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ABSTRACT Serological correlates of protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection after vaccination (“vaccine breakthrough”) have been described. However, T cell correlates of protection against breakthrough are incompletely defined, especially the specific contributions of CD4+ and CD8+ T cells. Here, 279 volunteers in the Protective Immunity from T Cells in Healthcare Workers (PITCH) UK cohort study were enrolled in a nested case-control study. Cases were those who tested SARS-CoV-2 PCR or lateral flow device (LFD) positive after two vaccine doses during the Delta-predominant era (n = 32), while controls were those who did not report a positive test or undergo anti-nucleocapsid immunoglobulin G (IgG) seroconversion during this period (n = 247). Previous SARS-CoV-2 infection prior to vaccination was associated with reduced odds of vaccine breakthrough. Using samples from 28 d after the second vaccine dose, before all breakthroughs occurred, we observed future cases had lower ancestral spike (S)- and receptor binding domain-specific IgG titers and S1- and S2-specific T cell interferon gamma (IFNγ) responses compared with controls, although these differences did not persist when individuals were stratified according to previous infection status before vaccination. In a subset of matched infection-naïve cases and controls, vaccine breakthrough cases had lower CD4+ and CD8+ IFNγ and tumor necrosis factor (TNF) responses to Delta S peptides compared with controls. For CD8+ responses, this difference appeared to be driven by reduced responses to Delta compared with ancestral peptides among cases; this reduced response to Delta peptides was not observed in controls. Our findings support a protective role for T cells against Delta breakthrough infection. IMPORTANCE Defining correlates of protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine breakthrough infection informs vaccine policy for booster doses and future vaccine designs. Existing studies demonstrate humoral correlates of protection, but the role of T cells in protection is still unclear. In this study, we explore antibody and T cell immune responses associated with protection against Delta variant vaccine breakthrough infection in a well-characterized cohort of UK Healthcare Workers (HCWs). We demonstrate evidence to support a role for CD4+ and CD8+ T cells as well as antibodies against Delta vaccine breakthrough infection. In addition, our results suggest a potential role for cross-reactive T cells in vaccine breakthrough.

Human MAIT cells sit at the interface between innate and adaptive immunity, are polyfunctional and are capable of killing pathogen infected cells via recognition of the Class IB molecule MR1. MAIT cells have recently been shown to possess an antiviral protective role in vivo and we therefore sought to explore this in relation to HIV-1 infection. There was marked activation of MAIT cells in vivo in HIV-1-infected individuals, which decreased following ART. Stimulation of THP1 monocytes with R5 tropic HIVBAL potently activated MAIT cells in vitro. This activation was dependent on IL-12 and IL-18 but was independent of the TCR. Upon activation, MAIT cells were able to upregulate granzyme B, IFNγ and HIV-1 restriction factors CCL3, 4, and 5. Restriction factors produced by MAIT cells inhibited HIV-1 infection of primary PBMCs and immortalized target cells in vitro. These data reveal MAIT cells to be an additional T cell population responding to HIV-1, with a potentially important role in controlling viral replication at mucosal sites.

Cytolytic CD4+ T cells play a prominent role in chronic viral infection. CD4+ CTLs clones specific for HIV-1 Nef and Gag are capable of killing HIV-1 infected CD4+ T cells and macrophages. Additionally, HIV-specific cytolytic CD4+ T cell responses in acute HIV infection are predictive of disease progression. CD57 expression on CD4s identifies cytolytic cells. These cells were dramatically increased in chronic HIV infection. CD57 expression correlated with cytolytic granules, granzyme B and perforin expression. They express lower CCR5 compared to CD57– cells, have less HIV total DNA, and were a minor component of the HIV reservoir. A small percentage of CD57+ CD4+ CTLs from EC were HIV-specific, could upregulate IFNγ with Gag peptide stimulation, express cytolytic granule markers and maintain TbethighEomes+ transcription factor phenotype. This was not observed in viraemic controllers. The maintenance of HIV-specific CD4 cytolytic function in Elite controllers together with CD8 CTLs may be important for the control of HIV viraemia and of potential relevance to cure strategies.

