scholarly journals Serological profiles of pan-coronavirus-specific responses in COVID-19 patients using a multiplexed electro-chemiluminescence-based testing platform

PLoS ONE ◽  
2021 ◽  
Vol 16 (6) ◽  
pp. e0252628
Author(s):  
Sidhartha Chaudhury ◽  
Jack Hutter ◽  
Jessica S. Bolton ◽  
Shilpa Hakre ◽  
Evelyn Mose ◽  
...  
Keyword(s):  
Nucleocapsid Protein ◽  
Cross Reactivity ◽  
Multiplex Assay ◽  
Spike Protein ◽  
Contact Tracing ◽  
Epitope Specificity ◽  
Army Hospital ◽  
Protein Receptor ◽  
Healthcare Administrators ◽  
Human Coronaviruses

Serological assessment of SARS-CoV-2 specific responses are an essential tool for determining the prevalence of past SARS-CoV-2 infections in the population especially when testing occurs after symptoms have developed and limited contact tracing is in place. The goal of our study was to test a new 10-plex electro-chemiluminescence-based assay to measure IgM and IgG responses to the spike proteins from multiple human coronaviruses including SARS-CoV-2, assess the epitope specificity of the SARS-CoV-2 antibody response against full-length spike protein, receptor-binding domain and N-terminal domain of the spike protein, and the nucleocapsid protein. We carried out the assay on samples collected from three sample groups: subjects diagnosed with COVID-19 from the U.S. Army hospital at Camp Humphreys in Pyeongtaek, South Korea; healthcare administrators from the same hospital but with no reported diagnosis of COVID-19; and pre-pandemic samples. We found that the new CoV-specific multiplex assay was highly sensitive allowing plasma samples to be diluted 1:30,000 with a robust signal. The reactivity of IgG responses to SARS-CoV-2 nucleocapsid protein and IgM responses to SARS-CoV-2 spike protein could distinguish COVID-19 samples from non-COVID-19 and pre-pandemic samples. The data from the three sample groups also revealed a unique pattern of cross-reactivity between SARS-CoV-2 and SARS-CoV-1, MERS-CoV, and seasonal coronaviruses HKU1 and OC43. Our findings show that the CoV-2 IgM response is highly specific while the CoV-2 IgG response is more cross-reactive across a range of human CoVs and also showed that IgM and IgG responses show distinct patterns of epitope specificity. In summary, this multiplex assay was able to distinguish samples by COVID-19 status and characterize distinct trends in terms of cross-reactivity and fine-specificity in antibody responses, underscoring its potential value in diagnostic or serosurveillance efforts.

scholarly journals Serological profiles of pan-coronavirus-specific responses in COVID-19 patients using a multiplexed electro-chemiluminescence-based testing platform

2021 ◽  
Author(s):  
Sidhartha Chaudhury ◽  
Jack Hutter ◽  
Jessica S Bolton ◽  
Shilpa Hakre ◽  
Evelyn Mose ◽  
...  
Keyword(s):  
Nucleocapsid Protein ◽  
Cross Reactivity ◽  
Multiplex Assay ◽  
Spike Protein ◽  
Contact Tracing ◽  
Epitope Specificity ◽  
Army Hospital ◽  
Protein Receptor ◽  
Healthcare Administrators ◽  
Human Coronaviruses

AbstractSerological assessment of SARS-CoV-2 specific responses are an essential tool for determining the prevalence of past SARS-CoV-2 infections in the population especially when testing occurs after symptoms have developed and limited contact tracing is in place. The goal of our study was to test a new 10-plex electro-chemiluminescence-based assay to measure IgM and IgG responses to the spike proteins from multiple human coronaviruses including SARS-CoV-2, assess the epitope specificity of the SARS-CoV-2 antibody response against full-length spike protein, receptor-binding domain and N-terminal domain of the spike protein, and the nucleocapsid protein. We carried out the assay on samples collected from three sample groups: subjects diagnosed with COVID-19 from the U.S. Army hospital at Camp Humphreys in Pyeongtaek, South Korea; healthcare administrators from the same hospital but with no reported diagnosis of COVID-19; and pre-pandemic samples. We found that the new CoV-specific multiplex assay was highly sensitive allowing plasma samples to be diluted 1:30,000 with a robust signal. The reactivity of IgG responses to SARS-CoV-2 nucleocapsid protein and IgM responses to SARS-CoV-2 spike protein could distinguish COVID-19 samples from non-COVID-19 and pre-pandemic samples. The data from the three sample groups also revealed a unique pattern of cross-reactivity between SARS-CoV-2 and SARS-CoV-1, MERS-CoV, and seasonal coronaviruses HKU1 and OC43. Our findings show that the CoV-2 IgM response is highly specific while the CoV-2 IgG response is more cross-reactive across a range of human CoVs and also showed that IgM and IgG responses show distinct patterns of epitope specificity. In summary, this multiplex assay was able to distinguish samples by COVID-19 status and characterize distinct trends in terms of cross-reactivity and fine-specificity in antibody responses, underscoring its potential value in diagnostic or serosurveillance efforts.

