Ancestral SARS-CoV-2-specific T cells cross-recognize Omicron (B.1.1.529)

Ancestral SARS-CoV-2-specific T cells cross-recognize Omicron (B.1.1.529) | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Short Report Ancestral SARS-CoV-2-specific T cells cross-recognize Omicron (B.1.1.529) Yu Gao, Curtis Cai, Alba Grifoni, Thomas Müller, Julia Niessl, and 15 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-1217466/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract The emergence of the SARS-CoV-2 variant-of-concern Omicron (B.1.1.529) has destabilized global efforts to control the impact of COVID-19. Recent data have suggested that B.1.1.529 can readily infect people with naturally acquired or vaccine-induced immunity, facilitated in some cases by viral escape from antibodies that neutralize ancestral SARS-CoV-2. However, severe disease appears to be relatively uncommon in such individuals, highlighting a potential role for other components of the adaptive immune system. We report here that SARS-CoV-2 spike-specific CD4 + and CD8 + T cells induced by prior infection and, more extensively, by mRNA vaccination provide comprehensive heterologous immune reactivity against B.1.1.529. Pairwise comparisons across groups further revealed that SARS-CoV-2 spike-reactive CD4 + and CD8 + T cells exhibited similar functional attributes, memory distributions, and phenotypic traits in response to the ancestral strain or B.1.1.529. Our data indicate that established SARS-CoV-2 spike-specific CD4 + and CD8 + T cell responses, especially after mRNA vaccination, remain largely intact against B.1.1.529. Immunology Infectious Diseases B.1.1.529 Omicron COVID-19 SARS-CoV-2 T cells Figures Figure 1 Figure 2 Main Natural infection with SARS-CoV-2 and vaccination with mRNA constructs encoding the viral spike protein typically generate effective immunity against COVID-19. However, the current pandemic has been fueled by the continual emergence of variants-of-concern (VOCs), such as Omicron (B.1.1.529). Emerging data indicate that B.1.1.529 is more transmissible than previous VOCs 1 . This phenotype can be explained by key mutations in the receptor-binding domain (RBD), which confer enhanced affinity for the ACE2 receptor 2 . Another major concern is that B.1.1.529 harbors a large number of additional mutations in the spike protein that could feasibly subvert immune recognition 3 . In line with this possibility, emerging reports have shown that neutralizing antibodies elicited against the ancestral Wuhan reference strain, either in the context of infection or vaccination, are less able to combat B.1.1.529 2,4 . These observations likely align with the propensity of B.1.1.529 to cause breakthrough infections, also in persons who have been vaccinated with two doses of mRNA vaccine or experienced previous SARS-CoV-2 infection 5 , 6 . Preliminary data suggest that breakthrough infections with B.1.1.529 might be associated with a lower risk of hospitalization and less often cause severe disease compared with the Delta VOC (B.1.617.2) 7 , 8 . One possible inference from the less aggressive clinical spectrum is that additional immune components beyond the antibody response attenuate the course of infection with B.1.1.529. Previous studies have demonstrated that robust CD4 + and CD8 + T cell responses are induced following SARS-CoV-2 infection or vaccination 9 – 15 . Several lines of evidence further suggest that CD4 + and CD8 + T cell responses can modulate disease severity in humans and suppress viral replication in animal models 16 – 19 . However, it has remained unclear to what extent ancestral SARS-CoV-2-specific CD4 + and CD8 + T cells cross-recognize B.1.1.529, especially given the unprecedented number of mutations in the spike protein, which likely shift the antigenic landscape more profoundly in relation to antecedent VOCs 20 . To address this question, we collected peripheral blood mononuclear cells from mRNA-vaccinated individuals 6 months after the second dose of Pfizer/BioNTech BNT162b2 ( n = 40), individuals in the convalescent phase 9 months after mild or severe COVID-19 ( n = 48), and seronegative individuals ( n = 48). Cells were stimulated in parallel with overlapping peptide pools spanning the entire spike protein sequences of the Wuhan reference strain (wildtype) or B.1.1.529. Activation-induced marker assays were used to quantify spike-specific CD4 + T cell responses via the upregulation of CD69 and CD40L (CD154) and spike-specific CD8 + T cell responses via the upregulation of CD69 and 4-1BB (CD137) (Extended Data Fig. 1a). The overall magnitude of the SARS-CoV-2 spike-specific CD4 + T cell response against B.1.1.529 showed a median reduction of 9% in mRNA-vaccinated individuals and a median reduction of 16% in convalescent individuals relative to the wildtype response (Fig. 1a, b). The corresponding response frequencies were also slightly lower for B.1.1.529 (Fig. 1c). Individual comparisons further revealed maximum reductions in magnitude of 58% in mRNA-vaccinated individuals, 56% in convalescent individuals, and 75% in seronegative individuals, comparing SARS-CoV-2 spike-specific CD4 + T cell responses against B.1.1.529 versus wildtype (Fig. 1d). These findings were validated using independently synthesized peptide pools spanning the entire spike protein (Extended Data Fig. 1b). To extend these findings, we investigated the phenotypic characteristics of SARS-CoV-2 spike-specific CD4 + T cells that cross-recognized B.1.1.529, with a particular focus on markers of T helper polarization (CCR4, CCR6, CXCR3, CXCR5) and memory differentiation (CCR7, CD45RA). No significant differences in T helper polarization were detected across intragroup comparisons of SARS-CoV-2 spike-specific CD4 + T cell responses against B.1.1.529 versus wildtype (Fig. 1e). Central memory T (T CM ) cells predominated among SARS-CoV-2 spike-specific CD4 + T cells in mRNA-vaccinated, convalescent, and seronegative individuals, but again, the subset composition of SARS-CoV-2 spike-specific CD4 + T cell responses against B.1.1.529 versus wildtype were similar across all groups (Fig. 1f). We also assessed the functionality of SARS-CoV-2 spike-specific CD4 + T cells in mRNA-vaccinated individuals, measuring the intracellular expression of IFN-γ, TNF, and IL-2 alongside CD69 and CD154. No significant differences in the ability of SARS-CoV-2 spike-specific CD4 + T cells to deploy multiple functions were apparent in response to stimulation with peptides representing B.1.1.529 versus wildtype (Fig. 1g). The overall magnitude of the SARS-CoV-2 spike-specific CD8 + T cell response against B.1.1.529 showed a median reduction of 8% in mRNA-vaccinated individuals and a median reduction of 30% in convalescent individuals relative to the wildtype response (Fig. 2a, b). These differences were mirrored in the corresponding response frequencies for B.1.1.529 versus wildtype (Fig. 2c). Individual comparisons further revealed maximum reductions in magnitude of 55% in mRNA-vaccinated individuals, 63% in convalescent individuals, and 60% in seronegative individuals, comparing SARS-CoV-2 spike-specific CD8 + T cell responses against B.1.1.529 versus wildtype (Fig. 2d). These findings were again validated using independently synthesized peptide pools spanning the entire spike protein (Extended Data Fig. 1c). In further experiments, we investigated the phenotypic characteristics of SARS-CoV-2 spike-specific CD8 + T cells that cross-recognized B.1.1.529, focusing on classic markers of memory differentiation (CCR7, CD45RA). Late effector memory T (T EMRA ) cells predominated among SARS-CoV-2 spike-specific CD8 + T cells in mRNA-vaccinated, convalescent, and seronegative individuals, but no significant differences in subset composition were detected across intragroup comparisons of SARS-CoV-2 spike-specific CD8 + T cell responses against B.1.1.529 versus wildtype (Fig. 2e). We also assessed the functionality of SARS-CoV-2 spike-specific CD8 + T cells in mRNA-vaccinated individuals, measuring the intracellular expression of granzyme B, IFN-γ, TNF, and IL-2 alongside CD69 and CD137. Akin to the corresponding analyses of SARS-CoV-2 spike-specific CD4 + T cells, no significant differences in the ability of SARS-CoV-2 spike-specific CD8 + T cells to deploy multiple functions were apparent in response to stimulation with peptides representing B.1.1.529 versus wildtype (Fig. 2f). In a final set of analyses, we merged all SARS-CoV-2 spike-specific CD4 + and CD8 + T cell data points together and found a significantly lower total T cell response against B.1.1.529 versus wildtype in convalescent, but not vaccinated subjects (Extended Data Fig. 1d). These analyses suggest that SARS-CoV-2 spike-specific T cell responses remain less intact against B.1.1.529 in convalescent individuals. The current global pandemic has been destabilized by the recent emergence of B.1.1.529, which continues to spread rapidly and supersede other VOCs. Our collective data indicate that SARS-CoV-2 spike-specific CD4 + and CD8 + T cells elicited by mRNA vaccination or prior infection remain largely intact against B.1.1.529. Alongside intrinsic viral factors, such as altered tropism and decreased replication in the lower respiratory tract 21 , such heterologous immune reactivity may explain why severe disease appears to be relatively uncommon after infection with this particular VOC. Moreover, the degree of cross-reactivity varied to some extent among individuals, most likely as a consequence of genetically encoded differences in antigen presentation, which could further modulate clinical outcomes associated with B.1.1.529. It should be noted that our evaluations were confined to peripheral blood samples, which do not necessarily reflect the entirety of the cellular immune response against SARS-CoV-2 22 . In addition, SARS-CoV-2 spike-specific CD4 + and CD8 + T cells cross-recognized B.1.1.529 less comprehensively in convalescent versus mRNA-vaccinated individuals, suggesting that booster immunization may provide benefits that extend beyond the induction of broadly neutralizing antibodies to enhance natural protection against recurrent episodes of COVID-19 2 . Declarations ACKNOWLEDGEMENTS This study was supported by grants from the SciLifeLab National COVID-19 Research Program, financed by the Knut and Alice Wallenberg Foundation, the Swedish Research Council, Nordstjernan AB, Region Stockholm, and the Karolinska Institutet. M.B. was supported by the Swedish Research Council, the Knut and Alice Wallenberg Foundation, the Karolinska Institutet, the Swedish Society of Medicine, the Swedish Cancer Society, the Swedish Childhood Cancer Fund, the Åke Wibergs Stiftelse, and the Jonas Söderquist Stiftelse. J.N. was supported by an EMBO Postdoctoral Fellowship (ALTF 1062-2020). This project has been funded in whole or in part with Federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under Contract No. 75N93021C00016 (to A.S.), Contract No. 75N9301900065 (to A.S, D.W.) and by the National Institute for Health Research via grant COV-LT2-0041 awarded (to D.A.P.). AUTHOR CONTRIBUTIONS Conceptualization: Y.G., C.C., J.K.S., D.A.P., H.G.L., A.C.K., A.S., S.A., M.B. Sample collection: Y.G., C.C., T.R.M., J.N., L.H., A.Ö., P.B., P.C., A.OLS., H.G.L., S.A., M.B. Investigation: Y.G., C.C., A.G., A.OLO., M.H., M.B. Formal analysis: Y.G., C.C. Visualization: Y.G., C.C., M.B. Resources: A.G., J.K.S., D.W., H.G.L, A.C.K., A.S., S.A., M.B. Funding acquisition: D.A.P., H.G.L., A.S., S.A., M.B. Supervision: J.K.S., H.G.L., S.A., M.B. Writing—original draft: Y.G., C.C., D.A.P., M.B. Writing—review and editing: Y.G., C.C., A.G., T.R.M., J.N., A.Ö., P.B., J.K.S., D.A.P., H.G.L., A.C.K., S.A., M.B. DECLARATION OF INTERESTS A.S. is a consultant for Gritstone Bio, Flow Pharma, Arcturus Therapeutics, ImmunoScape, CellCarta, Avalia, Moderna, Fortress and Repertoire., and the La Jolla Institute has filed patents to protect various aspects of the T cell epitope and vaccine design work. SA has received honoraria for lectures from Gilead, AbbVie, MSD, Biogen, and has received research grants from Gilead and AbbVie. M.B. is a consultant for Oxford Immunotec. The other authors declare no conflicts of interest. References 1. Dyer, O. Covid-19: Omicron is causing more infections but fewer hospital admissions than delta, South African data show. BMJ 375 , n3104 (2021). 2. Cameroni, E. , et al. Broadly neutralizing antibodies overcome SARS-CoV-2 Omicron antigenic shift. bioRxiv (2021). 3. https://www.ecdc.europa.eu/sites/default/files/documents/threat-assessment-covid-19-emergence-sars-cov-2-variant-omicron-december-2021.pdf . 4. Liu L. , et al. Striking antibody evasion manifested by the Omicron variant of SARS-CoV-2. Nature 23 December 2021 (2021). 5. Collie, S., Champion, J., Moultrie, H., Bekker, L.G. & Gray, G. Effectiveness of BNT162b2 Vaccine against Omicron Variant in South Africa. N Engl J Med (2021). 6. Goga, A. , et al. Breakthrough Covid-19 infections during periods of circulating Beta, Delta and Omicron variants of concern, among health care workers in the Sisonke Ad26.COV2.S vaccine trial, South Africa. MedRxiv (2021). 7. Neil Ferguson, A.G., Wes Hinsley and Erik Volz on behalf of the Imperial College COVID-19 response team. Report 50: Hospitalisation risk for Omicron cases in England. (2021). 8. Wolter N. , et al. Early assessment of the clinical severity of the SARS-CoV-2 Omicron variant in South Africa. bioRxiv (2021). 9. Braun, J. , et al. SARS-CoV-2-reactive T cells in healthy donors and patients with COVID-19. Nature 587 , 270-274 (2020). 10. Grifoni, A. , et al. Targets of T Cell Responses to SARS-CoV-2 Coronavirus in Humans with COVID-19 Disease and Unexposed Individuals. Cell 181 , 1489-1501 e1415 (2020). 11. Kalimuddin, S. , et al. Early T cell and binding antibody responses are associated with COVID-19 RNA vaccine efficacy onset. Med (N Y) 2 , 682-688 e684 (2021). 12. Le Bert, N. , et al. SARS-CoV-2-specific T cell immunity in cases of COVID-19 and SARS, and uninfected controls. Nature 584 , 457-462 (2020). 13. Oberhardt, V. , et al. Rapid and stable mobilization of CD8(+) T cells by SARS-CoV-2 mRNA vaccine. Nature 597 , 268-273 (2021). 14. Peng, Y. , et al. Broad and strong memory CD4(+) and CD8(+) T cells induced by SARS-CoV-2 in UK convalescent individuals following COVID-19. Nat Immunol 21 , 1336-1345 (2020). 15. Sekine, T. , et al. Robust T Cell Immunity in Convalescent Individuals with Asymptomatic or Mild COVID-19. Cell 183 , 158-168 e114 (2020). 16. Israelow, B. , et al. Adaptive immune determinants of viral clearance and protection in mouse models of SARS-CoV-2. Sci Immunol 6 , eabl4509 (2021). 17. McMahan, K. , et al. Correlates of protection against SARS-CoV-2 in rhesus macaques. Nature 590 , 630-634 (2021). 18. Rydyznski Moderbacher, C. , et al. Antigen-Specific Adaptive Immunity to SARS-CoV-2 in Acute COVID-19 and Associations with Age and Disease Severity. Cell 183 , 996-1012 e1019 (2020). 19. Tan, A.T. , et al. Early induction of functional SARS-CoV-2-specific T cells associates with rapid viral clearance and mild disease in COVID-19 patients. Cell Rep 34 , 108728 (2021). 20. Tarke, A. , et al. Impact of SARS-CoV-2 variants on the total CD4(+) and CD8(+) T cell reactivity in infected or vaccinated individuals. Cell Rep Med 2 , 100355 (2021). 21. Meng B. , et al. , SARS-CoV-2 Omicron spike mediated immune escape, infectivity and cell-cell fusion. bioRxiv (2021). 22. Niessl, J. , et al. Identification of resident memory CD8(+) T cells with functional specificity for SARS-CoV-2 in unexposed oropharyngeal lymphoid tissue. Sci Immunol 6 , eabk0894 (2021). 