For many years, the focus of prophylactic vaccines was to elicit neutralizing antibodies, but it has become increasingly evident that T cell-mediated immunity plays a central role in controlling persistent viral infections such as with human immunodeficiency virus, cytomegalovirus, and hepatitis C virus. Currently, various promising prophylactic vaccines, capable of inducing substantial vaccine-specific T cell responses, are investigated in preclinical and clinical studies. There is compelling evidence that protection by T cells is related to the magnitude and breadth of the T cell response, the type and homing properties of the memory T cell subsets, and their cytokine polyfunctionality and metabolic fitness. In this review, we evaluated these key factors that determine the qualitative and quantitative properties of CD4+ and CD8+ T cell responses in the context of chronic viral disease and prophylactic vaccine development. Elucidation of the mechanisms underlying T cell-mediated protection against chronic viral pathogens will facilitate the development of more potent, durable and safe prophylactic T cell-based vaccines.

Summary: The aim of this study was to investigate the pathogenesis of combination ipilimumab and nivolumab‐associated colitis (IN‐COL) by measuring gut‐derived and peripheral blood mononuclear cell (GMNC; PBMC) profiles. We studied GMNC and PBMC from patients with IN‐COL, IN‐treated with no adverse‐events (IN‐NAE), ulcerative colitis (UC) and healthy volunteers using flow cytometry. In the gastrointestinal‐derived cells we found high levels of activated CD8+ T cells and mucosal‐associated invariant T (MAIT) cells in IN‐COL, changes that were not evident in IN‐NAE or UC. UC, but not IN‐C, was associated with a high proportion of regulatory T cells (Treg). We sought to determine if local tissue responses could be measured in peripheral blood. Peripherally, checkpoint inhibition instigated a rise in activated memory CD4+ and CD8+ T cells, regardless of colitis. Low circulating MAIT cells at baseline was associated with IN‐COL patients compared with IN‐NAE in one of two cohorts. UC, but not IN‐COL, was associated with high levels of circulating plasmablasts. In summary, the alterations in T cell subsets measured in IN‐COL‐affected tissue, characterized by high levels of activated CD8+ T cells and MAIT cells and a low proportion of Treg, reflected a pathology distinct from UC. These tissue changes differed from the periphery, where T cell activation was a widespread on‐treatment effect, and circulating MAIT cell count was low but not reliably predictive of colitis. [ABSTRACT FROM AUTHOR]

Summary: High expression of the ATP‐binding cassette‐multi‐drug efflux protein 1 (MDR1) is a striking feature of mucosal‐associated invariant T (MAIT) cells, a prominent human innate‐like T cell subset. We demonstrate significantly higher MDR1 expression by CD8 + CD161 ++ Vα7.2 + MAIT cells than the phenotypically and functionally related CD8 + CD161 ++ Vα7.2‐subset and show MDR1 expression to be similarly high throughout MAIT CD4+ , CD8+ , double‐negative (DN) and double‐positive (DP) cell subsets. We demonstrate the MAIT cell‐predominant CD8+CD161++ subset to uniquely and efficiently efflux the cytotoxic anthracycline daunorubicin, retain function on daunorubicin exposure and demonstrate MDR1‐dependent protection from daunorubicin‐induced apoptosis. By contrast, CD8+CD161++Vα7.2+ MAIT cells were not protected from the anti‐proliferative and cytotoxic effects of the immunosuppressive MDR1 substrates tacrolimus and mycophenoic acid, although function following MAIT cell‐specific T cell receptor (TCR)‐dependent and ‐independent stimulation was preserved on in‐vitro exposure to these agents. Overall, our data further define MDR1 expression by CD161++ T and MAIT cells and demonstrate the potential for high MDR1 expression by MAIT cells to confer resistance to cytotoxic MDR1 substrates in vivo. As our understanding of the importance of MAIT cells in human immunity and immunopathology grows, this is an important observation for clinical contexts such as the treatment of malignancy, autoimmunity and post‐transplant immunosuppression. Mucosal Associated Invariant T (MAIT) cells express the multidrug efflux protein MDR1 across all subsets and at a significantly higher level than by other CD8+ T cells, including those that are CD161++ but Vα7.2α. This uniquely enabled MAIT cells to resist the cytotoxic effects of the anthracycline daunorubicin, but not the anti‐proliferative effects of immunosuppressants such as tacrolimus. Nonetheless, MAIT cell function was preserved upon exposure to both drug classes, which has important implications for the treatment of pathologies including malignancy, autoimmunity and transplant‐associated immunosuppression [ABSTRACT FROM AUTHOR]

