Establishment and recall of SARS-CoV-2 spike epitope-specific CD4+ T cell memory.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and vaccination elicit CD4+ T cell responses to the spike protein, including circulating follicular helper T (cTFH) cells that correlate with neutralizing antibodies. Using a novel HLA-DRB1*15:01/S751 tetramer to track spike-specific CD4+ T cells, we show that primary infection or vaccination induces robust S751-specific CXCR5- and cTFH cell memory responses. Secondary exposure induced recall of CD4+ T cells with a transitory CXCR3+ phenotype, and drove expansion of cTFH cells transiently expressing ICOS, CD38 and PD-1. In both contexts, cells exhibited a restricted T cell antigen receptor repertoire, including a highly public clonotype and considerable clonotypic overlap between CXCR5- and cTFH populations. Following a third vaccine dose, the rapid re-expansion of spike-specific CD4+ T cells contrasted with the comparatively delayed increase in antibody titers. Overall, we demonstrate that stable pools of cTFH and memory CD4+ T cells established by infection and/or vaccination are efficiently recalled upon antigen reexposure and may contribute to long-term protection against SARS-CoV-2.

Deficiency in the mitophagy mediator Parkin accelerates murine skin allograft rejection.

Alterations in mitochondrial function and associated quality control programs, including mitochondrial-specific autophagy, termed mitophagy, are gaining increasing recognition in the context of disease. However, the role of mitophagy in organ transplant rejection remains poorly understood. Using mice deficient in Parkin, a ubiquitin ligase that tags damaged or dysfunctional mitochondria for autophagic clearance, we assessed the impact of Parkin-dependent mitophagy on skin-graft rejection. We observed accelerated graft loss in Parkin-deficient mice across multiple skin graft models. Immune cell distributions posttransplant were largely unperturbed compared to wild-type; however, the CD8+ T cells of Parkin-deficient mice expressed more T-bet, IFNγ, and Ki67, indicating greater priming toward effector function. This was accompanied by increased circulating levels of IL-12p70 in Parkin-deficient mice. Using a mixed leukocyte reaction, we demonstrated that naïve Parkin-deficient CD4+ and CD8+ T cells exhibit enhanced activation marker expression and proliferative responses to alloantigen, which were attenuated with administration of a pharmacological mitophagy inducer (p62-mediated mitophagy inducer), known to increase mitophagy in the absence of a functional PINK1-Parkin pathway. These findings indicate a role for Parkin-dependent mitophagy in curtailing skin-graft rejection.

Cytokines enhance human Vγ9Vδ2 T-cell TCR-dependent and TCR-independent effector functions.

Vγ9Vδ2 T cells can recognize various molecules associated with cellular stress or transformation, providing a unique avenue for the treatment of cancers or infectious diseases. Nonetheless, Vγ9Vδ2 T-cell-based immunotherapies frequently achieve suboptimal efficacies in vivo. Enhancing the cytotoxic effector function of Vγ9Vδ2 T cells is one potential avenue through which the immunotherapeutic potential of this subset may be improved. We compared the use of four pro-inflammatory cytokines on the effector phenotype and functions of in vitro expanded Vγ9Vδ2 T cells, and demonstrated TCR-independent cytotoxicity mediated through CD26, CD16, and NKG2D, which could be further enhanced by IL-23, IL-18, and IL-15 stimulation throughout expansion. This work defines promising culture conditions that could improve Vγ9Vδ2 T-cell-based immunotherapies and furthers our understanding of how this subset might recognize and target transformed or infected cells.

Cutting Edge: SARS-CoV-2 Infection Induces Robust Germinal Center Activity in the Human Tonsil.

