With the emergence of highly transmissible variants of concern, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) still poses a global threat of coronavirus disease 2019 (COVID-19) resurgence. Cellular responses to novel variants are more robustly maintained than humoral responses, and therefore, cellular responses are of interest in assessing immune protection against severe disease in the population. We aimed to assess cellular responses to SARS-CoV-2 at the population level. IFNγ (interferon γ) responses to wild-type SARS-CoV-2 were analyzed using an ELISpot assay in vaccine-naive individuals with different humoral responses: Ig (IgM and/or IgG) seronegative (n = 90) and seropositive (n = 181) with low (<300 U/mL) or high (≥300 U/mL) humoral responses to the spike receptor binding domain (anti-S-RBD). Among the seropositive participants, 71.3% (129/181) were IFNγ ELISpot positive, compared to 15.6% (14/90) among the seronegative participants. Common COVID-19 symptoms such as fever and ageusia were associated with IFNγ ELISpot positivity in seropositive participants, whereas no participant characteristics were associated with IFNγ ELISpot positivity in seronegative participants. Fever and/or dyspnea and anti-S-RBD levels were associated with higher IFNγ responses. Symptoms of more severe disease and higher anti-S-RBD responses were associated with higher IFNγ responses. A significant proportion (15.6%) of seronegative participants had a positive IFNγ ELISpot. Assessment of cellular responses may improve estimates of the immune response to SARS-CoV-2 in the general population.
OBJECTIVE: Data on adaptive cellular immunity are of interest to define immune protection against severe acute respiratory syndrome coronavirus 2 in a population, which is important for decision-making on booster-vaccination strategies. This study provides data on associations between participant characteristics and cellular immune responses in vaccine-naive individuals with different humoral responses.

The safety and effectiveness of live virus vaccines, such as the varicella-zoster vaccine, are unknown in patients with inflammatory diseases receiving immunomodulatory therapy such as tumor necrosis factor inhibitors (TNFis).
To evaluate the safety and immunogenicity of the live attenuated zoster vaccine (ZVL) in patients receiving TNFis.
Randomized, blinded, placebo-controlled trial. (ClinicalTrials.gov: NCT02538341).
Academic and community-based rheumatology, gastroenterology, and dermatology practices.
Adults aged 50 years or older receiving TNFis for any indication.
Random assignment to ZVL versus placebo.
Glycoprotein enzyme-linked immunosorbent assay (gpELISA) and enzyme-linked immunosorbent spot (ELISpot) from serum and peripheral blood mononuclear cells measured at baseline and 6 weeks after vaccination. Suspected varicella infection or herpes zoster was clinically assessed using digital photographs and polymerase chain reaction on vesicular fluid.
Between March 2015 and December 2018, 617 participants were randomly assigned in a 1:1 ratio to receive ZVL (n = 310) or placebo (n = 307) at 33 centers. Mean age was 62.7 years (SD, 7.5); 66.1% of participants were female, 90% were White, 8.2% were Black, and 5.9% were Hispanic. The most common TNFi indications were rheumatoid arthritis (57.6%) and psoriatic arthritis (24.1%); TNFi medications were adalimumab (32.7%), infliximab (31.3%), etanercept (21.2%), golimumab (9.1%), and certolizumab (5.7%). Concomitant therapies included methotrexate (48.0%) and oral glucocorticoids (10.5%). Through week 6, no cases of confirmed varicella infection were found; cumulative incidence of varicella infection or shingles was 0.0% (95% CI, 0.0% to 1.2%). At 6 weeks, compared with baseline, the mean increases in geometric mean fold rise as measured by gpELISA and ELISpot were 1.33 percentage points (CI, 1.17 to 1.51 percentage points) and 1.39 percentage points (CI, 1.07 to 1.82 percentage points), respectively.
Potentially limited generalizability to patients receiving other types of immunomodulators.
This trial informs safety concerns related to use of live virus vaccines in patients receiving biologics.
The National Institute of Arthritis and Musculoskeletal and Skin Diseases and the American College of Rheumatology.

