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The emergence of SARS-CoV-2 in 2019 led to a global pandemic with a significant impact on healthcare systems. Healthcare workers were particularly vulnerable due to frequent contact with COVID-19 patients. Despite vaccination, they remained at higher risk as the vaccines provided limited protection against infection with viral variants, like Delta or Omicron BA.1 and BA.5. Three years after the onset of the pandemic, we evaluated SARS-CoV-2 infection frequencies among healthcare workers with varying levels of patient contact: high-risk (frequent COVID-19 patient contact), intermediate-risk (non-COVID-19 patient contact), and low-risk (no patient contact). We assessed their cellular and humoral immune responses based on their vaccination status and number of prior infections. SARS-CoV-2-specific antibodies were measured by immunoglobulin ELISA, and neutralizing antibody titers were determined against the viral variants D614G, Delta, and Omicron BA.1 and BA.5. Cellular immune responses were analyzed using an interferon-γ ELISpot. Notably, three years into the pandemic, healthcare workers in daily contact with COVID-19 patients did not have higher infection rates compared to healthcare workers with non-COVID-19 patient contact or no patient contact. Immune responses were similar across all groups, highlighting the effectiveness of vaccination and current hygiene standards in preventing virus transmission from patients to staff.

Abstract Tools for predicting COVID-19 outcomes enable personalized healthcare, potentially easing the disease burden. This collaborative study by 15 institutions across Europe aimed to develop a machine learning model for predicting the risk of in-hospital mortality post-SARS-CoV-2 infection. Blood samples and clinical data from 1286 COVID-19 patients collected from 2020 to 2023 across four cohorts in Europe and Canada were analyzed, with 2906 long non-coding RNAs profiled using targeted sequencing. From a discovery cohort combining three European cohorts and 804 patients, age and the long non-coding RNA LEF1-AS1 were identified as predictive features, yielding an AUC of 0.83 (95% CI 0.82–0.84) and a balanced accuracy of 0.78 (95% CI 0.77–0.79) with a feedforward neural network classifier. Validation in an independent Canadian cohort of 482 patients showed consistent performance. Cox regression analysis indicated that higher levels of LEF1-AS1 correlated with reduced mortality risk (age-adjusted hazard ratio 0.54, 95% CI 0.40–0.74). Quantitative PCR validated LEF1-AS1’s adaptability to be measured in hospital settings. Here, we demonstrate a promising predictive model for enhancing COVID-19 patient management.

ABSTRACT A crucial step in lowering the risk of invasive pneumococcal illness in high-risk populations, such as individuals with plaque psoriasis, is pneumococcal vaccination. The serologic response to the sequential vaccination with Prevenar 13 (PCV13) and Pneumovax 23 (PPSV23) in psoriasis patients under immunosuppressive therapy is still poorly characterized despite national recommendations suggesting vaccination for immunocompromised patients. In this prospective study, we investigated the serological response in 57 patients under active systemic treatment for moderate to severe plaque psoriasis who underwent sequential vaccination with PCV13 followed by PPSV23. Our analysis focused on global and serotype-specific anti-pneumococcal antibody responses over a 7-month period post-vaccination. Our findings reveal a robust serological response in patients with plaque psoriasis under systemic therapy. When comparing our results with a cohort of kidney transplant recipients who completed a similar sequential vaccination protocol, psoriasis patients showed higher antibody concentrations. In psoriasis patients, the mean levels of all global antibody classes tested (IgG, IgG2, IgA, IgM) increased more than 4-fold (P < 0.0001) and serotype-specific antibodies more than 1.9-fold (P < 0.01). In addition to providing strong evidence of the safety and effectiveness of sequential pneumococcal vaccination in individuals with plaque psoriasis, our work sheds light on the complex interactions that exist between immunosuppressive treatment, vaccination schedule, and antibody responses in various risk groups.IMPORTANCETo protect against severe courses of infection with Streptococcus pneumoniae, the national guidelines recommend sequential vaccination for these patients. However, there are only studies on the efficacy of a single administration of these vaccines in this particular risk group. The immunological responses to the vaccine were correlated with clinical patient data. In summary, our study shows for the first time that sequential vaccination is immunogenic in patients with moderate to severe plaque psoriasis.

