies/immunology , Antibodies, Viral/immunology , COVID-19/etiology , Coronavirus Infections/immunology , Coronavirus OC43, Human/immunology , Coronavirus OC43, Human/pathogenicity , Cross Reactions/immunology , Female , Germany , Humans , Immunity, Humoral/immunology , Immunoglobulin G/immunology , Longitudinal Studies , Male , Middle Aged , Pandemics , SARS-CoV-2/pathogenicity , Seasons , Severity of Illness Index , Spike Glycoprotein, Coronavirus/immunology
2.
Cell Rep ; 37(13): 110167, 2021 12 28.
Article in English | MEDLINE | ID: covidwho-1596401

ABSTRACT

Cross-reactivity and direct killing of target cells remain underexplored for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-specific CD8+ T cells. Isolation of T cell receptors (TCRs) and overexpression in allogeneic cells allows for extensive T cell reactivity profiling. We identify SARS-CoV-2 RNA-dependent RNA polymerase (RdRp/NSP12) as highly conserved, likely due to its critical role in the virus life cycle. We perform single-cell TCRαß sequencing in human leukocyte antigen (HLA)-A∗02:01-restricted, RdRp-specific T cells from SARS-CoV-2-unexposed individuals. Human T cells expressing these TCRαß constructs kill target cell lines engineered to express full-length RdRp. Three TCR constructs recognize homologous epitopes from common cold coronaviruses, indicating CD8+ T cells can recognize evolutionarily diverse coronaviruses. Analysis of individual TCR clones may help define vaccine epitopes that can induce long-term immunity against SARS-CoV-2 and other coronaviruses.


Subject(s)
Coronavirus RNA-Dependent RNA Polymerase/immunology , HLA-A2 Antigen/immunology , SARS-CoV-2/immunology , CD8-Positive T-Lymphocytes/immunology , COVID-19/immunology , COVID-19/therapy , Cell Culture Techniques , Cross Reactions/immunology , Epitopes, T-Lymphocyte/immunology , HLA-A Antigens/immunology , HLA-A2 Antigen/genetics , Humans , Immunodominant Epitopes/immunology , Leukocytes, Mononuclear/immunology , Leukocytes, Mononuclear/metabolism , Leukocytes, Mononuclear/virology , RNA, Viral/genetics , Receptors, Antigen, T-Cell/immunology , Receptors, Antigen, T-Cell, alpha-beta/genetics , Receptors, Antigen, T-Cell, alpha-beta/immunology , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/immunology
3.
Nat Immunol ; 23(1): 40-49, 2022 01.
Article in English | MEDLINE | ID: covidwho-1585824

ABSTRACT

SARS-CoV-2 infection is generally mild or asymptomatic in children but a biological basis for this outcome is unclear. Here we compare antibody and cellular immunity in children (aged 3-11 years) and adults. Antibody responses against spike protein were high in children and seroconversion boosted responses against seasonal Beta-coronaviruses through cross-recognition of the S2 domain. Neutralization of viral variants was comparable between children and adults. Spike-specific T cell responses were more than twice as high in children and were also detected in many seronegative children, indicating pre-existing cross-reactive responses to seasonal coronaviruses. Importantly, children retained antibody and cellular responses 6 months after infection, whereas relative waning occurred in adults. Spike-specific responses were also broadly stable beyond 12 months. Therefore, children generate robust, cross-reactive and sustained immune responses to SARS-CoV-2 with focused specificity for the spike protein. These findings provide insight into the relative clinical protection that occurs in most children and might help to guide the design of pediatric vaccination regimens.


Subject(s)
Antibodies, Viral/immunology , Coronavirus 229E, Human/immunology , Coronavirus OC43, Human/immunology , Cross Protection/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Adaptive Immunity/immunology , Adult , Antibodies, Neutralizing/immunology , COVID-19/immunology , COVID-19 Vaccines/immunology , Child , Child, Preschool , Cross Reactions/immunology , Humans
4.
Front Immunol ; 12: 793953, 2021.
Article in English | MEDLINE | ID: covidwho-1572289