23. Bergman, P. , et al. Safety and efficacy of the mRNA BNT162b2 vaccine against SARS-CoV-2 in five groups of immunocompromised patients and healthy controls in a prospective open-label clinical trial. EBioMedicine 74 , 103705 (2021). Online Methods Samples Healthy individuals (n=40) were sampled 6 months after the second dose of the BNT162b2 vaccine (Comirnaty ® , Pfizer/BioNTech) as part of a clinical trial registered at EudraCT (2021-000175-37) and https://clinicaltrials.gov (2021-000175-37) at Karolinska University Hospital, Sweden 23 . The two doses of the vaccine were given 21 days apart in standard doses. The vaccinated cohort of healthy individuals consisted of 23 women (58%)/17 men (43%), and the median age (range) was 53 (22-79) years. The study was approved by the Swedish Medical Product Agency. Additional samples from 15 of the vaccinated cohort participants were collected 3 months after the second dose for validation purposes (Extended Data Figure 1). Convalescent individuals after previous COVID-19 were sampled 9 months after RT-PCR verified SARS-CoV-2 infection leading to mild (i.e. non-hospitalized, n=26) or severe (i.e. hospitalized, n=22) disease during the first wave of the pandemic in March-April of 2020, before the emergence of the Alpha, Beta and Delta variants. The cohort of convalescent individuals after mild COVID-19 consisted of 8 women (31%)/18 men (69%), and the median age (range) was 54 (44-68) years. The cohort of convalescent individuals after severe COVID-19 consisted of 3 women (14%)/19 men (86%), and the median age (range) was 58 (33-66) years. Fourteen of the patients with severe COVID-19 (64%) had received intensive care at the time of COVID-19. None of the convalescent individuals had received any COVID-19 vaccination at the time of sample collections. Seronegative samples were acquired from healthy blood donors in late 2020. The absence of spike-specific antibodies was confirmed using an Anti-SARS-CoV-2 S Immunoassay (Roche) for vaccinated healthy individuals and seronegative blood donors. All participants provided written informed consent in accordance with the principles of the Declaration of Helsinki. The study was approved by the Regional Ethics Committee in Stockholm, Sweden. Peripheral blood mononuclear cells (PBMCs) were isolated via standard density gradient centrifugation and cryopreserved in fetal bovine serum (FBS) containing 10% dimethyl sulfoxide (DMSO). Peptides Overlapping peptides were designed to span the entire spike protein sequence of SARS-CoV-2 corresponding to the ancestral Wuhan strain (wildtype) or B.1.1.529. Test peptides comprising 15mers overlapping by 10 amino acids were synthesized as crude material for functional screens (TC Peptide Lab). Validation peptides comprising 20mers overlapping by 10 amino acids were synthesized to an equivalent specification (Sigma-Aldrich). All peptides were reconstituted in DMSO, diluted to stock concentrations of 100 μg/ml in phosphate-buffered saline (PBS), and stored at −20 ºC. Activation-induced marker assays PBMCs were thawed quickly, resuspended in RPMI 1640 containing 10% FBS, 1% L-glutamine, and 1% penicillin/streptomycin (complete medium) in the presence of DNase I (10 U/ml; Sigma-Aldrich), and rested at 1 × 10 6 cells per well in 96-well U-bottom plates (Corning) for 4 h at 37 ºC. The medium was then supplemented with anti-CXCR5–BB515 and anti-CD40 (unconjugated), followed 15 min later by the relevant peptide pools (1 μg/ml/peptide). Negative control wells contained equivalent DMSO. After 12 h, cells were washed in PBS supplemented with 2% FBS and 2 mM EDTA (FACS buffer) and stained with anti-CCR4/CD194–BB700, anti-CCR6/CD196–BUV737, anti-CCR7–APC-Cy7, and anti-CXCR3–AF647 for 10 min at 37 ºC. Additional surface stains were performed for 30 min at room temperature in the presence of Brilliant Stain Buffer Plus (BD Biosciences). Viable cells were identified by exclusion using a LIVE/DEAD Fixable Aqua Dead Cell Stain Kit (Thermo Fisher Scientific). Stained cells were then washed in FACS buffer, fixed in PBS containing 1% paraformaldehyde (PFA; Biotium), and acquired using a FACSymphony A5 (BD Biosciences). The gating strategy is shown in Extended Data Figure 1. All flow cytometry reagents are detailed in Supplementary Table 1. Intracellular cytokine staining PBMCs were thawed quickly, resuspended in complete medium in the presence of DNase I (10 U/ml; Sigma-Aldrich), and rested at 1 × 10 6 cells/well in 96-well U-bottom plates (Corning) for 4 h at 37 ºC. The medium was then supplemented with anti-CXCR5–BB515, followed 15 min later by the relevant peptide pools (1 μg/ml/peptide), and a further 1 h later by brefeldin A (1 μg/ml; Sigma-Aldrich) and monensin (0.7 μg/ml; BD Biosciences). Negative control wells contained equivalent DMSO. After 9 h, cells were washed in FACS buffer and stained with anti-CCR4/CD194–BB700, anti-CCR6/CD196–BUV737, anti-CCR7–APC-Cy7, and anti-CXCR3–BV750 for 10 min at 37 ºC. Additional surface stains were performed for 30 min at room temperature in the presence of Brilliant Stain Buffer Plus (BD Biosciences). Viable cells were identified by exclusion using a LIVE/DEAD Fixable Aqua Dead Cell Stain Kit (Thermo Fisher Scientific). Cells were then washed in FACS buffer and fixed/permeabilized using a FoxP3 Transcription Factor Staining Buffer Set (Thermo Fisher Scientific). Intracellular stains were performed for 30 min at room temperature. Stained cells were washed in FACS buffer, fixed in PBS containing 1% PFA (Biotium), and acquired using a FACSymphony A5 (BD Biosciences). All flow cytometry reagents are detailed in Supplementary Table 1. Data analysis and statistics Flow cytometry data were analyzed using FlowJo version 10.8.0 (FlowJo LLC). Stimulation indices were calculated as fold change in frequency relative to the negative control (equivalent DMSO). Positive responses were identified using a threshold stimulation index >2. Statistical analyses were performed using Prism version 9 (GraphPad). Significance between two paired groups was assessed using the Wilcoxon signed rank test. Functional profiles were deconvoluted using Boolean gating in FlowJo version 10.8.0 (FlowJo LLC) followed by downstream analyses in SPICE version 6.1 (https://niaid.github.io/spice/). Data availability Requests for data should be addressed to the corresponding author and will be made available within the bounds of confidentiality and data protection obligations. Supplementary Files rssupplementarytable1.docx Supplementary Table 1. Flow cytometry reagents. FigS1.png Extended Data Fig. 1. Peptide validation experiments. a, Representative flow cytometry plots showing the gating strategy used to assess spike-specific CD4+ and CD8+ T cell responses to peptide pools representing wildtype SARS-CoV-2 (WT) or B.1.1.529. b,c, Frequencies of spike-specific CD4+ (b) and CD8+ T cells (c) from mRNA-vaccinated individuals, comparing test 15mer peptide pools versus validation 20mer peptide pools representing wildtype SARS-CoV-2 (WT) or B.1.1.529. Each dot represents one donor. Data in dot plots are shown as median ± IQR. d, Pairwise analysis of total spike-specific, CD4+ (red lines) and CD8+ (blue lines), T cell response in vaccinated and convalescent donors against wildtype SARS-CoV-2 (WT) or B.1.1.529. