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      Comparative Assessment of Sera from Individuals after S-Gene RNA-Based SARS-CoV-2 Vaccination with Spike-Protein-Based and Nucleocapsid-Based Serological Assays

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          Abstract

          Due to the beginning of vaccination against COVID-19, serological discrimination between vaccine-associated humoral response and serology-based surveillance of natural SARS-CoV-2 infections as well as breakthrough infections becomes an issue of relevance. Here, we assessed the differentiated effects of the application of an RNA vaccine using SARS-CoV-2 spike protein epitopes on the results of both anti-spike protein–based serology (EUROIMMUN) and anti-nucleocapsid-based serology (VIROTECH). A total of 80 serum samples from vaccinees acquired at different time points after vaccination was assessed. While positive or borderline serological response in the anti-spike protein assay was observed for all samples (90% both IgG and IgA, 6.3% IgA only, 3.8% borderline IgG only), only a single case of a falsely positive IgM was observed for the anti-nucleocapsid assay as expected due to this assay’s specificity. Positive anti-spike protein antibodies were already detectable in the second week after the first dose of vaccination, with higher titers after the second dose of the vaccine. In conclusion, the combined application of anti-spike protein–based serology and anti-nucleocapsid-based serology will provide a useful option for the discrimination of vaccination response and natural infection.

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          Safety and Immunogenicity of Two RNA-Based Covid-19 Vaccine Candidates

          Edward E Walsh, Robert W Frenck, Ann Falsey … (2021)
          Abstract Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections and the resulting disease, coronavirus disease 2019 (Covid-19), have spread to millions of persons worldwide. Multiple vaccine candidates are under development, but no vaccine is currently available. Interim safety and immunogenicity data about the vaccine candidate BNT162b1 in younger adults have been reported previously from trials in Germany and the United States. Methods In an ongoing, placebo-controlled, observer-blinded, dose-escalation, phase 1 trial conducted in the United States, we randomly assigned healthy adults 18 to 55 years of age and those 65 to 85 years of age to receive either placebo or one of two lipid nanoparticle–formulated, nucleoside-modified RNA vaccine candidates: BNT162b1, which encodes a secreted trimerized SARS-CoV-2 receptor–binding domain; or BNT162b2, which encodes a membrane-anchored SARS-CoV-2 full-length spike, stabilized in the prefusion conformation. The primary outcome was safety (e.g., local and systemic reactions and adverse events); immunogenicity was a secondary outcome. Trial groups were defined according to vaccine candidate, age of the participants, and vaccine dose level (10 μg, 20 μg, 30 μg, and 100 μg). In all groups but one, participants received two doses, with a 21-day interval between doses; in one group (100 μg of BNT162b1), participants received one dose. Results A total of 195 participants underwent randomization. In each of 13 groups of 15 participants, 12 participants received vaccine and 3 received placebo. BNT162b2 was associated with a lower incidence and severity of systemic reactions than BNT162b1, particularly in older adults. In both younger and older adults, the two vaccine candidates elicited similar dose-dependent SARS-CoV-2–neutralizing geometric mean titers, which were similar to or higher than the geometric mean titer of a panel of SARS-CoV-2 convalescent serum samples. Conclusions The safety and immunogenicity data from this U.S. phase 1 trial of two vaccine candidates in younger and older adults, added to earlier interim safety and immunogenicity data regarding BNT162b1 in younger adults from trials in Germany and the United States, support the selection of BNT162b2 for advancement to a pivotal phase 2–3 safety and efficacy evaluation. (Funded by BioNTech and Pfizer; ClinicalTrials.gov number, NCT04368728.)
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            Biochemical characterization of SARS-CoV-2 nucleocapsid protein

            Weihong Zeng, Guangfeng Liu, Huan Ma … (2020)
            The nucleocapsid (N) protein is an important antigen for coronavirus, which participate in RNA package and virus particle release. In this study, we expressed the N protein of SARS-CoV-2 and characterized its biochemical properties. Static light scattering, size exclusive chromatography, and small-angle X-ray scattering (SAXS) showed that the purified N protein is largely a dimer in solution. CD spectra showed that it has a high percentage of disordered region at room temperature while it was best structured at 55 °C, suggesting its structural dynamics. Fluorescence polarization assay showed it has non-specific nucleic acid binding capability, which raised a concern in using it as a diagnostic marker. Immunoblot assays confirmed the presence of IgA, IgM and IgG antibodies against N antigen in COVID-19 infection patients' sera, proving the importance of this antigen in host immunity and diagnostics.
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              Sensitivity in Detection of Antibodies to Nucleocapsid and Spike Proteins of Severe Acute Respiratory Syndrome Coronavirus 2 in Patients With Coronavirus Disease 2019