Understanding the generation of immunity to SARS-CoV-2 in lymphoid tissues draining the site of infection has implications for immunity to SARS-CoV-2. We performed tonsil biopsies under local anesthesia in 19 subjects who had recovered from SARS-CoV-2 infection 24-225 d previously. The biopsies yielded >3 million cells for flow cytometric analysis in 17 subjects. Total and SARS-CoV-2 spike-specific germinal center B cells, and T follicular helper cells, were readily detectable in human tonsils early after SARS-CoV-2 infection, as assessed by flow cytometry. Responses were higher in samples within 2 mo of infection but still detectable in some subjects out to 7 mo following infection. We conclude the tonsils are a secondary lymphoid organ that develop germinal center responses to SARS-CoV-2 infection and could play a role in the long-term development of immunity.

Coformulation with Tattoo Ink for Immunological Assessment of Vaccine Immunogenicity in the Draining Lymph Node.

Characterization of germinal center B and T cell responses yields critical insights into vaccine immunogenicity. Nonhuman primates are a key preclinical animal model for human vaccine development, allowing both lymph node (LN) and circulating immune responses to be longitudinally sampled for correlates of vaccine efficacy. However, patterns of vaccine Ag drainage via the lymphatics after i.m. immunization can be stochastic, driving uneven deposition between lymphoid sites and between individual LN within larger clusters. To improve the accurate isolation of Ag-exposed LN during biopsies and necropsies, we developed and validated a method for coformulating candidate vaccines with tattoo ink in both mice and pigtail macaques. This method allowed for direct visual identification of vaccine-draining LN and evaluation of relevant Ag-specific B and T cell responses by flow cytometry. This approach is a significant advancement in improving the assessment of vaccine-induced immunity in highly relevant nonhuman primate models.

Phenotypic and functional characteristics of highly differentiated CD57+NKG2C+ NK cells in HIV-1-infected individuals.

Natural killer (NK) cells are important anti-viral effector cells. The function and phenotype of the NK cells that constitute an individual's NK cell repertoire can be influenced by ongoing or previous viral infections. Indeed, infection with human cytomegalovirus (HCMV) drives the expansion of a highly differentiated NK cell population characterized by expression of CD57 and the activating NKG2C receptor. This NK cell population has also been noted to occur in HIV-1-infected individuals. We evaluated the NK cells of HIV-1-infected and HIV-1-uninfected individuals to determine the relative frequency of highly differentiated CD57+NKG2C+ NK cells and characterize these cells for their receptor expression and responsiveness to diverse stimuli. Highly differentiated CD57+NKG2C+ NK cells occurred at higher frequencies in HCMV-infected donors relative to HCMV-uninfected donors and were dramatically expanded in HIV-1/HCMV co-infected donors. The expanded CD57+NKG2C+ NK cell population in HIV-1-infected donors remained stable following antiretroviral therapy. CD57+NKG2C+ NK cells derived from HIV-1-infected individuals were robustly activated by antibody-dependent stimuli that contained anti-HIV-1 antibodies or therapeutic anti-CD20 antibody, and these NK cells mediated cytolysis through NKG2C. Lastly, CD57+NKG2C+ NK cells from HIV-1-infected donors were characterized by reduced expression of the inhibitory NKG2A receptor. The abundance of highly functional CD57+NKG2C+ NK cells in HIV-1-infected individuals raises the possibility that these NK cells could play a role in HIV-1 pathogenesis or serve as effector cells for therapeutic/cure strategies.

MAIT Cells Upregulate α4ß7 in Response to Acute Simian Immunodeficiency Virus/Simian HIV Infection but Are Resistant to Peripheral Depletion in Pigtail Macaques.