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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Price","email":"","orcid":"","institution":"Cardiff University","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"David","middleName":"A.","lastName":"Price","suffix":""},{"id":73377168,"identity":"b19e5f0e-bd8b-4147-bb41-8a9665fc31b5","order_by":15,"name":"Hans-Gustaf Ljunggren","email":"","orcid":"","institution":"Karolinska Institutet","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Hans-Gustaf","middleName":"","lastName":"Ljunggren","suffix":""},{"id":73377169,"identity":"92392d23-fc14-4282-b113-73fae0d2a5c8","order_by":16,"name":"Annika C. Karlsson","email":"","orcid":"","institution":"Karolinska Institutet","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Annika","middleName":"C.","lastName":"Karlsson","suffix":""},{"id":73377170,"identity":"e093c378-674b-4673-8fd7-b1d3d44961a1","order_by":17,"name":"Alessandro Sette","email":"","orcid":"","institution":"La Jolla Institute for Immunology","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Alessandro","middleName":"","lastName":"Sette","suffix":""},{"id":73377171,"identity":"55f35049-d5b2-4df4-ab4b-b5076ffdef9f","order_by":18,"name":"Soo Aleman","email":"","orcid":"","institution":"Karolinska Institutet","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Soo","middleName":"","lastName":"Aleman","suffix":""},{"id":73395917,"identity":"f9a242a9-a138-4181-8613-43311c4b7cc5","order_by":19,"name":"Marcus Buggert","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA40lEQVRIiWNgGAWjYBCDBChtY8BGopaENNK1HDYgqFS3/ezDzzx/GPIMzi+/+Ljyx3ljPgb2hw/waTE7k24szdvGUGxw402x4ZmE22ZsDDzGeK0yO5DGxszbwJC44caZNMmGhNs2QC1sEni1nH/Gxgx0GEhL+s+GhHNALezPf+DVcgNoCw8bUMv59mOMDQkHgA5jMMOnA6jlGbPk3DaJYskbPMySDWnJxkATjAk4LI3xw5s/Nnl8548//NhgY2c4v7394Qe81kAA0FiJHGg4MROhHgL4jz8gWu0oGAWjYBSMLAAAfMRHnyGCI3gAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0003-0633-1719","institution":"Karolinska Institutet","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Marcus","middleName":"","lastName":"Buggert","suffix":""}],"badges":[],"createdAt":"2021-12-30 17:15:27","currentVersionCode":1,"declarations":{"humanSubjects":true,"vertebrateSubjects":false,"conflictsOfInterestStatement":true,"humanSubjectEthicalGuidelines":true,"humanSubjectConsent":true,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":false,"coiExplicitlySet":false},"doi":"10.21203/rs.3.rs-1217466/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-1217466/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":16946079,"identity":"c3129e86-11d2-4461-9458-c775857cd6d0","added_by":"auto","created_at":"2022-01-03 20:08:24","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":1458364,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eCross-reactive CD4+\u0026nbsp;T cell responses\u0026nbsp;against\u0026nbsp;B.1.1.529.\u003c/strong\u003e\u0026nbsp;\u0026nbsp;\u003c/p\u003e\u003cp\u003e\u003cstrong\u003ea\u003c/strong\u003e, Representative\u0026nbsp;flow cytometry\u0026nbsp;plots showing spike-specific CD4+\u0026nbsp;T cell responses\u0026nbsp;(CD69+CD154+)\u0026nbsp;to peptide pools representing\u0026nbsp;wildtype\u0026nbsp;SARS-CoV-2 (WT) or\u0026nbsp;B.1.1.529.\u0026nbsp;\u003cstrong\u003eb\u003c/strong\u003e, Frequencies of spike-specific CD4+\u0026nbsp;T cells\u0026nbsp;from\u0026nbsp;mRNA-vaccinated, convalescent, and seronegative individuals.\u0026nbsp;Each dot represents one donor.\u0026nbsp;Numbers indicate\u0026nbsp;median reduction\u0026nbsp;in the frequency of detected responses.\u0026nbsp;\u003cstrong\u003ec\u003c/strong\u003e, Stimulation indices calculated as fold change\u0026nbsp;in frequency relative to the\u0026nbsp;negative control.\u0026nbsp;Numbers indicate\u0026nbsp;the percentage of individuals with\u0026nbsp;a\u0026nbsp;detectable response.\u0026nbsp;\u003cstrong\u003ed\u003c/strong\u003e, Cross-reactive responses\u0026nbsp;depicted\u0026nbsp;on an individual basis\u0026nbsp;as\u0026nbsp;percent\u0026nbsp;B.1.1.529/WT.\u0026nbsp;\u003cstrong\u003ee\u003c/strong\u003e, Helper\u0026nbsp;polarization\u0026nbsp;of\u0026nbsp;spike-specific CD4+\u0026nbsp;T cells\u0026nbsp;with representative gating and\u0026nbsp;dot\u0026nbsp;plots showing the distribution\u0026nbsp;of subsets\u0026nbsp;across individuals with detectable responses. Pie charts show the mean frequency of each subset across all individuals\u0026nbsp;in each group.\u0026nbsp;\u003cstrong\u003ef\u003c/strong\u003e, Canonical memory\u0026nbsp;differentiation\u0026nbsp;profiles\u0026nbsp;of spike-specific CD4+\u0026nbsp;T cells\u0026nbsp;with representative gating and\u0026nbsp;dot\u0026nbsp;plots showing the distribution\u0026nbsp;of subsets\u0026nbsp;across individuals with detectable responses.\u0026nbsp;\u003cstrong\u003eg\u003c/strong\u003e,\u0026nbsp;Functional\u0026nbsp;profiles\u0026nbsp;of spike-specific CD4+\u0026nbsp;T cell responses from\u0026nbsp;mRNA-vaccinated individuals with representative gating and pie charts\u0026nbsp;showing\u0026nbsp;the\u0026nbsp;mean frequency\u0026nbsp;for each combination. Polyfunctional responses were compared\u0026nbsp;using\u0026nbsp;a\u0026nbsp;permutation test. Data in dot\u0026nbsp;plots are\u0026nbsp;shown\u0026nbsp;as median ±\u0026nbsp;IQR.\u0026nbsp;\u003c/p\u003e","description":"","filename":"Fig1.png","url":"https://assets-eu.researchsquare.com/files/rs-1217466/v1/fe457db49944d19141bf6549.png"},{"id":16945931,"identity":"45e9aed1-f16d-4dcf-aacb-f1f387c749fa","added_by":"auto","created_at":"2022-01-03 20:05:24","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":1054335,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eCross-reactive CD8+\u0026nbsp;T cell responses\u0026nbsp;against\u0026nbsp;B.1.1.529.\u0026nbsp;\u003c/strong\u003e\u0026nbsp;\u003c/p\u003e\u003cp\u003e\u003cstrong\u003ea\u003c/strong\u003e, Representative\u0026nbsp;flow cytometry\u0026nbsp;plots showing spike-specific CD8+\u0026nbsp;T cell responses (CD69+CD137+)\u0026nbsp;to peptide pools representing\u0026nbsp;wildtype\u0026nbsp;SARS-CoV-2 (WT) or\u0026nbsp;B.1.1.529.\u0026nbsp;\u003cstrong\u003eb\u003c/strong\u003e, Frequencies of spike-specific CD8+\u0026nbsp;T cells from mRNA-vaccinated, convalescent, and seronegative individuals. Each dot represents one donor.\u0026nbsp;Numbers indicate\u0026nbsp;median reduction in the frequency of detected responses.\u0026nbsp;\u003cstrong\u003ec\u003c/strong\u003e, Stimulation indices calculated as fold change\u0026nbsp;in frequency relative to the\u0026nbsp;negative control.\u0026nbsp;Numbers indicate\u0026nbsp;the percentage of individuals with\u0026nbsp;a\u0026nbsp;detectable response.\u0026nbsp;\u003cstrong\u003ed\u003c/strong\u003e, Cross-reactive responses\u0026nbsp;depicted on an individual basis as percent B.1.1.529/WT.\u0026nbsp;\u003cstrong\u003ee\u003c/strong\u003e,\u0026nbsp;Canonical memory differentiation\u0026nbsp;profiles\u0026nbsp;of spike-specific CD8+\u0026nbsp;T cells\u0026nbsp;with representative gating and\u0026nbsp;dot\u0026nbsp;plots showing the distribution\u0026nbsp;of subsets\u0026nbsp;across individuals with\u0026nbsp;detectable responses.\u0026nbsp;\u003cstrong\u003ef\u003c/strong\u003e,\u0026nbsp;Functional profiles\u0026nbsp;of spike-specific CD8+\u0026nbsp;T cells\u0026nbsp;from\u0026nbsp;mRNA-vaccinated individuals with representative gating and pie charts\u0026nbsp;showing\u0026nbsp;the mean frequency\u0026nbsp;for each combination. Polyfunctional responses were compared\u0026nbsp;using a\u0026nbsp;permutation test.\u0026nbsp;GrzB, granzyme B.\u0026nbsp;Data in bar charts are shown as mean\u0026nbsp;±\u0026nbsp;95% confidence intervals, and data\u0026nbsp;in dot\u0026nbsp;plots are\u0026nbsp;shown\u0026nbsp;as median ±\u0026nbsp;IQR.\u003c/p\u003e","description":"","filename":"Fig2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-1217466/v1/431623e9c8b5cb17645f58e8.jpg"},{"id":16978773,"identity":"e001e937-218c-4641-8e66-338301450e71","added_by":"auto","created_at":"2022-01-04 14:36:22","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":775345,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1217466/v1/f86c2011-b7a2-4d4c-ac2d-553aaa5bea26.pdf"},{"id":16945930,"identity":"a0109b62-da34-479a-a8c2-fe56815a4367","added_by":"auto","created_at":"2022-01-03 20:05:24","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":55076,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Table 1.\u003c/strong\u003e Flow cytometry reagents.