ABSTRACT

Durability of SARS-CoV-2 Spike antibody responses after infection provides information relevant to understanding protection against COVID-19 in humans. We report the results of a sequential evaluation of anti-SARS-CoV-2 antibodies in convalescent patients with a median follow-up of 14 months (range 12.4-15.4) post first symptom onset. We report persistence of antibodies for all four specificities tested [Spike, Spike Receptor Binding Domain (Spike-RBD), Nucleocapsid, Nucleocapsid RNA Binding Domain (N-RBD)]. Anti-Spike antibodies persist better than anti-Nucleocapsid antibodies. The durability analysis supports a bi-phasic antibody decay with longer half-lives of antibodies after 6 months and antibody persistence for up to 14 months. Patients infected with the Wuhan (WA1) strain maintained strong cross-reactive recognition of Alpha and Delta Spike-RBD but significantly reduced binding to Beta and Mu Spike-RBD. Sixty percent of convalescent patients with detectable WA1-specific NAb also showed strong neutralization of the Delta variant, the prevalent strain of the present pandemic. These data show that convalescent patients maintain functional antibody responses for more than one year after infection, suggesting a strong long-lasting response after symptomatic disease that may offer a prolonged protection against re-infection. One patient from this cohort showed strong increase of both Spike and Nucleocapsid antibodies at 14 months post-infection indicating SARS-CoV-2 re-exposure. These antibodies showed stronger cross-reactivity to a panel of Spike-RBD including Beta, Delta and Mu and neutralization of a panel of Spike variants including Beta and Gamma. This patient provides an example of strong anti-Spike recall immunity able to control infection at an asymptomatic level. Together, the antibodies from SARS-CoV-2 convalescent patients persist over 14 months and continue to maintain cross-reactivity to the current variants of concern and show strong functional properties.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Adult , Aged , Antibodies, Neutralizing/metabolism , Antibodies, Viral/metabolism , Binding Sites, Antibody/immunology , COVID-19/virology , Cohort Studies , Cross Reactions/immunology , Female , Humans , Male , Middle Aged , Neutralization Tests/methods , Nucleocapsid/immunology , Nucleocapsid/metabolism , Protein Binding/immunology , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/metabolism , Time Factors
5.
J Med Virol ; 93(12): 6544-6550, 2021 12.
Article in English | MEDLINE | ID: covidwho-1544302

ABSTRACT

We developed a rapid and simple magnetic chemiluminescence enzyme immunoassay on the Real Express-6 analyzer, which could simultaneously detect immunoglobulin G and immunoglobulin M antibodies against SARS-CoV-2 virus in human blood within 18 min, and which could be used to detect clinical studies to verify its clinical efficacy. We selected blood samples from 185 COVID-19 patients confirmed by polymerase chain reaction and 271 negative patients to determine the clinical detection sensitivity, specificity, stability, and precision of this method. Meanwhile, we also surveyed the dynamic variance of viral antibodies during SARS-CoV-2 infection. This rapid immunoassay test has huge potential benefits for rapid screening of SARS-CoV-2 infection and may help clinical drug and vaccine development.


Subject(s)
Antibodies, Viral/blood , COVID-19 Serological Testing/methods , COVID-19/diagnosis , Immunoglobulin G/blood , Immunoglobulin M/blood , SARS-CoV-2/immunology , Adolescent , Adult , Aged , Aged, 80 and over , Cross Reactions/immunology , Early Diagnosis , Female , Humans , Immunoassay/methods , Luminescent Measurements , Male , Mass Screening/methods , Middle Aged , Polymerase Chain Reaction , Sensitivity and Specificity , Young Adult
6.
Microbiol Spectr ; 9(2): e0141621, 2021 10 31.
Article in English | MEDLINE | ID: covidwho-1495015

ABSTRACT

The rapid worldwide spread of SARS-CoV-2 has accelerated research and development for controlling the COVID-19 pandemic. A multi-coronavirus protein microarray was created containing full-length proteins, overlapping protein fragments of various lengths, and peptide libraries from SARS-CoV-2 and four other human coronaviruses. Sera from confirmed COVID-19 patients as well as unexposed individuals were applied to multicoronavirus arrays to identify specific antibody reactivity. High-level IgG, IgM, and IgA reactivity to structural proteins S, M, and N of SARS-CoV-2, as well as accessory proteins such as ORF3a and ORF7a, were observed that were specific to COVID-19 patients. Antibody reactivity against overlapping 100-, 50-, and 30-amino acid fragments of SARS-CoV-2 proteins was used to identify antigenic regions. Numerous proteins of SARS-CoV, Middle East respiratory syndrome coronavirus (MERS-CoV), and the endemic human coronaviruses HCoV-NL63 and HCoV-OC43 were also more reactive with IgG, IgM, and IgA in COVID-19 patient sera than in unexposed control sera, providing further evidence of immunologic cross-reactivity between these viruses. Whereas unexposed individuals had minimal reactivity against SARS-CoV-2 proteins that poorly correlated with reactivity against HCoV-NL63 and HCoV-OC43 S2 and N proteins, COVID-19 patient sera had higher correlation between SARS-CoV-2 and HCoV responses, suggesting that de novo antibodies against SARS-CoV-2 cross-react with HCoV epitopes. Array responses were compared with validated spike protein-specific IgG enzyme-linked immunosorbent assays (ELISAs), showing agreement between orthologous methods. SARS-CoV-2 microneutralization titers were low in the COVID-19 patient sera but correlated with array responses against S and N proteins. The multi-coronavirus protein microarray is a useful tool for mapping antibody reactivity in COVID-19 patients. IMPORTANCE With novel mutant SARS-CoV-2 variants of concern on the rise, knowledge of immune specificities against SARS-CoV-2 proteins is increasingly important for understanding the impact of structural changes in antibody-reactive protein epitopes on naturally acquired and vaccine-induced immunity, as well as broader topics of cross-reactivity and viral evolution. A multi-coronavirus protein microarray used to map the binding of COVID-19 patient antibodies to SARS-CoV-2 proteins and protein fragments as well as to the proteins of four other coronaviruses that infect humans has shown specific regions of SARS-CoV-2 proteins that are highly reactive with patient antibodies and revealed cross-reactivity of these antibodies with other human coronaviruses. These data and the multi-coronavirus protein microarray tool will help guide further studies of the antibody response to COVID-19 and to vaccination against this worldwide pandemic.