A serologic response to hepatitis B virus ( HBV) defined as 'anti- HBc alone' is commonly observed, but its significance remains unclear. This study aimed to define the relationship between 'anti- HBc alone' serostatus and HBV infection, including HBV-specific T- and B-cell memory responses. We enrolled 31 'anti- HBc alone' patients. Total HBV DNA and ccc DNA were tested by nested polymerase chain reaction ( PCR) analysis in liver samples from 22 'anti- HBc alone' patients vs controls (chronic or resolved HBV infection), followed by HBsAg/ HBcAg immunohistochemical (IHC) staining. IFN- γ secretion by HBV-specific T cells was compared in individuals who were 'anti- HBc alone' ( n = 27), resolved HBV ( n = 21), chronic HBV ( n = 24) and 12 healthy controls using enzyme-linked immunospot ( ELISpot) assays. An HBsAg-IgG B-cell ELISpot assay was performed in 'anti- HBc alone' patients before and after one dose of recombinant HBsAg vaccine. The majority (23/31, 74.2%) of the 'anti- HBc alone' individuals were co-infected with HCV. Infrequent intrahepatic total HBV DNA (2/22, 9.1%) and ccc DNA (1/22, 4.5%) were detected in biopsies; HBsAg and HBcAg IHC staining was negative. HBV-specific T-cell responses were similar between 'anti- HBc alone' individuals and HBV resolvers. Circulating HBV-memory B-cell responses were detected in all 'anti- HBc alone' individuals, consistent with an HBsAg-specific memory pool. After one HBV vaccine dose, increased anti- HBs antibody levels were observed, accompanied by an expansion of HBsAg-specific memory B cells ( P = 0.0226). 'Anti- HBc alone' individuals showed HBV-specific T-cell and memory B-cell responses typical of previous viral exposure and protective memory, suggesting a resolved infection. [ABSTRACT FROM AUTHOR]

Antigen-specific T-cells are highly variable, spanning potent antiviral efficacy and damaging auto-reactivity. In virus infections, identifying the most efficacious responses is critical to vaccine design. However, current methods depend on indirect measures or on ex vivo expanded CTL clones. We here describe a novel application of cytotoxic saporin-conjugated tetramers to kill antigen-specific T-cells without significant off-target effects. The relative efficacy of distinct antiviral CD8+ T-cell specificity can be directly assessed via antigen-specific CD8+ T-cell depletion. The utility of these reagents is demonstrated here in identifying the CD8+ T-cell specificity most effective in preventing HIV progression in HIV-infected HLA-B*27-positive immune controllers.

Mutations in the X-linked inhibitor of apoptosis ( XIAP) gene have been associated with XLP-like disease, including recurrent Epstein-Barr virus ( EBV)-related haemophagocytic lymphohystiocytosis (HLH), but the immunopathogenic bases of EBV-related disease in XIAP deficiency is unknown. We present the first analysis of EBV-specific T cell responses in functional XIAP deficiency. In a family of patients with a novel mutation in XIAP ( G466 X) leading to a late-truncated protein and varying clinical features, we identified gradual hypogammaglobulinaemia and large expansions of T cell subsets, including a prominent CD4+ CD8+ population. Extensive ex-vivo analyses showed that the expanded T cell subsets were dominated by EBV-specific cells with conserved cytotoxic, proliferative and interferon ( IFN)-γ secretion capacity. The EBV load in blood fluctuated and was occasionally very high, indicating that the XIAPG466X mutation could impact upon EBV latency. XIAP deficiency may unravel a new immunopathogenic mechanism in EBV-associated disease. [ABSTRACT FROM AUTHOR]