Mucosal-associated invariant T (MAIT) cells are nonconventional T lymphocytes that recognize bacterial metabolites presented by MR1. Whereas gut bacterial translocation and the loss/dysfunction of peripheral MAIT cells in HIV infection is well described, MAIT cells in nonhuman primate models are poorly characterized. We generated a pigtail macaque (PTM)-specific MR1 tetramer and characterized MAIT cells in serial samples from naive and SIV- or simian HIV-infected PTM. Although PTM MAIT cells generally resemble the phenotype and transcriptional profile of human MAIT cells, they exhibited uniquely low expression of the gut-homing marker α4ß7 and were not enriched at the gut mucosa. PTM MAIT cells responded to SIV/simian HIV infection by proliferating and upregulating α4ß7, coinciding with increased MAIT cell frequency in the rectum. By 36 wk of infection, PTM MAIT cells were activated and exhibited a loss of Tbet expression but were not depleted as in HIV infection. Our data suggest the following: 1) MAIT cell activation and exhaustion is uncoupled from the hallmark depletion of MAIT cells during HIV infection; and 2) the lack of PTM MAIT cell enrichment at the gut mucosa may prevent depletion during chronic infection, providing a model to assess potential immunotherapeutic approaches to modify MAIT cell trafficking during HIV infection.

Influenza Virus Infection Enhances Antibody-Mediated NK Cell Functions via Type I Interferon-Dependent Pathways.

Natural killer (NK) cells are an important component in the control of influenza virus infection, acting to both clear virus-infected cells and release antiviral cytokines. Engagement of CD16 on NK cells by antibody-coated influenza virus-infected cells results in antibody-dependent cellular cytotoxicity (ADCC). Increasing the potency of antibody-mediated NK cell activity could ultimately lead to improved control of influenza virus infection. To understand if NK cells can be functionally enhanced following exposure to influenza virus-infected cells, we cocultured human peripheral blood mononuclear cells (PBMCs) with influenza virus-infected human alveolar epithelial (A549) cells and evaluated the capacity of NK cells to mediate antibody-dependent functions. Preincubation of PBMCs with influenza virus-infected cells markedly enhanced the ability of NK cells to respond to immune complexes containing hemagglutinin (HA) and anti-HA antibodies or transformed allogeneic cells in the presence or absence of a therapeutic monoclonal antibody. Cytokine multiplex, RNA sequencing, supernatant transfer, Transwell, and cytokine-blocking/cytokine supplementation experiments showed that type I interferons released from PBMCs were primarily responsible for the influenza virus-induced enhancement of antibody-mediated NK cell functions. Importantly, the influenza virus-mediated increase in antibody-dependent NK cell functionality was mimicked by the type I interferon agonist poly(I·C). We conclude that the type I interferon secretion induced by influenza virus infection enhances the capacity of NK cells to mediate ADCC and that this pathway could be manipulated to alter the potency of anti-influenza virus therapies and vaccines.IMPORTANCE Protection from severe influenza may be assisted by antibodies that engage NK cells to kill infected cells through ADCC. Studies have primarily focused on antibodies that have ADCC activity, rather than the capacity of NK cells to become activated and mediate ADCC during an influenza virus infection. We found that type I interferon released in response to influenza virus infection primes NK cells to become highly reactive to anti-influenza virus ADCC antibodies. Enhancing the capacity of NK cells to mediate ADCC could assist in controlling influenza virus infections.

Modulation of the CCR5 Receptor/Ligand Axis by Seminal Plasma and the Utility of In Vitro versus In Vivo Models.