\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"rssupplementarytable1.docx","url":"https://assets-eu.researchsquare.com/files/rs-1217466/v1/ddd8df6b9f53bd9bd6234f35.docx"},{"id":16946268,"identity":"ff47245c-278a-4ba0-bfe3-c208cf4c24be","added_by":"auto","created_at":"2022-01-03 20:11:24","extension":"png","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":1127687,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eExtended Data\u0026nbsp;Fig. 1.\u0026nbsp;Peptide validation experiments.\u003c/strong\u003e\u0026nbsp;\u003c/p\u003e\u003cp\u003e\u003cstrong\u003ea\u003c/strong\u003e, Representative\u0026nbsp;flow cytometry\u0026nbsp;plots showing\u0026nbsp;the gating strategy used to assess\u0026nbsp;spike-specific CD4+\u0026nbsp;and CD8+\u0026nbsp;T cell responses\u0026nbsp;to peptide pools representing\u0026nbsp;wildtype\u0026nbsp;SARS-CoV-2 (WT) or\u0026nbsp;B.1.1.529.\u0026nbsp;\u003cstrong\u003eb\u003c/strong\u003e,\u003cstrong\u003ec\u003c/strong\u003e,\u0026nbsp;Frequencies of spike-specific CD4+\u0026nbsp;(\u003cstrong\u003eb\u003c/strong\u003e) and CD8+\u0026nbsp;T cells (\u003cstrong\u003ec\u003c/strong\u003e) from mRNA-vaccinated individuals, comparing test\u0026nbsp;15mer peptide pools\u0026nbsp;\u003cem\u003eversus\u003c/em\u003e\u0026nbsp;validation 20mer\u0026nbsp;peptide pools representing\u0026nbsp;wildtype\u0026nbsp;SARS-CoV-2 (WT) or\u0026nbsp;B.1.1.529.\u0026nbsp;Each dot represents one donor.\u0026nbsp;Data in dot\u0026nbsp;plots are\u0026nbsp;shown\u0026nbsp;as median ±\u0026nbsp;IQR.\u0026nbsp;\u003cstrong\u003ed\u003c/strong\u003e,\u0026nbsp;Pairwise\u0026nbsp;analysis of total\u0026nbsp;spike-specific,\u0026nbsp;CD4+\u0026nbsp;(red lines)\u0026nbsp;and CD8+\u0026nbsp;(blue lines),\u0026nbsp;T cell\u0026nbsp;response\u0026nbsp;in vaccinated and convalescent donors against\u0026nbsp;wildtype\u0026nbsp;SARS-CoV-2 (WT) or\u0026nbsp;B.1.1.529. \u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"FigS1.png","url":"https://assets-eu.researchsquare.com/files/rs-1217466/v1/59f6c2a70bac929917cbe331.png"}],"financialInterests":"","formattedTitle":"\u003cp\u003eAncestral SARS-CoV-2-specific T cells cross-recognize Omicron (B.1.1.529)\u003c/p\u003e","fulltext":[{"header":"Main","content":"\u003cp\u003eNatural infection with SARS-CoV-2 and vaccination with mRNA constructs encoding the viral spike protein typically generate effective immunity against COVID-19. However, the current pandemic has been fueled by the continual emergence of variants-of-concern (VOCs), such as Omicron (B.1.1.529). Emerging data indicate that B.1.1.529 is more transmissible than previous VOCs\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e. This phenotype can be explained by key mutations in the receptor-binding domain (RBD), which confer enhanced affinity for the ACE2 receptor\u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e. Another major concern is that B.1.1.529 harbors a large number of additional mutations in the spike protein that could feasibly subvert immune recognition\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e. In line with this possibility, emerging reports have shown that neutralizing antibodies elicited against the ancestral Wuhan reference strain, either in the context of infection or vaccination, are less able to combat B.1.1.529\u003csup\u003e2,4\u003c/sup\u003e. These observations likely align with the propensity of B.1.1.529 to cause breakthrough infections, also in persons who have been vaccinated with two doses of mRNA vaccine or experienced previous SARS-CoV-2 infection\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e,\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003ePreliminary data suggest that breakthrough infections with B.1.1.529 might be associated with a lower risk of hospitalization and less often cause severe disease compared with the Delta VOC (B.1.617.2)\u003csup\u003e\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e,\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e. One possible inference from the less aggressive clinical spectrum is that additional immune components beyond the antibody response attenuate the course of infection with B.1.1.529. Previous studies have demonstrated that robust CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e T cell responses are induced following SARS-CoV-2 infection or vaccination\u003csup\u003e\u003cspan additionalcitationids=\"CR10 CR11 CR12 CR13 CR14\" citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e. Several lines of evidence further suggest that CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e T cell responses can modulate disease severity in humans and suppress viral replication in animal models\u003csup\u003e\u003cspan additionalcitationids=\"CR17 CR18\" citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u003c/sup\u003e. However, it has remained unclear to what extent ancestral SARS-CoV-2-specific CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e T cells cross-recognize B.1.1.529, especially given the unprecedented number of mutations in the spike protein, which likely shift the antigenic landscape more profoundly in relation to antecedent VOCs\u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eTo address this question, we collected peripheral blood mononuclear cells from mRNA-vaccinated individuals 6 months after the second dose of Pfizer/BioNTech BNT162b2 (\u003cem\u003en\u003c/em\u003e = 40), individuals in the convalescent phase 9 months after mild or severe COVID-19 (\u003cem\u003en\u003c/em\u003e = 48), and seronegative individuals (\u003cem\u003en\u003c/em\u003e = 48). Cells were stimulated in parallel with overlapping peptide pools spanning the entire spike protein sequences of the Wuhan reference strain (wildtype) or B.1.1.529. Activation-induced marker assays were used to quantify spike-specific CD4\u003csup\u003e+\u003c/sup\u003e T cell responses via the upregulation of CD69 and CD40L (CD154) and spike-specific CD8\u003csup\u003e+\u003c/sup\u003e T cell responses via the upregulation of CD69 and 4-1BB (CD137) (Extended Data Fig.\u0026nbsp;1a).\u003c/p\u003e \u003cp\u003eThe overall magnitude of the SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e T cell response against B.1.1.529 showed a median reduction of 9% in mRNA-vaccinated individuals and a median reduction of 16% in convalescent individuals relative to the wildtype response (Fig.\u0026nbsp;1a, b). The corresponding response frequencies were also slightly lower for B.1.1.529 (Fig.\u0026nbsp;1c). Individual comparisons further revealed maximum reductions in magnitude of 58% in mRNA-vaccinated individuals, 56% in convalescent individuals, and 75% in seronegative individuals, comparing SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e T cell responses against B.1.1.529 \u003cem\u003eversus\u003c/em\u003e wildtype (Fig.\u0026nbsp;1d). These findings were validated using independently synthesized peptide pools spanning the entire spike protein (Extended Data Fig.\u0026nbsp;1b).\u003c/p\u003e \u003cp\u003eTo extend these findings, we investigated the phenotypic characteristics of SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e T cells that cross-recognized B.1.1.529, with a particular focus on markers of T helper polarization (CCR4, CCR6, CXCR3, CXCR5) and memory differentiation (CCR7, CD45RA). No significant differences in T helper polarization were detected across intragroup comparisons of SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e T cell responses against B.1.1.529 \u003cem\u003eversus\u003c/em\u003e wildtype (Fig.\u0026nbsp;1e). Central memory T (T\u003csub\u003eCM\u003c/sub\u003e) cells predominated among SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e T cells in mRNA-vaccinated, convalescent, and seronegative individuals, but again, the subset composition of SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e T cell responses against B.1.1.529 \u003cem\u003eversus\u003c/em\u003e wildtype were similar across all groups (Fig.