Seasonal human coronavirus antibodies are boosted upon SARS-CoV-2 infection but not associated with protection

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly spread within the human population. Although SARS-CoV-2 is a novel coronavirus, most humans had been previously exposed to other antigenically distinct common seasonal human coronaviruses (hCoVs) before the coronavirus disease 2019 (COVID-19) pandemic. Here, we quantified levels of SARS-CoV-2-reactive antibodies and hCoV-reactive antibodies in serum samples collected from 431 humans before the COVID-19 pandemic. We then quantified pre-pandemic antibody levels in serum from a separate cohort of 251 individuals who became PCR-confirmed infected with SARS-CoV-2. Finally, we longitudinally measured hCoV and SARS-CoV-2 antibodies in the serum of hospitalized COVID-19 patients. Our studies indicate that most individuals possessed hCoV-reactive antibodies before the COVID-19 pandemic. We determined that ∼20% of these individuals possessed non-neutralizing antibodies that cross-reacted with SARS-CoV-2 spike and nucleocapsid proteins. These antibodies were not associated with protection against SARS-CoV-2 infections or hospitalizations, but they were boosted upon SARS-CoV-2 infection.

Keywords: SARS-CoV-2; antibodies; coronavirus; pre-existing immunity.

Graphical abstract

Figure 1

Identification of pre-existing cross-reactive SARS-CoV-2 antibodies in human serum prior to the pandemic (A–C) ELISAs were completed to quantify levels of serum antibodies binding to the SARS-CoV-2 full-length S protein (A), the S-RBD (B), and the N protein (C). Dashed line denotes lower limit of detection (LOD = 50); dotted line represents a threshold set 2-fold above LOD (>100). We tested samples collected from 431 individuals in the summer of 2017, prior to the global pandemic. We also tested samples collected from 15 individuals after confirmed SARS-CoV-2 infections as well as from recovered adults. (D) The relationship between antibody titers in donors with detectable IgG against the S-RBD and/or full-length S is shown. (E) SARS-CoV-2 pseudotype neutralization assays were completed with pre-pandemic serum samples with (n = 14) and without (n = 29) cross-reactive SARS-CoV-2 antibodies, as well as serum samples from individuals after confirmed SARS-CoV-2 infections (n = 15); one-way ANOVA Tukey’s multiple comparisons of log2 transformed antibody titers ^∗∗∗∗p < 0.0001; dotted line denotes lower LOD (=10). (F–H) ELISAs were completed to quantify levels of serum antibodies binding to the full-length S proteins from 229E, NL63, and OC43 with pre-pandemic serum samples with (n = 17) and without (n = 17) cross-reactive SARS-CoV-2 antibodies. Unpaired t tests of log2 transformed antibody titers ^∗∗p < 0.002. Horizontal lines indicate geometric mean and error bars represent standard deviation. See also Figure S1, Figure S2, Figure S3, Figure S4, and S5.

Figure S1

There are no obvious age-related differences in pre-pandemic SARS-CoV-2 and hCoV reactive antibodies, related to Figure 1 ELISAs were completed to measure levels of serum antibodies binding to the SARS-CoV-2 full-length spike (S) protein (A), SARS-CoV-2 receptor binding domain (S-RBD) of S (B), SARS-CoV-2 nucleocapsid (N) protein (C), 229E S protein (D), NL63 S protein (E), and OC43 S protein (F). Serum samples collected from 431 individuals in the summer of 2017 were tested. Reciprocal titer from serially-diluted serum samples (A-C) and optical densities at 450nm wavelength (OD₄₅₀) of 1:500 dilution of serum (D-F) are shown. Dashed line denotes lower limit of detection (LOD = 50), dotted line represents a threshold set 2-fold above LOD (> 100).

Figure S2

Comparison of ELISA data using unpurified and purified serum IgG antibodies, related to Figure 1 and 2 IgG was purified from sera samples from individuals without (A; n = 5) and with (B; n = 11) pre-pandemic cross-reactive antibodies. IgG was also purified from serum samples from individuals who had recovered from a confirmed SARS-CoV-2 infection (C; n = 5). ELISAs were completed to quantify levels of serum antibodies binding to SARS-CoV-2 full length S, S-RBD, and N protein with and without IgG magnetic bead purification. The dotted line represents a threshold set 2-fold above the limit of detection (> 100).

