T-cell and antibody responses to first BNT162b2 vaccine dose in previously infected and SARS-CoV-2-naive UK health-care workers: a multicentre prospective cohort study

Abstract

Background: Previous infection with SARS-CoV-2 affects the immune response to the first dose of the SARS-CoV-2 vaccine. We aimed to compare SARS-CoV-2-specific T-cell and antibody responses in health-care workers with and without previous SARS-CoV-2 infection following a single dose of the BNT162b2 (tozinameran; Pfizer-BioNTech) mRNA vaccine.

Methods: We sampled health-care workers enrolled in the PITCH study across four hospital sites in the UK (Oxford, Liverpool, Newcastle, and Sheffield). All health-care workers aged 18 years or older consenting to participate in this prospective cohort study were included, with no exclusion criteria applied. Blood samples were collected where possible before vaccination and 28 (±7) days following one or two doses (given 3-4 weeks apart) of the BNT162b2 vaccine. Previous infection was determined by a documented SARS-CoV-2-positive RT-PCR result or the presence of positive anti-SARS-CoV-2 nucleocapsid antibodies. We measured spike-specific IgG antibodies and quantified T-cell responses by interferon-γ enzyme-linked immunospot assay in all participants where samples were available at the time of analysis, comparing SARS-CoV-2-naive individuals to those with previous infection.

Findings: Between Dec 9, 2020, and Feb 9, 2021, 119 SARS-CoV-2-naive and 145 previously infected health-care workers received one dose, and 25 SARS-CoV-2-naive health-care workers received two doses, of the BNT162b2 vaccine. In previously infected health-care workers, the median time from previous infection to vaccination was 268 days (IQR 232-285). At 28 days (IQR 27-33) after a single dose, the spike-specific T-cell response measured in fresh peripheral blood mononuclear cells (PBMCs) was higher in previously infected (n=76) than in infection-naive (n=45) health-care workers (median 284 [IQR 150-461] vs 55 [IQR 24-132] spot-forming units [SFUs] per 10⁶ PBMCs; p<0·0001). With cryopreserved PBMCs, the T-cell response in previously infected individuals (n=52) after one vaccine dose was equivalent to that of infection-naive individuals (n=19) after receiving two vaccine doses (median 152 [IQR 119-275] vs 162 [104-258] SFUs/10⁶ PBMCs; p=1·00). Anti-spike IgG antibody responses following a single dose in 142 previously infected health-care workers (median 270 373 [IQR 203 461-535 188] antibody units [AU] per mL) were higher than in 111 infection-naive health-care workers following one dose (35 001 [17 099-55 341] AU/mL; p<0·0001) and higher than in 25 infection-naive individuals given two doses (180 904 [108 221-242 467] AU/mL; p<0·0001).

Interpretation: A single dose of the BNT162b2 vaccine is likely to provide greater protection against SARS-CoV-2 infection in individuals with previous SARS-CoV-2 infection, than in SARS-CoV-2-naive individuals, including against variants of concern. Future studies should determine the additional benefit of a second dose on the magnitude and durability of immune responses in individuals vaccinated following infection, alongside evaluation of the impact of extending the interval between vaccine doses.

Funding: UK Department of Health and Social Care, and UK Coronavirus Immunology Consortium.

Figure 1

T-cell responses following BNT162b2 vaccine in SARS-CoV-2-naive and previously infected individuals (A) IFNγ ELISpot responses to a single dose in 45 infection-naive and 76 previously infected individuals (Sheffield cohort) with freshly isolated PBMCs. Summed responses from four overlapping peptide pools spanning the entire spike, nucleocapsid, and membrane protein pool responses are shown. (B) Comparison of IFNγ ELISpot responses with cryopreserved PBMCs in 58 infection-naive and 52 previously infected individuals a median of 28 days (IQR 26–33) following a single dose, along with 19 infection-naive individuals a median of 28 days (27–32) following two doses. Responses to peptide pools representing the S1 and S2 subunits of the spike protein, and a combined pool presenting membrane and nucleocapsid proteins, are shown. Pooled data from the Liverpool, Newcastle, and Oxford cohorts are shown, with centre-stratified data shown in appendix 1 (p 11). (C–E) Intracellular cytokine staining in individuals who received one dose of vaccine. Expression levels of IFNγ, IL-2, and TNFα in CD4+ T cells of 31 previously infected and 32 infection-naive individuals to the S1 protein (C) and S2 protein (D). (E) The proportion of IFNγ production from CD4+ T cells, calculated by dividing the proportion of IFNγ-positive CD4+ T cells by the total IFNγ-positive CD4+ and CD8+ T cells. Bars represent medians and IQRs. ELISpot=enzyme-linked immunospot. IFNγ=interferon-γ. IL-2=interleukin-2. PBMCs=peripheral blood mononuclear cells. SFUs=spot-forming units. TNFα=tumour necrosis factor-α.

