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High-risk groups, including Indigenous people, are at risk of severe COVID-19. Here we found that Australian First Nations peoples elicit effective immune responses to COVID-19 BNT162b2 vaccination, including neutralizing antibodies, receptor-binding domain (RBD) antibodies, SARS-CoV-2 spike-specific B cells, and CD4+ and CD8+ T cells. In First Nations participants, RBD IgG antibody titers were correlated with body mass index and negatively correlated with age. Reduced RBD antibodies, spike-specific B cells and follicular helper T cells were found in vaccinated participants with chronic conditions (diabetes, renal disease) and were strongly associated with altered glycosylation of IgG and increased interleukin-18 levels in the plasma. These immune perturbations were also found in non-Indigenous people with comorbidities, indicating that they were related to comorbidities rather than ethnicity. However, our study is of a great importance to First Nations peoples who have disproportionate rates of chronic comorbidities and provides evidence of robust immune responses after COVID-19 vaccination in Indigenous people.

Significance Indigenous populations worldwide are highly susceptible to influenza virus infections. Vaccination with inactivated virus is highly recommended to protect Indigenous populations, including Indigenous Australians. There is no study to date that assessed immune responses induced by the inactivated seasonal influenza vaccine in the Indigenous population. Vaccine recommendations are thus based on data generated for non-Indigenous populations and might not be representative for Indigenous people. We found robust antibody responses to influenza vaccination induced in Indigenous Australians, with activation profiles of cTFH1 cells at the acute response strongly correlating with total change of antibody vaccine titers induced by vaccination. Our work strongly supports the recommendation of influenza vaccination to protect Indigenous populations from severe seasonal influenza virus infections and subsequent complications., Morbidity and mortality rates from seasonal and pandemic influenza occur disproportionately in high-risk groups, including Indigenous people globally. Although vaccination against influenza is recommended for those most at risk, studies on immune responses elicited by seasonal vaccines in Indigenous populations are largely missing, with no data available for Indigenous Australians and only one report published on antibody responses in Indigenous Canadians. We recruited 78 Indigenous and 84 non-Indigenous Australians vaccinated with the quadrivalent influenza vaccine into the Looking into InFluenza T cell immunity - Vaccination cohort study and collected blood to define baseline, early (day 7), and memory (day 28) immune responses. We performed in-depth analyses of T and B cell activation, formation of memory B cells, and antibody profiles and investigated host factors that could contribute to vaccine responses. We found activation profiles of circulating T follicular helper type-1 cells at the early stage correlated strongly with the total change in antibody titers induced by vaccination. Formation of influenza-specific hemagglutinin-binding memory B cells was significantly higher in seroconverters compared with nonseroconverters. In-depth antibody characterization revealed a reduction in immunoglobulin G3 before and after vaccination in the Indigenous Australian population, potentially linked to the increased frequency of the G3m21* allotype. Overall, our data provide evidence that Indigenous populations elicit robust, broad, and prototypical immune responses following immunization with seasonal inactivated influenza vaccines. Our work strongly supports the recommendation of influenza vaccination to protect Indigenous populations from severe seasonal influenza virus infections and their subsequent complications.

Indigenous people worldwide are at high risk of developing severe influenza disease. HLA-A*24:02 allele, highly prevalent in Indigenous populations, is associated with influenza-induced mortality, although the basis for this association is unclear. Here, we define CD8+ T-cell immune landscapes against influenza A (IAV) and B (IBV) viruses in HLA-A*24:02-expressing Indigenous and non-Indigenous individuals, human tissues, influenza-infected patients and HLA-A*24:02-transgenic mice. We identify immunodominant protective CD8+ T-cell epitopes, one towards IAV and six towards IBV, with A24/PB2550–558-specific CD8+ T cells being cross-reactive between IAV and IBV. Memory CD8+ T cells towards these specificities are present in blood (CD27+CD45RA− phenotype) and tissues (CD103+CD69+ phenotype) of healthy individuals, and effector CD27−CD45RA−PD-1+CD38+CD8+ T cells in IAV/IBV patients. Our data show influenza-specific CD8+ T-cell responses in Indigenous Australians, and advocate for T-cell-mediated vaccines that target and boost the breadth of IAV/IBV-specific CD8+ T cells to protect high-risk HLA-A*24:02-expressing Indigenous and non-Indigenous populations from severe influenza disease.

How innate and adaptive immune responses work in concert to resolve influenza disease is yet to be fully investigated in one single study. Here, we utilize longitudinal samples from patients hospitalized with acute influenza to understand these immune responses. We report the dynamics of 18 important immune parameters, related to clinical, genetic and virological factors, in influenza patients across different severity levels. Influenza disease correlates with increases in IL-6/IL-8/MIP-1α/β cytokines and lower antibody responses. Robust activation of circulating T follicular helper cells correlates with peak antibody-secreting cells and influenza heamaglutinin-specific memory B-cell numbers, which phenotypically differs from vaccination-induced B-cell responses. Numbers of influenza-specific CD8+ or CD4+ T cells increase early in disease and retain an activated phenotype during patient recovery. We report the characterisation of immune cellular networks underlying recovery from influenza infection which are highly relevant to other infectious diseases.

