Detection of Anti-SARS-CoV-2-S2 IgG Is More Sensitive Than Anti-RBD IgG in Identifying Asymptomatic COVID-19 Patients.

Characterizing the serologic features of asymptomatic SARS-CoV-2 infection is imperative to improve diagnostics and control of SARS-CoV-2 transmission. In this study, we evaluated the antibody profiles in 272 plasma samples collected from 59 COVID-19 patients, consisting of 18 asymptomatic patients, 33 mildly ill patients and 8 severely ill patients. We measured the IgG against five viral structural proteins, different isotypes of immunoglobulins against the Receptor Binding Domain (RBD) protein, and neutralizing antibodies. The results showed that the overall antibody response was lower in asymptomatic infections than in symptomatic infections throughout the disease course. In contrast to symptomatic patients, asymptomatic patients showed a dominant IgG-response towards the RBD protein, but not IgM and IgA. Neutralizing antibody titers had linear correlations with IgA/IgM/IgG levels against SARS-CoV-2-RBD, as well as with IgG levels against multiple SARS-CoV-2 structural proteins, especially with anti-RBD or anti-S2 IgG. In addition, the sensitivity of anti-S2-IgG is better in identifying asymptomatic infections at early time post infection compared to anti-RBD-IgG. These data suggest that asymptomatic infections elicit weaker antibody responses, and primarily induce IgG antibody responses rather than IgA or IgM antibody responses. Detection of IgG against the S2 protein could supplement nucleic acid testing to identify asymptomatic patients. This study provides an antibody detection scheme for asymptomatic infections, which may contribute to epidemic prevention and control.

Delayed neutralizing antibody response in the acute phase correlates with severe progression of COVID-19.

Adaptive immunity to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) dynamics remain largely unknown. The neutralizing antibody (NAb) levels in patients with coronavirus disease 2019 (COVID-19) are helpful for understanding the pathology. Using SARS-CoV-2 pseudotyped virus, serum sample neutralization values in symptomatic COVID-19 patients were measured using the chemiluminescence reduction neutralization test (CRNT). At least two sequential serum samples collected during hospitalization were analyzed to assess NAbs neutralizing activity dynamics at different time points. Of the 11 patients, four (36.4%), six (54.5%), and one (9.1%) had moderate, severe, and critical disease, respectively. Fifty percent neutralization (N50%-CRNT) was observed upon admission in 90.9% (10/11); all patients acquired neutralizing activity 2-12 days after onset. In patients with moderate disease, neutralization was observed at earliest within two days after symptom onset. In patients with severe-to-critical disease, neutralization activity increased, plateauing 9-16 days after onset. Neutralization activity on admission was significantly higher in patients with moderate disease than in patients with severe-to-critical disease (relative % of infectivity, 6.4% vs. 41.1%; P = .011). Neutralization activity on admission inversely correlated with disease severity. The rapid NAb response may play a crucial role in preventing the progression of COVID-19.

COVID-19 mRNA vaccine induced antibody responses against three SARS-CoV-2 variants.

As SARS-CoV-2 has been circulating for over a year, dozens of vaccine candidates are under development or in clinical use. The BNT162b2 mRNA COVID-19 vaccine induces spike protein-specific neutralizing antibodies associated with protective immunity. The emergence of the B.1.1.7 and B.1.351 variants has raised concerns of reduced vaccine efficacy and increased re-infection rates. Here we show, that after the second dose, the sera of BNT162b2-vaccinated health care workers (n = 180) effectively neutralize the SARS-CoV-2 variant with the D614G substitution and the B.1.1.7 variant, whereas the neutralization of the B.1.351 variant is five-fold reduced. Despite the reduction, 92% of the seronegative vaccinees have a neutralization titre of >20 for the B.1.351 variant indicating some protection. The vaccinees' neutralization titres exceeded those of recovered non-hospitalized COVID-19 patients. Our work provides evidence that the second dose of the BNT162b2 vaccine induces cross-neutralization of at least some of the circulating SARS-CoV-2 variants.

Previous SARS-CoV-2 Infection Increases B.1.1.7 Cross-Neutralization by Vaccinated Individuals.