Sexual HIV-1 transmission occurs primarily in the presence of semen. Although data from macaque studies suggest that CCR5+ CD4+ T cells are initial targets for HIV-1 infection, the impact of semen on T cell CCR5 expression and ligand production remains inconclusive. To determine if semen modulates the lymphocyte CCR5 receptor/ligand axis, primary human T cell CCR5 expression and natural killer (NK) cell anti-HIV-1 antibody-dependent beta chemokine production was assessed following seminal plasma (SP) exposure. Purified T cells produce sufficient quantities of RANTES to result in a significant decline in CCR5bright T cell frequency following 16 h of SP exposure (P = 0.03). Meanwhile, NK cells retain the capacity to produce limited amounts of MIP-1α/MIP-1ß in response to anti-HIV-1 antibody-dependent stimulation (median, 9.5% MIP-1α+ and/or MIP-1ß+), despite the immunosuppressive nature of SP. Although these in vitro experiments suggest that SP-induced CCR5 ligand production results in the loss of surface CCR5 expression on CD4+ T cells, the in vivo implications are unclear. We therefore vaginally exposed five pigtail macaques to SP and found that such exposure resulted in an increase in CCR5+ HIV-1 target cells in three of the animals. The in vivo data support a growing body of evidence suggesting that semen exposure recruits target cells to the vagina that are highly susceptible to HIV-1 infection, which has important implications for HIV-1 transmission and vaccine design.IMPORTANCE The majority of HIV-1 vaccine studies do not take into consideration the impact that semen exposure might have on the mucosal immune system. In this study, we demonstrate that seminal plasma (SP) exposure can alter CCR5 expression on T cells. Importantly, in vitro studies of T cells in culture cannot replicate the conditions under which immune cells might be recruited to the genital mucosa in vivo, leading to potentially erroneous conclusions about the impact of semen on mucosal HIV-1 susceptibility.

γδ T cells mediate robust anti-HIV functions during antiretroviral therapy regardless of immune checkpoint expression.

Objectives: Although antiretroviral therapy (ART) efficiently suppresses HIV viral load, immune dysregulation and dysfunction persist in people living with HIV (PLWH). γδ T cells are functionally impaired during untreated HIV infection, but the extent to which they are reconstituted upon ART is currently unclear. Methods: Utilising a cohort of ART-treated PLWH, we assessed the frequency and phenotype, characterised in vitro functional responses and defined the impact of immune checkpoint marker expression on effector functions of both Vδ1 and Vδ2 T cells. We additionally explore the in vitro expansion of Vδ2 T cells from PLWH on ART and the mechanisms by which such expanded cells may sense and kill HIV-infected targets. Results: A matured NK cell-like phenotype was observed for Vδ1 T cells among 25 ART-treated individuals (PLWH/ART) studied compared to 17 HIV-uninfected controls, with heightened expression of 2B4, CD160, TIGIT and Tim-3. Despite persistent phenotypic perturbations, Vδ1 T cells from PLWH/ART exhibited strong CD16-mediated activation and degranulation, which were suppressed upon Tim-3 and TIGIT crosslinking. Vδ2 T cell degranulation responses to the phosphoantigen (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate at concentrations up to 2 ng mL-1 were significantly impaired in an immune checkpoint-independent manner among ART-treated participants. Nonetheless, expanded Vδ2 T cells from PLWH/ART retained potent anti-HIV effector functions, with the NKG2D receptor contributing substantially to the elimination of infected cells. Conclusion: Our findings highlight that although significant perturbations remain within the γδ T cell compartment throughout ART-treated HIV, both subsets retain the capacity for robust anti-HIV effector functions.

Clonally expanded memory CD8+ T cells accumulate in atherosclerotic plaques and are pro-atherogenic in aged mice.

Aging is a strong risk factor for atherosclerosis and induces accumulation of memory CD8+ T cells in mice and humans. Biological changes that occur with aging lead to enhanced atherosclerosis, yet the role of aging on CD8+ T cells during atherogenesis is unclear. In this study, using femle mice, we found that depletion of CD8+ T cells attenuated atherogenesis in aged, but not young, animals. Furthermore, adoptive transfer of splenic CD8+ T cells from aged wild-type, but not young wild-type, donor mice significantly enhanced atherosclerosis in recipient mice lacking CD8+ T cells. We also characterized T cells in healthy and atherosclerotic young and aged mice by single-cell RNA sequencing. We found specific subsets of age-associated CD8+ T cells, including a Granzyme K+ effector memory subset, that accumulated and was clonally expanded within atherosclerotic plaques. These had transcriptomic signatures of T cell activation, migration, cytotoxicity and exhaustion. Overall, our study identified memory CD8+ T cells as therapeutic targets for atherosclerosis in aging.