As establishment of SARS-CoV-2-specific T cell memory in children remains largely unexplored, we recruited convalescent COVID-19 children and adults to define their circulating memory SARS-CoV-2-specific CD4+ and CD8+ T cells prior to vaccination. We analysed epitope-specific T cells directly ex vivo using seven HLA class-I and class-II tetramers presenting SARS-CoV-2 epitopes, together with Spike-specific B cells. Unvaccinated children who seroconverted had comparable spike-specific, but lower ORF1a- and N-specific memory T cell responses compared to adults. This agreed with our TCR sequencing data showing reduced clonal expansion in children. A strong stem cell memory phenotype and common T cell receptor motifs were detected within tetramer-specific T cells in seroconverted children. Conversely, children who did not seroconvert had tetramer-specific T cells of predominantly naïve phenotypes and diverse TCRαβ repertoires. Our study demonstrates generation of SARS-CoV-2-specific T cell memory with common TCRαβ motifs in unvaccinated seroconverted children after their first virus encounter.

Children are at lower risk of developing severe COVID-19, yet the underlying immune mechanisms are understudied. While children’s innate immunity can drive rapid resolution of SARS-CoV-2 infection, the establishment of SARS-CoV-2-specific T-cell and B-cell memory in COVID-19 children remains unexplored. We recruited a household cohort to understand SARS-CoV-2-specific CD4+ and CD8+ T-cell immune responses at one month after mild or asymptomatic SARS-CoV-2 infection in PCR-positive children in comparison to those found in their mothers. We analysed SARS-CoV-2-specific T-cell responses, together with B-cells, directly ex vivo using six HLA class-I tetramers and one class-II tetramer presenting SARS57 CoV-2 T-cell epitopes (A1/ORF1a1637, A2/S269, A3/N361, A24/S1208, B7/N105, B40/N322 and DPB4/S167), and Spike- and Receptor Binding Domain (RBD)-specific B-cell probes. Our in depth profiling of epitope-specific T-cell responses at quantitative, phenotypic and clonal levels found that only children who seroconverted had prominent memory T-cell and B-cell profiles. These children had a high magnitude of SARS-CoV-2-specific T-cells displaying memory phenotypes and prevalent T-cell receptor motifs, which were not observed in PCR+ RBD but IgG-negative children. This suggests that seroconversion but not PCR-positivity defines establishment of adaptive SARS-CoV-2-specific immunological memory in children. Our study suggests that COVID-19 vaccination of children could be a major advantage in terms of establishing T-cell and B-cell immunological memory, especially in children who did not seroconvert after SARS-CoV-2 infection.

Background: Morbidity and mortality rates from seasonal and pandemic influenza occur disproportionately in high-risk groups, including Indigenous people globally. Although vaccination against influenza is recommended for those most at risk, studies on immune responses elicited by seasonal vaccines in Indigenous populations are largely missing, with no data available for Indigenous Australians and only one report published on antibody responses in Indigenous Canadians. Methods: We recruited 78 Indigenous and 84 non-Indigenous Australians vaccinated with the quadrivalent influenza vaccine into the LIFT-V cohort, and collected blood to define baseline, early (day 7) and memory (day 28) immune responses. We performed in-depth analyses of T and B cell activation, formation of memory B cells and antibody profiles, and investigated host factors that could contribute to vaccine responses. Findings:We found robust antibody responses to influenza vaccination induced in Indigenous Australians, with activation profiles of circulating T follicular helper type-1 (cT FH 1) cells at the acute response strongly correlating with total change of antibody vaccine titres induced by vaccination. Formation of influenza-specific haemagglutinin (HA)-binding memory B cells was significantly higher in seroconverters compared to non-seroconverters. Back-boosting of antibodies towards previously circulating influenza strains was observed following influenza vaccination. In-depth antibody characterisation revealed a reduction in IgG3 pre- and post-vaccination in the Indigenous Australian population, potentially linked to the increased frequency of G3m21* allotype. Interpretation: Overall, our data provide the first evidence that Indigenous populations elicit robust, broad and prototypical immune responses following immunisation with seasonal inactivated influenza vaccines. Our work strongly supports the recommendation of influenza vaccination to protect Indigenous populations from severe seasonal influenza virus infections and its subsequent complications. Funding: This work was funded by the Australian National Health and Medical Research Council (NHMRC) Program Grant (#1071916) to KK, NHMRC Project Grant (#1122524) to KK, ST, AM, NHMRC Project Grant (#1141840) to MP, KLF, Clifford Craig Foundation Grant (#145) to KLF, and NHMRC Investigator Grant (#1173871) to KK supported this work. EBC is a NHMRC Peter Doherty Fellow. ST is a NHMRC Career Development Fellow (#1145033). This work is supported by the NHMRC program grant 111 #1149990 (SJK), NHMRC project grant #1162760 (AKW). KK is supported by a NHMRC Senior Research Fellowship (#1102792) and SJK by a NHMRC Senior Principal Research Fellowship (#1136322). LH is supported by the Melbourne International Research Scholarship the Melbourne International Fee Remission Scholarship from The University of Melbourne. CES has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No 792532. The Melbourne WHO Collaborating Centre for Reference Research on Influenza is supported by the Australian Government Department of Health. Declaration of Interest: None to declare. Ethical Approval: All experiments were conducted according to the Declaration of Helsinki principles and NHMRC Code of practice and approved by the University of Melbourne Human Research Ethics Committee (#1955465 and #1441452), the Human Research Ethics Committee of the Northern Territory Department of Health and Menzies School of Health Research (#2012-1928), and the Tasmanian Human Research Ethics Committee (#H0015460).

