Phenotypes and Functions of SARS-CoV-2-Reactive T Cells.

Infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19), which is an ongoing pandemic disease. SARS-CoV-2-specific CD4+ and CD8+ T-cell responses have been detected and characterized not only in COVID-19 patients and convalescents, but also unexposed individuals. Here, we review the phenotypes and functions of SARS-CoV-2-specific T cells in COVID-19 patients and the relationships between SARS-CoV-2-specific T-cell responses and COVID-19 severity. In addition, we describe the phenotypes and functions of SARS-CoV-2-specific memory T cells after recovery from COVID-19 and discuss the presence of SARS-CoV-2-reactive T cells in unexposed individuals and SARS-CoV-2-specific T-cell responses elicited by COVID-19 vaccines. A better understanding of T-cell responses is important for effective control of the current COVID-19 pandemic.

Structural assessment of HLA-A2-restricted SARS-CoV-2 spike epitopes recognized by public and private T-cell receptors.

T cells play a vital role in combatting SARS-CoV-2 and forming long-term memory responses. Whereas extensive structural information is available on neutralizing antibodies against SARS-CoV-2, such information on SARS-CoV-2-specific T-cell receptors (TCRs) bound to their peptide-MHC targets is lacking. Here we determine the structures of a public and a private TCR from COVID-19 convalescent patients in complex with HLA-A2 and two SARS-CoV-2 spike protein epitopes (YLQ and RLQ). The structures reveal the basis for selection of particular TRAV and TRBV germline genes by the public but not the private TCR, and for the ability of the TCRs to recognize natural variants of RLQ but not YLQ. Neither TCR recognizes homologous epitopes from human seasonal coronaviruses. By elucidating the mechanism for TCR recognition of an immunodominant yet variable epitope (YLQ) and a conserved but less commonly targeted epitope (RLQ), this study can inform prospective efforts to design vaccines to elicit pan-coronavirus immunity.

Assessment of SARS-CoV-2 Immunity in Convalescent Children and Adolescents.

Persistence of protective immunity for SARS-CoV-2 is important against reinfection. Knowledge on SARS-CoV-2 immunity in pediatric patients is currently lacking. We opted to assess the SARS-CoV-2 adaptive immunity in recovered children and adolescents, addressing the pediatrics specific immunity towards COVID-19. Two independent assays were performed to investigate humoral and cellular immunological memory in pediatric convalescent COVID-19 patients. Specifically, RBD IgG, CD4+, and CD8+ T cell responses were identified and quantified in recovered children and adolescents. SARS-CoV-2-specific RBD IgG detected in recovered patients had a half-life of 121.6 days and estimated duration of 7.9 months compared with baseline levels in controls. The specific T cell response was shown to be independent of days after diagnosis. Both CD4+ and CD8+ T cells showed robust responses not only to spike (S) peptides (a main target of vaccine platforms) but were also similarly activated when stimulated by membrane (M) and nuclear (N) peptides. Importantly, we found the differences in the adaptive responses were correlated with the age of the recovered patients. The CD4+ T cell response to SARS-CoV-2 S peptide in children aged <12 years correlated with higher SARS-CoV-2 RBD IgG levels, suggesting the importance of a T cell-dependent humoral response in younger children under 12 years. Both cellular and humoral immunity against SARS-CoV-2 infections can be induced in pediatric patients. Our important findings provide fundamental knowledge on the immune memory responses to SARS-CoV-2 in recovered pediatric patients.

Immunoinformatic Analysis Reveals Antigenic Heterogeneity of Epstein-Barr Virus Is Immune-Driven.