Coinfection with HIV adversely impacts every stage of hepatitis C ( HCV) infection. Liver damage in HCV infection results from host antiviral responses rather than direct viral pathogenesis. Despite depressed cellular immunity, coinfected patients show accelerated hepatic fibrosis compared with HCV monoinfected patients. This paradox is poorly understood. T-regulatory (Treg) cells ( CD4+ and FOXP3+) are hypothesized to limit hepatic damage in HCV. Our hypothesis was that reduced frequency of hepatic Treg in HIV/ HCV coinfection compared with HCV monoinfection may explain poorer outcomes. We quantified FOXP3+, CD4+, CD8+ and CD20+ cells in liver biopsies of 35 male subjects matched by age and ISHAK fibrosis score, 12 HIV monoinfected, 11 HCV monoinfected and 12 HIV/ HCV coinfected. Cell counts were performed using indirect immunohistochemical staining and light microscopy. HIV/ HCV coinfected subjects had fewer hepatic FOXP3+ ( P = 0.031) and CD4+ cells ( P = 0.001) than HCV monoinfected subjects. Coinfected subjects had more hepatic CD8+ cells compared with HCV monoinfected ( P = 0.023), and a lower ratio of FOXP3+ to CD8+ cells (0.08 vs 0.27, P < 0.001). Multivariate analysis showed number of CD4+ cells controlled for differences in number of FOXP3+ cells. Fewer hepatic FOXP3+ and CD4+ cells in HIV/ HCV coinfection compared with HCV monoinfection suggests lower Treg activity, driven by an overall loss of CD4+ cells. Higher number of CD8+ cells in HIV/ HCV coinfection suggests higher cytotoxic activity. This may explain poorer outcomes in HIV/ HCV coinfected patients and suggests a potential mechanism by which highly active antiretroviral therapy may benefit these patients. [ABSTRACT FROM AUTHOR]

Cytomegalovirus (CMV) has been shown to induce large populations of CD8 T-effector memory cells that unlike central memory persist in large quantities following infection, a phenomenon commonly termed “memory inflation”. Although murine models to date have shown very large and persistent CMV-specific T-cell expansions following infection, there is considerable variability in CMV-specific T-memory responses in humans. Historically such memory inflation in humans has been assumed a consequence of reactivation events during the life of the host. Because basic information about CMV infection/re-infection and reactivation in immune competent humans is not available, we used a murine model to test how primary infection, reinfection, and reactivation stimuli influence memory inflation. We show that low titer infections induce “partial” memory inflation of both mCMV specific CD8 T-cells and antibody. We show further that reinfection with different strains can boost partial memory inflation. Finally, we show preliminary results suggesting that a single strong reactivation stimulus does not stimulate memory inflation. Altogether, our results suggest that while high titer primary infections can induce memory inflation, reinfections during the life of a host may be more important than previously appreciated.

The importance of CD8+ T cells in the control of viral infections is well established. However, what differentiates CD8+ T cell responses in individuals who control infection and those who do not is not well understood. ‘Functional sensitivity’ describes an important quality of the T cell response and is determined in part by the affinity of the T cell receptor for antigen. A more sensitive T cell response is generally believed to be more efficient and associated with better control of viral infection, yet may also drive viral mutation and immune escape. Various in vitro techniques have been used to measure T cell sensitivity; however, rapid ex vivo analysis of this has been made possible by the application of the ‘magic’ tetramer technology. Such tools have potentially important applications in the design and evaluation of vaccines. [ABSTRACT FROM AUTHOR]

Background. Human immunodeficiency virus (HIV) may influence the outcome and natural history of hepatitis C virus (HCV) infection through an impact on acute HCV-specific T cell responses. Methods. Fifty-five HIV-positive males with acute HCV infection were identified; monoinfected individuals (n = 8) were used for peripheral blood mononuclear cell comparison. In 14 coinfected and 8 HCV-monoinfected patients, HCV-specific T cell responses against a range of HCV antigens were assessed using interferon (IFN)-γenzyme-linked immunospot (ELISpot) and proliferation assays. E1/E2 region genetic diversity and the selection pressure on the virus were measured in 8 coinfected patients by use of cloned sequences over time. Results. HCV persisted in 52 (95%) coinfected individuals. HCV/HIV coinfection significantly reduced IFN-γELISpot responses versus those in HCV-monoinfected individuals, especially against nonstructural proteins (1/10 vs. 5/8; P = .008). In coinfected patients, increasedHCVgenetic diversity was observed between the first and subsequent time points, with no evidence for positive selection in the E1/E2 region sequenced. Conclusion. HIV coinfection is associated with increased rates of HCV persistence and a lack of critical CD4T cell responses, with no evidence of immune selection pressure during early HCV infection. Loss of key cellular immune responses against HCV during acute disease may contribute to the failure of early host control of HCV in HCV/HIVcoinfected patients. [ABSTRACT FROM AUTHOR]

