PMC8515133_01
Normal generation of SARS-CoV-2-specific CD4+ and CD8+ T cells in the HVDRR patient and her parents. (A) SARS-CoV-2-specific CD4+ T cells measured as percentage of AIM+ (CD137+OX40+) CD4+ T cells after stimulation of PBMC with peptide pools encompassing the first half of the spike protein (S1), the second half of the spike protein (S2), the membrane protein (M) and the nucleoprotein (N). (-) represents control PBMC treated with the vehicle DMSO. Data are shown as mean +- SEM; *p < 0.05. Samples were from the HVDRR patient (filled circles), pater (half-filled squares) and mater (half-filled circles). (B) FACS dot plots showing the fraction of AIM+ (CD137+OX40+) CD4+ T cells. (C) SARS-CoV-2-specific CD8+ T cells measured as percentage of AIM+ (CD137+CD69+) CD8+ T cells after stimulation of PBMC with peptide pools as described in (A). Data are shown as mean +- SEM. *p < 0.05; ***p < 0.005. Symbols indicate the individuals as described in (A). (D) FACS dot plots shoving the fraction AIM+ (CD137+CD69+) CD8+ T cells.
fimmu-12-758154-g001.jpg
PMC8515133_01_fimmu-12-758154-g001.jpg
f1
In all three family members, we found a clear induction of both CD4+AIM+ ( Figures 1A, B , for gating strategy please see  Supplementary Figure 1A ) and CD8+AIM+ cells ( Figures 1C, D ) with frequencies comparable to those previously reported for SARS-CoV-2-specific CD4+ and CD8+ memory T cells in COVID-19 convalescent individuals. SARS-CoV-2-specific T cell responses, we determined the T cell cytokine profile in PBMC stimulated with SARS-CoV-2 peptides for 24 h. In line with other studies, we found a clear induction of IFNgamma+CD4+ T cells in PBMC stimulated with the spike peptide pools and membrane peptides ( Figure 2A , for gating strategy please see  Supplementary    Figure 1B ). Stimulation with peptides from the nucleoprotein did not result in any significant induction of IFNgamma+CD4+ T cells.
Normal cytokine profile of SARS-CoV-2-specific T cells from the HVDRR patient and her parents. Frequency of IFNgamma+
(A) CD4+ and (B) CD8+ T cells in PBMC treated with DMSO (-) or peptide pools encompassing the first half of the spike protein (S1), the second half of the spike protein (S2), the membrane protein (M) and the nucleoprotein (N). Samples were from the HVDRR patient (filled circles), pater (half-filled squares) and mater (half-filled circles). Data are shown as mean +- SEM. (C-H) Cytokine mRNA and cytokine concentration in the supernatants of PBMC obtained from the HVDRR patient in 2018 and from the HVDRR patient and her parents in 2021 and stimulated as described above. The mRNA expression levels of (C) IFNgamma, (E) IL-2 and (G) TNF are shown as geometric mean +- SD. Concentrations of (D) IFNgamma, (F) IL-2 and (H) TNF in the supernatants are shown as mean +- SEM; *p < 0.05; **p < 0.01; ***p < 0.005; ****p < 0.001.
fimmu-12-758154-g002.jpg
PMC8515133_01_fimmu-12-758154-g002.jpg
f2
SARS-CoV-2-specific T cell responses, we determined the T cell cytokine profile in PBMC stimulated with SARS-CoV-2 peptides for 24 h. In line with other studies, we found a clear induction of IFNgamma+CD4+ T cells in PBMC stimulated with the spike peptide pools and membrane peptides ( Figure 2A , for gating strategy please see  Supplementary    Figure 1B ). Stimulation with peptides from the nucleoprotein did not result in any significant induction of IFNgamma+CD4+ T cells. T cells in PBMC stimulated with the spike peptide pools and membrane peptides but not with the peptides from the nucleoprotein ( Figure 2B ). Thus, SARS-CoV-2-specific T cells from the HVDRR patient and her parents were clearly able to produce IFNgamma. Figures 2C, D ). Similar results were seen for TNF ( Figures 2E-H ). Furthermore, we determined the secretion of the Th2 cytokines IL-4 and IL-13 following activation with the SARS-CoV-2 peptides. In contrast to IFNgamma, IL-2 and TNF, the SARS-CoV-2 peptides did not induce production of IL-4 and IL-13 ( Supplementary Figures 2A, B ). Thus, the post-infectious SARS-CoV-2-specific T cells isolated from the HVDRR patient, and her parents had a Th1-like cytokine profile as previously described for SARS-CoV-2-specific T cells.
Normal SARS-CoV-2 RBD-specific IgG antibody titers in the plasma of the HVDRR patient and her parents. SARS-CoV-2 RBD-specific IgG antibody titers in the plasma from unexposed subjects (n = 200), subjects with a mild course of COVID-19 (n = 72) and from the HVDRR patient and her parent from 2018 and 21 days after COVID-19 in 2021. The dashed line represents the cut-off for the ELISA. Antibody titers are shown for each individual and as mean +- SEM; ***p < 0.005, ****p < 0.001. HVDRR patient (filled circles), pater (half-filled squares) and mater (half-filled circles).
fimmu-12-758154-g003.jpg
PMC8515133_01_fimmu-12-758154-g003.jpg
f3
IgG titers in plasma from 2018 from the HVDRR patient and her parents were similar to the anti-RBD IgG titers in plasma from unexposed healthy individuals ( Figure 3 ). Importantly, post-infection plasma from the HVDRR patient and her parents showed similar levels of IgG antibodies against the RBD as plasma from SARS-CoV-2 convalescent in the general population that had had a mild course of COVID-19 ( Figure 3 ).