Background: Vaccination against Streptococcus pneumoniae is advised for transplant recipients to reduce morbidity and mortality associated with invasive pneumococcal disease. However, data on alloantibodies after sequential vaccination (with a pneumococcal conjugate vaccine followed by a polysaccharide vaccine) are still lacking. Methods: In the current study, we determined HLA class I and II and major histocompatibility class I-related chain A (MICA) antibodies in 41 clinically stable kidney transplant recipients. These antibodies were measured prior to and post sequential pneumococcal vaccination over a period of 12 months. Alloantibodies were measured by Luminex bead-based assays, and pneumococcal IgG antibodies were measured by ELISA. Results: Over a 12-month period, the sequential analysis revealed no significant change in alloantibodies. One patient developed de novo donor-specific antibodies (DSA) 1.5 months after the first vaccination, with mean fluorescence intensities of up to 2300. These DSA became undetectable in the follow-up, and the patient showed no signs of allograft rejection. Another patient experienced a biopsy-proven borderline rejection 7 months after the first vaccination but did not develop de novo DSA. Both maintained stable kidney function. As expected, the pneumococcal antibodies increased significantly after vaccination (p < 0.0001). Conclusions: Given the overall risk of alloimmune responses in transplant recipients, we would not attribute the two noticeable patient courses to vaccination. Thus, we consider sequential vaccination immunologically safe.

Abstract Cross-reactive cellular and humoral immunity can substantially contribute to antiviral defense against SARS-CoV-2 variants of concern (VOC). While the adult SARS-CoV-2 cellular and humoral immunity and its cross-recognition potential against VOC is broadly analyzed, similar data regarding the pediatric population are missing. In this study, we perform an analysis of the humoral and cellular SARS-CoV-2 response immune of 32 convalescent COVID-19 children (children), 27 convalescent vaccinated adults(C + V+) and 7 unvaccinated convalescent adults (C + V-). Similarly to adults, a significant reduction of cross-reactive neutralizing capacity against delta and omicron VOC was observed 6 months after SARS-CoV-2 infection. While SAR-CoV-2 neutralizing capacity was comparable among children and C + V- against all VOC, children demonstrated as expected an inferior humoral response when compared to C + V+. Nevertheless, children generated SARS-CoV-2 reactive T cells with broad cross-recognition potential. When compared to V + C+, children presented even comparable frequencies of WT-reactive CD4 + and CD8 + T cells with high avidity and functionality. Taking into consideration the limitations of study - unknown disease onset for 53% of the asymptomatic pediatric subjects, serological detection of SARS-CoV-2 infection-, our results suggest that following SARS-CoV-2 infection children generate a humoral SARS-CoV-2 response with neutralizing potential comparable to unvaccinated COVID-19 convalescent adults as well a sustained SARS-CoV-2 cellular response cross-reactive to VOC.

While SARS-CoV-2 has transitioned to an endemic phase, infections caused by newly emerged variants continue to result in severe, and sometimes fatal, outcomes or lead to long-term COVID-19 symptoms. Vulnerable populations, such as PLWH, face an elevated risk of severe illness. Emerging variants of SARS-CoV-2, including numerous Omicron subvariants, are increasingly associated with breakthrough infections. Adapting mRNA vaccines to these new variants may offer improved protection against Omicron for vulnerable individuals. In this study, we examined humoral and cellular immune responses before and after administering adapted booster vaccinations to PLWH, alongside a control group of healthy individuals. Four weeks following booster vaccination, both groups exhibited a significant increase in neutralizing antibodies and cellular immune responses. Notably, there was no significant difference in humoral immune response between PLWH and the healthy controls. Immune responses declined rapidly in both groups three months post vaccination. However, PLWH still showed significantly increased neutralizing antibody titers even after three months. These findings demonstrate the efficacy of the adapted vaccination regimen. The results suggest that regular booster immunizations may be necessary to sustain protective immunity.