CD4+ T-cell responses are important for the outcome of hepatitis C virus (HCV) infection. However, the functional status of HCV-specific CD4+ T cells in persistent infection is poorly understood. It is generally recognized that proliferative capacity of HCV-specific CD4+ T cells is weak or absent in persistent infection, but whether this results from deletion of antigen-specific cells or represents maintenance of antigen-specific but poorly proliferative populations is not defined. We used a set of ex vivo assays to evaluate the functionality of HCV specific CD4+ T cells in persistent and resolved infection. Peripheral blood mononuclear cells (PBMC) from 24 prospectively recruited HCV polymerase chain reaction (PCR) positive individuals, 12 spontaneously resolved individuals (i.e. anti-HCV+, PCR−) and 11 healthy controls were analysed for interferon- γ (IFN- γ) and interleukin 2 (IL-2) secretion by enzyme linked immunospot assays (ELISpot). HCV-specific CD4+ proliferative responses of carboxy fluorescein succinimidyl ester-labelled PBMC were assessed using a sensitive single cell flow cytometric assay. Sustained IFN- γ ELISpot responses were observed in the PCR+ group. However, proliferation of HCV-specific CD4+ T cells in the PCR+ group was substantially reduced on a per cell basis, in parallel to IL-2 secretion, compared with responses in the PCR− group. In PCR− individuals, a strong relationship between cytokine secretion and proliferative capacity was seen. However, in PCR+ individuals, IFN- γ secretion far exceeded proliferative capacity. During persistent HCV infection, some CD4+ T-cell specificities appear to be lost, as measured using a range of techniques, but others, potentially important, are maintained as IFN- γ secretors but with low proliferative capacity even using a highly sensitive assay. Such subsets may yet play a significant role in vivo and also provide a template for modulation in immunotherapeutic interventions. [ABSTRACT FROM AUTHOR]

Background: Human immunodeficiency virus/hepatitis C virus (HIV/HCV) coinfection is a common and complex clinical problem in which loss of immunological control of HCV occurs, with increased HCV viral load and more aggressive liver disease. Cellular immune responses, particularly secretion of interferon γ (IFN-γ) appear to be important in the control of HCV, and a detectable HCV specific CD4 response is associated with clearance of the virus. HCV specific CD8+ T cell responses, weak in chronic HCV infection, have been shown to be further impaired in HIV coinfection and this CD8+ T cell deficiency is related to the decline in CD4 T cell count. Aims: To compare the CD4 T cell response to HCV in HIV/HCV coinfected and HCV monoinfected individuals and to determine the relationship of responses with declining CD4 count. Patients: The study subjects were a cohort of 68 HCV monoinfected and 67 HCV/HIV coinfected haemophiliac children and adolescents (the Hemophilia Growth and Development Study) who were followed for a seven year period. Methods: We analysed IFN-γ secreting CD4+ responses to HCV proteins and peptides and HIV p24 antigen using an ELISpot assay. Results: We found a significant decrease in HCV specific responses among those who were HIV coinfected (10/67 v 36/68; p<0.0001 ) both in numbers of responders and frequency of specific cells. This did not appear to be closely related to CD4 count. Conclusions: The reduction in HCV specific CD4 T cells in coinfection provide a cellular mechanism for the loss of control of HCV in coinfected individuals, even in those with relatively preserved CD4+ T cell counts and CD4+ T cell responses to HIV. [ABSTRACT FROM AUTHOR]

Hepatitis C virus (HCV) readily sets up persistence after acute infection. Cellular immune responses are thought to play a major role in control of the virus. Failure of CD4+ T-cell responses in acute disease is associated with viral persistence but the dynamics of this are poorly understood. We aimed to assess such responses using a novel set of Class II tetrameric complexes (tetramers) to study helper T-cells ex vivo in acute disease. We analysed the HCV-specific CD4+ T-cell response in a patient with acute hepatitis c infection. We were able to track the virus-specific CD4+ T-cells directly ex vivo with HLA DR4 tetramers. Proliferative responses were absent initially, recovered as viral load dropped and were lost again during relapse. Longitudinal tetramer analyses showed expanded populations of antiviral CD4+ T-cells throughout acute infection despite lack of proliferation. A pattern of transient CD4+ T-cell proliferative responses as HCV is partially controlled is observed. Failure to control virus is associated with emergence of 'dysfunctional' CD4+ T-cell populations. Failure to control HCV in acute disease may relate to the capacity to sustain efficient immune responses as virus attempts to 'bounce back' after partial control. [ABSTRACT FROM AUTHOR]