The diagnosis of tuberculous pericarditis (TBP) remains challenging. This prospective study evaluated the diagnostic value of Xpert MTB/RIF (Xpert) and T-SPOT.TB and adenosine deaminase (ADA) for TBP in a high burden setting. A total of 123 HIV-negative patients with suspected TBP were enrolled at a tertiary referral hospital in China. Pericardial fluids were collected and subjected to the three rapid tests, and the results were compared with the final confirmed diagnosis. Of 105 patients in the final analysis, 39 (37.1%) were microbiologically, histopathologically or clinically diagnosed with TBP. The sensitivity, specificity, positive predictive value, negative predictive value, positive likelihood ratio, negative likelihood ratio and diagnostic odds ratio (DOR) for Xpert were 66.7%, 98.5%, 96.3%, 83.3%, 44.0, 0.338, and 130.0, respectively, compared to 92.3%, 87.9%, 81.8%, 95.1%, 7.6, 0.088, and 87.0, respectively, for T-SPOT.TB, and 82.1%, 92.4%, 86.5%, 89.7%, 10.8, 0.194, and 55.8, respectively, for ADA (≥ 40 U/L). ROC curve analysis revealed a cut-off point of 48.5 spot-forming cells per million pericardial effusion mononuclear cells for T-SPOT.TB, which had a DOR value of 183.8, while a cut-off point of 41.5 U/L for ADA had a DOR value of 70.9. Xpert (Step 1: rule-in) followed by T-SPOT.TB [cut-off point] (Step 2: rule-out) showed the highest DOR value of 252.0, with only 5.7% (6/105) of patients misdiagnosed. The two-step algorithm consisting of Xpert and T-SPOT.TB could offer rapid and accurate diagnosis of TBP.

Prostate cancer (PCa) has been under investigation as a target for antigen-specific immunotherapies in metastatic disease settings for the last two decades leading to a licensure of the first therapeutic cancer vaccine, Sipuleucel-T, in 2010. However, neither Sipuleucel-T nor other experimental PCa vaccines that emerged later induce strong T-cell immunity.
In this first-in-man study, VANCE, we evaluated a novel vaccination platform based on two replication-deficient viruses, chimpanzee adenovirus (ChAd) and MVA (Modified Vaccinia Ankara), targeting the oncofetal self-antigen 5T4 in early stage PCa. Forty patients, either newly diagnosed with early-stage PCa and scheduled for radical prostatectomy or patients with stable disease on an active surveillance protocol, were recruited to the study to assess the vaccine safety and T-cell immunogenicity. Secondary and exploratory endpoints included immune infiltration into the prostate, prostate-specific antigen (PSA) change, and assessment of phenotype and functionality of antigen-specific T cells.
The vaccine had an excellent safety profile. Vaccination-induced 5T4-specific T-cell responses were measured in blood by ex vivo IFN-γ ELISpot and were detected in the majority of patients with a mean level in responders of 198 spot-forming cells per million peripheral blood mononuclear cells. Flow cytometry analysis demonstrated the presence of both CD8+ and CD4+ polyfunctional 5T4-specific T cells in the circulation. 5T4-reactive tumor-infiltrating lymphocytes were isolated from post-treatment prostate tissue. Some of the patients had a transient PSA rise 2-8 weeks following vaccination, possibly indicating an inflammatory response in the target organ.
An excellent safety profile and T-cell responses elicited in the circulation and also detected in the prostate gland support the evaluation of the ChAdOx1-MVA 5T4 vaccine in efficacy trials. It remains to be seen if this vaccination strategy generates immune responses of sufficient magnitude to mediate clinical efficacy and whether it can be effective in late-stage PCa settings, as a monotherapy in advanced disease or as part of multi-modality PCa therapy. To address these questions, the phase I/II trial, ADVANCE, is currently recruiting patients with intermediate-risk PCa, and patients with advanced metastatic castration-resistant PCa, to receive this vaccine in combination with nivolumab.
The trial was registered with the U.S. National Institutes of Health (NIH) Clinical Trials Registry (ClinicalTrials.gov identifier NCT02390063).