Summary: Background: Optimal initial tacrolimus dosing and early exposure of tacrolimus after renal transplantation is not well studied. Methods: In this open-label, 6 months, multicenter, randomized controlled, non-inferiority study, we randomly assigned 432 renal allograft recipients to receive basiliximab induction, mycophenolate and steroids and either standard prolonged-release tacrolimus (trough levels: 7–9 ng/ml; Standard Care arm), or an initial 7-day fixed 5 mg/day dose of prolonged-release tacrolimus followed by lower tacrolimus predose levels (trough levels: 5–7 ng/ml; Slow & Low arm). The primary end point was the combined incidence rate of biopsy-proven acute rejections (BPAR; including borderline), graft failure, or death at 6 months with a non-inferiority margin of 12.5%. (EudraCT-Nr: 2013-001770-19. Findings: The combined primary endpoint in the Slow & Low arm was non-inferior compared to the Standard Care arm (22.1% versus 20.7%; difference: 1.4%, 90% CI −5.5% to 8.3%). The overall rate of BPAR including borderlines was similar (Slow & Low 17.4% versus Standard Care 16.6%). Safety parameters such as delayed graft function, kidney function, donor specific HLA-antibodies, infections, or post-transplantation diabetes mellitus did not differ. Interpretation: This is the first study to show that an initial fixed dose of 5 mg per day followed by lower tacrolimus exposure is non-inferior compared to standard tacrolimus therapy and equally efficient and safe within 6 months after renal transplantation. These data suggest that therapeutic drug monitoring for prolonged release tacrolimus can be abandoned until start of the second week after transplantation. Funding: Investigator-initiated trial, financial support by Astellas Pharma GmbH.

Abstract It has recently been shown that von Willebrand factor (VWF) multimers contribute to immunothrombosis in Coronavirus disease 2019 (COVID-19). Since COVID-19 is associated with an increased risk of autoreactivity, the present study investigates, whether the generation of autoantibodies to ADAMTS13 contributes to this finding. In this observational prospective controlled multicenter study blood samples and clinical data of patients hospitalized for COVID-19 were collected from April to November 2020. The study included 156 individuals with 90 patients having confirmed COVID-19 of mild to critical severity. 30 healthy individuals and 36 critically ill ICU patients without COVID-19 served as controls. ADAMTS13 antibodies occurred in 31 (34.4%) COVID-19 patients. Antibodies occurred more often in critically ill COVID-19 patients (55.9%) than non-COVID-19 ICU patients and healthy controls (5.6% and 6.7%; p < 0.001), respectively. Generation of ADAMTS13 antibodies in COVID-19 was associated with lower ADAMTS13 activity (56.5%, interquartile range (IQR) 21.25 vs. 71.5%, IQR 24.25, p = 0.0041), increased disease severity (severe or critical in 90% vs. 62.3%, p = 0.019), and a trend to higher mortality (35.5% vs. 18.6%, p = 0.077). Median time to antibody development was 11 days after first positive SARS-CoV-2-PCR specimen. Gel analysis of VWF multimers resembled the constellation in patients with TTP. The present study demonstrates for the first time, that generation of ADAMTS13 antibodies is frequent in COVID-19, associated with lower ADAMTS13 activity and increased risk of an adverse disease course. These findings provide a rationale to include ADAMTS13 antibodies in the diagnostic workup of SARS-CoV-2 infections.