Whole genome sequencing of Epstein-Barr virus (EBV) isolates from around the world has uncovered pervasive strain heterogeneity, but the forces driving strain diversification and the impact on immune recognition remained largely unknown. Using a data mining approach, we analyzed more than 300 T-cell epitopes in 168 published EBV strains. Polymorphisms were detected in approximately 65% of all CD8+ and 80% of all CD4+ T-cell epitopes and these numbers further increased when epitope flanking regions were included. Polymorphisms in CD8+ T-cell epitopes often involved MHC anchor residues and resulted in changes of the amino acid subgroup, suggesting that only a limited number of conserved T-cell epitopes may represent generic target antigens against different viral strains. Although considered the prototypic EBV strain, the rather low degree of overlap with most other viral strains implied that B95.8 may not represent the ideal reference strain for T-cell epitopes. Instead, a combinatorial library of consensus epitopes may provide better targets for diagnostic and therapeutic purposes when the infecting strain is unknown. Polymorphisms were significantly enriched in epitope versus non-epitope protein sequences, implicating immune selection in driving strain diversification. Remarkably, CD4+ T-cell epitopes in EBNA2, EBNA-LP, and the EBNA3 family appeared to be under negative selection pressure, hinting towards a beneficial role of immune responses against these latency type III antigens in virus biology. These findings validate this immunoinformatics approach for providing novel insight into immune targets and the intricate relationship of host defense and virus evolution that may also pertain to other pathogens.

Elucidating T Cell and B Cell Responses to SARS-CoV-2 in Humans: Gaining Insights into Protective Immunity and Immunopathology.

The SARS-CoV-2 pandemic is an unprecedented epochal event on at least two fronts. Firstly, in terms of the rapid spread and the magnitude of the outbreak, and secondly, on account of the equally swift response of the scientific community that has galvanized itself into action and has successfully developed, tested and deployed highly effective and novel vaccines in record time to combat the virus. The sophistication and diversification of the scientific toolbox we now have at our disposal has enabled us to interrogate both the breadth and the depth of the immune response to a degree that is unparalleled in recent memory. In terms of our understanding of what is critical to contain the virus and mitigate the effects the pandemic, neutralizing antibodies to SARS-CoV-2 garner most of the attention, however, it is essential to recognize that it is the quality and the fitness of the virus-specific T cell and B cell response that lays the foundation and the backdrop for an effective neutralizing antibody response. In this report, we will review some of the key findings that have helped define and delineate some of the essential attributes of T and B cell responses in the setting of SARS-CoV-2 infection.

Analysis of TCR Repertoire by High-Throughput Sequencing Indicates the Feature of T Cell Immune Response after SARS-CoV-2 Infection.

Coronavirus disease 2019 (COVID-19) is a global infectious disease caused by the SARS-CoV-2 coronavirus. T cells play an essential role in the body's fighting against the virus invasion, and the T cell receptor (TCR) is crucial in T cell-mediated virus recognition and clearance. However, little has been known about the features of T cell response in convalescent COVID-19 patients. In this study, using 5'RACE technology and PacBio sequencing, we analyzed the TCR repertoire of COVID-19 patients after recovery for 2 weeks and 6 months compared with the healthy donors. The TCR clustering and CDR3 annotation were exploited to discover groups of patient-specific TCR clonotypes with potential SARS-CoV-2 antigen specificities. We first identified CD4+ and CD8+ T cell clones with certain clonal expansion after infection, and then observed the preferential recombination usage of V(D) J gene segments in CD4+ and CD8+ T cells of COVID-19 patients with different convalescent stages. More important, the TRBV6-5-TRBD2-TRBJ2-7 combination with high frequency was shared between CD4+ T and CD8+ T cells of different COVID-19 patients. Finally, we found the dominant characteristic motifs of the CDR3 sequence between recovered COVID-19 and healthy control. Our study provides novel insights on TCR in COVID-19 with different convalescent phases, contributing to our understanding of the immune response induced by SARS-CoV-2.

Circulating integrin α4 ß7+ CD4 T cells are enriched for proliferative transcriptional programs in HIV infection.