scholarly journals Immunological imprinting of the antibody response in COVID-19 patients

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Teresa Aydillo ◽  
Alexander Rombauts ◽  
Daniel Stadlbauer ◽  
Sadaf Aslam ◽  
Gabriela Abelenda-Alonso ◽  
...  
Keyword(s):  
Immune Response ◽  
Severe Acute Respiratory Syndrome ◽  
Antibody Response ◽  
Humoral Immune Response ◽  
Nucleocapsid Protein ◽  
Spike Protein ◽  
Ongoing Research ◽  
Variable Regions ◽  
Humoral Immune ◽  
Human Coronaviruses

AbstractIn addition to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), humans are also susceptible to six other coronaviruses, for which consecutive exposures to antigenically related and divergent seasonal coronaviruses are frequent. Despite the prevalence of COVID-19 pandemic and ongoing research, the nature of the antibody response against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is unclear. Here we longitudinally profile the early humoral immune response against SARS-CoV-2 in hospitalized coronavirus disease 2019 (COVID-19) patients and quantify levels of pre-existing immunity to OC43, HKU1 and 229E seasonal coronaviruses, and find a strong back-boosting effect to conserved but not variable regions of OC43 and HKU1 betacoronaviruses spike protein. However, such antibody memory boost to human coronaviruses negatively correlates with the induction of IgG and IgM against SARS-CoV-2 spike and nucleocapsid protein. Our findings thus provide evidence of immunological imprinting by previous seasonal coronavirus infections that can potentially modulate the antibody profile to SARS-CoV-2 infection.

scholarly journals Production of specific antibodies against SARS-coronavirus nucleocapsid protein without cross reactivity with human coronaviruses 229E and OC43

2010 ◽  
Vol 11 (2) ◽  
pp. 165 ◽  
Author(s):  
Hyun-Kyoung Lee ◽  
Byoung-Hee Lee ◽  
Seung-Hyeok Seok ◽  
Min-Won Baek ◽  
Hui-Young Lee ◽  
...  
Keyword(s):  
Nucleocapsid Protein ◽  
Cross Reactivity ◽  
Sars Coronavirus ◽  
Specific Antibodies ◽  
Human Coronaviruses

scholarly journals Systematic evaluation of SARS-CoV-2 spike protein derived peptides for diagnosis of COVID-19 patients

2020 ◽  
Author(s):  
Yang Li ◽  
Danyun Lai ◽  
Qing Lei ◽  
Zhaowei Xu ◽  
Hongyan Hou ◽  
...  
Keyword(s):  
Serological Test ◽  
Protein S ◽  
Cross Reactivity ◽  
Spike Protein ◽  
Systematic Evaluation ◽  
Peptide Microarray ◽  
Serological Tests ◽  
S Protein ◽  
Asymptomatic Patients ◽  
Human Coronaviruses

Serological test plays an essential role in monitoring and combating COVID-19 pandemic. Recombinant spike protein (S protein), especially S1 protein is one of the major reagents for serological tests. However, the high cost in production of S protein, and the possible cross-reactivity with other human coronaviruses poses unneglectable challenges. Taking advantage of a peptide microarray of full spike protein coverage, we analyzed 2,434 sera from 858 COVID-19 patients, sera from 63 asymptomatic patients and 610 controls collected from multiple clinical centers. Based on the results of the peptide microarray, we identified several S protein derived 12-mer peptides that have high diagnosis performance. Particularly, for monitoring IgG response, one peptide (aa 1148-1159 or S2-78) has a comparable sensitivity (95.5%, 95% CI 93.7-96.9%) and specificity (96.7%, 95% CI 94.8-98.0%) to that of S1 protein for detection of both COVID-19 patients and asymptomatic infections. Furthermore, the performance of S2-78 IgG for diagnosis was successfully validated by ELISA with an independent sample cohort. By combining S2-78/ S1 with other peptides, a two-step strategy was proposed to ensure both the sensitivity and specificity of S protein based serological assay. The peptide/s identified in this study could be applied independently or in combination with S1 protein for accurate, affordable, and accessible COVID-19 diagnosis.

