Immunity

The COVID-19 pandemic is an ongoing global health threat, yet our understanding of the cellular disease dynamics remains limited. In our unique COVID-19 human challenge study we used single cell genomics of nasopharyngeal swabs and blood to temporally resolve abortive, transient and sustained infections in 16 seronegative individuals challenged with preAlpha-SARS-CoV-2. Our analyses revealed rapid changes in cell type proportions and dozens of highly dynamic cellular response states in epithelial and immune cells associated with specific timepoints or infection status. We observed that the interferon response in blood precedes the nasopharynx, and that nasopharyngeal immune infiltration occurred early in transient but later in sustained infection, and thus correlated with preventing sustained infection. Ciliated cells showed an acute response phase, upregulated MHC class II while infected, and were most permissive for viral replication, whilst nasal T cells and macrophages were infected non-productively. We resolve 54 T cell states, including acutely activated T cells that clonally expanded while carrying convergent SARS-CoV-2 motifs. Our novel computational pipeline (Cell2TCR) identifies activated antigen-responding clonotype groups and motifs in any dataset. Together, we show that our detailed time series data (covid19cellatlas.org) can serve as a "Rosetta stone" for the epithelial and immune cell responses, and reveals early dynamic responses associated with protection from infection.

The durability of infection-induced SARS-CoV-2 immunity has major implications for public health mitigation and vaccine development. Animal studies1,2 and the scarcity of confirmed re-infection3 suggests immune protection is likely, although the durability of this protection is debated. Lasting immunity following acute viral infection requires maintenance of both serum antibody and antigen-specific memory B and T lymphocytes and is notoriously pathogen specific, ranging from life-long for smallpox or measles4, to highly transient for common cold coronaviruses (CCC)5. Neutralising antibody responses are a likely correlate of protective immunity and exclusively recognise the viral spike (S) protein, predominantly targeting the receptor binding domain (RBD) within the S1 sub-domain6. Multiple reports describe waning of S-specific antibodies in the first 2-3 months following infection7-12. However, extrapolation of early linear trends in decay might be overly pessimistic, with several groups reporting that serum neutralisation is stable over time in a proportion of convalescent subjects8,12-17. While SARS-CoV-2 specific B and T cell responses are readily induced by infection6,13,18-24, the longitudinal dynamics of these key memory populations remains poorly resolved. Here we comprehensively profiled antibody, B and T cell dynamics over time in a cohort recovered from mild-moderate COVID-19. We find that binding and neutralising antibody responses, together with individual serum clonotypes, decay over the first 4 months post-infection, as expected, with a similar decline in S-specific CD4+ and circulating T follicular helper (cTFH) frequencies. In contrast, S-specific IgG+ memory B cells (MBC) consistently accumulate over time, eventually comprising a significant fraction of circulating MBC. Modelling of the concomitant immune kinetics predicts maintenance of serological neutralising activity above a titre of 1:40 in 50% of convalescent subjects to 74 days, with probable additive protection from B and T cells. Overall, our study suggests SARS-CoV-2 immunity after infection is likely to be transiently protective at a population level. SARS-CoV-2 vaccines may require greater immunogenicity and durability than natural infection to drive long-term protection.

T cells play key protective but also pathogenic roles in COVID-19. We studied expression of long non-coding RNAs (lncRNAs) in COVID-19 T cell transcriptomes by integrating previously published single-cell RNA sequencing datasets. The long intergenic non-coding RNA MALAT1 was the most highly transcribed lncRNA in T cells, with Th1 cells demonstrating the lowest and CD8+ resident memory cells the highest MALAT1 expression, amongst CD4+ and CD8+ T cells populations, respectively. We then identified gene signatures that covaried with MALAT1 in single T cells. A significantly higher number of transcripts correlated negatively with MALAT1 than those that correlated. Enriched functional annotations of the MALAT1-anti-correlating gene signature included processes associated with T cell activation such as cell division, oxidative phosphorylation and response to cytokine. The MALAT1 anti-correlating gene signature shared by both CD4+ and CD8+ T cells marked dividing T cells in both lung and blood of COVID-19 patients. Focussing on the tissue, we used an independent patient cohort of post-mortem COVID-19 lung samples and demonstrated that MALAT1 suppression was indeed a marker of MKI67+ proliferating CD8+ T cells. Our results reveal MALAT1 suppression and its associated gene signature are a hallmark of human proliferating T cells.