Hepatitis C virus (HCV) is a variable RNA virus that can readily establish persistent infection. Cellular immune responses are important in the early control of the virus. Evidence from animal models suggests that mutation in epitopes recognized by CD8+ T lymphocytes may play an important role in the establishment of persistence but in human persistent infection, equivalent evidence is lacking. We investigated this by analysing a unique resource: viruses from a set of chronically HCV-infected individuals in whom the CD8+ T-cell responses in liver had previously been accurately mapped. Virus was sequenced in seven individuals at 10 epitopes restricted by 10 human leucocyte antigen (HLA) molecules. Two main patterns emerged: in the majority of epitopes sequenced, no variation was seen. In three epitopes, mutations were identified which were compatible with immune escape as assessed using phylogenetic and/or functional studies. These data suggest that – even where specific intrahepatic T cells are detectable – many epitopes do not undergo mutation in chronic human infection. On the contrary, virus may escape from intrahepatic CD8+ T-cell responses in a ‘patchy’ manner in certain specific epitopes. Furthermore, longitudinal studies to identify the differences between ‘selecting’ and ‘nonselecting’ intrahepatic CD8+ T-cell responses are needed in HCV infection. [ABSTRACT FROM AUTHOR]

Cellular immunity plays an important role in the control of persistent virus infections such as hepatitis C virus (HCV). Antiviral CD4+ T cell responses have been shown to accompany resolution of acute disease and there is also a consistent association between HLA Class II genes, notably HLADRB1*1101 (and the closely linked HLADQB1*0301) and disease resolution. We initially mapped longitudinal CD4+ T cell responses in an individual after spontaneous resolution of acute HCV, and identified three HLA-DR11-restricted responses which vary in immunodominance over time. Functional assays and HLA Class II tetramer staining revealed one to be a response to a commonly recognized epitope, NS31248--1261, although cytokine capture assays showed these specific cells to be at a very low frequency. In this patient, and in others reported, this most frequently recognized HLA-DR11 restricted epitope is not immunodominant. We analysed whether sequence variability within and between genotypes might account for differences in recognition of HLA-DR11 restricted epitopes. We found that a limited number, including NS31248--1261, showed extreme sequence conservation. Within NS3, the ability of peptides to accept amino acid substitutions was clearly related to the structure of the protein. Overall the data provide a deeper understanding of the relationship between protein structure and variability of HLA-DR11 restricted peptides and may explain the apparent dominance of responses to NS31248--1261 across studies but not within an individual immune response. [ABSTRACT FROM AUTHOR]

SUMMARY CD4+ T lymphocyte responses are thought to play a major role in control of the hepatitis C virus (HCV). Few, however, have been mapped down to the level of peptide and HLA restriction. Furthermore, the ability of such T cells to respond to viruses which differ in genotype has not been addressed in detail. In most cases of persistent infection with HCV, CD4 proliferative responses are weak or absent. From a large cohort of persistently infected patients, we identified an individual with unusually robust and persistent responses in the face of chronic infection. We firstly mapped two peptide epitopes to regions of the nonstructural protein NS4 (aa1686–1705 and aa 1746–1765). However, in contrast to the genotype 1a derived antigens used for mapping, the infecting virus was identified as genotype 3a. Strikingly, the patient's CD4 response to these epitopes were specific only for the genotype 1a sequence, and did not recognize genotype 3a synthetic peptides. Serologic assays indicated that prior exposure to HCV of genotype 1 had occurred. This patient therefore maintains strong CD4 proliferative responses which are genotype specific and not cross-reactive. The apparent ‘misdirection’ of these nonprotective responses has important implications for the role of natural and vaccine induced CD4 responses in the face of variable viruses. [ABSTRACT FROM AUTHOR]