BACKGROUND: The prevalence of cytomegalovirus in China is high and the infection could result in disastrous consequences. Cellular immunity is the main mechanism for virus controlling. We explored the performance of cytomegalovirus antigen-specific enzyme-linked immunospot assay in patients with different infection states in endemic area.
METHODS: Twenty-eight patients with reliable results were included in statistical analysis. Peripheral blood mononuclear cells were extracted and were stimulated by cytomegalovirus phosphoprotein 65 or immediate early protein-1 antigen respectively. Spot forming cells (SFCs) were counted to evaluate the cellular immune response elicited by antigens.
RESULTS: Spots could be clearly displayed, and evenly dispersed with a clean background. The numbers of SFCs were 0 [0-0], 426 [210-601] and 230 [57-513] for uninfected individuals (n = 7), latently infected patients (n = 11) and actively infected patients (n = 10) respectively, which were statistically different. The number of SFCs stimulated by phosphoprotein 65 was significantly higher than that by immediate early protein-1.
CONCLUSIONS: The number of SFCs was significantly different among patients with different infection state. The stimulatory effect of phosphoprotein 65 was better than that of immediate early protein-1.

A conditionally replication-defective human cytomegalovirus (CMV) vaccine (V160) derived from AD169 and genetically engineered to express CMV pentameric complex (gH/gL/pUL128/pUL130/pUL131) was developed and evaluated for phase 1 vaccine safety and immunogenicity in CMV-seronegative and CMV-seropositive adults.
Subjects received 3 doses of V160 or placebo on day 1, month 1, and month 6. Four vaccine dose levels, formulated with or without aluminum phosphate adjuvant, were evaluated. Injection-site and systemic adverse events (AEs) and vaccine viral shedding were monitored. CMV-specific cellular and humoral responses were measured by interferon-gamma ELISPOT and virus neutralization assay up to 12 months after last dose.
V160 was generally well-tolerated, with no serious AEs observed. Transient, mild-to-moderate injection-site and systemic AEs were reported more frequently in vaccinated subjects than placebo. Vaccine viral shedding was not detected in any subject, confirming the nonreplicating feature of V160. Robust neutralizing antibody titers were elicited and maintained through 12 months postvaccination. Cellular responses to structural and nonstructural viral proteins were observed, indicating de novo expression of viral genes postvaccination.
V160 displayed an acceptable safety profile. Levels of neutralizing antibodies and T-cell responses in CMV-seronegative subjects were within ranges observed following natural CMV infection.
. NCT01986010.

Antibodies against foreign antigens are a critical component of the overall immune response and can facilitate pathogen clearance during a primary infection and also protect against subsequent infections. Dysregulation of the antibody response can lead to an autoimmune disease, malignancy, or enhanced infection. Since the experimental delineation of a distinct B cell lineage in 1965, various methods have been developed to understand antigen-specific B cell responses in the context of autoimmune diseases, primary immunodeficiencies, infection, and vaccination. In this review, we summarize the established techniques and discuss new and emerging technologies for probing the B cell response in vitro and in vivo by taking advantage of the specificity of B cell receptor (BCR)-associated and secreted antibodies. These include ELISPOT, flow cytometry, mass cytometry, and fluorescence microscopy to identify and/or isolate primary antigen-specific B cells. We also present our approach to identify rare antigen-specific B cells using magnetic enrichment followed by flow cytometry. Once these cells are isolated, in vitro proliferation assays and adoptive transfer experiments in mice can be used to further characterize antigen-specific B cell activation, function, and fate. Transgenic mouse models of B cells targeting model antigens and of B cell signaling have also significantly advanced our understanding of antigen-specific B cell responses in vivo.