The pandemic spread of the novel coronavirus SARS-CoV-2 is due, in part, to the immunological properties of the host-viral interaction. The clinical presentation varies greatly from individual to individual, with asymptomatic carriers, mild to moderate-presenting patients and severely affected patients. Variation in immune response to SARS-CoV-2 may underlie this clinical variation. Using a high dimensional systems immunology platform, we have analyzed the peripheral blood compartment of 6 healthy individuals, 23 mild-to-moderate COVID-19 patients and 20 severe COVID-19 patients. We identify distinct immunological signatures in the peripheral blood of the mild-to-moderate and severe COVID-19 patients, including T cell lymphopenia, more consistent with peripheral hypo-than hyper-immune activation. Unique to the severe COVID-19 cases was a large increase in the proportion of IL-10-secreting regulatory T cells, a lineage known to possess anti-inflammatory properties in the lung. Annotated data is openly available (https://flowrepository.ors/experiments/2713) with clinical correlates, as a systems immunology resource for the COVID-19 research community.

Pronounced immune escape by the SARS-CoV-2 Omicron variant has resulted in large numbers of individuals with hybrid immunity, generated through a combination of vaccination and infection. Based primarily on circulating neutralizing antibody (NAb) data, concerns have been raised that omicron breakthrough infections in triple-vaccinated individuals result in poor induction of omicron-specific immunity, and that a history of prior SARS-CoV-2 in particular is associated with profound immune dampening. Taking a broader and comprehensive approach, we characterized mucosal and blood immunity to both spike and non-spike antigens following BA.1/BA.2 infections in triple mRNA-vaccinated individuals, with and without a history of previous SARS-CoV-2 infection. We find that the majority of individuals increase BA.1/BA.2/BA.5-specific NAb following infection, but confirm that the magnitude of increase and post-omicron titres are indeed higher in those who were infection-naive. In contrast, significant increases in nasal antibody responses are seen regardless of prior infection history, including neutralizing activity against BA.5 spike. Spike-specific T cells increase only in infection-naive vaccinees; however, post-omicron T cell responses are still significantly higher in previously-infected individuals, who appear to have maximally induced responses with a CD8+ phenotype of high cytotoxic potential after their 3rd mRNA vaccine dose. Antibody and T cell responses to non-spike antigens also increase significantly regardless of prior infection status, with a boost seen in previously-infected individuals to immunity primed by their first infection. These findings suggest that hybrid immunity induced by omicron breakthrough infections is highly dynamic, complex, and compartmentalised, with significant immune enhancement that can help protect against COVID-19 caused by future omicron variants.

In previously unvaccinated and uninfected individuals, non-RBD SARS-CoV-2 spike-specific B cells were prominent in two distinct, durable, resting, cross-reactive, "pre-existing" switched memory B cell compartments. While pre-existing RBD-specific B cells were extremely rare in uninfected and unvaccinated individuals, these two pre-existing switched memory B cell compartments were molded by vaccination and infection to become the primary source of RBD-specific B cells that are triggered by vaccine boosting. The frequency of wild-type RBD-binding memory B cells that cross-react with the Omicron variant RBD did not alter with boosting. In contrast, after a boost, B cells recognizing the full-length Omicron variant spike protein expanded, with pre-existing resting memory B cells differentiating almost quantitatively into effector B cell populations. B cells derived from "ancient" pre-existing memory cells and that recognize the full-length wild-type spike with the highest avidity after boosting are the B cells that also bind the Omicron variant spike protein. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/21268554v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@1de97acorg.highwire.dtl.DTLVardef@b7ab7forg.highwire.dtl.DTLVardef@5c38dcorg.highwire.dtl.DTLVardef@99106c_HPS_FORMAT_FIGEXP M_FIG C_FIG

Dengue virus (DENV) poses a major global health burden, with limited vaccine availability and concerns that immunization of dengue-naive individuals may exacerbate disease severity due to antibody-dependent enhancement. This challenge highlights the need for deeper insight into primary immune responses. Using a controlled human challenge model, we conducted a longitudinal analysis of B-cell receptor repertoire dynamics during primary DENV1 infection. Our study reveals that the acute-phase response is dominated by memory-derived B-cell clones, indicating pre-existing cross-reactive immunity. Concurrently, we identified highly public, convergent B-cell clones arising from naive B cells, characterized by shared sequence features across individuals. These clones exhibit atypical maturation kinetics: while they switch to the IgG isotype during the convalescent phase, they retain germline-like sequences with limited somatic hypermutation. This suggests that affinity maturation is delayed compared to canonical responses. Our findings provide mechanistic context for previous reports of natural antibody responses in dengue and refine current models of how neutralizing and potentially enhancing antibodies emerge following primary infection. By resolving the timing and origin of early B-cell responses, this work contributes to a more precise framework for understanding flavivirus immunity. These insights may guide vaccine design strategies that aim to elicit protective immunity without enhancing the risk of severe disease. One Sentence SummaryPrimary DENV1 infection elicits convergent clones that undergo atypical maturation.

