Persistent symptoms after COVID-19 are not associated with differential SARS-CoV-2 antibody or T cell immunity

Abstract

Among the unknowns in decoding the pathogenesis of SARS-CoV-2 persistent symptoms in Long Covid is whether there is a contributory role of abnormal immunity during acute infection. It has been proposed that Long Covid is a consequence of either an excessive or inadequate initial immune response. Here, we analyze SARS-CoV-2 humoral and cellular immunity in 86 healthcare workers with laboratory confirmed mild or asymptomatic SARS-CoV-2 infection during the first wave. Symptom questionnaires allow stratification into those with persistent symptoms and those without for comparison. During the period up to 18-weeks post-infection, we observe no difference in antibody responses to spike RBD or nucleoprotein, virus neutralization, or T cell responses. Also, there is no difference in the profile of antibody waning. Analysis at 1-year, after two vaccine doses, comparing those with persistent symptoms to those without, again shows similar SARS-CoV-2 immunity. Thus, quantitative differences in these measured parameters of SARS-CoV-2 adaptive immunity following mild or asymptomatic acute infection are unlikely to have contributed to Long Covid causality. ClinicalTrials.gov (NCT04318314).

Fig. 1. Longitudinal antibody and T cell responses to SARS-CoV-2 during the first 16–18 weeks after infection in HCW that either recovered or reported persistent symptoms.

A cohort of health care workers (HCW) recruited in March 2020 with laboratory confirmed SARS-CoV-2 infection (n = 86, 33% male) were followed up weekly. Serum was assayed for A S1 RBD and B Nucleocapsid antibody titers. Twenty-five HCW reported persistent symptoms at 12 months post-infection (open circle, 28% male) whilst the remaining 61 HCW fully recovered (closed circle, 34% male). In both panels, peak antibody titer during the 24-week follow-up and longitudinal data is plotted for each HCW. C S1 RBD and nucleocapsid antibody titers plotted longitudinally relative to timing of positive SARS-CoV-2 PCR (Recovered, black, n = 15, 40% male; Persistent symptoms, red, n = 9, 44% male). Thirty-seven of Recovered HCW (closed circle, 30% male) and 21 of the HCW with persistent symptoms (open circle, 24% male) were assayed for D Neutralizing antibody (IC50) against SARS-CoV-2 ancestral (Wuhan Hu-1) strain pseudovirus at 16–18 weeks; E T cell response against spike and nucleocapsid recombinant proteins and F peptide pools containing mapped epitopes from spike, nucleocapsid, membrane and ORF3a/7a proteins. G T cell response against peptide pool containing epitopes from Cytomegalovirus, Epstein Barr virus and Influenza (Recovered, n = 34, closed circle, 29% male; Persistent symptoms, n = 19, open circle, 26% male). For A–G, numbers of HCW showing a positive response for each assay are shown at the top of each plot and the proportion of each group with a high (black), low (grey) or no response (white) are represented by doughnut plots below. Longitudinal T cell responses to spike H and non-spike I peptide megapools assayed by ELISpot (in Recovered HCW, black circle, n = 14, 36% male; and HCW reporting Persistent symptoms, red circle, n = 8, 50% male) at 12 months after SARS-CoV-2 infection. Data is plotted relative to when HCW had a SARS-CoV-2 positive PCR. Two-tailed Mann-Whitney U tests (Graphpad Prism version 8.0) were used to test for significant differences between recovered HCW and those reporting persistent symptoms. Source data are provided as a Source Data file. Error bars shown are geometric mean ± 1 geometric SD. Ab antibody, COI cut-off index, N nucleocapsid, ORF open reading frame, PBMC peripheral blood mononuclear cells, RBD receptor binding domain, SD standard deviation, SFC spot forming cells, U units.

Fig. 2. Antibody and T cell responses against SARS-CoV-2 at 12 months after laboratory confirmed infection in HCW that either Recovered or reported Persistent symptoms.

A S1 RBD (Recovered, closed circle, n = 53, 40% male); Persistent symptoms, open circle, n = 25, 28% male) and B Nucleocapsid antibody (Recovered, closed circle, n = 61, 34% male; Persistent symptoms, open circle, n = 25, 28% male) titers in HCW 12 months after laboratory confirmed SARS-CoV-2 infection. S1 RBD data is plotted for HCW who had received 2 doses of COVID-19 vaccine at the time of the 12-month blood sample. Numbers of HCW with a positive antibody titer are shown at the top of each plot. The proportion of HCW with a high (black), low (grey) or no antibody (white) titer is shown in doughnut plots below. Fifteen Recovered HCW (closed circle, 57% male) and 7 HCW with Persistent symptoms (open circle, 57% male) were assayed for C Neutralizing antibody IC50 against live ancestral (Wuhan Hu-1) SARS-CoV-2 virus and D T cell responses against spike and nucleocapsid mapped epitope peptide pools, 12 months after laboratory confirmed SARS-CoV-2 infection. Numbers of HCW showing a positive neutralizing antibody or T cell response for each assay are shown at the top of each plot. The proportion of HCW in each group with a high (black), low (grey) or no response (white) (as defined in each key) are represented by doughnut plots below. Two-tailed Mann-Whitney U test (Graphpad Prism version 8.0) was used to test for significant differences between Recovered HCW and those reporting Persistent symptoms. Source data are provided as a Source Data file. HCW with serological evidence of a subsequent SARS-CoV-2 re-infection between 6 and 12 months are annotated in green. HCW who reported persistent shortness of breath at 12 months are shown as purple open circles. Error bars shown are geometric mean + 1 geometric SD. Ab antibody, COI cut-off index, HCW health care workers, N nucleocapsid, PBMC peripheral blood mononuclear cells, RBD receptor binding domain, SD standard deviation, SFC spot forming cells, U units.