Phenotypic and functional characterization of pharmacologically expanded Vγ9Vδ2 T cells in pigtail macaques.

While gaining interest as treatment for cancer and infectious disease, the clinical efficacy of Vγ9Vδ2 T cell-based immunotherapeutics has to date been limited. An improved understanding of γδ T cell heterogeneity across lymphoid and non-lymphoid tissues, before and after pharmacological expansion, is required. Here, we describe the phenotype and tissue distribution of Vγ9Vδ2 T cells at steady state and following in vivo pharmacological expansion in pigtail macaques. Intravenous phosphoantigen administration with subcutaneous rhIL-2 drove robust expansion of Vγ9Vδ2 T cells in blood and pulmonary mucosa, while expansion was confined to the pulmonary mucosa following intratracheal antigen administration. Peripheral blood Vγ9Vδ2 T cell expansion was polyclonal, and associated with a significant loss of CCR6 expression due to IL-2-mediated receptor downregulation. Overall, we show the tissue distribution and phenotype of in vivo pharmacologically expanded Vγ9Vδ2 T cells can be altered based on the antigen administration route, with implications for tissue trafficking and the clinical efficacy of Vγ9Vδ2 T cell immunotherapeutics.

Immune profiling of SARS-CoV-2 infection during pregnancy reveals NK cell and γδ T cell perturbations.

Pregnancy poses a greater risk for severe COVID-19; however, underlying immunological changes associated with SARS-CoV-2 during pregnancy are poorly understood. We defined immune responses to SARS-CoV-2 in unvaccinated pregnant and nonpregnant women with acute and convalescent COVID-19, quantifying 217 immunological parameters. Humoral responses to SARS-CoV-2 were similar in pregnant and nonpregnant women, although our systems serology approach revealed distinct antibody and FcγR profiles between pregnant and nonpregnant women. Cellular analyses demonstrated marked differences in NK cell and unconventional T cell activation dynamics in pregnant women. Healthy pregnant women displayed preactivated NK cells and γδ T cells when compared with healthy nonpregnant women, which remained unchanged during acute and convalescent COVID-19. Conversely, nonpregnant women had prototypical activation of NK and γδ T cells. Activation of CD4+ and CD8+ T cells and T follicular helper cells was similar in SARS-CoV-2-infected pregnant and nonpregnant women, while antibody-secreting B cells were increased in pregnant women during acute COVID-19. Elevated levels of IL-8, IL-10, and IL-18 were found in pregnant women in their healthy state, and these cytokine levels remained elevated during acute and convalescent COVID-19. Collectively, we demonstrate perturbations in NK cell and γδ T cell activation in unvaccinated pregnant women with COVID-19, which may impact disease progression and severity during pregnancy.

Nanoparticles reduce monocytes within the lungs to improve outcomes after influenza virus infection in aged mice.

Older people exhibit dysregulated innate immunity to respiratory viral infections, including influenza and SARS-CoV-2, and show an increase in morbidity and mortality. Nanoparticles are a potential practical therapeutic that could reduce exaggerated innate immune responses within the lungs during viral infection. However, such therapeutics have not been examined for effectiveness during respiratory viral infection, particular in aged hosts. Here, we employed a lethal model of influenza viral infection in vulnerable aged mice to examine the ability of biodegradable, cargo-free nanoparticles, designated ONP-302, to resolve innate immune dysfunction and improve outcomes during infection. We administered ONP-302 via i.v. injection to aged mice at day 3 after infection, when the hyperinflammatory innate immune response was already established. During infection, we found that ONP-302 treatment reduced the numbers of inflammatory monocytes within the lungs and increased their number in both the liver and spleen, without impacting viral clearance. Importantly, cargo-free nanoparticles reduced lung damage, reduced histological lung inflammation, and improved gas exchange and, ultimately, the clinical outcomes in influenza-infected aged mice. In conclusion, ONP-302 improves outcomes in influenza-infected aged mice. Thus, our study provides information concerning a practical therapeutic, which, if translated clinically, could improve disease outcomes for vulnerable older patients suffering from respiratory viral infections.

