Dynamics of Neutralizing Antibody Responses Following Natural SARS-CoV-2 Infection and Correlation with Commercial Serologic Tests. A Reappraisal and Indirect Comparison with Vaccinated Subjects

Abstract

Neutralising antibodies (NAbs) represent the real source of protection against SARS-CoV-2 infections by preventing the virus from entering target cells. The gold standard in the detection of these antibodies is the plaque reduction neutralization test (PRNT). As these experiments must be done in a very secure environment, other techniques based on pseudoviruses: pseudovirus neutralization test (pVNT) or surrogate virus neutralization test (sVNT) have been developed. Binding assays, on the other hand, measure total antibodies or IgG, IgM, and IgA directed against one epitope of the SARS-CoV-2, independently of their neutralizing capacity. The aim of this study is to compare the performance of six commercial binding assays to the pVNT and sVNT. In this study, we used blood samples from a cohort of 62 RT-PCR confirmed COVID-19 patients. Based on the results of the neutralizing assays, adapted cut-offs for the binding assays were calculated. The use of these adapted cut-offs does not permit to improve the accuracy of the serological assays and we did not find an adapted cut-off able to improve the capacity of these tests to detect NAbs. For a part of the population, a longitudinal follow-up with at least two samples for the same patient was performed. From day 14 to day 291, more than 75% of the samples were positive for NAbs (n = 87/110, 79.1%). Interestingly, 6 months post symptoms onset, the majority of the samples (N = 44/52, 84.6%) were still positive for NAbs. This is in sharp contrast with the results we obtained 6 months post-vaccination in our cohort of healthcare workers who have received the two-dose regimens of BNT162b2. In this cohort of vaccinated subjects, 43% (n = 25/58) of the participants no longer exhibit NAbs activity 180 days after the administration of the first dose of BNT162b2.

Keywords: COVID-19; SARS-CoV-2; neutralizing antibody.

Figure 1

Evolution of NAbs titer in pVNT (A) and sVNT (B) as a function of days post symptoms onset. Green squared and purple dots are concordant results. Squares framed in red are those for which there is a discordant result between pVNT result and at least one of the six non-neutralizing immunoassays. Dots circled in orange are those for which there is a discordant result between sVNT and at least one of the six non-neutralizing immunoassays. Dots circled in red are those for which only sVNT was discordant. Red dots are those for which sVNT is discordant from pVNT. The details of these discrepancies are reported in Table 2.

Figure 2

Roc curve analyses of all serological tests in this study and sVNT technique in comparison with pVNT based on the manufacturer’s cut-offs.

Figure 3

(A) Kinetics of NAbs since the symptom onset in non-hospitalized and hospitalized patients. Blue plain lines represent non-hospitalized patients and red plain lines represent hospitalized patients. Samples obtained at different times for the same patient are connected by dotted lines. (B) Kinetics of NAbs since the day of first dose vaccination in seronegative and seropositive individuals. Blue plain lines represent seronegative individuals and red plain lines represent seronegative individuals. Samples obtained at different times for the same individual are connected by dotted lines.

Figure 4

Comparison of NAbs titers between COVID-19 patients, seropositive vaccinated individuals, and seronegative vaccinated individuals at three different timeframes. The median is presented on top of each timeframe for each population. Bars represent the mean and standard error or mean.