Indigenous people worldwide are at high-risk of developing severe influenza disease. HLA-A*24:02 allele, highly prevalent in Indigenous populations, is associated with influenza-induced mortality, although the basis for this association is unclear. We defined CD8+ T-cell immune landscapes against influenza A (IAV) and B (IBV) viruses in HLA-A*24:02-expressing Indigenous and non-Indigenous individuals, human tissues, influenza-infected patients and HLA-A*24:02-transgenic mice. We identified immunodominant protective CD8+ T-cell epitopes, one towards IAV and six towards IBV, with A24/PB2550-558-specific CD8+ T-cells cells being cross-reactive between IAV and IBV. Memory CD8+ T-cells towards these specificities were present in blood (CD27+CD45RA- phenotype) and tissues (CD103+CD69+ phenotype) of healthy subjects, and effector CD27-CD45RA-PD-1+CD38+CD8+ T-cells in IAV/IBV patients. Our data present the first evidence of influenza-specific CD8+ T-cell responses in Indigenous Australians, and advocate for T-cell-mediated vaccines that target and boost the breadth of IAV/IBV-specific CD8+ T-cells to protect high-risk HLA-A*24:02-expressing Indigenous and non-Indigenous populations from severe influenza disease.One Sentence SummaryInfluenza-specific CD8+ T-cell specificities restricted by HLA-A*24:02.

Indigenous people worldwide are at high risk of developing severe influenza disease. HLA-A*24:02 allele, highly prevalent in Indigenous populations, is associated with influenza-induced mortality, although the basis for this association is unclear. Here, we define CD8+ T-cell immune landscapes against influenza A (IAV) and B (IBV) viruses in HLA-A*24:02-expressing Indigenous and non-Indigenous individuals, human tissues, influenza-infected patients and HLA-A*24:02-transgenic mice. We identify immunodominant protective CD8+ T-cell epitopes, one towards IAV and six towards IBV, with A24/PB2550–558-specific CD8+ T cells being cross-reactive between IAV and IBV. Memory CD8+ T cells towards these specificities are present in blood (CD27+CD45RA− phenotype) and tissues (CD103+CD69+ phenotype) of healthy individuals, and effector CD27−CD45RA−PD-1+CD38+CD8+ T cells in IAV/IBV patients. Our data show influenza-specific CD8+ T-cell responses in Indigenous Australians, and advocate for T-cell-mediated vaccines that target and boost the breadth of IAV/IBV-specific CD8+ T cells to protect high-risk HLA-A*24:02-expressing Indigenous and non-Indigenous populations from severe influenza disease., The immunology of Indigenous populations is generally understudied outside the context of diseases that are prevalent in these communities. Here the authors identify prevalence of influenza CD8+ T cell epitopes in an Indigenous Australian population expressing the susceptibility allomorph HLA A*24:02 and validate immunodominance of some of these epitopes in mice.

We report in-concert dynamics of 18 key immune parameters, related to clinical, genetic and virological factors, in patients hospitalized with influenza across different severity levels. Influenza disease was associated with correlated increases in IL6/IL-8/MIP-1α/β cytokines and lower antibody responses. Robust activation of circulating T follicular helper cells (cTfhs) correlated with peak antibody-secreting cells (ASC) and influenza heamaglutinin-specific memory B-cell numbers, which phenotypically differed from vaccination-induced B-cell responses. Influenza-specific CD8+/CD4+ T-cells increased early in disease and remained activated during patient recovery. Here, we describe the broadest to-date immune cellular networks underlying recovery from influenza infection, highly relevant to other infectious diseases.