Figure S3

Correlation between N, S, and S-RBD antibody titers in pre-pandemic samples, related to Figure 1 Shown are the relationships between serum IgG antibody titers against the SARS-CoV-2 N protein and S-RBD (A) or full length S (B) from 431 individuals whose samples were collected prior to the pandemic in the summer of 2017. Dotted line represents a threshold set 2-fold above the limit of detection (> 100).

Figure S4

SARS-CoV-2 pseudotype neutralization curves, related to Figure 1 Raw neutralization curves for data from Figure 1E are shown, including samples from individuals who did not have pre-pandemic cross reactive SARS-CoV-2 antibodies (A), individuals who possessed pre-pandemic cross reactive SARS-CoV-2 antibodies (B), and individuals following confirmed SARS-CoV-2 infection (C). Mean and error bars are shown for each replicate; dotted line denotes the cut-off for foci reduction neutralization of 50% (FRNT50).

Figure S5

Pre-pandemic cross-reactive antibodies do not neutralize SARS-CoV-2 in bonafide BSL3-level neutralization assays, related to Figure 1 Neutralization assays with live SARS-CoV-2 were completed using 9 pre-pandemic samples with cross-reactive SARS-CoV-2 antibodies, 7 pre-pandemic samples without cross-reactive SARS-CoV-2 antibodies, and 5 samples from individuals who recovered from a PCR-confirmed SARS-CoV-2 infection. The pre-pandemic samples for these experiments were collected in 2019 and are different from those shown in Figure 1 (which were collected in 2017).

Figure 2

Pre-pandemic SARS-CoV-2 and OC43-reactive antibodies are not associated with protection from SARS-CoV-2 infection (A and B) We quantified antibody levels in pre-pandemic serum samples collected from individuals who later became SARS-CoV-2 infected (cases; n = 251) and those who did not become SARS-CoV-2 infected (controls; n = 251). ELISAs were completed to quantify levels of antibodies reactive to SARS-CoV-2 proteins (S, S-RBD, and N) and the OC43 S protein. Shown are data using samples collected from the entire cohort between August 2013 and March 2020 (A) and samples from a smaller subset of individuals collected between April 2019 and March 2020 (B). Antibody titers between cases and controls were not significantly different as determined by unpaired t tests of log2 transformed antibody titers. Dashed line denotes lower limit of detection (LOD = 50), dotted line represents a threshold set 2-fold above LOD (>100). See also Tables S1 and S2.

Figure 3

SARS-CoV-2 infections boost antibodies that react to OC43 S protein (A–D) We quantified antibody levels in serum collected from 27 individuals 0 and 7 days after hospitalization for COVID-19. ELISAs were completed to quantify levels of antibodies reactive to the S proteins of 229E, NL63, OC43, and SARS-CoV-2. IgG titers (A) and titer fold change (B) are shown. Levels of OC43 S-reactive antibodies (C) and fold change in OC43 S-reactive antibodies (D) were not associated with disease outcome. Paired t test of log2 transformed antibody titers, ^∗∗∗∗p < 0.0001. One-way ANOVA Tukey’s multiple comparisons fold-change in antibody titers, ^∗p < 0.04. Horizontal lines indicate the median and error bars show interquartile ranges. See also Figure S6.

Figure S6

Antibodies directed to the S2 region of OC43 spike are boosted during SARS-CoV-2 infection, related to Figure 3 We quantified antibody levels in serum collected from 27 individuals 0 and 7 days after hospitalization for COVID-19. ELISAs were completed to measure levels of serum antibodies binding to the OC43 full-length spike (FL) protein and the individual S1 and S2 subunits of the OC43 spike. (A) IgG titers and (B) titer fold change are shown. Paired t test of log2 transformed antibody titers, ^∗∗∗∗p < 0.0001. One-way ANOVA Tukey’s multiple comparisons fold-change in antibody titers, * p < 0.02 ^∗∗p < 0.005. Horizontal lines indicate the median and error bars show interquartile range.