SARS-CoV-2 infection induces sustained humoral immune responses in convalescent patients following symptomatic COVID-19

Abstract

medRxiv preprint doi: https://doi.org/10.1101/2020.07.21.20159178 Long-term antibody responses and neutralizing activities following SARS-CoV-2 infections have not yet been elucidated. We quantified immunoglobulin M (IgM) and G (IgG) antibodies recognizing the SARS-CoV-2 receptor-binding domain (RBD) of the spike (S) or the nucleocapsid (N) protein, and neutralizing antibodies during a period of six months following COVID-19 disease onset in 349 symptomatic COVID-19 patients, which were among the first world-wide being infected. The positivity rate and magnitude of IgM-S and IgG-N responses increased rapidly. High levels of IgM-S/N and IgG-S/N at 2-3 weeks after disease onset were associated with virus control and IgG-S titers correlated closely with the capacity to neutralize SARS-CoV-2. While specific IgM-S/N became undetectable 12 weeks after disease onset in most patients, IgG-S/N titers showed an intermediate contraction phase, but stabilized at relatively high levels over the six months observation period. At late time points the positivity rates for binding and neutralizing SARS-CoV-2-specific antibodies was still over 70%. Taken together, our data indicate sustained humoral immunity in recovered patients who suffer from symptomatic COVID-19, suggesting prolonged immunity. Fig 1. Longitudinal analyses of IgM and IgG responses recognizing the RBD of spike or the nucleoprotein of SARS-CoV-2 in confirmed COVID-19 patients. IgM and IgG against the RBD of the spike protein (‘S’) and the nucleoprotein (‘N’) of SARS-CoV-2 were detected by capture chemiluminescence immunoassays (CLIA). (A) Positive rate of individual antibodies tested at the indicated dates following onset of symptoms. (B-C) The plasma antibody levels (IgM-S, IgM-N, IgG-S, and IgG-N) in patients with different disease courses are presented. The boxes in Fig. 1C show medians (middle line), 75% (upper line) and 25% quartiles (lower line). (D) Sequential sampling and analyses of antibody titers in 35 COVID-19 cases. Characteristics of two patients with low IgG antibody levels. Patient 13: a 46-year-old female with fever, cough, dizziness, and fatigue for 6 days; patient 17: a 38-year-old female with fever and chest tightness for 4 days. The cut-off value for IgM-S detection was 0.7 AU/ml. The cut-off value for IgM-N, IgG-S, and IgG-N were 1 (shown on the left Y axis). Fig 2: Correlation of antibody titers with virus control and severity of illness in hospitalized COVID-19 patients. (A) S- and N- specific CLIA-reactive IgM/IgG were compared in COVID-19 patients who were virus RNA-negative (red) versus those who were virus RNA-positive (blue) at the time-point of sampling at different periods after the disease onset. Each antibody detection value is either classified into the RNA-negative group or the RNA-positive group according to the simultaneous RNA detection result. A total of 343 results were acquired for this analysis. (B) Comparison of S and N-specific CLIA-reactive IgM/IgG titers between severe (red) and non-severe (blue) patients. (C) Comparison of 64 severe and non-severe patients (69 samples) at different S- and N- specific CLIA-reactive IgM/IgG levels at the 4th week after symptoms onset. LMM was used for statistical analysis. *p<0.05; **p <0.01; ***p<0.001. Fig 3: N- and S- specific CLIA-reactive IgG responses have different predictive values for neutralization capacities. A total of 186 samples from 137 symptomatic COVID-19 patients were assessed concerning SARS-CoV-2 neutralization titers and grouped according to the weeks after symptom onset. (A) Proportions of plasma neutralization activity were stratified in two-week intervals. (B) Correlation analysis of neutralization titer with S- and N- specific CLIA-reactive IgM/IgG in COVID-19 patients. A non-parametric Spearman’s correlation test was used for the statistical analyses. In the graphs, p, r, and N indicate the p-value, correlation coefficient, and sample size, respectively. (C) Distribution of neutralizing activity at different S- and N-specific CLIA-reactive IgM/IgGs. (D) Based on the predicted cutoff value and IgG-S titer, the neutralizing activity of all confirmed patients at different time points was calculated. Figure S1. Positive rate of combined antibodies tested in a time series following the onset of symptoms. IgM and IgG against the RBD of the spike protein and nucleoproteins of SARS-CoV-2 were detected by capture chemiluminescence immunoassays (CLIA). Positive rate of IgM-S + IgM-N, IgG-S + IgG-N, IgM-S + IgM-N + IgG-S + IgG-N tested in time series after the onset of symptoms. Table S1. Clinical and laboratory characteristics of hospitalized COVID-19 patients Figure S2. Comparison of differences in antibody titers according to severity of disease, gender, and age by a generalized linear model. Comparison of S and N-specific CLIA-reactive IgM/IgG titers between RNA-positive and -negative cases (A), severe and non-severe patients (B), female and male patients (C-D) and the young (<65 years old) and elder (≥65 years old) patients (E-F). The boxes in Fig. S2C and S2E show medians (middle line), 75% (upper line) and 25% quartiles (lower line). Figure S3. Receiver operating characteristic (ROC) curve of IgG-S titers to predict neutralizing activity of COVID-19 patients. The AUC of IgG-S was 0.865 (95% CI 0.777–0.953; p<0.0001). The optimal cutoff value was 4.99 AU/ml.