Anti-PEG Antibodies Boosted in Humans by SARS-CoV-2 Lipid Nanoparticle mRNA Vaccine.

Humans commonly have low level antibodies to poly(ethylene) glycol (PEG) due to environmental exposure. Lipid nanoparticle (LNP) mRNA vaccines for SARS-CoV-2 contain small amounts of PEG, but it is not known whether PEG antibodies are enhanced by vaccination and what their impact is on particle-immune cell interactions in human blood. We studied plasma from 130 adults receiving either the BNT162b2 (Pfizer-BioNTech) or mRNA-1273 (Moderna) mRNA vaccines or no SARS-CoV-2 vaccine for PEG-specific antibodies. Anti-PEG IgG was commonly detected prior to vaccination and was significantly boosted a mean of 13.1-fold (range 1.0-70.9) following mRNA-1273 vaccination and a mean of 1.78-fold (range 0.68-16.6) following BNT162b2 vaccination. Anti-PEG IgM increased 68.5-fold (range 0.9-377.1) and 2.64-fold (0.76-12.84) following mRNA-1273 and BNT162b2 vaccination, respectively. The rise in PEG-specific antibodies following mRNA-1273 vaccination was associated with a significant increase in the association of clinically relevant PEGylated LNPs with blood phagocytes ex vivo. PEG antibodies did not impact the SARS-CoV-2 specific neutralizing antibody response to vaccination. However, the elevated levels of vaccine-induced anti-PEG antibodies correlated with increased systemic reactogenicity following two doses of vaccination. We conclude that PEG-specific antibodies can be boosted by LNP mRNA vaccination and that the rise in PEG-specific antibodies is associated with systemic reactogenicity and an increase of PEG particle-leukocyte association in human blood. The longer-term clinical impact of the increase in PEG-specific antibodies induced by lipid nanoparticle mRNA vaccines should be monitored. It may be useful to identify suitable alternatives to PEG for developing next-generation LNP vaccines to overcome PEG immunogenicity in the future.

Lung-resident memory B cells established after pulmonary influenza infection display distinct transcriptional and phenotypic profiles.

Recent studies have established that memory B cells, largely thought to be circulatory in the blood, can take up long-term residency in inflamed tissues, analogous to widely described tissue-resident T cells. The dynamics of recruitment and retention of memory B cells to tissues and their immunological purpose remains unclear. Here, we characterized tissue-resident memory B cells (BRM) that are stably maintained in the lungs of mice after pulmonary influenza infection. Influenza-specific BRM were localized within inducible bronchus-associated lymphoid tissues (iBALTs) and displayed transcriptional signatures distinct from classical memory B cells in the blood or spleen while showing partial overlap with memory B cells in lung-draining lymph nodes. We identified lung-resident markers, including elevated expression of CXCR3, CCR6, and CD69, on hemagglutinin (HA)- and nucleoprotein (NP)-specific lung BRM. We found that CCR6 facilitates increased recruitment and/or retention of BRM in lungs and differentiation into antibody-secreting cells upon recall. Although expression of CXCR3 and CCR6 was comparable in total and influenza-specific memory B cells isolated across tissues of human donors, CD69 expression was higher in memory B cells from lung and draining lymph nodes of human organ donors relative to splenic and PBMC-derived populations, indicating that mechanisms underpinning BRM localization may be evolutionarily conserved. Last, we demonstrate that human memory B cells in lungs are transcriptionally distinct to populations in lung-draining lymph nodes or PBMCs. These data suggest that BRM may constitute a discrete component of B cell immunity, positioned at the lung mucosa for rapid humoral response against respiratory viral infections.

Serological and cellular inflammatory signatures in end-stage kidney disease and latent tuberculosis.