scholarly journals Identification of Conserved Epitopes in SARS-CoV-2 Spike and Nucleocapsid Protein

2021 ◽  
Vol 23 ◽  
Author(s):  
Sergio Forcelloni ◽  
Anna Benedetti ◽  
Maddalena Dilucca ◽  
Andrea Giansanti
Keyword(s):  
Nucleocapsid Protein ◽  
Rna Binding ◽  
High Rate ◽  
Spike Protein ◽  
Binding Motif ◽  
Homologous Proteins ◽  
Nucleocapsid Proteins ◽  
Rate Of Evolution ◽  
Human Coronaviruses ◽  
Heptad Repeats

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a novel virus that first occurred in Wuhan in December 2019. The spike glycoproteins and nucleocapsid proteins are the most common targets for the development of vaccines and antiviral drugs. Objective: We herein analyze the rate of evolution along with the sequences of spike and nucleocapsid proteins in relation to the spatial locations of their epitopes, previously suggested to contribute to the immune response caused by SARS-CoV-2 infections. Methods: We compare homologous proteins of seven human coronaviruses: HCoV-229E, HCoV-OC43, SARS-CoV, HCoV-NL63, HCoV-HKU1, MERS-CoV, and SARS-CoV-2. We then focus on the local, structural order-disorder propensity of the protein regions where the SARS-CoV-2 epitopes are located. Results : We show that most of nucleocapsid protein epitopes overlap the RNA-binding and dimerization domains, and some of them are characterized by a low rate of evolutions. Similarly, spike protein epitopes are preferentially located in regions that are predicted to be ordered and well-conserved, in correspondence of the heptad repeats 1 and 2. Interestingly, both the receptor-binding motif to ACE2 and the fusion peptide of spike protein are characterized by a high rate of evolution. Conclusion: Our results provide evidence for conserved epitopes that might help develop broad-spectrum SARS-CoV-2 vaccines.

scholarly journals Identification of conserved epitopes in SARS-CoV-2 spike and nucleocapsid protein

2020 ◽  
Author(s):  
Sergio Forcelloni ◽  
Anna Benedetti ◽  
Maddalena Dilucca ◽  
Andrea Giansanti
Keyword(s):  
Nucleocapsid Protein ◽  
Rna Binding ◽  
Spike Protein ◽  
Disease Spread ◽  
Binding Motif ◽  
T Cell Epitopes ◽  
Nucleocapsid Proteins ◽  
Structural Order ◽  
Rates Of Evolution ◽  
Human Coronaviruses

AbstractSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which first occurred in Wuhan (China) in December 2019, is a novel virus that causes a severe acute respiratory disease. The virus spike glycoproteins and nucleocapsid proteins are the main targets for the development of vaccines and antiviral drugs, to control the disease spread. We herein study the structural order-disorder propensity and the rates of evolution of these two proteins to characterize their B cell and T cell epitopes, previously suggested to contribute to immune response caused by SARS-CoV-2 infections. We first analyzed the rates of evolution along the sequences of spike and nucleocapsid proteins in relation to the spatial locations of their epitopes. For this purpose, we compared orthologs from seven human coronaviruses: SARS-CoV-2, SARS-CoV, MERS-CoV, HCoV-229E, HCoV-OC43, HCoV-NL63, and HCoV-HKU1. We then focus on the local, structural order-disorder propensities of the protein regions where the SARS-CoV-2 epitopes are located. We show that the vast majority of nucleocapsid protein epitopes overlap the RNA-binding and dimerization domains and some of them are characterized by low rates of evolutions. Similarly, spike protein epitopes are preferentially located in regions that are predicted to be ordered and well-conserved, in correspondence of the heptad repeats 1 and 2. Interestingly, both the receptor-binding motif to ACE2 and the fusion peptide of spike protein are characterized by high rates of evolution, probably to overcome host immunity. In conclusion, our results provide evidence for conserved epitopes that may help to develop long-lasting, broad-spectrum SARS-CoV-2 vaccines.

scholarly journals Isolation of cross-reactive monoclonal antibodies against divergent human coronaviruses that delineate a conserved and vulnerable site on the spike protein

2020 ◽  
Author(s):  
Chunyan Wang ◽  
Rien van Haperen ◽  
Javier Gutiérrez-Álvarez ◽  
Wentao Li ◽  
Nisreen M.A. Okba ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Distinct Species ◽  
Cross Reactivity ◽  
Spike Protein ◽  
Protective Antibodies ◽  
Therapeutic Measures ◽  
Neutralizing Potential ◽  
Immunoglobulin Repertoire ◽  
Human Coronaviruses ◽  
Virion Surface