Even within a single protein, antibody binding can have beneficial, neutral, or harmful effects during the response to infection. Resolving a polyclonal antibody repertoire across a pathogens proteome to specific epitopes may therefore explain much of the heterogeneity in susceptibility to infectious disease. However, the three-dimensional nature of antibody-epitope interactions makes the discovery of non-obvious targets challenging. We implemented a novel computational method and synthetic biology pipeline for identifying epitopes that are functionally important in the SARS-CoV-2 proteome and identified an IgM-dominant response to an exposed Membrane protein epitope which to our knowledge is the strongest correlate of severe disease identified to date (adjusted OR 72.14, 95% CI: 9.71 - 1300.15), stronger even than the exponential association of severe disease with age. We also identify persistence (> 2 years) of this IgM response in individuals with longCOVID, and a correlation with fatigue and depression symptom burden. The repetitive arrangement of this epitope and the pattern of isotype class switching is consistent with this being a previously unrecognized T independent antigen. These findings point to a coronavirus host-pathogen interaction characteristic of severe virus driven immune pathology. This epitope is a promising vaccine and therapeutic target as it is highly conserved through SARS-CoV-2 variant evolution in humans to date and in related coronaviruses (e.g. SARS-CoV), showing far less evolutionary plasticity than targets on the Spike protein. This provides a promising biomarker for longCOVID and a target to complement Spike-directed vaccination which could broaden humoral protection from severe or persistent disease or novel coronavirus spillovers. One-Sentence SummaryUsing a novel protein-structure-based B cell epitope discovery method with a wide range of possible applications, we have identified a simple to measure host-pathogen antibody signature associated with severe COVID-19 and longCOVID and suggest the viral Membrane protein contains an epitope that acts as a T independent antigen during infection triggering extrafollicular B cell activation.

Vaccine-induced immunity may impact subsequent de novo responses to drifted epitopes in SARS-CoV-2 variants, but this has been difficult to quantify due to the challenges in recruiting unvaccinated control groups whose first exposure to SARS-CoV-2 is a primary infection. Through local, statewide, and national SARS-CoV-2 testing programs, we were able to recruit cohorts of individuals who had recovered from either primary or post-vaccination infections by either the Delta or Omicron BA.1 variants. Regardless of variant, we observed greater Spike-specific and neutralizing antibody responses in post-vaccination infections than in those who were infected without prior vaccination. Through analysis of variant-specific memory B cells as markers of de novo responses, we observed that Delta and Omicron BA.1 infections led to a marked shift in immunodominance in which some drifted epitopes elicited minimal responses, even in primary infections. Prior immunity through vaccination had a small negative impact on these de novo responses, but this did not correlate with cross-reactive memory B cells, arguing against competitive inhibition of naive B cells. We conclude that dampened de novo B cell responses against drifted epitopes are mostly a function of altered immunodominance hierarchies that are apparent even in primary infections, with a more modest contribution from pre-existing immunity, perhaps due to accelerated antigen clearance.

Severe COVID-19 disproportionately impacts older individuals and those with comorbidities. It is estimated that approximately 80% of COVID-19 deaths are observed among individuals >65 years of age. However, the immunological underpinnings of severe COVID-19 in the aged have yet to be defined. This study captures the longitudinal immune response to SARS-CoV-2 infection in a cohort of young and aged patients with varying disease severity. Phenotypic transcriptional and functional examination of the peripheral mononuclear cells revealed age-, time, and disease severity-specific adaptations. Gene expression signatures within memory B cells suggest qualitative differences in the antibody responses in aged patients with severe disease. Examination of T cells showed profound lymphopenia, that worsened over time and correlated with lower levels of plasma cytokines important for T cell survival in aged patients with severe disease. Single cell RNA sequencing revealed augmented signatures of activation, exhaustion, cytotoxicity, and type-I interferon signaling in memory T cells and NK cells. Although hallmarks of a cytokine storm were evident in both groups, older individuals exhibited elevated levels of chemokines that mobilize inflammatory myeloid cells, notably in those who succumbed to disease. Correspondingly, we observed a re-distribution of DC and monocytes with severe disease that was accompanied by a rewiring towards a more regulatory phenotype. Several of these critical changes, such as the reduction of surface HLA-DR on myeloid cells, were reversed in young but not aged patients over time. In summary, the data presented here provide novel insights into the impact of aging on the host response to SARS-CoV2 infection.