With the spread of new variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), there is a need to assess the protection conferred by both previous infections and current vaccination. Here we tested the neutralizing activity of infected and/or vaccinated individuals against pseudoviruses expressing the spike of the original SARS-CoV-2 isolate Wuhan-Hu-1 (WH1), the D614G mutant and the B.1.1.7 variant. Our data show that parameters of natural infection (time from infection and nature of the infecting variant) determined cross-neutralization. Uninfected vaccinees showed a small reduction in neutralization against the B.1.1.7 variant compared to both the WH1 strain and the D614G mutant. Interestingly, upon vaccination, previously infected individuals developed more robust neutralizing responses against B.1.1.7, suggesting that vaccines can boost the neutralization breadth conferred by natural infection.

Testing-on-a-probe biosensors reveal association of early SARS-CoV-2 total antibodies and surrogate neutralizing antibodies with mortality in COVID-19 patients.

The association of mortality with the early humoral response to SARS-CoV-2 infection within the first few days after onset of symptoms (DAOS) has not been thoroughly investigated partly due to a lack of sufficiently sensitive antibody testing methods. Here we report two sensitive and automated testing-on-a-probe (TOP) biosensor assays for SARS-CoV-2 viral specific total antibodies (TAb) and surrogate neutralizing antibodies (SNAb), which are suitable for clinical use. The TOP assays employ an RBD-coated quartz probe using a Cy5-Streptavidin-polysacharide conjugate to improve sensitivity and minimize interference. Disposable cartridges containing pre-dispensed reagents require no liquid manipulation or fluidics during testing. The TOP-TAb assay exhibited higher sensitivity in the 0-7 DAOS window than a widely used FDA-EUA assay. The rapid and automated TOP-SNAb correlated well with two well-established SARS-CoV-2 virus neutralization tests. The clinical utility of the TOP assays was demonstrated by evaluating early antibody responses in 120 SARS-CoV-2 RT-PCR positive adult hospitalized patients. Higher TAb and SNAb positivity rates and more robust antibody responses at patient's initial hospital presentation were seen in inpatients who survived COVID-19 than those who died in the hospital. Survival analysis using the Cox Proportional Hazards Model showed that patients who had negative TAb and/or SNAb at initial hospital presentation were at a higher risk of in-hospital mortality. Furthermore, TAb and SNAb levels at presentation were inversely associated with SARS-CoV-2 viral load based on concurrent RT-PCR testing. Overall, the sensitive and automated TAb and SNAb assays allow the detection of early SARS-CoV-2 antibodies which associate with mortality.

Correlation of SARS-CoV-2 Neutralizing Antibodies to an Automated Chemiluminescent Serological Immunoassay.

INTRODUCTION: Neutralizing antibodies (NAbs) are capable of binding to a virus to render it incapable of infection. The ability of commercially available SARS-CoV-2 serological tests to detect NAbs has not been widely reported. We sought to correlate the antibodies detected by an automated chemiluminescent immunoassay with NAbs. METHODS: Residual serum samples from 35 patients that had a positive antibody test using the LIAISON® SARS-CoV-2 S1/S2 IgG chemiluminescent immunoassay and 2 antibody-negative control sera were tested for NAbs using a plaque reduction neutralization test (PRNT). RESULTS: NAbs were detected in 66% (23/35) of the antibody-positive samples. The immunoassay signal value ranged from 21.7 to 131.3 AU/mL (median, 90.5) with significant correlation between it and the PRNT (r = 0.61, P = 0.002). In the samples without NAbs, the immunoassay signal ranged from 16.3 to 66.2 AU/mL (median, 27.2). An immunoassay signal cutoff of >41 AU/mL was 91% sensitive and 92% specific for the detection of NAbs. DISCUSSION: It is important that correlates of immunity to SARS-CoV-2 be identified and NAbs are considered to be central indicators of such. PRNT is the gold-standard test for identifying NAbs but it cannot be used for large-scale testing of populations. It is necessary to establish relationships between it and widely used commercial serological assays for SARS-CoV-2.