HIV preferentially infects α4 ß7+ CD4 T cells, forming latent reservoirs that contribute to HIV persistence during antiretroviral therapy. However, the properties of α4 ß7+ CD4 T cells in blood and mucosal compartments remain understudied. Employing two distinct models of HIV infection, HIV-infected humans and simian-human immunodeficiency virus (SHIV)-infected rhesus macaques, we show that α4 ß7+ CD4 T cells in blood are enriched for genes regulating cell cycle progression and cellular metabolism. Unlike their circulating counterparts, rectal α4 ß7+ CD4 T cells exhibited a core tissue-residency gene expression program. These features were conserved across primate species, indicating that the environment influences memory T-cell transcriptional networks. Our findings provide an important molecular foundation for understanding the role of α4 ß7 in HIV infection.

Does immune recognition of SARS-CoV2 epitopes vary between different ethnic groups?

The SARS-CoV2 mediated Covid-19 pandemic has impacted humankind at an unprecedented scale. While substantial research efforts have focused towards understanding the mechanisms of viral infection and developing vaccines/ therapeutics, factors affecting the susceptibility to SARS-CoV2 infection and manifestation of Covid-19 remain less explored. Given that the Human Leukocyte Antigen (HLA) system is known to vary among ethnic populations, it is likely to affect the recognition of the virus, and in turn, the susceptibility to Covid-19. To understand this, we used bioinformatic tools to probe all SARS-CoV2 peptides which could elicit T-cell response in humans. We also tried to answer the intriguing question of whether these potential epitopes were equally immunogenic across ethnicities, by studying the distribution of HLA alleles among different populations and their share of cognate epitopes. Results indicate that the immune recognition potential of SARS-CoV2 epitopes tend to vary between different ethnic groups. While the South Asians are likely to recognize higher number of CD8-specific epitopes, Europeans are likely to identify higher number of CD4-specific epitopes. We also hypothesize and provide clues that the newer mutations in SARS-CoV2 are unlikely to alter the T-cell mediated immunogenic responses among the studied ethnic populations. The work presented herein is expected to bolster our understanding of the pandemic, by providing insights into differential immunological response of ethnic populations to the virus as well as by gaging the possible effects of mutations in SARS-CoV2 on efficacy of potential epitope-based vaccines through evaluating ∼40,000 viral genomes.

Robust SARS-CoV-2-specific T cell immunity is maintained at 6 months following primary infection.

The immune response to SARS-CoV-2 is critical in controlling disease, but there is concern that waning immunity may predispose to reinfection. We analyzed the magnitude and phenotype of the SARS-CoV-2-specific T cell response in 100 donors at 6 months following infection. T cell responses were present by ELISPOT and/or intracellular cytokine staining analysis in all donors and characterized by predominant CD4+ T cell responses with strong interleukin (IL)-2 cytokine expression. Median T cell responses were 50% higher in donors who had experienced a symptomatic infection, indicating that the severity of primary infection establishes a 'set point' for cellular immunity. T cell responses to spike and nucleoprotein/membrane proteins were correlated with peak antibody levels. Furthermore, higher levels of nucleoprotein-specific T cells were associated with preservation of nucleoprotein-specific antibody level although no such correlation was observed in relation to spike-specific responses. In conclusion, our data are reassuring that functional SARS-CoV-2-specific T cell responses are retained at 6 months following infection.

Yeast display of MHC-II enables rapid identification of peptide ligands from protein antigens (RIPPA).

CD4+ T cells orchestrate adaptive immune responses via binding of antigens to their receptors through specific peptide/MHC-II complexes. To study these responses, it is essential to identify protein-derived MHC-II peptide ligands that constitute epitopes for T cell recognition. However, generating cells expressing single MHC-II alleles and isolating these proteins for use in peptide elution or binding studies is time consuming. Here, we express human MHC alleles (HLA-DR4 and HLA-DQ6) as native, noncovalent αß dimers on yeast cells for direct flow cytometry-based screening of peptide ligands from selected antigens. We demonstrate rapid, accurate identification of DQ6 ligands from pre-pro-hypocretin, a narcolepsy-related immunogenic target. We also identify 20 DR4-binding SARS-CoV-2 spike peptides homologous to SARS-CoV-1 epitopes, and one spike peptide overlapping with the reported SARS-CoV-2 epitope recognized by CD4+ T cells from unexposed individuals carrying DR4 subtypes. Our method is optimized for immediate application upon the emergence of novel pathogens.