Advances in multimodal single cell analysis can empower high-resolution dissection of human vaccination responses. The resulting data capture multiple layers of biological variations, including molecular and cellular states, vaccine formulations, inter- and intra-subject differences, and responses unfolding over time. Transforming such data into biological insight remains a major challenge. Here we present a systematic framework applied to multimodal single cell data obtained before and after influenza vaccination without adjuvants or pandemic H5N1 vaccination with the AS03 adjuvant. Our approach pinpoints responses shared across or unique to specific cell types and identifies adjuvant specific signatures, including pro-survival transcriptional states in B lymphocytes that emerged one day after vaccination. We also reveal that high antibody responders to the unadjuvanted vaccine have a distinct baseline involving a rewired network of cell type specific transcriptional states. Remarkably, the status of certain innate immune cells in this network in high responders of the unadjuvanted vaccine appear "naturally adjuvanted": they resemble phenotypes induced early in the same cells only by vaccination with AS03. Furthermore, these cell subsets have elevated frequency in the blood at baseline and increased cell-intrinsic phospho-signaling responses after LPS stimulation ex vivo in high compared to low responders. Our findings identify how variation in the status of multiple immune cell types at baseline may drive robust differences in innate and adaptive responses to vaccination and thus open new avenues for vaccine development and immune response engineering in humans.

Evaluation of host-response blood transcriptional signatures of viral infection have so far failed to test whether these biomarkers reflect different biological processes that may be leveraged for distinct translational applications. We addressed this question in the SARS-CoV-2 human challenge model. We found differential time profiles for interferon (IFN) stimulated blood transcriptional responses represented by measurement of single genes. MX1 transcripts correlated with a rapid and transient wave of type 1 IFN stimulated genes (ISG) across all cell types, which may precede PCR detection of replicative infection. Another ISG, IFI27, showed a delayed but sustained response restricted to myeloid peripheral blood mononuclear cells, attributable to gene and cell-specific epigenetic regulation. These findings were reproducible in diverse respiratory virus challenges, and in natural infection with SARS-CoV-2 or unselected respiratory viruses. The MX1 response achieved superior diagnostic accuracy in early infection, correlation with viral load and identification of virus culture positivity, with potential to stratify patients for time sensitive antiviral treatment. IFI27 achieved superior diagnostic accuracy across the time course of symptomatic infection. Compared to blood, measurement of these responses in nasal mucosal samples was less sensitive and did not discriminate between early and late phases of infection.

Coronavirus disease 2019 (COVID-19) displays high clinical variability but the parameters that determine disease severity are still unclear. Pre-existing T cell memory has been hypothesized as a protective mechanism but conclusive evidence is lacking. Here we demonstrate that all unexposed individuals harbor SARS-CoV-2-specific memory T cells with marginal cross-reactivity to common cold corona and other unrelated viruses. They display low functional avidity and broad protein target specificities and their frequencies correlate with the overall size of the CD4+ memory compartment reflecting the "immunological age" of an individual. COVID-19 patients have strongly increased SARS-CoV-2-specific inflammatory T cell responses that are correlated with severity. Strikingly however, patients with severe COVID-19 displayed lower TCR functional avidity and less clonal expansion. Our data suggest that a low avidity pre-existing T cell memory negatively impacts on the T cell response quality against neoantigens such as SARS-CoV-2, which may predispose to develop inappropriate immune reactions especially in the elderly. We propose the immunological age as an independent risk factor to develop severe COVID-19. Key points- Pre-existing SARS-CoV-2-reactive memory T cells are present in all humans, but have low functional avidity and broad target specificities - Pre-existing memory T cells show only marginal cross-reactivity to common cold corona viruses - Frequencies of pre-existing memory T cells increase with the size of the CD4+ memory compartment reflecting the "immunological age" of the individual - Low-avidity and polyclonal, but strongly enhanced SARS-CoV-2 specific T cell responses develop in severe COVID-19, suggesting their origin from pre-existing memory - The immunological age may represent a risk factor to develop severe COVID-19