Subject(s)
Antibodies, Viral/immunology , Coronavirus NL63, Human/immunology , Coronavirus OC43, Human/immunology , Epitopes/immunology , Middle East Respiratory Syndrome Coronavirus/immunology , SARS-CoV-2/immunology , Antibodies, Viral/blood , Binding Sites, Antibody/immunology , COVID-19/immunology , Coronavirus Nucleocapsid Proteins/immunology , Cross Reactions/immunology , Enzyme-Linked Immunosorbent Assay , Humans , Immunoglobulin A/immunology , Immunoglobulin G/immunology , Immunoglobulin M/immunology , Phosphoproteins/immunology , Protein Array Analysis , Spike Glycoprotein, Coronavirus/immunology , Viral Proteins/immunology , Viroporin Proteins/immunology
7.
Sci Rep ; 11(1): 20363, 2021 10 13.
Article in English | MEDLINE | ID: covidwho-1467138

ABSTRACT

COVID-19 diagnostics was quickly ramped up worldwide early 2020 based on the detection of viral RNA. However, based on the scientific knowledge for pre-existing coronaviruses, it was expected that the SARS-CoV-2 RNA will be detected from symptomatic and at significant rates also from asymptomatic individuals due to persistence of non-infectious RNA. To increase the efficacy of diagnostics, surveillance, screening and pandemic control, rapid methods, such as antigen tests, are needed for decentralized testing and to assess infectiousness. A novel automated mariPOC SARS-CoV-2 test was developed for the detection of conserved structural viral nucleocapsid proteins. The test utilizes sophisticated optical laser technology for two-photon excitation and individual detection of immunoassay solid-phase particles. We validated the new method against qRT-PCR. Sensitivity of the test was 100.0% (13/13) directly from nasopharyngeal swab specimens and 84.4% (38/45) from swab specimens in undefined transport mediums. Specificity of the test was 100.0% (201/201). The test's limit of detection was 2.7 TCID50/test. It showed no cross-reactions. Our study shows that the new test can detect infectious individuals already in 20 min with clinical sensitivity close to qRT-PCR. The mariPOC is a versatile platform for syndromic testing and for high capacity infection control screening of infectious individuals.


Subject(s)
COVID-19 Serological Testing/methods , COVID-19/diagnosis , Adult , Aged , Antigens, Viral/analysis , COVID-19/immunology , Cross Reactions/immunology , Female , Finland/epidemiology , Humans , Immunoassay/methods , Male , Middle Aged , Nasopharynx/virology , RNA, Viral/genetics , Reproducibility of Results , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Sensitivity and Specificity
8.
Front Immunol ; 12: 727989, 2021.
Article in English | MEDLINE | ID: covidwho-1450808

ABSTRACT

Background: A growing number of experiments have suggested potential cross-reactive immunity between severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and previous human coronaviruses. We conducted the present retrospective cohort study to investigate the relationship between previous Middle East respiratory syndrome-coronavirus (MERS-CoV) infection and the risk of SARS-CoV-2 infection as well as the relationship between previous MERS-CoV and COVID-19-related hospitalization and mortality. Methods: Starting in March 2020, we prospectively followed two groups of individuals who tested negative for COVID-19 infection. The first group had a previously confirmed MERS-CoV infection, which was compared to a control group of MERS-negative individuals. The studied cohort was then followed until November 2020 to track evidence of contracting COVID-19 infection. Findings: A total of 82 (24%) MERS-positive and 260 (31%) MERS-negative individuals had COVID-19 infection. Patients in the MERS-positive group had a lower risk of COVID-19 infection than those in the MERS-negative group (Risk ratio [RR] 0.696, 95% confidence interval [CI] 0.522-0.929; p =0.014). The risk of COVID-19-related hospitalization in the MERS-positive group was significantly higher (RR 4.036, 95% CI 1.705-9.555; p =0.002). The case fatality rate (CFR) from COVID-19 was 4.9% in the MERS-positive group and 1.2% in the MERS-negative group (p =0.038). The MERS-positive group had a higher risk of death than the MERS-negative group (RR 6.222, 95% CI 1.342-28.839; p =0.019). However, the risk of mortality was similar between the two groups when death was adjusted for age (p =0.068) and age and sex (p =0.057). After controlling for all the independent variables, only healthcare worker occupation and >1 comorbidity were independent predictors of SARS-CoV-2 infection. Interpretation: Individuals with previous MERS-CoV infection can exhibit a cross-reactive immune response to SARS-CoV-2 infection. Our study demonstrated that patients with MERS-CoV infection had higher risks of COVID-19-related hospitalization and death than MERS-negative individuals.