Special AT‐rich binding protein‐1 (SATB1) is a global chromatin organizer capable of activating or repressing gene transcription in mice and humans. The role of SATB1 is pivotal for T‐cell development, with SATB1‐knockout mice being neonatally lethal, although the exact mechanism is unknown. Moreover, SATB1 is dysregulated in T‐cell lymphoma and proposed to suppress transcription of the Pdcd1 gene, encoding the immune checkpoint programmed cell death protein 1 (PD‐1). Thus, SATB1 expression in T‐cell subsets across different tissue compartments in humans is of potential importance for targeting PD‐1. Here, we comprehensively analyzed SATB1 expression across different human tissues and immune compartments by flow cytometry and correlated this with PD‐1 expression. We investigated SATB1 protein levels in pediatric and adult donors and assessed expression dynamics of this chromatin organizer across different immune cell subsets in human organs, as well as in antigen‐specific T cells directed against acute and chronic viral infections. Our data demonstrate that SATB1 expression in humans is the highest in T‐cell progenitors in the thymus, and then becomes downregulated in mature T cells in the periphery. Importantly, SATB1 expression in peripheral mature T cells is not static and follows fine‐tuned expression dynamics, which appear to be tissue‐ and antigen‐dependent. Furthermore, SATB1 expression negatively correlates with PD‐1 expression in virus‐specific CD8+ T cells. Our study has implications for understanding the role of SATB1 in human health and disease and suggests an approach for modulating PD‐1 in T cells, highly relevant to human malignancies or chronic viral infections.

Influenza epidemics lead to severe illness, life-threatening complications, and deaths, especially in the elderly. As CD8+ T cells are associated with rapid recovery from influenza, we investigated the effects of aging on antigen-specific CD8+ T cells across the universal influenza epitopes in humans. We show that aging is characterized by altered frequencies in T cell subsets, with naive T cells being partially replaced by activated effector/memory populations. Although we observed no striking differences in TCR signaling capacity, T cells in the elderly had increased expression of transcription factors Eomes and T-bet, and such changes were most apparent in CD8+ T cells. Strikingly, the numbers of antigen-specific CD8+ T cells across universal influenza epitopes were reduced in the elderly, although their effector/memory phenotypes remained stable. To understand whether diminished numbers of influenza-specific CD8+ T cells in the elderly resulted from alteration in TCR clonotypes, we dissected the TCRαβ repertoire specific for the prominent HLA-A*02:01-restricted-M158–66 (A2/M158) influenza epitope. We provide the first ex vivo data on paired antigen-specific TCRαβ clonotypes in the elderly, showing that influenza-specific A2/M158+ TCRαβ repertoires in the elderly adults varied from those in younger adults, with the main features being a reduction in the frequency of the public TRAV27–TRBV19 TCRαβ clonotype, increased proportion of private TCRαβ signatures, broader use of TRAV and TRBV gene segments, and large clonal expansion of private TCRαβ clonotypes with longer CDR3 loops. Our study supports the development of T cell-targeted influenza vaccines that would boost the T cell compartment during life and maintain the numbers and optimal TCRαβ signatures in the elderly.

T cells are defined by a heterodimeric surface receptor, the T cell receptor (TCR), that mediates recognition of pathogen-associated epitopes through interactions with peptide and major histocompatibility complexes (pMHCs). TCRs are generated by genomic rearrangement of the germline TCR locus, a process termed V(D)J recombination, that has the potential to generate marked diversity of TCRs (estimated to range from 1015 (ref. 1) to as high as 1061 (ref. 2) possible receptors). Despite this potential diversity, TCRs from T cells that recognize the same pMHC epitope often share conserved sequence features, suggesting that it may be possible to predictively model epitope specificity. Here we report the in-depth characterization of ten epitope-specific TCR repertoires of CD8+ T cells from mice and humans, representing over 4,600 in-frame single-cell-derived TCRαβ sequence pairs from 110 subjects. We developed analytical tools to characterize these epitope-specific repertoires: a distance measure on the space of TCRs that permits clustering and visualization, a robust repertoire diversity metric that accommodates the low number of paired public receptors observed when compared to single-chain analyses, and a distance-based classifier that can assign previously unobserved TCRs to characterized repertoires with robust sensitivity and specificity. Our analyses demonstrate that each epitope-specific repertoire contains a clustered group of receptors that share core sequence similarities, together with a dispersed set of diverse 'outlier' sequences. By identifying shared motifs in core sequences, we were able to highlight key conserved residues driving essential elements of TCR recognition. These analyses provide insights into the generalizable, underlying features of epitope-specific repertoires and adaptive immune recognition.