\u0026nbsp;1f). We also assessed the functionality of SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e T cells in mRNA-vaccinated individuals, measuring the intracellular expression of IFN-γ, TNF, and IL-2 alongside CD69 and CD154. No significant differences in the ability of SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e T cells to deploy multiple functions were apparent in response to stimulation with peptides representing B.1.1.529 \u003cem\u003eversus\u003c/em\u003e wildtype (Fig.\u0026nbsp;1g).\u003c/p\u003e \u003cp\u003eThe overall magnitude of the SARS-CoV-2 spike-specific CD8\u003csup\u003e+\u003c/sup\u003e T cell response against B.1.1.529 showed a median reduction of 8% in mRNA-vaccinated individuals and a median reduction of 30% in convalescent individuals relative to the wildtype response (Fig.\u0026nbsp;2a, b). These differences were mirrored in the corresponding response frequencies for B.1.1.529 \u003cem\u003eversus\u003c/em\u003e wildtype (Fig.\u0026nbsp;2c). Individual comparisons further revealed maximum reductions in magnitude of 55% in mRNA-vaccinated individuals, 63% in convalescent individuals, and 60% in seronegative individuals, comparing SARS-CoV-2 spike-specific CD8\u003csup\u003e+\u003c/sup\u003e T cell responses against B.1.1.529 \u003cem\u003eversus\u003c/em\u003e wildtype (Fig.\u0026nbsp;2d). These findings were again validated using independently synthesized peptide pools spanning the entire spike protein (Extended Data Fig.\u0026nbsp;1c).\u003c/p\u003e \u003cp\u003eIn further experiments, we investigated the phenotypic characteristics of SARS-CoV-2 spike-specific CD8\u003csup\u003e+\u003c/sup\u003e T cells that cross-recognized B.1.1.529, focusing on classic markers of memory differentiation (CCR7, CD45RA). Late effector memory T (T\u003csub\u003eEMRA\u003c/sub\u003e) cells predominated among SARS-CoV-2 spike-specific CD8\u003csup\u003e+\u003c/sup\u003e T cells in mRNA-vaccinated, convalescent, and seronegative individuals, but no significant differences in subset composition were detected across intragroup comparisons of SARS-CoV-2 spike-specific CD8\u003csup\u003e+\u003c/sup\u003e T cell responses against B.1.1.529 \u003cem\u003eversus\u003c/em\u003e wildtype (Fig.\u0026nbsp;2e). We also assessed the functionality of SARS-CoV-2 spike-specific CD8\u003csup\u003e+\u003c/sup\u003e T cells in mRNA-vaccinated individuals, measuring the intracellular expression of granzyme B, IFN-γ, TNF, and IL-2 alongside CD69 and CD137. Akin to the corresponding analyses of SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e T cells, no significant differences in the ability of SARS-CoV-2 spike-specific CD8\u003csup\u003e+\u003c/sup\u003e T cells to deploy multiple functions were apparent in response to stimulation with peptides representing B.1.1.529 \u003cem\u003eversus\u003c/em\u003e wildtype (Fig.\u0026nbsp;2f). In a final set of analyses, we merged all SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e T cell data points together and found a significantly lower total T cell response against B.1.1.529 \u003cem\u003eversus\u003c/em\u003e wildtype in convalescent, but not vaccinated subjects (Extended Data Fig.\u0026nbsp;1d). These analyses suggest that SARS-CoV-2 spike-specific T cell responses remain less intact against B.1.1.529 in convalescent individuals.\u003c/p\u003e \u003cp\u003eThe current global pandemic has been destabilized by the recent emergence of B.1.1.529, which continues to spread rapidly and supersede other VOCs. Our collective data indicate that SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e T cells elicited by mRNA vaccination or prior infection remain largely intact against B.1.1.529. Alongside intrinsic viral factors, such as altered tropism and decreased replication in the lower respiratory tract\u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e, such heterologous immune reactivity may explain why severe disease appears to be relatively uncommon after infection with this particular VOC. Moreover, the degree of cross-reactivity varied to some extent among individuals, most likely as a consequence of genetically encoded differences in antigen presentation, which could further modulate clinical outcomes associated with B.1.1.529. It should be noted that our evaluations were confined to peripheral blood samples, which do not necessarily reflect the entirety of the cellular immune response against SARS-CoV-2\u003csup\u003e22\u003c/sup\u003e. In addition, SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e T cells cross-recognized B.1.1.529 less comprehensively in convalescent \u003cem\u003eversus\u003c/em\u003e mRNA-vaccinated individuals, suggesting that booster immunization may provide benefits that extend beyond the induction of broadly neutralizing antibodies to enhance natural protection against recurrent episodes of COVID-19\u003csup\u003e2\u003c/sup\u003e.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch1\u003eACKNOWLEDGEMENTS\u003c/h1\u003e\n\u003cp\u003eThis study was supported by grants from the SciLifeLab National COVID-19 Research Program, financed by the Knut and Alice Wallenberg Foundation, the Swedish Research Council, Nordstjernan AB, Region Stockholm, and the Karolinska Institutet. M.B. was supported by the Swedish Research Council, the Knut and Alice Wallenberg Foundation, the Karolinska Institutet, the Swedish Society of Medicine, the Swedish Cancer Society, the Swedish Childhood Cancer Fund, the \u0026Aring;ke Wibergs Stiftelse, and the Jonas S\u0026ouml;derquist Stiftelse. J.N. was supported by an EMBO Postdoctoral Fellowship (ALTF 1062-2020). This project has been funded in whole or in part with Federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under Contract No. 75N93021C00016 (to A.S.), Contract No. 75N9301900065 (to A.S, D.W.) and by the National Institute for Health Research via grant COV-LT2-0041 awarded (to D.A.P.).\u003c/p\u003e\n\u003ch1\u003eAUTHOR CONTRIBUTIONS\u003c/h1\u003e\n\u003cp\u003eConceptualization: Y.G., C.C., J.K.S., D.A.P., H.G.L., A.C.K., A.S., S.A., M.B.\u003c/p\u003e\n\u003cp\u003eSample collection: Y.G., C.C., T.R.M., J.N., L.H., A.\u0026Ouml;., P.B., P.C., A.OLS., H.G.L., S.A., M.B.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eInvestigation: Y.G., C.C., A.G., A.OLO., M.H., M.B.\u003c/p\u003e\n\u003cp\u003eFormal analysis: Y.G., C.C.\u003c/p\u003e\n\u003cp\u003eVisualization: Y.G., C.C., M.B.\u003c/p\u003e\n\u003cp\u003eResources: A.G., J.K.S., D.W., H.G.L, A.C.K., A.S., S.A., M.B.\u003c/p\u003e\n\u003cp\u003eFunding acquisition: D.A.P., H.G.L., A.S., S.A., M.B.\u003c/p\u003e\n\u003cp\u003eSupervision: J.K.S., H.G.L., S.A., M.B.\u003c/p\u003e\n\u003cp\u003eWriting\u0026mdash;original draft: Y.G., C.C., D.A.P., M.B.\u003c/p\u003e\n\u003cp\u003eWriting\u0026mdash;review and editing: Y.G., C.C., A.G., T.R.M., J.N., A.\u0026Ouml;., P.B., J.K.S., D.A.P., H.G.L., A.C.K., S.A., M.B.\u003c/p\u003e\n\u003ch1\u003eDECLARATION OF INTERESTS\u003c/h1\u003e\n\u003cp\u003eA.S. is a consultant for Gritstone Bio, Flow Pharma, Arcturus Therapeutics, ImmunoScape, CellCarta, Avalia, Moderna, Fortress and Repertoire., and the La Jolla Institute\u0026nbsp;has filed patents to protect various aspects of the T cell epitope and vaccine design work.\u0026nbsp;SA has received honoraria for lectures from Gilead, AbbVie, MSD, Biogen, and has received research grants from Gilead and AbbVie.\u0026nbsp;M.B. is a consultant for Oxford Immunotec.\u0026nbsp;The other authors declare no conflicts of interest.\u003cbr\u003e\u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003cp\u003e1.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Dyer, O. 