Subject(s)
COVID-19/epidemiology , COVID-19/mortality , Cross Reactions/immunology , Middle East Respiratory Syndrome Coronavirus/immunology , SARS-CoV-2/immunology , Adolescent , Adult , Aged , Aged, 80 and over , Child , Child, Preschool , Female , Hospitalization/statistics & numerical data , Humans , Infant , Infant, Newborn , Male , Middle Aged , Prospective Studies , Retrospective Studies , Saudi Arabia/epidemiology , Young Adult
9.
PLoS Pathog ; 17(9): e1009958, 2021 09.
Article in English | MEDLINE | ID: covidwho-1440996

ABSTRACT

Cross-reactive epitopes (CREs) are similar epitopes on viruses that are recognized or neutralized by same antibodies. The S protein of SARS-CoV-2, similar to type I fusion proteins of viruses such as HIV-1 envelope (Env) and influenza hemagglutinin, is heavily glycosylated. Viral Env glycans, though host derived, are distinctly processed and thereby recognized or accommodated during antibody responses. In recent years, highly potent and/or broadly neutralizing human monoclonal antibodies (bnAbs) that are generated in chronic HIV-1 infections have been defined. These bnAbs exhibit atypical features such as extensive somatic hypermutations, long complementary determining region (CDR) lengths, tyrosine sulfation and presence of insertions/deletions, enabling them to effectively neutralize diverse HIV-1 viruses despite extensive variations within the core epitopes they recognize. As some of the HIV-1 bnAbs have evolved to recognize the dense viral glycans and cross-reactive epitopes (CREs), we assessed if these bnAbs cross-react with SARS-CoV-2. Several HIV-1 bnAbs showed cross-reactivity with SARS-CoV-2 while one HIV-1 CD4 binding site bnAb, N6, neutralized SARS-CoV-2. Furthermore, neutralizing plasma antibodies of chronically HIV-1 infected children showed cross neutralizing activity against SARS-CoV-2 pseudoviruses. Collectively, our observations suggest that human monoclonal antibodies tolerating extensive epitope variability can be leveraged to neutralize pathogens with related antigenic profile.


Subject(s)
Broadly Neutralizing Antibodies/immunology , HIV Antibodies/immunology , HIV-1/immunology , SARS-CoV-2/immunology , Antibodies, Monoclonal/immunology , COVID-19/immunology , Cross Reactions/immunology , Humans , Plasma/immunology
10.
Cell Rep ; 37(2): 109823, 2021 10 12.
Article in English | MEDLINE | ID: covidwho-1433047

ABSTRACT

Although both infections and vaccines induce memory B cell (MBC) populations that participate in secondary immune responses, the MBCs generated in each case can differ. Here, we compare SARS-CoV-2 spike receptor binding domain (S1-RBD)-specific primary MBCs that form in response to infection or a single mRNA vaccination. Both primary MBC populations have similar frequencies in the blood and respond to a second S1-RBD exposure by rapidly producing plasmablasts with an abundant immunoglobulin (Ig)A+ subset and secondary MBCs that are mostly IgG+ and cross-react with the B.1.351 variant. However, infection-induced primary MBCs have better antigen-binding capacity and generate more plasmablasts and secondary MBCs of the classical and atypical subsets than do vaccine-induced primary MBCs. Our results suggest that infection-induced primary MBCs have undergone more affinity maturation than vaccine-induced primary MBCs and produce more robust secondary responses.


Subject(s)
COVID-19 Vaccines/immunology , Plasma Cells/immunology , SARS-CoV-2/immunology , Adult , Animals , Antibodies, Viral/immunology , B-Lymphocyte Subsets/immunology , B-Lymphocytes/immunology , COVID-19/immunology , COVID-19/metabolism , Cross Reactions/immunology , Female , HEK293 Cells , Humans , Immunization/methods , Immunologic Memory , Male , Mice , Mice, Inbred C57BL , Middle Aged , RNA, Messenger/immunology , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism , Vaccination/methods , Vaccines/immunology
11.
Emerg Microbes Infect ; 10(1): 1931-1946, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1429140