Influenza A, B and C viruses (IAV, IBV and ICV, respectively) circulate globally and infect humans, with IAV and IBV causing the most severe disease. CD8+ T cells confer cross-protection against IAV strains, however the responses of CD8+ T cells to IBV and ICV are understudied. We investigated the breadth of CD8+ T cell cross-recognition and provide evidence of CD8+ T cell cross-reactivity across IAV, IBV and ICV. We identified immunodominant CD8+ T cell epitopes from IBVs that were protective in mice and found memory CD8+ T cells directed against universal and influenza-virus-type-specific epitopes in the blood and lungs of healthy humans. Lung-derived CD8+ T cells displayed tissue-resident memory phenotypes. Notably, CD38+Ki67+CD8+ effector T cells directed against novel epitopes were readily detected in IAV- or IBV-infected pediatric and adult subjects. Our study introduces a new paradigm whereby CD8+ T cells confer unprecedented cross-reactivity across all influenza viruses, a key finding for the design of universal vaccines.

Memory CD8(+)T lymphocytes (CTLs) specific for antigenic peptides derived from internal viral proteins confer broad protection against distinct strains of influenza A virus (IAV). However, immune efficacy can be undermined by the emergence of escape mutants. To determine how T-cell receptor (TCR) composition relates to IAV epitope variability, we used ex vivo peptide-HLA tetramer enrichment and single-cell multiplex analysis to compare TCRs targeted to the largely conserved HLA-A*0201-M158and the hypervariable HLA-B*3501-NP418antigens. The TCRαβs for HLA-B*3501-NP418 (+)CTLs varied among individuals and across IAV strains, indicating that a range of mutated peptides will prime different NP418-specific CTL sets. Conversely, a dominant public TRAV27/TRBV19(+)TCRαβ was selected in HLA-A*0201(+)donors responding to M158 This public TCR cross-recognized naturally occurring M158variants complexed with HLA-A*0201. Ternary structures showed that induced-fit molecular mimicry underpins TRAV27/TRBV19(+)TCR specificity for the WT and mutant M158peptides, suggesting the possibility of universal CTL immunity in HLA-A*0201(+)individuals. Combined with the high population frequency of HLA-A*0201, these data potentially explain the relative conservation of M158 Moreover, our results suggest that vaccination strategies aimed at generating broad protection should incorporate variant peptides to elicit cross-reactive responses against other specificities, especially those that may be relatively infrequent among IAV-primed memory CTLs.

Influenza A viruses (IAVs) cause significant morbidity and mortality worldwide, despite new strain-specific vaccines being available annually. As IAV-specific CD8(+) T cells promote viral control in the absence of neutralizing antibodies, and can mediate cross-reactive immunity toward distinct IAVs to drive rapid recovery from both mild and severe influenza disease, there is great interest in developing a universal T cell vaccine. However, despite detailed studies in mouse models of influenza virus infection, there is still a paucity of data on human epitope-specific CD8(+) T cell responses to IAVs. This review focuses on our current understanding of human CD8(+) T cell immunity against distinct IAVs and discusses the possibility of achieving a CD8(+) T cell mediated-vaccine that protects against multiple, distinct IAV strains across diverse human populations. We also review the importance of CD8(+) T cell immunity in individuals highly susceptible to severe influenza infection, including those hospitalised with influenza, the elderly and Indigenous populations.

Influenza A, B and C viruses (IAV, IBV and ICV, respectively) circulate globally and infect humans, with IAV and IBV causing the most severe disease. CD8

Next-generation vaccines that utilize T cells could potentially overcome the limitations of current influenza vaccines that rely on antibodies to provide narrow subtype-specific protection and are prone to antigenic mismatch with circulating strains. Evidence from animal models shows that T cells can provide heterosubtypic protection and are crucial for immune control of influenza virus infections. This has provided hope for the design of a universal vaccine able to prime against diverse influenza virus strains and subtypes. However, multiple hurdles exist for the realisation of a universal T cell vaccine. Overall primary concerns are: extrapolating human clinical studies, seeding durable effective T cell resident memory (Trm), population human leucocyte antigen (HLA) coverage, and the potential for T cell-mediated immune escape. Further comprehensive human clinical data is needed during natural infection to validate the protective role T cells play during infection in the absence of antibodies. Furthermore, fundamental questions still exist regarding the site, longevity and duration, quantity, and phenotype of T cells needed for optimal protection. Standardised experimental methods, and eventually simplified commercial assays, to assess peripheral influenza-specific T cell responses are needed for larger-scale clinical studies of T cells as a correlate of protection against influenza infection. The design and implementation of a T cell-inducing vaccine will require a consensus on the level of protection acceptable in the community, which may not provide sterilizing immunity but could protect the individual from severe disease, reduce the length of infection, and potentially reduce transmission in the community. Therefore, increasing the standard of care potentially offered by T cell vaccines should be considered in the context of pandemic preparedness and zoonotic infections, and in combination with improved antibody vaccine targeting methods. Current pandemic vaccine preparedness measures and ongoing clinical trials under-utilise T cell-inducing vaccines, reflecting the myriad questions that remain about how, when, where, and which T cells are needed to fight influenza virus infection. This review aims to bring together basic fundamentals of T cell biology with human clinical data, which need to be considered for the implementation of a universal vaccine against influenza that harnesses the power of T cells.