AbstractThe coronavirus spike glycoprotein, located on the virion surface, is the key mediator of cell entry. As such, it is an attractive target for the development of protective antibodies and vaccines. Here we describe two human monoclonal antibodies, 1.6C7 and 28D9, that display a remarkable cross-reactivity against distinct species from three Betacoronavirus subgenera, capable of binding the spike proteins of SARS-CoV and SARS-CoV-2, MERS-CoV and the endemic human coronavirus HCoV-OC43. Both antibodies, derived from immunized transgenic mice carrying a human immunoglobulin repertoire, blocked MERS-CoV infection in cells, whereas 28D9 also showed weak cross-neutralizing potential against HCoV-OC43, SARS-CoV and SARS-CoV-2 in a neutralization-sensitive virus pseudotyping system, but not against authentic virus. Both cross-reactive monoclonal antibodies were found to target the stem helix in the spike protein S2 fusion subunit which, in the prefusion conformation of trimeric spike, forms a surface exposed membrane-proximal helical bundle, that is antibody-accessible. We demonstrate that administration of these antibodies in mice protects from a lethal MERS-CoV challenge in both prophylactic and/or therapeutic models. Collectively, these antibodies delineate a conserved, immunogenic and vulnerabe site on the spike protein which spurs the development of broad-range diagnostic, preventive and therapeutic measures against coronaviruses.

scholarly journals Systematic evaluation of IgG responses to SARS-CoV-2 spike protein-derived peptides for monitoring COVID-19 patients

2021 ◽  
Author(s):  
Yang Li ◽  
Dan-yun Lai ◽  
Qing Lei ◽  
Zhao-wei Xu ◽  
Feng Wang ◽  
...  
Keyword(s):  
Diagnostic Performance ◽  
Protein Production ◽  
Protein S ◽  
Cross Reactivity ◽  
Spike Protein ◽  
Systematic Evaluation ◽  
Serological Tests ◽  
S Protein ◽  
Asymptomatic Patients ◽  
Human Coronaviruses

AbstractSerological tests play an essential role in monitoring and combating the COVID-19 pandemic. Recombinant spike protein (S protein), especially the S1 protein, is one of the major reagents used for serological tests. However, the high cost of S protein production and possible cross-reactivity with other human coronaviruses pose unavoidable challenges. By taking advantage of a peptide microarray with full spike protein coverage, we analyzed 2,434 sera from 858 COVID-19 patients, 63 asymptomatic patients and 610 controls collected from multiple clinical centers. Based on the results, we identified several S protein-derived 12-mer peptides that have high diagnostic performance. In particular, for monitoring the IgG response, one peptide (aa 1148–1159 or S2–78) exhibited a sensitivity (95.5%, 95% CI 93.7–96.9%) and specificity (96.7%, 95% CI 94.8–98.0%) comparable to those of the S1 protein for the detection of both symptomatic and asymptomatic COVID-19 cases. Furthermore, the diagnostic performance of the S2–78 (aa 1148–1159) IgG was successfully validated by ELISA in an independent sample cohort. A panel of four peptides, S1–93 (aa 553–564), S1–97 (aa 577–588), S1–101 (aa 601–612) and S1–105 (aa 625–636), that likely will avoid potential cross-reactivity with sera from patients infected by other coronaviruses was constructed. The peptides identified in this study may be applied independently or in combination with the S1 protein for accurate, affordable, and accessible COVID-19 diagnosis.

scholarly journals COVID-19 and human milk: SARS-CoV-2, antibodies, and neutralizing capacity

2020 ◽  
Author(s):  
Ryan M Pace ◽  
Janet E Williams ◽  
Kirsi M Järvinen ◽  
Mandy B Belfort ◽  
Christina DW Pace ◽  
...  
Keyword(s):  
Milk Sample ◽  
Nucleocapsid Protein ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Strong Correlations ◽  
Child Transmission ◽  
Milk Samples ◽  
Protein Receptor ◽  
Neutralizing Capacity ◽  
Before And After