ABSTRACT

Identification of relevant epitopes is crucial for the development of subunit peptide vaccines inducing neutralizing and cellular immunity against SARS-CoV-2. Our aim was the characterization of epitopes in the receptor-binding domain (RBD) of SARS-CoV-2 spike (S) protein to generate a peptide vaccine. Epitope mapping using a panel of 10 amino acid overlapped 15-mer peptides covering region 401-515 from RBD did not identify linear epitopes when tested with sera from infected individuals or from RBD-immunized mice. However, immunization of mice with these 15-mer peptides identified four peptides located at region 446-480 that induced antibodies recognizing the peptides and RBD/S1 proteins. Immunization with peptide 446-480 from S protein formulated with Freund's adjuvant or with CpG oligodeoxinucleotide/Alum induced polyepitopic antibody responses in BALB/c and C56BL/6J mice, recognizing RBD (titres of 3 × 104-3 × 105, depending on the adjuvant) and displaying neutralizing capacity (80-95% inhibition capacity; p < 0.05) against SARS-CoV-2. Murine CD4 and CD8T-cell epitopes were identified in region 446-480 and vaccination experiments using HLA transgenic mice suggested the presence of multiple human T-cell epitopes. Antibodies induced by peptide 446-480 showed broad recognition of S proteins and S-derived peptides belonging to SARS-CoV-2 variants of concern. Importantly, vaccination with peptide 446-480 or with a cyclic version of peptide 446-488 containing a disulphide bridge between cysteines 480 and 488, protected humanized K18-hACE2 mice from a lethal dose of SARS-CoV-2 (62.5 and 75% of protection; p < 0.01 and p < 0.001, respectively). This region could be the basis for a peptide vaccine or other vaccine platforms against Covid-19.


Subject(s)
Antibodies, Neutralizing/immunology , COVID-19 Vaccines/immunology , COVID-19/prevention & control , Immunity, Cellular , Immunity, Humoral , SARS-CoV-2/immunology , Adjuvants, Immunologic/administration & dosage , Animals , Antibodies, Neutralizing/blood , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , COVID-19/immunology , COVID-19 Vaccines/standards , Cross Reactions/immunology , Epitope Mapping , Epitopes, B-Lymphocyte , Epitopes, T-Lymphocyte/immunology , Humans , Immunization , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Transgenic , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , Vaccines, Subunit/immunology , Vaccines, Synthetic/immunology
13.
Front Immunol ; 12: 635942, 2021.
Article in English | MEDLINE | ID: covidwho-1389176

ABSTRACT

SARS-CoV-2 infection takes a mild or clinically inapparent course in the majority of humans who contract this virus. After such individuals have cleared the virus, only the detection of SARS-CoV-2-specific immunological memory can reveal the exposure, and hopefully the establishment of immune protection. With most viral infections, the presence of specific serum antibodies has provided a reliable biomarker for the exposure to the virus of interest. SARS-CoV-2 infection, however, does not reliably induce a durable antibody response, especially in sub-clinically infected individuals. Consequently, it is plausible for a recently infected individual to yield a false negative result within only a few months after exposure. Immunodiagnostic attention has therefore shifted to studies of specific T cell memory to SARS-CoV-2. Most reports published so far agree that a T cell response is engaged during SARS-CoV-2 infection, but they also state that in 20-81% of SARS-CoV-2-unexposed individuals, T cells respond to SARS-CoV-2 antigens (mega peptide pools), allegedly due to T cell cross-reactivity with Common Cold coronaviruses (CCC), or other antigens. Here we show that, by introducing irrelevant mega peptide pools as negative controls to account for chance cross-reactivity, and by establishing the antigen dose-response characteristic of the T cells, one can clearly discern between cognate T cell memory induced by SARS-CoV-2 infection vs. cross-reactive T cell responses in individuals who have not been infected with SARS-CoV-2.


Subject(s)
COVID-19/immunology , COVID-19/virology , Host-Pathogen Interactions/immunology , SARS-CoV-2/immunology , T-Lymphocytes/immunology , Antigens, Viral/immunology , Biomarkers , COVID-19/metabolism , Cross Reactions/immunology , Cytokines/metabolism , Epitopes, T-Lymphocyte/immunology , Humans , Immunodominant Epitopes/immunology , Immunologic Memory , Peptides/immunology , Protein Binding , Spike Glycoprotein, Coronavirus/immunology , T-Lymphocytes/metabolism
14.
Nature ; 584(7821): 457-462, 2020 08.
Article in English | MEDLINE | ID: covidwho-1373437

ABSTRACT

Memory T cells induced by previous pathogens can shape susceptibility to, and the clinical severity of, subsequent infections1. Little is known about the presence in humans of pre-existing memory T cells that have the potential to recognize severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here we studied T cell responses against the structural (nucleocapsid (N) protein) and non-structural (NSP7 and NSP13 of ORF1) regions of SARS-CoV-2 in individuals convalescing from coronavirus disease 2019 (COVID-19) (n = 36). In all of these individuals, we found CD4 and CD8 T cells that recognized multiple regions of the N protein. Next, we showed that patients (n = 23) who recovered from SARS (the disease associated with SARS-CoV infection) possess long-lasting memory T cells that are reactive to the N protein of SARS-CoV 17 years after the outbreak of SARS in 2003; these T cells displayed robust cross-reactivity to the N protein of SARS-CoV-2. We also detected SARS-CoV-2-specific T cells in individuals with no history of SARS, COVID-19 or contact with individuals who had SARS and/or COVID-19 (n = 37). SARS-CoV-2-specific T cells in uninfected donors exhibited a different pattern of immunodominance, and frequently targeted NSP7 and NSP13 as well as the N protein. Epitope characterization of NSP7-specific T cells showed the recognition of protein fragments that are conserved among animal betacoronaviruses but have low homology to 'common cold' human-associated coronaviruses. Thus, infection with betacoronaviruses induces multi-specific and long-lasting T cell immunity against the structural N protein. Understanding how pre-existing N- and ORF1-specific T cells that are present in the general population affect the susceptibility to and pathogenesis of SARS-CoV-2 infection is important for the management of the current COVID-19 pandemic.