Profound Treg perturbations correlate with COVID-19 severity.

The hallmark of severe COVID-19 is an uncontrolled inflammatory response, resulting from poorly understood immunological dysfunction. We hypothesized that perturbations in FoxP3+ T regulatory cells (Treg), key enforcers of immune homeostasis, contribute to COVID-19 pathology. Cytometric and transcriptomic profiling revealed a distinct Treg phenotype in severe COVID-19 patients, with an increase in Treg proportions and intracellular levels of the lineage-defining transcription factor FoxP3, correlating with poor outcomes. These Tregs showed a distinct transcriptional signature, with overexpression of several suppressive effectors, but also proinflammatory molecules like interleukin (IL)-32, and a striking similarity to tumor-infiltrating Tregs that suppress antitumor responses. Most marked during acute severe disease, these traits persisted somewhat in convalescent patients. A screen for candidate agents revealed that IL-6 and IL-18 may individually contribute different facets of these COVID-19-linked perturbations. These results suggest that Tregs may play nefarious roles in COVID-19, by suppressing antiviral T cell responses during the severe phase of the disease, and by a direct proinflammatory role.

Reduction and exhausted features of T lymphocytes under serological changes, and prognostic factors in COVID-19 progression.

AIMS: Coronavirus disease 2019 (COVID-19) is a novel viral infection threatening worldwide health as currently there exists no effective treatment strategy and vaccination programs are not publicly available yet. T lymphocytes play an important role in antiviral defenses. However, T cell frequency and functionality may be affected during the disease. MATERIAL AND METHODS: Total blood samples were collected from patients with mild and severe COVID-19, and the total lymphocyte number, as well as CD4+ and CD8 + T cells were assessed using flowcytometry. Besides, the expression of exhausted T cell markers was evaluated. The levels of proinflammatory cytokines were also investigated in the serum of all patients using enzyme-linked immunesorbent assay (ELISA). Finally, the obtained results were analyzed along with laboratory serological reports. RESULTS: COVID-19 patients showed lymphopenia and reduced CD4+ and CD8 + T cells, as well as high percentage of PD-1 expression by T cells, especially in severe cases. Serum secretion of TNF-α, IL-1ß, and IL-2 receptor (IL-2R) were remarkably increased in patients with severe symptoms, as compared with healthy controls. Moreover, high levels of triglyceride (TG) and low density lipoprotein cholesterol (LDL-C), were correlated with the severity of the disease. CONCLUSION: Reduced number and function of T cells were observed in COVID-19 patients, especially in severe patients. Meanwhile, the secretion of proinflammatory cytokines was increased as the disease developed. High level of serum IL-2R was also considered as a sign of lymphopenia. Additionally, hypercholesterolemia and hyperlipidemia could be important prognostic factors in determining the severity of the infection.

Single-cell RNA sequencing reveals ex vivo signatures of SARS-CoV-2-reactive T cells through 'reverse phenotyping'.