Human cytomegalovirus (CMV) is a ubiquitous pathogen which sets up a lifelong persistent infection and which can lead to significant disease in the immunosuppressed. The immunological mechanisms controlling CMV in the long term are not defined completely, but CD8+ T lymphocytes are thought to play an important role. Antiviral CD8+ T lymphocytes may exist in very large pools in healthy individuals. Although the detailed composition of these pools is not completely understood, there is known to be heterogeneity, in particular of CD45 isoform expression. We have therefore investigated the CD8+ T-lymphocyte response against CMV directly ex vivo using Class I tetramers combined with stains for a range of phenotypic markers followed by four-colour flow cytometric analysis. In particular, we examined expression of these phenotypic markers in relation to the expression of CD45 isoforms. We found that a spectrum of phenotypes exists stably, from CD45R0high/RAlow through CD45RAhigh/R0low, and that expression of other surface markers such as CD28 and CD62L, and also TCR usage, may vary in parallel with CD45 isoform expression. In some individuals, expansions of antigen-specific CD8+ T lymphocytes bearing specific TCR Vβ chains were restricted to cells of particular CD45 isoforms. Immunity against CMV comprises a large population of CD8+ T lymphocytes with heterogeneous potential, a spectrum in which CD45 isoform expression may play a central role. [ABSTRACT FROM AUTHOR]

Summary This paper examines the numerical and functional consequences of various stimuli on antiviral CD8+ T-cell memory using a mathematical model. The model is based upon biological evidence from the murine model of infection with lymphocytic choriomeningitis virus (LCMV) that the phenotype of immunological memory represents low-level responses driven by various stimuli, and the memory CTL population is partitioned between resting, cycling and effector cells. These subpopulations differ in their lifespan, their potential to mediate antiviral protection and in the stimuli needed for their maintenance. Three types of maintenance stimuli are examined: non-antigen-specific (bystander) stimulation, persisting antigen stimulation and reinfection-mediated stimulation. The modelling predicts that: (i) stable persistence of CTL memory requires the presence of either bystander or antigen-specific stimulation above a certain threshold depending on the sensitivity of memory CTL to stimulation and their life-span; (ii) a relatively low level of stimuli (approximately 104 fold less on a per CTL basis compared to acute infection) is needed to stabilize the expanded memory CTL population; (iii) the presence of CTL subsets in the memory pool of different activation states and lifespans ensures the robustness of memory persistence in the face of temporal variation in the low-level stimuli and; (iv) an ‘optimal’ population structure of the memory CTL pool, in terms of immediate protection, requires the presence of both activated cycling and effector CTL. For this, persisting antigen alone or synergistically with bystander signals provide the appropriate stimulation, so that the stimuli equivalent to approximately 30 p.f.u. of LCMV in the spleen are sufficient to maintain approximately 105–106 specific CTL in the memory pool. These observations are relevant both to our understanding of natural protective immunity... [ABSTRACT FROM AUTHOR]

Background: High expression of CD161 on CD8+ T cells is associated with a population of cells thought to play a role in mucosal immunity. We wished to investigate this subset in an HIV and Mycobacterium tuberculosis (MTB) endemic African setting. Methods: A flow cytometric approach was used to assess the frequency and phenotype of CD161++CD8+ T cells. 80 individuals were recruited for cross-sectional analysis: controls (n = 18), latent MTB infection (LTBI) only (n = 16), pulmonary tuberculosis (TB) only (n = 8), HIV only (n = 13), HIV and LTBI co-infection (n = 15) and HIV and TB co-infection (n = 10). The impact of acute HIV infection was assessed in 5 individuals recruited within 3 weeks of infection. The frequency of CD161++CD8+ T cells was assessed prior to and during antiretroviral therapy (ART) in 14 HIV-positive patients. Results: CD161++CD8+ T cells expressed high levels of the HIV co-receptor CCR5, the tissue-homing marker CCR6, and the Mucosal-Associated Invariant T (MAIT) cell TCR Vα7.2. Acute and chronic HIV were associated with lower frequencies of CD161++CD8+ T cells, which did not correlate with CD4 count or HIV viral load. ART was not associated with an increase in CD161++CD8+ T cell frequency. There was a trend towards lower levels of CD161++CD8+ T cells in HIV-negative individuals with active and latent TB. In those co-infected with HIV and TB, CD161++CD8+ T cells were found at low levels similar to those seen in HIV mono-infection. Conclusions: The frequencies and phenotype of CD161++CD8+ T cells in this South African cohort are comparable to those published in European and US cohorts. Low-levels of this population were associated with acute and chronic HIV infection. Lower levels of the tissue-trophic CD161++ CD8+ T cell population may contribute to weakened mucosal immune defense, making HIV-infected subjects more susceptible to pulmonary and gastrointestinal infections and detrimentally impacting on host defense against TB.