Subject(s)
Betacoronavirus/immunology , Coronavirus Infections/immunology , Pneumonia, Viral/immunology , Severe Acute Respiratory Syndrome/immunology , T-Lymphocytes/immunology , Betacoronavirus/chemistry , COVID-19 , Case-Control Studies , Coronavirus Infections/virology , Coronavirus Nucleocapsid Proteins , Cross Reactions/immunology , Humans , Immunodominant Epitopes/immunology , Nucleocapsid Proteins/chemistry , Nucleocapsid Proteins/immunology , Pandemics , Phosphoproteins , Pneumonia, Viral/virology , SARS-CoV-2
15.
mBio ; 12(4): e0141521, 2021 08 31.
Article in English | MEDLINE | ID: covidwho-1370889

ABSTRACT

Severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) possesses a discriminative polybasic cleavage motif in its spike protein that is recognized by the host furin protease. Proteolytic cleavage activates the spike protein, thereby affecting both the cellular entry pathway and cell tropism of SARS-CoV-2. Here, we investigated the impact of the furin cleavage site on viral growth and pathogenesis using a hamster animal model infected with SARS-CoV-2 variants bearing mutations at the furin cleavage site (S gene mutants). In the airway tissues of hamsters, the S gene mutants exhibited low growth properties. In contrast to parental pathogenic SARS-CoV-2, hamsters infected with the S gene mutants showed no body weight loss and only a mild inflammatory response, thereby indicating the attenuated variant nature of S gene mutants. This transient infection was sufficient for inducing protective neutralizing antibodies that cross-react with different SARS-CoV-2 lineages. Consequently, hamsters inoculated with S gene mutants showed resistance to subsequent infection with both the parental strain and the currently emerging SARS-CoV-2 variants belonging to lineages B.1.1.7 and P.1. Taken together, our findings revealed that the loss of the furin cleavage site causes attenuation in the airway tissues of hamsters and highlighted the potential benefits of S gene mutants as potential immunogens. IMPORTANCE SARS-CoV-2 uses its spike protein to enter target cells. The spike protein is cleaved by a host protease, and this event facilitates viral entry and broadens cell tropism. In this study, we employed SARS-CoV-2 mutants lacking the S protein cleavage site and characterized their growth and pathogenicity using hamsters, a laboratory animal model for SARS-CoV-2 infection. These mutants exerted low pathogenicity but induced sufficient levels of neutralizing antibodies in hamsters, which protected hamsters from rechallenge with pathogenic clinical SARS-CoV-2 strains. These virus mutants may be used as protective immunogens against SARS-CoV-2 infection.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/pathology , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/genetics , Animals , Cell Line , Chlorocebus aethiops , Cross Reactions/immunology , Furin/metabolism , Humans , SARS-CoV-2/genetics , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Vaccines, Attenuated/immunology , Vero Cells , Virulence/genetics
16.
Immunity ; 54(10): 2385-2398.e10, 2021 10 12.
Article in English | MEDLINE | ID: covidwho-1370548

ABSTRACT

Potent neutralizing SARS-CoV-2 antibodies often target the spike protein receptor-binding site (RBS), but the variability of RBS epitopes hampers broad neutralization of multiple sarbecoviruses and drifted viruses. Here, using humanized mice, we identified an RBS antibody with a germline VH gene that potently neutralized SARS-related coronaviruses, including SARS-CoV and SARS-CoV-2 variants. X-ray crystallography revealed coordinated recognition by the heavy chain of non-RBS conserved sites and the light chain of RBS with a binding angle mimicking the angiotensin-converting enzyme 2 (ACE2) receptor. The minimum footprints in the hypervariable region of RBS contributed to the breadth of neutralization, which was enhanced by immunoglobulin G3 (IgG3) class switching. The coordinated binding resulted in broad neutralization of SARS-CoV and emerging SARS-CoV-2 variants of concern. Low-dose therapeutic antibody treatment in hamsters reduced the virus titers and morbidity during SARS-CoV-2 challenge. The structural basis for broad neutralizing activity may inform the design of a broad spectrum of therapeutics and vaccines.