Immunization with the inactivated influenza vaccine (IIV) remains the most effective strategy to combat seasonal influenza infections. IIV activates B cells and T follicular helper (TFH) cells and thus engenders antibody-secreting cells and serum antibody titers. However, the cellular events preceding generation of protective immunity in humans are inadequately understood. We undertook an in-depth analysis of B cell and T cell immune responses to IIV in 35 healthy adults. Using recombinant hemagglutinin (rHA) probes to dissect the quantity, phenotype, and isotype of influenza-specific B cells against A/California09-H1N1, A/Switzerland-H3N2, and B/Phuket, we showed that vaccination induced a three-pronged B cell response comprising a transient CXCR5-CXCR3+ antibody-secreting B cell population, CD21hiCD27+ memory B cells, and CD21loCD27+ B cells. Activation of circulating TFH cells correlated with the development of both CD21lo and CD21hi memory B cells. However, preexisting antibodies could limit increases in serum antibody titers. IIV had no marked effect on CD8+, mucosal-associated invariant T, γδ T, and natural killer cell activation. In addition, vaccine-induced B cells were not maintained in peripheral blood at 1 year after vaccination. We provide a dissection of rHA-specific B cells across seven human tissue compartments, showing that influenza-specific memory (CD21hiCD27+) B cells primarily reside within secondary lymphoid tissues and the lungs. Our study suggests that a rational design of universal vaccines needs to consider circulating TFH cells, preexisting serological memory, and tissue compartmentalization for effective B cell immunity, as well as to improve targeting cellular T cell immunity.

Suppressor of cytokine signaling (SOCS) proteins are key regulators of innate and adaptive immunity. Mice lacking functional SOCS4 are hypersusceptible to primary infection with influenza A virus (IAV), displaying dysregulated pro-inflammatory cytokine and chemokine production in the lungs, delayed viral clearance and impaired trafficking of influenza-specific CD8(+) T cells to the site of infection. Therefore, we postulated that SOCS4 is a critical regulator of anti-viral immunity. Unexpectedly, SOCS4 was not required for CD8(+) T-cell memory generation, nor was it required to efficiently recall those cells in response to secondary IAV infection. Wild-type or SOCS4-deficient mice primed and re-challenged with serologically different influenza strains, did not show differences in susceptibility to IAV and cleared the virus from the lungs at the same rate. We have not observed differences in trafficking or numbers of IAV-specific cells, numbers of resident memory T cells or in cytokine profiles in lungs of infected animals. Our data show that despite an impaired primary immune response in Socs4(R108X/R108X) mice, SOCS4 is dispensable for an efficient recall response to influenza virus infection.

The characteristics of the TCR repertoire expressed by epitope-specific CD8+ T cells can be an important determinant of the quality of immune protection against virus infection. Most studies of epitope-specific TCR repertoires focus solely on an analysis of TCR β-chains, rather than the combined TCRαβ heterodimers that confer specificity. Hence, the importance of complementary α- and β-chain pairing in determining TCR specificity and T cell function is not well understood. Our earlier study of influenza-specific TCR repertoires in a C57BL/6J mouse model described a structural basis for preferred TCRαβ pairing that determined exquisite specificity for the DbPA224 epitope from influenza A virus. We have now extended this analysis using retrogenic mice engineered to express single TCR α- or β-chains specific for the DbNP366 or DbPA224 epitopes derived from influenza A virus. We found that particular TCRαβ combinations were selected for recognition of these epitopes following infection, indicating that pairing of certain α- and β-chain sequences is key for determining TCR specificity. Furthermore, we demonstrated that some TCRαβ heterodimers were preferentially expanded from the naive repertoire in response to virus infection, suggesting that appropriate αβ pairing confers optimal T cell responsiveness to Ag.

The ability to decode antigen specificities encapsulated\ud in the sequences of rearranged T-cell receptor\ud (TCR) genes is critical for our understanding of\ud the adaptive immune system and promises signifi-\ud cant advances in the field of translational medicine.\ud Recent developments in high-throughput sequencing\ud methods (immune repertoire sequencing technology,\ud or RepSeq) and single-cell RNA sequencing\ud technology have allowed us to obtain huge numbers\ud of TCR sequences from donor samples and link them\ud to T-cell phenotypes. However, our ability to annotate\ud these TCR sequences still lags behind, owing\ud to the enormous diversity of the TCR repertoire and\ud the scarcity of available data on T-cell specificities. In\ud this paper, we present VDJdb, a database that stores\ud and aggregates the results of published T-cell speci-\ud ficity assays and provides a universal platform that\ud couples antigen specificities with TCR sequences.\ud We demonstrate that VDJdb is a versatile instrument\ud for the annotation of TCR repertoire data, enabling a\ud concatenated view of antigen-specific TCR sequence\ud motifs. VDJdb can be accessed at https://vdjdb.cdr3.\ud net and https://github.com/antigenomics/vdjdb-db.