Mucosal-associated invariant T (MAIT) cells are an innate-like T-cell population restricted by the non-polymorphic, major histocompatibility complex class I-related protein 1, MR1. MAIT cells are activated by a broad range of bacteria through detection of riboflavin metabolites bound by MR1, but their direct cytolytic capacity upon recognition of cognate target cells remains unclear. We show that resting human MAIT cells are uniquely characterized by a lack of granzyme (Gr) B and low perforin expression, key granule proteins required for efficient cytotoxic activity, but high levels of expression of GrA and GrK. Bacterial activation of MAIT cells rapidly induced GrB and perforin, licensing these cells to kill their cognate target cells. Using a novel flow cytometry-based killing assay, we show that licensed MAIT cells, but not ex vivo MAIT cells from the same donors, can efficiently kill Escherichia coli-exposed B-cell lines in an MR1- and degranulation-dependent manner. Finally, we show that MAIT cells are highly proliferative in response to antigenic and cytokine stimulation, maintaining high expression of GrB, perforin, and GrA, but reduced expression of GrK following antigenic proliferation. The tightly regulated cytolytic capacity of MAIT cells may have an important role in the control of intracellular bacterial infections, such as Mycobacterium tuberculosis. [ABSTRACT FROM AUTHOR]

We have recently shown that latent murine cytomegalovirus (MCMV) can influence murine transplant allograft acceptance. During these studies we became aware that vivarium-housed control mice can acquire occult MCMV infection. The purpose of this investigation was to confirm occult MCMV transmission and determine the timing, vehicle, and possible consequences of transmission. Mice arriving from a commercial vendor were negative for MCMV both by commercial serologic testing and by our nested PCR. Mice housed in our vivarium became positive for MCMV DNA 30-60 days after arrival, but remained negative for MCMV by commercial serologic testing. To confirm MCMV we sequenced PCR products for several genes and showed >99% homology to MCMV. Further sequence analyses show that the occult MCMV is similar to a laboratory strain of MCMV, but the vehicle of transmission remains unclear. Control tissues from historical experiments with unexplained graft losses were evaluated for occult MCMV, and mice with unexplained allograft losses showed significantly higher incidence of occult MCMV than did allograft acceptors. Deliberate infection with very low titer MCMV confirmed that viral transmission can occur without measurable virus specific antibody or T-cell responses. These data suggest that vivarium-housed mice can develop occult MCMV that is missed by currently available commercial serologic testing, and that these infections may influence transplant allograft acceptance. Copyright (c) 2010 Elsevier B.V. All rights reserved. [ABSTRACT FROM AUTHOR]

Abstract: There are few studies that have examined the frequencies of epitope-specific CD4+ T cells following the use of a highly effective vaccine, yet such data would potentially be of value for the development of novel vaccination strategies. In this study we tracked human epitope-specific CD4+ T cell responses over time after immunisation with a live attenuated varicella zoster virus vaccine by MHC Class II tetrameric complexes and functional assays. We show that the peptide-specific responses reflect those against whole virus antigens, and are similar in both frequency and phenotype to those found in healthy volunteers, despite a highly attenuated and clinically inapparent infection. [Copyright &y& Elsevier]

Six of seven HLA-A*2402-positive individuals with acute parvovirus B19 infections made vigorous CD8-positive cytotoxic T-cell (CTL) responses to the viral epitope FYTPLADQF. All responders showed highly focused T-cell receptor (TCR) usage, using almost exclusively BV5.1. The BV5.1 TCR dominated the acute response, was maintained over time, and was also used by a remotely infected individual. Nine CTL clones and two oligoclonal lines obtained from three unrelated individuals used BV5.1, BJ2.1, and a conserved TCR CDR3 of nine amino acids. This commonly recognized epitope is likely important in long-term protective immunity and should be included in vaccine design. [ABSTRACT FROM AUTHOR]