OBJECTIVES: Tuberculosis comorbidity with chronic diseases including diabetes, HIV and chronic kidney disease is of rising concern. In particular, latent tuberculosis infection (LTBI) comorbidity with end-stage kidney disease (ESKD) is associated with up to 52.5-fold increased risk of TB reactivation to active tuberculosis infection (ATBI). The immunological mechanisms driving this significant rise in TB reactivation are poorly understood. To contribute to this understanding, we performed a comprehensive assessment of soluble and cellular immune features amongst a unique cohort of patients comorbid with ESKD and LTBI. METHODS: We assessed the plasma and cellular immune profiles from patients with and without ESKD and/or LTBI (N = 40). We characterised antibody glycosylation, serum complement and cytokine levels. We also assessed classical and non-classical monocytes and T cells with flow cytometry. Using a systems-based approach, we identified key immunological features that discriminate between the different disease states. RESULTS: Individuals with ESKD exhibited a highly inflammatory plasma profile and an activated cellular state compared with those without ESKD, including higher levels of inflammatory antibody Fc glycosylation structures and activated CX3CR1+ monocytes that correlate with increased inflammatory plasma cytokines. Similar elevated inflammatory signatures were also observed in ESKD+/LTBI+ compared with ESKD-/LTBI+, suggesting that ESKD induces an overwhelming inflammatory immune state. In contrast, no significant inflammatory differences were observed when comparing LTBI+ and LTBI- individuals. CONCLUSION: Our study highlights the highly inflammatory state induced by ESKD. We hypothesise that this inflammatory state could contribute to the increased risk of TB reactivation in ESKD patients.

Immunogenicity of prime-boost protein subunit vaccine strategies against SARS-CoV-2 in mice and macaques.

SARS-CoV-2 vaccines are advancing into human clinical trials, with emphasis on eliciting high titres of neutralising antibodies against the viral spike (S). However, the merits of broadly targeting S versus focusing antibody onto the smaller receptor binding domain (RBD) are unclear. Here we assess prototypic S and RBD subunit vaccines in homologous or heterologous prime-boost regimens in mice and non-human primates. We find S is highly immunogenic in mice, while the comparatively poor immunogenicity of RBD is associated with limiting germinal centre and T follicular helper cell activity. Boosting S-primed mice with either S or RBD significantly augments neutralising titres, with RBD-focussing driving moderate improvement in serum neutralisation. In contrast, both S and RBD vaccines are comparably immunogenic in macaques, eliciting serological neutralising activity that generally exceed levels in convalescent humans. These studies confirm recombinant S proteins as promising vaccine candidates and highlight multiple pathways to achieving potent serological neutralisation.

Adaptive immunity to human coronaviruses is widespread but low in magnitude.

OBJECTIVES: Endemic human coronaviruses (hCoVs) circulate worldwide but cause minimal mortality. Although seroconversion to hCoV is near ubiquitous during childhood, little is known about hCoV-specific T-cell memory in adults. METHODS: We quantified CD4 T-cell and antibody responses to hCoV spike antigens in 42 SARS-CoV-2-uninfected individuals. Antigen-specific memory T cells and circulating T follicular helper (cTFH) cells were identified using an activation-induced marker assay and characterised for memory phenotype and chemokine receptor expression. RESULTS: T-cell responses were widespread within conventional memory and cTFH compartments but did not correlate with IgG titres. SARS-CoV-2 cross-reactive T cells were observed in 48% of participants and correlated with HKU1 memory. hCoV-specific T cells exhibited a CCR6+ central memory phenotype in the blood, but were enriched for frequency and CXCR3 expression in human lung-draining lymph nodes. CONCLUSION: Overall, hCoV-specific humoral and cellular memory are independently maintained, with a shared phenotype existing among coronavirus-specific CD4 T cells. This understanding of endemic coronavirus immunity provides insight into the homeostatic maintenance of immune responses that are likely to be critical components of protection against SARS-CoV-2.