ObjectiveThe pathogenetic mechanisms by which HLA-DRB1 alleles are associated with anticitrullinated peptide antibody (ACPA)-positive rheumatoid arthritis (RA) are incompletely understood. RA high-risk HLA-DRB1 alleles are known to share a common motif, the ‘shared susceptibility epitope (SE)’. Here, the electropositive P4 pocket of HLA-DRB1 accommodates self-peptide residues containing citrulline but not arginine. HLA-DRB1 His/Phe13β stratifies with ACPA-positive RA, while His13βSer polymorphisms stratify with ACPA-negative RA and RA protection. Indigenous North American (INA) populations have high risk of early-onset ACPA-positive RA, whereby HLA-DRB1*04:04 and HLA-DRB1*14:02 are implicated as risk factors for RA in INA. However, HLA-DRB1*14:02 has a His13βSer polymorphism. Therefore, we aimed to verify this association and determine its molecular mechanism.MethodsHLA genotype was compared in 344 INA patients with RA and 352 controls. Structures of HLA-DRB1*1402-class II loaded with vimentin-64Arg59-71, vimentin-64Cit59-71 and fibrinogen β−74Cit69-81 were solved using X-ray crystallography. Vimentin-64Cit59-71-specific and vimentin59-71-specific CD4+ T cells were characterised by flow cytometry using peptide-histocompatibility leukocyte antigen (pHLA) tetramers. After sorting of antigen-specific T cells, TCRα and β-chains were analysed using multiplex, nested PCR and sequencing.ResultsACPA+ RA in INA was independently associated with HLA-DRB1*14:02. Consequent to the His13βSer polymorphism and altered P4 pocket of HLA-DRB1*14:02, both citrulline and arginine were accommodated in opposite orientations. Oligoclonal autoreactive CD4+ effector T cells reactive with both citrulline and arginine forms of vimentin59-71 were observed in patients with HLA-DRB1*14:02+ RA and at-risk ACPA- first-degree relatives. HLA-DRB1*14:02-vimentin59-71-specific and HLA-DRB1*14:02-vimentin-64Cit59-71-specific CD4+ memory T cells were phenotypically distinct populations.ConclusionHLA-DRB1*14:02 broadens the capacity for citrullinated and native self-peptide presentation and T cell expansion, increasing risk of ACPA+ RA.

Severe influenza A virus (IAV) infection is associated with immune dysfunction. Here, we show circulating CD8+ T-cell profiles from patients hospitalized with avian H7N9, seasonal IAV, and influenza vaccinees. Patient survival reflects an early, transient prevalence of highly activated CD38+HLA-DR+PD-1+ CD8+ T cells, whereas the prolonged persistence of this set is found in ultimately fatal cases. Single-cell T cell receptor (TCR)-αβ analyses of activated CD38+HLA-DR+CD8+ T cells show similar TCRαβ diversity but differential clonal expansion kinetics in surviving and fatal H7N9 patients. Delayed clonal expansion associated with an early dichotomy at a transcriptome level (as detected by single-cell RNAseq) is found in CD38+HLA-DR+CD8+ T cells from patients who succumbed to the disease, suggesting a divergent differentiation pathway of CD38+HLA-DR+CD8+ T cells from the outset during fatal disease. Our study proposes that effective expansion of cross-reactive influenza-specific TCRαβ clonotypes with appropriate transcriptome signatures is needed for early protection against severe influenza disease., Virus-specific CD8+ T cells are crucial during H7N9 influenza infection, but CD8+ T cell dysfunction is associated with poor prognosis. Here, the authors use molecular and phenotypic analysis to establish persistence of clonally diverse CD8+ T cell populations during fatal infection.

Immunization with the inactivated influenza vaccine (IIV) remains the most effective strategy to combat seasonal influenza infections. IIV activates B cells and T follicular helper (T

Influenza epidemics lead to severe illness, life-threatening complications, and deaths, especially in the elderly. As CD8

CD27 is a co-stimulatory immune-checkpoint receptor, constitutively expressed on a broad range of T-cells (αβ and γδ), NK-cells and B-cells. Ligation of CD27 with CD70 results in potent co-stimulatory effects. In mice, co-stimulation of CD8+ T-cells through CD27 promotes immune activation and enhances primary, secondary, memory and recall responses towards viral infections. Limited in vitro human studies support mouse experiments and show that CD27 co-stimulation enhances antiviral T-cell immunity. Given the potent co-stimulatory effects of CD27, manipulating CD27 signalling is of interest for viral, autoimmune and anti-tumour immunotherapies. This review focuses on the role of CD27 co-stimulation in anti-viral T-cell immunity and discusses clinical studies utilising the CD27 co-stimulation pathway for anti-viral, anti-tumour and autoimmune immunotherapy.