Lack of virus-specific cell-mediated immunity (CMI) is associated with worse viral infection outcome in hematopoietic stem cell transplantation (HSCT). We aimed to evaluate the role of immunological monitoring of Epstein-Barr virus (EBV) infection in addition to virological one in 33 adult and 18 pediatric allogeneic HSCT recipients. Virological monitoring of infection was performed on whole blood samples by a quantitative real-time PCR assay. Immunological monitoring was performed by Enzyme-linked ImmunoSPOT assay, evaluating EBV-specific CMI, at fixed time-points and when EBV DNAemia was ≥ 10,000 copies/mL. Fifty-one percent of patients developed a post-transplant EBV infection and reduced-intensity conditioning regimen was the only factor associated to infection (P = 0.023). Lack of EBV-specific CMI during active EBV infection was associated with a greater severity of infection. Patients without EBV-specific CMI showed higher median peak level of EBV DNAemia than patients with EBV-specific CMI (P = 0.014), and consequently received more frequently, at EBV DNAemia peak, anti-CD20 therapy (0 versus 54.5%, P = 0.002). No patients with EBV-specific CMI versus 27.2% without EBV-specific CMI developed EBV-related complications (P = 0.063), including two lethal EBV-related post-transplant lymphoproliferative disorders. Combined immunological and virological measurements could improve EBV infection management in HSCT, anticipating the beginning of preemptive treatment from the EBV DNAemia peak to the finding of the lack of EBV-specific CMI.

BACKGROUND: Zoster vaccination is recommended for people with a history of herpes zoster (HZ), but the most effective timing of vaccine administration after zoster illness is unresolved. This prospective observational study compared the immunogenicity and safety of administering HZ vaccine at 6-12 months and 1-5 years after zoster illness.
METHODS: Blood samples were collected before the administration of live zoster vaccine and 6 weeks after vaccination. Varicella-zoster virus (VZV) IgG concentrations and T-cell responses were assessed by glycoprotein enzyme-linked immunosorbent assay and interferon-γ enzyme-linked immunospot assay (ELISPOT), respectively.
RESULTS: The baseline geometric mean value (GMV) of VZV IgG was higher in the 6-12 months group than in the 1-5 years group (245.5 IU/mL vs. 125.9 IU/mL; P = 0.021). However, the GMV increased significantly in both groups (P = 0.002 in the 6-12 months group; P <0.001 in the 1-5 years group). The results of the ELISPOT assay were not significant for differences of the GMV between baseline and 6-week post-vaccination groups, while the GMV increased significantly in both groups (P = 0.001 in the 6-12 months group; P <0.001 in the 1-5 years group).
CONCLUSIONS: The immunogenicity of zoster vaccine may be similar whether administered 6-12 months, or >1 year after zoster illness.
BACKGROUND: ClinicalTrials.gov Identifier: NCT02704572.

To determine whether immunogenicity of the herpes zoster live-attenuated vaccine (ZVL) in elderly individuals is influenced by frailty status and cytokine levels, individuals aged at least 65 years were prospectively administered ZVL after their frailty status and cytokine levels were assessed. Humoral and cellular immunogenicity were assessed using enzyme-linked immunosorbent assays for the varicella-zoster virus-specific IgG antibody and varicella-zoster virus-specific enzyme-linked immunospot assays, before and 6 weeks after vaccination. We enrolled 69 participants (mean age, 74.3 years); 31 were non-robust. Thirty-seven participants had a greater than 2 geometric mean fold rise in IgG antibodies, 22 more participants had at least 10 spot-forming cells/106 peripheral blood mononuclear cells (from 35 to 57 participants), and 42 experienced a twofold geometric mean fold rise increase of spot-forming cells 6 weeks after vaccination. The geometric mean fold rises of the antibody titers were similar in the robust and non-robust groups; moreover, the baseline enzyme-linked immunospot assay geometric mean values were not significantly different before or 6 weeks after vaccination. We observed no significant correlations between baseline cytokine levels and immunogenicity. Approximately half of the community-dwelling elderly participants in this study experienced a twofold rise in antibodies and one-third more participants showed at least 10 spot-forming cells after ZVL administration, suggesting that the ZVL can boost immunity in non-robust (especially pre-frail) and robust elderly individuals.