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Effectiveness of BNT162b2 Vaccine against Omicron Variant in South Africa. \u003cem\u003eN Engl J Med\u003c/em\u003e (2021).\u003c/p\u003e\n\u003cp\u003e6.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Goga, A.\u003cem\u003e, et al.\u003c/em\u003e Breakthrough Covid-19 infections during periods of circulating Beta, Delta and Omicron variants of concern, among health care workers in the Sisonke Ad26.COV2.S vaccine trial, South Africa. \u003cem\u003eMedRxiv\u003c/em\u003e (2021).\u003c/p\u003e\n\u003cp\u003e7.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Neil Ferguson, A.G., Wes Hinsley and Erik Volz on behalf of the Imperial College COVID-19 response team. Report 50: Hospitalisation risk for Omicron cases in England. \u0026nbsp;(2021).\u003c/p\u003e\n\u003cp\u003e8.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Wolter N.\u003cem\u003e, et al.\u003c/em\u003e Early assessment of the clinical severity of the SARS-CoV-2 Omicron variant in South Africa. \u003cem\u003ebioRxiv\u003c/em\u003e (2021).\u003c/p\u003e\n\u003cp\u003e9.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Braun, J.\u003cem\u003e, et al.\u003c/em\u003e SARS-CoV-2-reactive T cells in healthy donors and patients with COVID-19. \u003cem\u003eNature\u003c/em\u003e \u003cstrong\u003e587\u003c/strong\u003e, 270-274 (2020).\u003c/p\u003e\n\u003cp\u003e10.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Grifoni, A.\u003cem\u003e, et al.\u003c/em\u003e Targets of T Cell Responses to SARS-CoV-2 Coronavirus in Humans with COVID-19 Disease and Unexposed Individuals. \u003cem\u003eCell\u003c/em\u003e \u003cstrong\u003e181\u003c/strong\u003e, 1489-1501 e1415 (2020).\u003c/p\u003e\n\u003cp\u003e11.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Kalimuddin, S.\u003cem\u003e, et al.\u003c/em\u003e Early T cell and binding antibody responses are associated with COVID-19 RNA vaccine efficacy onset. \u003cem\u003eMed (N Y)\u003c/em\u003e \u003cstrong\u003e2\u003c/strong\u003e, 682-688 e684 (2021).\u003c/p\u003e\n\u003cp\u003e12.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Le Bert, N.\u003cem\u003e, et al.\u003c/em\u003e SARS-CoV-2-specific T cell immunity in cases of COVID-19 and SARS, and uninfected controls. \u003cem\u003eNature\u003c/em\u003e \u003cstrong\u003e584\u003c/strong\u003e, 457-462 (2020).\u003c/p\u003e\n\u003cp\u003e13.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Oberhardt, V.\u003cem\u003e, et al.\u003c/em\u003e Rapid and stable mobilization of CD8(+) T cells by SARS-CoV-2 mRNA vaccine. \u003cem\u003eNature\u003c/em\u003e \u003cstrong\u003e597\u003c/strong\u003e, 268-273 (2021).\u003c/p\u003e\n\u003cp\u003e14.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Peng, Y.\u003cem\u003e, et al.\u003c/em\u003e Broad and strong memory CD4(+) and CD8(+) T cells induced by SARS-CoV-2 in UK convalescent individuals following COVID-19. \u003cem\u003eNat Immunol\u003c/em\u003e \u003cstrong\u003e21\u003c/strong\u003e, 1336-1345 (2020).\u003c/p\u003e\n\u003cp\u003e15.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Sekine, T.\u003cem\u003e, et al.\u003c/em\u003e Robust T Cell Immunity in Convalescent Individuals with Asymptomatic or Mild COVID-19. \u003cem\u003eCell\u003c/em\u003e \u003cstrong\u003e183\u003c/strong\u003e, 158-168 e114 (2020).\u003c/p\u003e\n\u003cp\u003e16.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Israelow, B.\u003cem\u003e, et al.\u003c/em\u003e Adaptive immune determinants of viral clearance and protection in mouse models of SARS-CoV-2. \u003cem\u003eSci Immunol\u003c/em\u003e \u003cstrong\u003e6\u003c/strong\u003e, eabl4509 (2021).\u003c/p\u003e\n\u003cp\u003e17.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;McMahan, K.\u003cem\u003e, et al.\u003c/em\u003e Correlates of protection against SARS-CoV-2 in rhesus macaques. \u003cem\u003eNature\u003c/em\u003e \u003cstrong\u003e590\u003c/strong\u003e, 630-634 (2021).\u003c/p\u003e\n\u003cp\u003e18.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Rydyznski Moderbacher, C.\u003cem\u003e, et al.\u003c/em\u003e Antigen-Specific Adaptive Immunity to SARS-CoV-2 in Acute COVID-19 and Associations with Age and Disease Severity. \u003cem\u003eCell\u003c/em\u003e \u003cstrong\u003e183\u003c/strong\u003e, 996-1012 e1019 (2020).\u003c/p\u003e\n\u003cp\u003e19.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Tan, A.T.\u003cem\u003e, et al.\u003c/em\u003e Early induction of functional SARS-CoV-2-specific T cells associates with rapid viral clearance and mild disease in COVID-19 patients. \u003cem\u003eCell Rep\u003c/em\u003e \u003cstrong\u003e34\u003c/strong\u003e, 108728 (2021).\u003c/p\u003e\n\u003cp\u003e20.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Tarke, A.\u003cem\u003e, et al.\u003c/em\u003e Impact of SARS-CoV-2 variants on the total CD4(+) and CD8(+) T cell reactivity in infected or vaccinated individuals. \u003cem\u003eCell Rep Med\u003c/em\u003e \u003cstrong\u003e2\u003c/strong\u003e, 100355 (2021).\u003c/p\u003e\n\u003cp\u003e21.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Meng B.\u003cem\u003e, et al.\u003c/em\u003e, SARS-CoV-2 Omicron spike mediated immune escape, infectivity and cell-cell fusion. \u003cem\u003ebioRxiv\u003c/em\u003e (2021).\u003c/p\u003e\n\u003cp\u003e22.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Niessl, J.\u003cem\u003e, et al.\u003c/em\u003e Identification of resident memory CD8(+) T cells with functional specificity for SARS-CoV-2 in unexposed oropharyngeal lymphoid tissue. \u003cem\u003eSci Immunol\u003c/em\u003e \u003cstrong\u003e6\u003c/strong\u003e, eabk0894 (2021).\u003c/p\u003e\n\u003cp\u003e23.\u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Bergman, P.\u003cem\u003e, et al.\u003c/em\u003e Safety and efficacy of the mRNA BNT162b2 vaccine against SARS-CoV-2 in five groups of immunocompromised patients and healthy controls in a prospective open-label clinical trial. \u003cem\u003eEBioMedicine\u003c/em\u003e \u003cstrong\u003e74\u003c/strong\u003e, 103705 (2021).\u003c/p\u003e"},{"header":"Online Methods","content":"\u003cp\u003e\u003cstrong\u003e\u003cem\u003eSamples\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eHealthy individuals (n=40) were sampled 6 months after the second dose of the BNT162b2 vaccine\u0026nbsp;(Comirnaty\u003csup\u003e\u0026reg;\u003c/sup\u003e, Pfizer/BioNTech)\u0026nbsp;as part of a clinical trial registered at EudraCT (2021-000175-37) and https://clinicaltrials.gov (2021-000175-37) at Karolinska University Hospital, Sweden\u003csup\u003e23\u003c/sup\u003e. The two doses of the vaccine were given 21 days apart in standard doses. The vaccinated cohort of healthy individuals consisted of 23 women (58%)/17 men (43%), and the median age (range) was 53 (22-79) years. The study was approved by the Swedish Medical Product Agency. Additional samples from 15 of the vaccinated cohort participants were collected 3 months after the second dose for validation purposes (Extended Data Figure 1). Convalescent individuals after previous COVID-19 were sampled 9 months after RT-PCR verified SARS-CoV-2 infection leading to mild (i.e. non-hospitalized, n=26) or severe (i.e. hospitalized, n=22) disease during the first wave of the pandemic in March-April of 2020, before the emergence of the Alpha, Beta and Delta variants. The cohort of convalescent individuals after mild COVID-19 consisted of 8 women (31%)/18 men (69%), and the median age (range) was 54 (44-68) years. The cohort of convalescent individuals after severe COVID-19 consisted of 3 women (14%)/19 men (86%), and the median age (range) was 58 (33-66) years. Fourteen of the patients with severe COVID-19 (64%) had received intensive care at the time of COVID-19. None of the convalescent individuals had received any COVID-19 vaccination at the time of sample collections. Seronegative samples were acquired from healthy blood donors in late 2020. The absence of spike-specific antibodies was confirmed using an Anti-SARS-CoV-2 S Immunoassay (Roche) for vaccinated healthy individuals and seronegative blood donors. All participants provided written informed consent in accordance with the principles of the Declaration of Helsinki. The study was approved by the Regional Ethics Committee in Stockholm, Sweden. Peripheral blood mononuclear cells (PBMCs) were isolated via standard density gradient centrifugation and cryopreserved in fetal bovine serum (FBS) containing 10% dimethyl sulfoxide (DMSO).