Linear B-cell epitopes in the spike and nucleocapsid proteins as markers of SARS-CoV-2 exposure and disease severity

Abstract

Background Given the unceasing worldwide surge in COVID-19 cases, there is an imperative need to develop highly specific and sensitive serology assays to define exposure to Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). Methods Pooled plasma samples from PCR positive COVID-19 patients were used to identify linear B-cell epitopes from a SARS-CoV-2 peptide library of spike (S), envelope (E), membrane (M), and nucleocapsid (N) structural proteins by peptide-based ELISA. Hit epitopes were further validated with 79 COVID-19 patients with different disease severity status, 13 seasonal human CoV, 20 recovered SARS patients and 22 healthy donors. Findings Four immunodominant epitopes, S14P5, S20P2, S21P2 and N4P5, were identified on the S and N viral proteins. IgG responses to all identified epitopes displayed a strong detection profile, with N4P5 achieving the highest level of specificity (100%) and sensitivity (>96%) against SARS-CoV-2. Furthermore, the magnitude of IgG responses to S14P5, S21P2 and N4P5 were strongly associated with disease severity. Interpretation IgG responses to the peptide epitopes can serve as useful indicators for the degree of immunopathology in COVID-19 patients, and function as higly specific and sensitive sero-immunosurveillance tools for recent or past SARS-CoV-2 infections. The flexibility of these epitopes to be used alone or in combination will allow for the development of improved point-of-care-tests (POCTs). Funding Biomedical Research Council (BMRC), the A*ccelerate GAP-funded project (ACCL/19-GAP064-R20H-H) from Agency of Science, Technology and Research (A*STAR), and National Medical Research Council (NMRC) COVID-19 Research fund (COVID19RF-001) and CCGSFPOR20002. ATR is supported by the Singapore International Graduate Award (SINGA), A*STAR.

Epitope-Based Immunoinformatics Approach on Nucleocapsid Protein of Severe Acute Respiratory Syndrome-Coronavirus-2

“…As a consequence, our study predicted several B cell epitopes that were in line with those identified by Grifoni et al (Table S2). Additionally, one of the predicted B-cell epitope from amino acid residues 154-166 was in agreement with the study from Amrun et al (Table S2) [74]. Moreover, several studies have reported the characterization of B-cell epitopes from the N protein of many viruses from humans and animals [23,[75][76][77].…”

Sensitive detection of total anti-Spike antibodies and isotype switching in asymptomatic and symptomatic individuals with COVID-19

“…We previously developed an ELISA-based serological assay based on the four immunodominant IgG linear epitopes on the S and N proteins. 21 To capture a wider repertoire of antibodies against the S protein, in this study we developed a flow cytometry assay based on the full-length S protein. The assay is more time efficient; results are available within 2 h. The assay is also well suited to detect anti-S protein antibodies in symptomatic and asymptomatic individuals.…”

“… 41 Demographic data, clinical and laboratory parameters during the hospitalisation period were retrieved from patient records ( Table S1 ). 21 Whole blood of patients was collected into BD Vacutainer® CPT tubes and centrifuged at 1700 g for 20 min to obtain plasma fractions. Plasma samples were categorised according to three time points: median 5 days post-illness onset (pio), median 10 days pio, and median 23 days pio.…”

SARS-CoV-2 Epitope Mapping on Microarrays Highlights Strong Immune-Response to N Protein Region

“…Full-length recombinant antigens, however, in addition to high costs and storage constraints, may suffer from stability problems, batch-to-batch variations and in some cases may give rise to cross-reactivity leading to ambiguous detection outcomes. This might be the case for SARS-CoV-2 when using proteins sharing high homology with genetically similar, co-circulating, human coronaviruses (hCoV) that are responsible for common cold [ 6 , 7 ]. Studies towards fine dissection of the specific regions on the viral proteins recognized by virus-specific antibodies have been recently published focusing on the S protein [ 6 , 8 , 9 , 10 , 11 ].…”