Given the rapid spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and the recent implementation of SARS-CoV-2 vaccination, we have much to learn about the duration of immune protection and the interface between the immune responses to infection and to vaccination. To address these questions, we monitored immune responses to SARS-CoV-2 infection in convalescent individuals over seven months and following mRNA vaccination. Spike Receptor-Binding-Domain (RBD)-specific circulating antibodies and plasma neutralizing activity generally decreased over time, whereas RBD-specific memory B cells persisted. Additionally, using antibody depletion techniques, we showed that the neutralizing activity of plasma specifically resides in the anti-RBD antibodies. More vigorous antibody and B cell responses to vaccination were observed in previously infected subjects relative to uninfected comparators, presumably due to immune priming by infection. SARS-CoV-2 infection also led to increased numbers of double negative B memory cells, which are described as a dysfunctional B cell subset. This effect was reversed by SARS-CoV-2 vaccination, providing a potential mechanistic explanation for the vaccination-induced reduction in symptoms in patients with "Long-COVID".

Blood transcriptional profiling is a powerful tool to evaluate immune responses to infection; however, blood collection via traditional phlebotomy remains a barrier to precise characterization of the immune response in dynamic infections (e.g., respiratory viruses). Here we present an at-home self-collection methodology, homeRNA, to study the host transcriptional response during acute SARS-CoV-2 infections. This method uniquely enables high frequency measurement of the host immune kinetics in non-hospitalized adults during the acute and most dynamic stage of their infection. COVID-19+ and healthy participants self-collected blood every other day for two weeks with daily nasal swabs and symptom surveys to track viral load kinetics and symptom burden, respectively. While healthy uninfected participants showed remarkably stable immune kinetics with no significant dynamic genes, COVID-19+ participants, on the contrary, depicted a robust response with over 418 dynamic genes associated with interferon and innate viral defense pathways. When stratified by vaccination status, we detected distinct response signatures between unvaccinated and breakthrough (vaccinated) infection subgroups; unvaccinated individuals portrayed a response repertoire characterized by higher innate antiviral responses, interferon signaling, and cytotoxic lymphocyte responses while breakthrough infections portrayed lower levels of interferon signaling and enhanced early cell-mediated response. Leveraging cross-platform longitudinal sampling (nasal swabs and blood), we observed that IFI27, a key viral response gene, tracked closely with SARS-CoV-2 viral clearance in individual participants. Taken together, these results demonstrate that at-home sampling can capture key host antiviral responses and facilitate frequent longitudinal sampling to detect transient host immune kinetics during dynamic immune states. One Sentence SummarySelf-blood collection using homeRNA captures temporal dynamics in host transcriptional immune response during acute SARS-CoV-2 infection.

The incredible variety of proteins associated with immune responses presents a major challenge in immune monitoring. When combinations of these proteins are measured, cell types that influence disease can be precisely identified. Here, we introduce TerraFlow, a novel data analysis tool that performs an exhaustive search of disease-associated cell populations from high-parameter flow cytometry experiments. Using a newly generated dataset, from 24-color immune checkpoint-focused and 18-color immune function-focused experiments, we apply TerraFlow to classical Hodgkin lymphoma (cHL), where systemic T-cell immunity has not been investigated in detail. We reveal novel immune perturbations in newly diagnosed cHL, as well as persistent immune perturbations after treatment. Newly diagnosed patients have elevated levels of activated (CD278+), exhausted (e.g., CD366+ and CD152+ phenotypes), and IL17-expressing cells, along with diminished levels of naive and central memory (CD127+) T-cells and fewer IFN{gamma}+ and TNF+ T-cells. Exhaustion signatures are reduced with treatment, but compared to healthy individuals, treated patients still exhibit more activated (CD278+ phenotypes), exhausted (CD366+), and IL17-expressing cells. Notably, TerraFlow identifies more phenotypic differences between patient groups than FlowSOM and CellCNN, often with better predictive power. Finally, we introduce a new non-gating approach for data analysis that obviates the need for time-consuming and subjective setting of fluorescence thresholds. Our results benchmark TerraFlow against common methods, provide mechanistic support for past reports of immune deficiency in cHL, and provide a roadmap for future immunotherapy and biomarker studies.