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003ePeptides\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eOverlapping peptides were designed to span the entire spike protein sequence of SARS-CoV-2 corresponding to the ancestral Wuhan strain (wildtype) or B.1.1.529. Test peptides comprising 15mers overlapping by 10 amino acids were synthesized as crude material for functional screens (TC Peptide Lab). Validation peptides comprising 20mers overlapping by 10 amino acids were synthesized to an equivalent specification (Sigma-Aldrich). All peptides were reconstituted in DMSO, diluted to stock concentrations of 100 \u0026mu;g/ml in phosphate-buffered saline (PBS), and stored at \u0026minus;20 \u0026ordm;C.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eActivation-induced marker assays\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePBMCs were thawed quickly, resuspended in\u0026nbsp;RPMI 1640 containing 10% FBS, 1% L-glutamine, and 1% penicillin/streptomycin\u0026nbsp;(complete medium) in the presence of DNase I (10 U/ml; Sigma-Aldrich), and rested at 1 \u0026times; 10\u003csup\u003e6\u003c/sup\u003e cells per well in 96-well U-bottom plates (Corning) for 4 h at 37 \u0026ordm;C. The medium was then supplemented with anti-CXCR5\u0026ndash;BB515 and anti-CD40 (unconjugated), followed 15 min later by the relevant peptide pools (1 \u0026mu;g/ml/peptide). Negative control wells contained equivalent DMSO. After 12 h, cells were washed in PBS supplemented with 2% FBS and 2 mM EDTA (FACS buffer) and stained with anti-CCR4/CD194\u0026ndash;BB700, anti-CCR6/CD196\u0026ndash;BUV737, anti-CCR7\u0026ndash;APC-Cy7, and anti-CXCR3\u0026ndash;AF647 for 10 min at 37 \u0026ordm;C. Additional surface stains were performed for 30 min at room temperature in the presence of Brilliant Stain Buffer Plus (BD Biosciences). Viable cells were identified by exclusion using a LIVE/DEAD Fixable Aqua Dead Cell Stain Kit (Thermo Fisher Scientific). Stained cells were then washed in FACS buffer, fixed in PBS containing 1% paraformaldehyde (PFA; Biotium), and acquired using a FACSymphony A5 (BD Biosciences). The gating strategy is shown in Extended Data Figure 1. All flow cytometry reagents are detailed in Supplementary Table 1.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eIntracellular cytokine staining\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePBMCs were thawed quickly, resuspended in complete medium in the presence of DNase I (10 U/ml; Sigma-Aldrich), and rested at 1 \u0026times; 10\u003csup\u003e6\u003c/sup\u003e cells/well in 96-well U-bottom plates (Corning) for 4 h at 37 \u0026ordm;C. The medium was then supplemented with anti-CXCR5\u0026ndash;BB515, followed 15 min later by the relevant peptide pools (1 \u0026mu;g/ml/peptide), and a further 1 h later by brefeldin A (1 \u0026mu;g/ml; Sigma-Aldrich) and monensin (0.7 \u0026mu;g/ml; BD Biosciences). Negative control wells contained equivalent DMSO. After 9 h, cells were washed in FACS buffer and stained with anti-CCR4/CD194\u0026ndash;BB700, anti-CCR6/CD196\u0026ndash;BUV737, anti-CCR7\u0026ndash;APC-Cy7, and anti-CXCR3\u0026ndash;BV750 for 10 min at 37 \u0026ordm;C. Additional surface stains were performed for 30 min at room temperature in the presence of Brilliant Stain Buffer Plus (BD Biosciences). Viable cells were identified by exclusion using a LIVE/DEAD Fixable Aqua Dead Cell Stain Kit (Thermo Fisher Scientific). Cells were then washed in FACS buffer and fixed/permeabilized using a FoxP3 Transcription Factor Staining Buffer Set (Thermo Fisher Scientific). Intracellular stains were performed for 30 min at room temperature. Stained cells were washed in FACS buffer, fixed in PBS containing 1% PFA (Biotium), and acquired using a FACSymphony A5 (BD Biosciences). All flow cytometry reagents are detailed in Supplementary Table 1.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eData analysis and statistics\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFlow cytometry data were analyzed using FlowJo version 10.8.0 (FlowJo LLC). Stimulation indices were calculated as fold change in frequency relative to the negative control (equivalent DMSO). Positive responses were identified using a threshold stimulation index \u0026gt;2. Statistical analyses were performed using Prism version 9 (GraphPad). Significance between two paired groups was assessed using the Wilcoxon signed rank test. Functional profiles were deconvoluted using Boolean gating in FlowJo version 10.8.0 (FlowJo LLC) followed by downstream analyses in SPICE version 6.1 (https://niaid.github.io/spice/).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eData availability\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eRequests for data should be addressed to the corresponding author and will be made available within the bounds of confidentiality and data protection obligations.\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"Karolinska Institute","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"B.1.1.529, Omicron, COVID-19, SARS-CoV-2, T cells","lastPublishedDoi":"10.21203/rs.3.rs-1217466/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-1217466/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe emergence of the SARS-CoV-2 variant-of-concern Omicron (B.1.1.529) has destabilized global efforts to control the impact of COVID-19. Recent data have suggested that B.1.1.529 can readily infect people with naturally acquired or vaccine-induced immunity, facilitated in some cases by viral escape from antibodies that neutralize ancestral SARS-CoV-2. However, severe disease appears to be relatively uncommon in such individuals, highlighting a potential role for other components of the adaptive immune system. We report here that SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e T cells induced by prior infection and, more extensively, by mRNA vaccination provide comprehensive heterologous immune reactivity against B.1.1.529. Pairwise comparisons across groups further revealed that SARS-CoV-2 spike-reactive CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e T cells exhibited similar functional attributes, memory distributions, and phenotypic traits in response to the ancestral strain or B.1.1.529. Our data indicate that established SARS-CoV-2 spike-specific CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e T cell responses, especially after mRNA vaccination, remain largely intact against B.1.1.529.\u003c/p\u003e","manuscriptTitle":"Ancestral SARS-CoV-2-specific T cells cross-recognize Omicron (B.1.1.529)","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2022-01-03 20:05:22","doi":"10.21203/rs.3.rs-1217466/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"950596c4-4d88-4de0-92fd-1a9e1cebd229","owner":[],"postedDate":"January 3rd, 2022","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":9445584,"name":"Immunology"},{"id":9445585,"name":"Infectious Diseases"}],"tags":[],"updatedAt":"2022-01-03T20:05:22+00:00","versionOfRecord":[],"versionCreatedAt":"2022-01-03 20:05:22","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-1217466","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-1217466","identity":"rs-1217466","version":["v1"]},"buildId":"FbvkV6FR0MCFSLy54lSbu","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}