              Peter D. Burbelo, Francis X. Riedo, Chihiro Morishima … (2020)
              Abstract Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the cause of coronavirus disease 2019 (COVID-19), is associated with respiratory-related disease and death. Assays to detect virus-specific antibodies are important to understand the prevalence of infection and the course of the immune response. Methods Quantitative measurements of plasma or serum antibodies to the nucleocapsid and spike proteins were analyzed using luciferase immunoprecipitation system assays in 100 cross-sectional or longitudinal samples from patients with SARS-CoV-2 infection. A subset of samples was tested both with and without heat inactivation. Results At >14 days after symptom onset, antibodies against SARS-CoV-2 nucleocapsid protein showed 100% sensitivity and 100% specificity, whereas antibodies to spike protein were detected with 91% sensitivity and 100% specificity. Neither antibody levels nor the rate of seropositivity were significantly reduced by heat inactivation of samples. Analysis of daily samples from 6 patients with COVID-19 showed anti-nucleocapsid and spike protein antibodies appearing between days 8 and 14 after initial symptoms. Immunocompromised patients generally had a delayed antibody response to SARS-CoV-2, compared with immunocompetent patients. Conclusions Antibody to the nucleocapsid protein of SARS-CoV-2 is more sensitive than spike protein antibody for detecting early infection. Analyzing heat-inactivated samples with a luciferase immunoprecipitation system assay is a safe and sensitive method for detecting SARS-CoV-2 antibodies.
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                Author and article information

                Contributors
                Chao-Min Cheng: Role: Academic Editor
                Journal
                Journal ID (nlm-ta): Diagnostics (Basel)
                Journal ID (iso-abbrev): Diagnostics (Basel)
                Journal ID (publisher-id): diagnostics
                Title: Diagnostics
                Publisher: MDPI
                ISSN (Electronic): 2075-4418
                Publication date (Electronic): 03 March 2021
                Publication date Collection: March 2021
                Volume: 11
                Issue: 3
                Electronic Location Identifier: 426
                Affiliations
                [1 ]Institute for Medical Microbiology, University Medical Center Göttingen, 37075 Göttingen, Germany; anja.doerschug@ 123456stud.uni-goettingen.de (A.D.); julian.schwanbeck@ 123456med.uni-goettingen.de (J.S.); kemal.mese@ 123456med.uni-goettingen.de (K.M.); ugross@ 123456gwdg.de (U.G.)
                [2 ]Institute for Medical Microbiology, Virology and Hygiene, University Medicine Rostock, 18057 Rostock, Germany; hagen.frickmann@ 123456med.uni-rostock.de (H.F.); andreas.hahn@ 123456uni-rostock.de (A.H.)
                [3 ]Department of Microbiology and Hospital Hygiene, Bundeswehr Hospital Hamburg, 20359 Hamburg, Germany
                [4 ]Department of Anesthesiology, University Medical Center Göttingen, 37075 Göttingen, Germany; elif.yilmaz@ 123456med.uni-goettingen.de
                Author notes
                [* ]Correspondence: azautne@ 123456gwde.de ; Tel.: +49-551-39-65927
                [†]

                Hagen Frickmann and Anja Dörschug contributed equally to this work.

                Author information
                https://orcid.org/0000-0002-8967-9528
                https://orcid.org/0000-0003-2803-0167
                https://orcid.org/0000-0002-5157-8369
                https://orcid.org/0000-0003-0984-013X
                Article
                Publisher ID: diagnostics-11-00426
                DOI: 10.3390/diagnostics11030426
                PMC ID: 7998789
                PubMed ID: 33802453
                SO-VID: c9063501-6aaa-40e7-8549-f2d880ae9d26
                Copyright © © 2021 by the authors.
                License:

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                Date received : 15 February 2021
                Date accepted : 26 February 2021
                Categories
                Subject: Communication

                Keywords: sars-cov-2,covid-19,serology,vaccination,surveillance,nucleocapsid,spike protein
                Data availability:
                Keywords: sars-cov-2, covid-19, serology, vaccination, surveillance, nucleocapsid, spike protein

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