SARS-CoV-2 vaccine-acquired immunity provides robust cross-variant recognition, while infection-acquired immunity can be heterogenous, with disease severity often modulating post-recovery responses. We assessed antibody waning dynamics between infection- and vaccination-acquired immunity across variants of concern (VOC). mRNA vaccination induced potent, cross-VOC Spike recognition and functional responses, but waned more rapidly for Omicron Spike. Hospitalized individuals developed more durable functional responses with lower peaks compared to mRNA vaccination, while outpatients exhibited slower decay than inactivated vaccine recipients. Humoral decay for the receptor binding domain tracked with neutralizing antibody titers, while S2-directed responses tracked with antibody-dependent myeloid cellular phagocytosis. Boosting the recovered patients with mRNA or inactivated vaccines expanded humoral breadth, durability, and restored functional responses, eliminating the severity- and platform-associated decay differences. Therefore, post-recovery hybrid immunization compensates for this distinction and broadens humoral breadth, highlighting the value of boosting immunity in previously infected individuals. One Sentence SummaryInfection- and vaccine-acquired immunity to COVID-19 exhibit different functional antibody profiles, each characterized by distinct kinetics of waning over time.

SARS-CoV-2 infection and vaccination elicit potent immune responses. Our study presents a comprehensive multimodal single-cell dataset of peripheral blood of patients with acute COVID-19 and of healthy volunteers before and after receiving the SARS-CoV-2 mRNA vaccine and booster. We compared host immune responses to the virus and vaccine using transcriptional profiling, coupled with B/T cell receptor repertoire reconstruction. COVID-19 patients displayed an enhanced interferon signature and cytotoxic gene upregulation, absent in vaccine recipients. These findings were validated in an independent dataset. Analysis of B and T cell repertoires revealed that, while the majority of clonal lymphocytes in COVID-19 patients were effector cells, clonal expansion was more evident among circulating memory cells in vaccine recipients. Furthermore, while clonal {beta} T cell responses were observed in both COVID-19 patients and vaccine recipients, dramatic expansion of clonal {gamma}{delta}T cells was found only in infected individuals. Our dataset enables comparative analyses of immune responses to infection versus vaccination, including clonal B and T cell responses. Integrating our data with publicly available datasets allowed us to validate our findings in larger cohorts. To our knowledge, this is the first dataset to include comprehensive profiling of longitudinal samples from healthy volunteers pre/post SARS-CoV-2 vaccine and booster.

The dynamics of innate and adaptive immunity to infection in infants remain obscure. Here, we used a multi-omics approach to perform a longitudinal analysis of immunity to SARS-CoV-2 infection in infants and young children in the first weeks and months of life by analyzing blood samples collected before, during, and after infection with Omicron and Non-Omicron variants. Infection stimulated robust antibody titers that, unlike in adults, were stably maintained for >300 days. Antigen-specific memory B cell (MCB) responses were durable for 150 days but waned thereafter. Somatic hypermutation of V-genes in MCB accumulated progressively over 9 months. The innate response was characterized by upregulation of activation markers on blood innate cells, and a plasma cytokine profile distinct from that seen in adults, with no inflammatory cytokines, but an early and transient accumulation of chemokines (CXCL10, IL8, IL-18R1, CSF-1, CX3CL1), and type I IFN. The latter was strongly correlated with viral load, and expression of interferon-stimulated genes (ISGs) in myeloid cells measured by single-cell transcriptomics. Consistent with this, single-cell ATAC-seq revealed enhanced accessibility of chromatic loci targeted by interferon regulatory factors (IRFs) and reduced accessibility of AP-1 targeted loci, as well as traces of epigenetic imprinting in monocytes, during convalescence. Together, these data provide the first snapshot of immunity to infection during the initial weeks and months of life.

The rapid global spread of SARS-CoV-2 and resultant mortality and social disruption have highlighted the need to better understand coronavirus immunity to expedite vaccine development efforts. Multiple candidate vaccines, designed to elicit protective neutralising antibodies targeting the viral spike glycoprotein, are rapidly advancing to clinical trial. However, the immunogenic properties of the spike protein in humans are unresolved. To address this, we undertook an in-depth characterisation of humoral and cellular immunity against SARS-CoV-2 spike in humans following mild to moderate SARS-CoV-2 infection. We find serological antibody responses against spike are routinely elicited by infection and correlate with plasma neutralising activity and capacity to block ACE2/RBD interaction. Expanded populations of spike-specific memory B cells and circulating T follicular helper cells (cTFH) were detected within convalescent donors, while responses to the receptor binding domain (RBD) constitute a minor fraction. Using regression analysis, we find high plasma neutralisation activity was associated with increased spike-specific antibody, but notably also with the relative distribution of spike-specific cTFH subsets. Thus both qualitative and quantitative features of B and T cell immunity to spike constitute informative biomarkers of the protective potential of novel SARS-CoV-2 vaccines.