The in vivo phenotypic profile of T cells reactive to severe acute respiratory syndrome (SARS)-CoV-2 antigens remains poorly understood. Conventional methods to detect antigen-reactive T cells require in vitro antigenic re-stimulation or highly individualized peptide-human leukocyte antigen (pHLA) multimers. Here, we use single-cell RNA sequencing to identify and profile SARS-CoV-2-reactive T cells from Coronavirus Disease 2019 (COVID-19) patients. To do so, we induce transcriptional shifts by antigenic stimulation in vitro and take advantage of natural T cell receptor (TCR) sequences of clonally expanded T cells as barcodes for 'reverse phenotyping'. This allows identification of SARS-CoV-2-reactive TCRs and reveals phenotypic effects introduced by antigen-specific stimulation. We characterize transcriptional signatures of currently and previously activated SARS-CoV-2-reactive T cells, and show correspondence with phenotypes of T cells from the respiratory tract of patients with severe disease in the presence or absence of virus in independent cohorts. Reverse phenotyping is a powerful tool to provide an integrated insight into cellular states of SARS-CoV-2-reactive T cells across tissues and activation states.

Human CD4+ T cells specific for dominant epitopes of SARS-CoV-2 Spike and Nucleocapsid proteins with therapeutic potential.

Since December 2019, Coronavirus disease-19 (COVID-19) has spread rapidly throughout the world, leading to a global effort to develop vaccines and treatments. Despite extensive progress, there remains a need for treatments to bolster the immune responses in infected immunocompromised individuals, such as cancer patients who recently underwent a haematopoietic stem cell transplantation. Immunological protection against COVID-19 is mediated by both short-lived neutralizing antibodies and long-lasting virus-reactive T cells. Therefore, we propose that T cell therapy may augment efficacy of current treatments. For the greatest efficacy with minimal adverse effects, it is important that any cellular therapy is designed to be as specific and directed as possible. Here, we identify T cells from COVID-19 patients with a potentially protective response to two major antigens of the SARS-CoV-2 virus, Spike and Nucleocapsid protein. By generating clones of highly virus-reactive CD4+ T cells, we were able to confirm a set of nine immunodominant epitopes and characterize T cell responses against these. Accordingly, the sensitivity of T cell clones for their specific epitope, as well as the extent and focus of their cytokine response was examined. Moreover, using an advanced T cell receptor (TCR) sequencing approach, we determined the paired TCR-αß sequences of clones of interest. While these data on a limited population require further expansion for universal application, the results presented here form a crucial first step towards TCR-transgenic CD4+ T cell therapy of COVID-19.

HIV/SARS-CoV-2 coinfection: A global perspective.

Since its first appearance in Wuhan, China, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly spread throughout the world and has become a global pandemic. Several medical comorbidities have been identified as risk factors for coronavirus disease 2019 (COVID-19). However, it remains unclear whether people living with human immunodefeciency virus (PLWH) are at an increased risk of COVID-19 and severe disease manifestation, with controversial suggestion that HIV-infected individuals could be protected from severe COVID-19 by means of antiretroviral therapy or HIV-related immunosuppression. Several cases of coinfection with HIV and SARS-CoV-2 have been reported from different parts of the globe. This review seeks to provide a holistic overview of SARS-CoV-2 infection in PLWH.

Immune responses and pathogenesis in persistently PCR-positive patients with SARS-CoV-2 infection.

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 emerged in China in December 2019 and then rapidly spread worldwide. Why COVID-19 patients with the same clinical condition have different outcomes remains unclear. This study aimed to examine the differences in the phenotype and functions of major populations of immune cells between COVID-19 patients with same severity but different outcomes. Four common type adult inpatients with laboratory confirmed COVID-19 from Beijing YouAn Hospital, Capital Medical University were included in this study. The patients were divided into two groups based on whether or not COVID-19 polymerase chain reaction (PCR)-negative conversion occurred within 3 weeks. Peripheral blood samples were collected to compare the differences in the phenotype and functions of major populations of immune cells between the two groups of patients. The result shows that the proportions of CD3+ CD8+ CD38+ HLA-DR+ CD27- effector T killer cells generally declined, whereas that of CD3+ CD4+ CD8+ double-positive T cells (DPTs) increased in the persistently PCR-positive patients. In summary, considering the imbalance between effector T killer cells/CD3+CD4+CD8+ DPTs was a possible key factor for PCR-negative conversion in patients with COVID-19.