Background: It is not known whether SARS-CoV-2 can be transmitted from mother to infant during breastfeeding, and if so whether the benefits of breastfeeding outweigh this risk. This study was designed to evaluate 1) if SARS-CoV-2 RNA can be detected in milk and on the breast of infected women, 2) concentrations of milk-borne anti-SARS-CoV-2 antibodies, and 3) the capacity of milk to neutralize SARS-CoV-2 infectivity. Methods: We collected 37 milk samples and 70 breast swabs (before and after breast washing) from 18 women recently diagnosed with COVID-19. Samples were analyzed for SARS-CoV-2 RNA using RT-qPCR. Milk was also analyzed for IgA and IgG specific for the nucleocapsid protein, receptor binding domain (RBD), S2 subunit of the spike protein of SARS-CoV-2, as well as 2 seasonal coronaviruses using ELISA; and for its ability to neutralize SARS-CoV-2. Results: We did not detect SARS-CoV-2 RNA in any milk sample. In contrast, SARS-CoV-2 RNA was detected on several breast swabs, although only one was considered conclusive. All milk contained SARS-CoV-2-specific IgA and IgG, and levels of anti-RBD IgA correlated with SARS-CoV-2 neutralization. Strong correlations between levels of IgA and IgG to SARS-CoV-2 and seasonal coronaviruses were noted. Conclusions: Our data do not support maternal-to-child transmission of SARS-CoV-2 via milk; however, risk of transmission via breast skin should be further evaluated. Importantly, milk produced by infected mothers is a source of anti-SARS-CoV-2 IgA and IgG and neutralizes SARS-CoV-2 activity. These results support recommendations to continue breastfeeding during mild-to-moderate maternal COVID-19 illness.

scholarly journals 146. Intact Sense of Taste and Smell During COVID-19 Infection Is Associated with Absence of of SARS-CoV-2 Spike Protein Antibody Responses within 3 Months of Symptomatic Illness

2021 ◽  
Vol 8 (Supplement_1) ◽  
pp. S88-S88
Author(s):  
James M Wilson ◽  
Sheena Gillani ◽  
Robert Bencshop ◽  
Josh Poorbaugh ◽  
Ajay Nirula ◽  
...  
Keyword(s):  
Immune Responses ◽  
Symptom Onset ◽  
Post Infection ◽  
Nucleocapsid Protein ◽  
Disease Duration ◽  
Cross Reactivity ◽  
Spike Protein ◽  
Clinical Factors ◽  
Series Completion ◽  
Taste And Smell

Abstract Background Although studies show most COVID-19 survivors have post-infection immunity against SARS-CoV-2 that could prevent re-infection, there is still a need to identify the breadth of antibody (Ab) responses associated with clinical phenotypes. We characterized Ab profiles at the estimated peak of Ab diversity among adults with recovered SARS-CoV-2 infections and determined their relationships with clinical factors. Methods From April-June 2020, 41 health system employees with PCR-confirmed symptomatic COVID-19 infection enrolled 8-10 weeks after symptom onset. Symptom questionnaires including baseline demographics, COVID-19 symptoms, disease severity, and disease duration were collected and plasma samples were assayed using a custom Luminex Multiplex platform (Figure 1) to measure the antibody response against 20 COVID-19 related antigens (Figure 2). Differences in Ab profile titers among different groups were tested using nonparametric t test and Benjamini-Hochberg adjustment for multiplicity. Associations were considered significant at FDR< 0.05. Figure 1: Description of the Luminex Serology Assay Figure 2: List of the COVID-19 Related Antigens and Controls Measured Results Mean age was 48 years (range 27-68), with 51% female, 37% White, 32% Black, 29% Asian, and 17% LatinX. Ab profiles (Figure 3) showed 100% cross-reactivity with related alpha and beta coronavirus, and 95% with SARS-CoV-1. 78% had Abs against SARS-CoV-2 nucleocapsid protein (NCP). However, 29% of patients had no immune response against the four spike protein epitopes. These participants also reported fewer symptoms, including no cases of anosmia/ageusia, suggesting mild illness. Anosmia/ageusia, fever, and cough associated significantly with higher Ab titers (Figure 4). Conclusion Broad immune responses to various SARS-CoV-2 and related antigens were found among a heterogeneous patient population. However, less than 3 months after symptom onset, protective Ab responses to SARS-CoV-2 spike proteins were not detected in nearly one-third of recovered patients, primarily with mild infection. Intact sense of smell and taste demonstrated the greatest association with loss of seroprotective SARS-CoV-2 Ab responses, which may be clinically useful to predict post-infection immunity. Next steps include comparing the magnitude of Ab responses following full series completion with mRNA vaccination among this cohort. Disclosures Robert Bencshop, PhD, Eli Lilly (Employee) Josh Poorbaugh, PhD, Eli Lilly (Employee) Ajay Nirula, MD/PhD, Eli Lilly (Employee, Shareholder) Lin Zhang, PhD, Eli Lilly and Company (Employee, Shareholder) Stephanie Beasley, BA, Eli Lilly (Employee)

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