Subject(s)
Broadly Neutralizing Antibodies/immunology , Cross Reactions/immunology , SARS-CoV-2/immunology , Animals , Betacoronavirus/immunology , Binding Sites, Antibody , Broadly Neutralizing Antibodies/chemistry , Broadly Neutralizing Antibodies/therapeutic use , COVID-19/prevention & control , COVID-19/therapy , COVID-19/virology , Cricetinae , Humans , Immunoglobulin Class Switching , Immunoglobulin Fab Fragments/chemistry , Immunoglobulin Fab Fragments/metabolism , Immunoglobulin G/chemistry , Immunoglobulin G/immunology , Mice , Protein Domains , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism
17.
Front Immunol ; 12: 696370, 2021.
Article in English | MEDLINE | ID: covidwho-1357528

ABSTRACT

The COVID-19 pandemic is caused by SARS-CoV-2, a novel zoonotic coronavirus. Emerging evidence indicates that preexisting humoral immunity against other seasonal human coronaviruses (HCoVs) plays a critical role in the specific antibody response to SARS-CoV-2. However, current work to assess the effects of preexisting and cross-reactive anti-HCoVs antibodies has been limited. To address this issue, we have adapted our previously reported multiplex assay to simultaneously and quantitatively measure anti-HCoV antibodies. The full mPlex-CoV panel covers the spike (S) and nucleocapsid (N) proteins of three highly pathogenic HCoVs (SARS-CoV-1, SARS-CoV-2, MERS) and four human seasonal strains (OC43, HKU1, NL63, 229E). Combining this assay with volumetric absorptive microsampling (VAMS), we measured the anti-HCoV IgG, IgA, and IgM antibodies in fingerstick blood samples. The results demonstrate that the mPlex-CoV assay has high specificity and sensitivity. It can detect strain-specific anti-HCoV antibodies down to 0.1 ng/ml with 4 log assay range and with low intra- and inter-assay coefficients of variation (%CV). We also estimate multiple strain HCoVs IgG, IgA and IgM concentration in VAMS samples in three categories of subjects: pre-COVID-19 (n=21), post-COVID-19 convalescents (n=19), and COVID-19 vaccine recipients (n=14). Using metric multidimensional scaling (MDS) analysis, HCoVs IgG concentrations in fingerstick blood samples were well separated between the pre-COVID-19, post-COVID-19 convalescents, and COVID-19 vaccine recipients. In addition, we demonstrate how multi-dimensional scaling analysis can be used to visualize IgG mediated antibody immunity against multiple human coronaviruses. We conclude that the combination of VAMS and the mPlex-Cov assay is well suited to performing remote study sample collection under pandemic conditions to monitor HCoVs antibody responses in population studies.


Subject(s)
Antibodies, Viral/blood , Coronavirus/immunology , Cross Reactions/immunology , Immunoassay/methods , Antibodies, Viral/immunology , Betacoronavirus/immunology , COVID-19/immunology , Coronavirus 229E, Human/immunology , Coronavirus NL63, Human/immunology , Coronavirus Nucleocapsid Proteins/immunology , Coronavirus OC43, Human/immunology , Humans , Immunoglobulin A/blood , Immunoglobulin A/immunology , Immunoglobulin G/blood , Immunoglobulin G/immunology , Immunoglobulin M/blood , Immunoglobulin M/immunology , SARS Virus/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology
18.
Int Immunopharmacol ; 99: 108020, 2021 Oct.
Article in English | MEDLINE | ID: covidwho-1330896

ABSTRACT

The spike protein of the SARS-CoV-2 virus is the foremost target for the designing of vaccines and therapeutic antibodies and also acts as a crucial antigen in the assessment of COVID-19 immune responses. The enveloped viruses; such as SARS-CoV-2, Human Immunodeficiency Virus-1 (HIV-1) and influenza, often hijack host-cell glycosylation pathways and influence pathobiology and immune selection. These glycan motifs can lead to either immune evasion or viral neutralization by the production of cross-reactive antibodies that can lead to antibody-dependent enhancement (ADE) of infection. Potential cross-protection from influenza vaccine has also been reported in COVID-19 infected individuals in several epidemiological studies recently; however, the scientific basis for these observations remains elusive. Herein, we show that the anti-SARS-CoV2 antibodies cross-reacts with the Hemagglutinin (HA) protein. This phenomenon is common to both the sera from convalescent SARS-CoV-2 donors and spike immunized mice, although these antibodies were unable to cross-neutralize, suggesting the presence of a non-neutralizing antibody response. Epitope mapping suggests that the cross-reactive antibodies are targeted towards glycan epitopes of the SARS-CoV-2 spike and HA. Overall, our findings address the cross-reactive responses, although non-neutralizing, elicited against RNA viruses and warrant further studies to investigate whether such non-neutralizing antibody responses can contribute to effector functions such as antibody-dependent cellular cytotoxicity (ADCC) or ADE.