Immune Memory in Mild COVID-19 Patients and Unexposed Donors Reveals Persistent T Cell Responses After SARS-CoV-2 Infection.

Understanding the causes of the diverse outcome of COVID-19 pandemic in different geographical locations is important for the worldwide vaccine implementation and pandemic control responses. We analyzed 42 unexposed healthy donors and 28 mild COVID-19 subjects up to 5 months from the recovery for SARS-CoV-2 specific immunological memory. Using HLA class II predicted peptide megapools, we identified SARS-CoV-2 cross-reactive CD4+ T cells in around 66% of the unexposed individuals. Moreover, we found detectable immune memory in mild COVID-19 patients several months after recovery in the crucial arms of protective adaptive immunity; CD4+ T cells and B cells, with a minimal contribution from CD8+ T cells. Interestingly, the persistent immune memory in COVID-19 patients is predominantly targeted towards the Spike glycoprotein of the SARS-CoV-2. This study provides the evidence of both high magnitude pre-existing and persistent immune memory in Indian population. By providing the knowledge on cellular immune responses to SARS-CoV-2, our work has implication for the development and implementation of vaccines against COVID-19.

Efficacy and safety of human papillomavirus vaccination in HIV-infected patients: a systematic review and meta-analysis.

The prophylactic vaccines available to protect against infections by HPV are well tolerated and highly immunogenic. People with HIV have a higher risk of developing HPV infection and HPV-associated cancers due to a lower immune response, and due to viral interactions. We performed a systematic review of RCTs to assess HPV vaccines efficacy and safety on HIV-infected people compared to placebo or no intervention in terms of seroconversion, infections, neoplasms, adverse events, CD4+ T-cell count and HIV viral load. The vaccine-group showed a seroconversion rate close to 100% for each vaccine and a significantly higher level of antibodies against HPV vaccine types, as compared to the placebo group (MD = 4333.3, 95% CI 2701.4; 5965.1 GMT EL.U./ml for HPV type 16 and MD = 1408.8, 95% CI 414.8; 2394.7 GMT EL.U./ml for HPV type 18). There were also no differences in terms of severe adverse events (RR = 0.6, 95% CI 0.2; 1.6) and no severe adverse events (RR = 0.6, 95% CI 0.9; 1.2) between vaccine and placebo groups. Secondary outcomes, such as CD4 + T-cell count and HIV viral load, did not differ between groups (MD = 14.8, 95% CI - 35.1; 64.6 cells/µl and MD = 0.0, 95% CI - 0.3; 0.3 log10 RNA copies/ml, respectively). Information on the remaining outcomes was scarce and that did not allow us to combine the data. The results support the use of the HPV vaccine in HIV-infected patients and highlight the need of further RCTs assessing the effectiveness of the HPV vaccine on infections and neoplasms.

Circulating CD4 T Cells Elicited by Endemic Coronaviruses Display Vast Disparities in Abundance and Functional Potential Linked to Antigen Specificity and Age.

Repeated infections with endemic human coronaviruses (hCoV) are thought to reflect lack of long-lasting protective immunity. We evaluated circulating human CD4 T cells collected prior to 2020 for reactivity towards hCoV spike proteins, probing for the ability to produce interferon-γ, interleukin-2, or granzyme B. We found robust reactivity to spike-derived epitopes, comparable to influenza, but highly variable abundance and functional potential across subjects, depending on age and viral antigen specificity. To explore potential of these memory cells to be recruited in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, we examined the subjects for cross-reactive recognition of epitopes from SARS-CoV-2 nucleocapsid, membrane/envelope, and spike. Functional potential of these cross-reactive CD4 T cells was highly variable; nucleocapsid-specific CD4 T cells but not spike-reactive cells showed exceptionally high levels of granzyme production upon stimulation. These results are considered in light of recruitment of hCoV-reactive cells into responses to SARS-CoV infections or vaccinations.