Figure 2

T-cell responses across spike protein following BNT162b2 vaccine with IFNγ ELISpot (A) Representation of spike protein showing positions of overlapping peptide pools used in T-cell assays in relation to S1 and S2 subunits of the spike protein, NTD, RBD, and transmembrane domain. (B, C) IFNγ ELISpot responses to a single dose in 76 previously infected (B) and 45 infection-naive individuals (C) with freshly isolated PBMCs, stratified as responses to peptide pools representing amino acids 1–330, 321–645, 636–960, and 950–1273 of the spike protein. Bars represent medians and IQRs. (D) Changes in the contribution from different peptide pools to the total spike T-cell response before and after a single dose in 76 previously infected individuals. Each column represents responses from a single individual, with responses from the same individual before and after the first dose aligned in the two plots. ELISpot=enzyme-linked immunospot. IFNγ=interferon-γ. NTD=N-terminal domain. PBMCs=peripheral blood mononuclear cells. RBD=receptor binding domain. SFUs=spot-forming units.

Figure 3

Antibody responses following BNT162b2 vaccine in infection-naive and previously infected individuals (A) Comparison of anti-spike antibody responses in 111 infection-naive and 142 previously infected individuals following a single dose and 25 infection-naive individuals following two doses. Bars represent geometric means and 95% CIs. The horizontal dotted line denotes the threshold for positivity in MesoScale Discovery SARS-CoV-2 spike assay based on mean plus 3 SD from 103 pre-pandemic negative controls (1160·3 AU/mL). Antibody titres calibrated to the WHO international standard for anti-SARS-CoV-2 immunoglobulin (National Institute for Biological Standards and Control 20/136) are shown in appendix 1 (p 14). (B) Correlation between antibody responses to the spike protein and the RBD following vaccination. Dotted lines denote the threshold for positivity in the MesoScale Discovery SARS-CoV-2 spike (1160·3 AU/mL) and RBD (1169·0 AU/mL) assay defined by pre-pandemic negative controls. (C) Relationship between time from positive SARS-CoV-2 PCR to first vaccine dose and post-vaccine anti-spike antibodies in 104 previously infected individuals. (D) Comparison of anti-spike T-cell responses measured by interferon-γ enzyme-linked immunospot and anti-spike antibodies following vaccination. Correlation coefficient (Spearman's ρ, r_s) between T-cell and antibody responses following a single dose of the BNT162b2 vaccine displayed separately for 97 infection-naive and 126 previously infected individuals. Pooled responses from the entire cohort are displayed. The same data stratified by T-cell assay (fresh vs cryopreserved PBMCs) are shown in appendix 1 (p 17). Antibody data are presented on a log₁₀ scaled axis for visualisation, with statistical comparisons done on untransformed data. AU=antibody units. PBMCs=peripheral blood mononuclear cells. RBD=receptor binding domain. SFUs=spot forming units.

Figure 4

Antibody responses following BNT162b2 vaccine in naive and previously infected individuals to seasonal human coronaviruses and SARS-CoV-2 variants of concern (A) Comparison of antibody responses to spike proteins from seasonal human coronaviruses 229E, NL63, HKU1, and OC43 before and after vaccination in 111 infection-naive and 142 previously infected individuals following a single dose and 25 infection-naive individuals following two doses. (B) Surrogate neutralisation activity before and after one vaccine dose in ten naive and ten previously infected individuals to spike proteins from SARS-CoV-2 variants, including variants of concern. Activity is expressed as units per mL with 1 unit per mL equivalent to 1 μg/mL of neutralising activity of the anti-spike monoclonal antibody standard. Thresholds for positivity, based on mean plus 3 SD from 23 pre-pandemic negative control samples, were 1·02 units per mL for D614G, 1·13 units per mL for the alpha (B.1.1.7) variant, 0·93 units per mL for the beta (B.1.351) variant, and 0·98 units per mL for the gamma (P.1) variant. The horizontal dashed line denotes 1 unit per mL. (C) Ability of plasma from ten infection-naive and ten previously infected individuals, before and after a single vaccine dose, to neutralise live virus, expressed as the reciprocal titre required for 50% reduction in infectious focus-forming units (NT50) of lineage B and the beta variant of SARS-CoV-2 in a microneutralisation assay. Microneutralisation data from the 25 naive individuals sampled at 7 days following two doses have been previously described and are presented here for comparison. (D) Reduction in neutralisation titres (NT50) for each plasma sample against the beta variant compared to a B lineage virus. Antibody data are presented on a log₁₀ scaled axis for visualisation, with statistical comparisons done on untransformed data. AU=antibody units.