Subject(s)
COVID-19/immunology , Cross Reactions/immunology , Hemagglutinin Glycoproteins, Influenza Virus/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Neutralizing , Antigen-Antibody Reactions , Binding Sites, Antibody/immunology , Cell Culture Techniques , Chlorocebus aethiops , Dogs , Epitope Mapping , Epitopes/immunology , Glycosylation , Humans , Influenza Vaccines/immunology , Madin Darby Canine Kidney Cells , Membrane Glycoproteins/metabolism , Mice , Mice, Inbred C57BL , SARS-CoV-2/immunology , Vero Cells
19.
Expert Rev Vaccines ; 20(9): 1051-1057, 2021 09.
Article in English | MEDLINE | ID: covidwho-1327291

ABSTRACT

INTRODUCTION: The COVID-19 pandemic is a globalized health concern caused by a beta-coronavirus named Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Since December 2019, when this outbreak flared in Wuhan, China, COVID-19 cases have been continuously rising all over the world. Due to the emergence of SARS-CoV-2 mutants, subsequent waves are flowing in a faster manner as compared to the primary wave, which is more contagious and causing higher mortality. Recently, India has emerged as the new epicenter of the second wave by mutants of SARS-CoV-2. After almost eighteen months of this outbreak, some COVID-19 dedicated therapeutics and vaccines are available, and a few are under trial, but the situation is still uncontrolled. AREA COVERED: This perspective article covers the repurposing of childhood vaccines like Bacille Calmette-Guerin (BCG), Measles, Mumps, Rubella (MMR), and Oral Polio Vaccine (OPV), which are live attenuated vaccines and have been shown the protective effect through 'trained immunity and 'crossreactivity.' EXPERT OPINION: This perspective article has suggested that combinatorial use of these childhood vaccines might exert a better protective effect along with the available COVID-19 therapeutic and vaccines which could be considered as a preventive option against SARS-CoV-2 infection as well as its subsequent waves.


Subject(s)
BCG Vaccine/immunology , COVID-19 Vaccines/immunology , COVID-19/prevention & control , Drug Repositioning/methods , SARS-CoV-2/immunology , Vaccines, Attenuated/immunology , Cross Reactions/immunology , Diphtheria-Tetanus-Pertussis Vaccine/immunology , Humans , Immunity, Innate/drug effects , Immunity, Innate/immunology , Measles-Mumps-Rubella Vaccine/immunology , Poliovirus Vaccine, Oral/immunology , Spike Glycoprotein, Coronavirus/immunology , Vaccination , Yellow Fever Vaccine/immunology
20.
J Immunol Methods ; 497: 113104, 2021 10.
Article in English | MEDLINE | ID: covidwho-1322209

ABSTRACT

Mitigation of the COVID-19 pandemic requires an understanding of the antibody response to SARS-CoV-2. However, throughout the development of SARS-CoV-2 IgG antibody assays during the past year, cross-reactivity to other coronaviruses remained a question. To address these issues, we evaluated IgG in COVID-19 convalescent plasma samples for reactivity against three SARS-CoV-2 antigens including full-length spike, receptor binding domain, and the proximal extracellular fusion domain, and spike antigens from other coronaviruses (SARS-CoV, MERS-CoV, hCoV-HKU1, hCoV-OC43, hCoV-229E and hCoV-NL63) using the VaxArray Coronavirus SeroAssay which is a multiplexed antigen assay developed by InDevR Inc. These results were compared to two commercial SARS-CoV-2 IgG ELISAs targeting either the SARS-CoV-2 nucleocapsid or spike antigens and a live virus focus reduction neutralizing antibody test (FRNT). The VaxArray platform showed high specificity for detection of SARS-CoV-2 IgG, evident from lack of reactivity to SARS-CoV-2 antigens despite significant reactivity to endemic coronavirus antigens in pre-pandemic samples. SARS-CoV-2 IgG positive samples reacted weakly to SARS-CoV spike but not to MERS-CoV. While the VaxArray platform had overall comparable results to the spike and nucleocapsid IgG ELISAs, results were more similar to the spike antigen ELISA and the platform displayed a higher sensitivity and specificity than both ELISAs. Samples with FRNT titers below 1/23 reported negative on VaxArray, while positive samples on VaxArray had significantly higher neutralizing antibody titers. These results suggest that the VaxArray Coronavirus SeroAssay performs with high sensitivity and specificity for the detection of SARS-CoV-2 IgG, and positive results on the platform indicate SARS-CoV-2 neutralizing activity.


Subject(s)
Antibodies, Viral/immunology , Antigens, Viral/immunology , COVID-19/diagnosis , Immunoassay/methods , Immunoglobulin G/immunology , SARS-CoV-2/immunology , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/immunology , Antibodies, Viral/blood , COVID-19/epidemiology , COVID-19/virology , Cross Reactions/immunology , Enzyme-Linked Immunosorbent Assay/methods , Humans , Immunoglobulin G/blood , Pandemics/prevention & control , Reproducibility of Results , SARS-CoV-2/physiology , Sensitivity and Specificity , Spike Glycoprotein, Coronavirus/immunology
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