Novel influenza viruses often cause differential infection patterns across different age groups, an effect that is defined as heterogeneous demographic susceptibility. This occurred during the A/H2N2 pandemic, when children experienced higher influenza attack rates than adults. Since the recognition of conserved epitopes across influenza subtypes by CD8 + cytotoxic T lymphocytes (CTLs) limit influenza disease, we hypothesized that conservation of CTL antigenic peptides (Ag-p) in viruses circulating before the pH2N2-1957 may have resulted in differential CTL immunity. We compared viruses isolated in the years preceding the pandemic (1941 to 1957) to which children and adults were exposed to viruses circulating decades earlier (1918 to 1940), which could infect adults only. Consistent with phylogenetic models, influenza viruses circulating from 1941 to 1957, which infected children, shared with pH2N2 the majority (∼89%) of the CTL peptides within the most immunogenic nucleoprotein, matrix 1, and polymerase basic 1, thus providing evidence for minimal pH2N2 CTL escape in children. Our study, however, identified potential CTL immune evasion from pH2N2 irrespective of age, within HLA-A*03:01 + individuals for PB1 471 -L473V/N476I variants and HLA-B*15:01 + population for NP 404–414 -V408I mutant. Further experiments using the murine model of B-cell-deficient mice showed that multiple influenza infections resulted in superior protection from influenza-induced morbidity, coinciding with accumulation of tissue-resident memory CD8 + T cells in the lung. Our study suggests that protection against H2N2-1957 pandemic influenza was most likely linked to the number of influenza virus infections prior to the pandemic challenge rather than differential preexisting CTL immunity. Thus, the regimen of a CTL-based vaccine/vaccine-component may benefit from periodic boosting to achieve fully protective, asymptomatic influenza infection. IMPORTANCE Due to a lack of cross-reactive neutralizing antibodies, children are particularly susceptible to influenza infections caused by novel viral strains. Preexisting T cell immunity directed at conserved viral regions, however, can provide protection against influenza viruses, promote rapid recovery and better clinical outcomes. When we asked whether high susceptibility of children (compared to adults) to the pandemic H2N2 influenza strain was associated with immune evasion from T-cell immunity, we found high conservation within T-cell antigenic regions in pandemic H2N2. However, the number of influenza infections prior to the challenge was linked to protective, asymptomatic infections and establishment of tissue-resident memory T cells. Our study supports development of vaccines that prime and boost T cells to elicit cross-strain protective T cells, especially tissue-resident memory T cells, for lifelong immunity against distinct influenza viruses.

Indigenous populations, including Indigenous Australians, are highly susceptible to severe influenza disease and the underlying mechanisms are unknown. We studied immune and genetic factors that could predicate severe influenza disease in Indigenous Australians enrolled in the LIFT study: looking into influenza T-cell immunity. To examine CD8(+) T-cell immunity, we characterised human leukocyte antigen (HLA) profiles. HLA typing confirmed previous studies showing predominant usage of HLA-A*02:01, 11:01, 24:02, 34:01 and HLA-B*13:01, 15:21, 40:01/02, 56:01/02 in Indigenous Australians. We identified two new HLA alleles (HLA-A*02:new and HLA-B*56:new). Modelling suggests that variations within HLA-A*02:new (but not HLA-B56:new) could affect peptide binding. There is a relative lack of known influenza epitopes for the majority of these HLAs, with the exception of a universal HLA-A*02:01-M158 epitope and proposed epitopes presented by HLA-A*11:01/HLA-A*24:02. To dissect universal CD8(+) T-cell responses, we analysed the magnitude, function and T-cell receptor (TCR) clonality of HLA-A*02:01-M158(+)CD8(+) T cells. We found comparable IFN-γ, TNF and CD107a and TCRαβ characteristics in Indigenous and non-Indigenous Australians, suggesting that the ~15% of Indigenous people that express HLA-A*02:01 have universal influenza-specific CD8(+) T-cell immunity. Furthermore, the frequency of an influenza host risk factor, IFITM3-C/C, was comparable between Indigenous Australians and Europeans, suggesting that expression of this allele does not explain increased disease severity at a population level. Our study indicates a need to identify novel influenza-specific CD8(+) T-cell epitopes restricted by HLA-A and HLA-B alleles prevalent in Indigenous populations for the rational design of universal T-cell vaccines.