SARS-CoV-2 memory B and T cell profiles in mild COVID-19 convalescent patients

Abstract

Objectives: Antiviral adaptive immunity involves memory B cells (MBC) and memory T cells (MTC). The dynamics of MBC and MTC in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) convalescents warrant further investigation.

Methods: In this cross-sectional and longitudinal study, blood-derived MBC and MTC responses were evaluated in 68 anti-spike IgG-positive mild coronavirus disease 2019 (COVID-19) convalescents at visit 1, between 1 and 7 months (median 4.1 months) after disease onset. SARS-CoV-2 anti-spike IgG was determined by ELISA, MBC by SARS-CoV-2-specific receptor binding domain (RBD) ELISpot, and interferon gamma (IFN-γ)-, interleukin 2 (IL2)-, and IFN-γ+IL2-secreting MTC by IFN-γ and IL2 SARS-CoV-2 FluoroSpot. For 24 patients sampled at the first visit, the IgG, MBC, and MTC analyses were also performed 3 months later at the second visit.

Results: Seventy-two percent of convalescents were both MBC- and MTC-positive, 18% were MBC-positive and MTC-negative, and 10% were MTC-positive and MBC-negative. The peak MBC response level was detected at 3 months after COVID-19 onset and persisted up to 7 months post infection. Significant MTC levels were detected 1 month after onset in response to S1, S2_N, and SNMO peptide pools. The frequency and magnitude of the MTC response to SNMO was higher than those to S1 and S2_N. Longitudinal analysis demonstrated that even when specific humoral immunity declined, the cellular immunity persisted.

Conclusions: The study findings demonstrate the durability of adaptive cellular immunity at least for 7 months after SARS-CoV-2 infection, suggesting long-lasting protection.

Keywords: Convalescents; IgG; Memory B cells; Memory T cells; SARS-CoV-2.

Figure 1

Study design. MBC, memory B cells; MTC, memory T cells.

Figure 2

Representative images for the MTC and MBC tests. (A) MTC test: PBMCs (250 000 cells/well) were stimulated for 48 h with S1, S2_N, and SNMO of the Mabtech SARS-CoV-2 peptide pools or αCD3 as positive control. The MTC were analyzed on the Mabtech IRIS reader using an IFN-γ or IL2 FluoroSpot assay. (B) MBC test: PBMCs were stimulated with R848 and IL2 for 96 h (S). After pre-stimulation, 250 000 cells/well were incubated on an anti-IgG FluoroSpot plate for 18 h. RBD-specific MBC were detected using RBD–streptactin-550 conjugate. The analysis was performed on the Mabtech IRIS reader. U, unstimulated cells; S, stimulated cells; MTC, memory T cells; MBC, memory B cells.

Figure 3

Dynamics of the humoral and adaptive immune response in convalescent patients. (A) Dynamics of anti-SARS-CoV-2 MBC. The number of subjects per group was as follows: HS, n = 16; 1 month, n = 13; 3 months, n = 21; 5 months, n = 19; 7 months, n = 15. The dashed line denotes the cutoff level in HS (6.8 SFU), calculated as 90% of confidence interval. (B) Dynamics of IFN-γ MTC. (C) Dynamics of IL2 MTC. (D) Dynamics of IFN-γ+IL2 MTC. For graphs B, C, and D, the number of subjects per group was as follows: HS, n = 12; 1 month, n = 9; 3 months, n = 9; 5 months, n = 13; 7 months, n = 10. The dashed line denotes the cutoff level in HS (0.0 SFU) calculated as 90% of confidence interval. HS, healthy subject controls; MBC, memory B cells; IFN-γ MTC, overall IFN-γ-producing memory T cells calculated as the sum of the positive SFU of IFN-γ-secreting cells in response to S1, S2_N, and SNMO; IL2 MTC, overall IL2-producing memory T cells calculated as the sum of the positive SFU of IL2-secreting cells in response to S1, S2_N, and SNMO; IFN-γ+IL2 MTC, overall IFN-γ+IL2-producing memory T cells calculated as the sum of the positive SFU of IFN-γ+IL2-secreting cells in response to S1, S2_N, and SNMO. *P < 0.03 by Mann–Whitney U-test as compared to HS. NS is non-significant as compared to HS. In graphs A, B, C, and D, no significant difference in SFU levels between the different time points was observed.

Figure 4

Correlation between adaptive immune memory components. (A) Correlation (by Spearman rank test) between anti-S1 spike IgG and MBC levels. (B) Correlation between overall IL2-secreting MTC and MBC levels. (C) Correlation between overall IFN-γ-secreting MTC and MBC. (D) Correlation between overall IFN-γ and IL2-secreting MTC. SFU, spot-forming units; MBC, memory B cells; IFN-γ MTC, overall IFN-γ-producing memory T cells calculated as the sum of the positive SFU of IFN-γ-secreting cells in response to S1, S2_N, and SNMO; IL2 MTC, overall IL2-producing memory T cells calculated as the sum of the positive SFU of IL2-secreting cells in response to S1, S2_N, and SNMO.

Figure 5

Dynamics of SARS-CoV-2 IFN-γ-, IL2-, and IFN-γ+IL2-secreting MTC cells in response to SNMO peptides. (A) Dynamics of IFN-γ-secreting MTC in response to the SNMO peptide pool. (B) Dynamics of IL2-secreting MTC in response to the SNMO peptide pool. (C) Dynamics of IFN-γ+IL2-secreting MTC in response to the SNMO peptide pool. The number of subjects per group was as follows: HS, n = 12; 1 month, n = 9; 3 months, n = 9; 5 months, n = 13; 7 months, n = 10. The dashed line denotes the cutoff level in the HS (0.0 SFU) calculated as 90% of confidence interval. HS, healthy subjects; SFU, spot-forming units; TFO, time from COVID-19 onset; SNMO IFN-γ MTC, IFN-γ-secreting memory T cells in response to SNMO; SNMO IL2 MTC, IL2-secreting memory T cells in response to SNMO; SNMO IFN-γ+IL2 MTC, IFN-γ+IL2-secreting memory T cells in response to SNMO. *P < 0.05 by Mann–Whitney U-test as compared to HS. NS is non-significant as compared to HS. In graphs A, B, and C, no significant difference in SFU levels between the different time points were observed.

Figure 6

MBC levels in patients with decreased anti-spike IgG. (A) Convalescent patients switching from positive to negative IgG level between visit 1 (black dots) and visit 2 (gray dots). (B) MBC changes in convalescent patients switching from positive to negative IgG level between visit 1 (black dots) and visit 2 (gray dots). (C) Patients with a decreased IgG level, yet remaining IgG-positive between the first (black dots) and second (gray dots) visits. (D) MBC changes in patients with a decreased IgG level, yet remaining IgG-positive. MBC, memory B cells.