ObjectivesFarnesyltransferase inhibitors (FTI), which inhibit the prenylation of Ras GTPases, were developed as anti-cancer drugs. As additional target proteins for prenylation were identified in the past, it is likely that FTI have potential value for therapeutic purposes beyond cancer. The effect of FTI on B-cells remains unclear. To address this issue, we investigated the effects of in vitro FTI treatment on effector and regulatory B-cells in healthy controls and renal transplant patients.MethodsFor this purpose, B-cells were isolated from the peripheral blood of healthy controls and renal transplant patients. Purified B-cells were stimulated via Toll-like-receptor 9 (TLR-9) in the presence or absence of FTI. Regulatory functions, such as IL-10 and Granzyme B (GrB) secretion, were assessed by flow cytometry. In addition, effector B-cell functions, such as plasma cell formation and IgG secretion, were studied.ResultsThe two FTI Lonafarnib and tipifarnib both suppressed TLR-9-induced B-cell proliferation. Maturation of IL-10 producing B-cells was suppressed by FTI at high concentrations as well as induction of GrB-secreting B-cells. Plasma blast formation and IgG secretion were potently suppressed by FTI. Moreover, purified B-cells from immunosuppressed renal transplant patients were also susceptible to FTI-induced suppression of effector functions, evidenced by diminished IgG secretion.ConclusionFTI suppress in vitro B-cell proliferation and plasma cell formation while partially preserving IL-10 as well as GrB production of B-cells. Thus, FTI may have immunosuppressive capacity encouraging further studies to investigate the potential immunomodulatory value of this agent.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused millions of COVID-19 cases and deaths worldwide. Severity of pulmonary pathologies and poor prognosis were reported to be associated with the activation non-virus-specific bystander T cells. In addition, high concentrations of the macrophage migration inhibitory factor (MIF) were found in serum of COVID-19 patients. We hypothesized that these two pathogenic factors might be related and analyzed the expression of receptors for MIF on T cells in COVID-19. T cells from PBMCs of hospitalized patients with mild and severe COVID-19 were characterized. A significantly higher proportion of CD4+ and CD8+ T cells from COVID-19 patients expressed CD74 on the cell surface compared to healthy controls. To induce intracellular signaling upon MIF binding, CD74 forms complexes with CD44, CXCR2, or CXCR4. The vast majority of CD74+ T cells expressed CD44, whereas expression of CXCR2 and CXCR4 was low in controls but increased upon SARS-CoV-2 infection. Hence, T cells in COVID-19 patients express receptors that render them responsive to MIF. A detailed analysis of CD74+ T cell populations revealed that most of them had a central memory phenotype early in infection, while cells with an effector and effector memory phenotype arose later during infection. Furthermore, CD74+ T cells produced more cytotoxic molecules and proliferation markers. Our data provide new insights into the MIF receptor and co-receptor repertoire of bystander T cells in COVID-19 and uncovers a novel and potentially druggable aspect of the immunological footprint of SARS-CoV-2.

Abstract Introduction Immunosuppressive therapy is associated with an increased risk of severe courses of SARS-CoV-2 infection, with frequently delayed viral clearance. We report a case of an acute kidney transplant failure in persistent SARS-CoV-2 infection in a patient with absolute B-cell depletion after administration of rituximab for AB0-incompatible living donor kidney transplantation. Case presentation A 34-year-old unvaccinated patient is diagnosed with SARS-CoV-2 infection four months after kidney transplantation. With only mild symptoms and an estimated glomerular filtration rate (eGFR) of 44 ml/min/1.73 m2, therapy with molnupiravir was initially given. Within the next eight weeks, transplant biopsies were performed for acute graft failure. These showed acute T-cell rejection with severe acute tubular epithelial damage with only mild interstitial fibrosis and tubular atrophy (BANFF cat. 4 IB), and borderline rejection (BANFF cat. 3). A therapy with prednisolone and intravenous immunoglobulins was performed twice. With unchanged graft failure, the third biopsy also formally showed BANFF cat. 4 IB. However, fluorescence in situ hybridization detected SARS-CoV-2 viruses in large portions of the distal tubules. After nine weeks of persistent COVID-19 disease neither anti-SARS-CoV-2 IgG nor a SARS-CoV-2-specific cellular immune response could be detected, leading to the administration of sotrovimab and remdesivir. Among them, SARS-CoV-2 clearance, detection of IgG, and improvement of graft function were achieved. Conclusion Lack of viral clearance can lead to complications of SARS-CoV-2 infection with atypical manifestations. In kidney transplant patients, before initiating therapy, the differential diagnoses of “rejection” and “virus infection” should be weighed against each other in an interdisciplinary team of nephrologists, infectious diseases specialists and pathologists.

BackgroundBreakthrough infections with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants are increasingly observed in vaccinated individuals. Immune responses towards SARS-CoV-2 variants, particularly Omicron-BA.5, are poorly understood. We investigated the humoral and cellular immune responses of hospitalized COVID-19 patients during Delta and Omicron infection waves.MethodsThe corresponding SARS-CoV-2 variant of the respective patients were identified by whole genome sequencing. Humoral immune responses were analyzed by ELISA and a cell culture-based neutralization assay against SARS-CoV-2 D614G isolate (wildtype), Alpha, Delta (AY.43) and Omicron (BA.1 and BA.5). Cellular immunity was evaluated with an IFN-γ ELISpot assay.ResultsOn a cellular level, patients showed a minor IFN-γ response after stimulating PBMCs with mutated regions of SARS-CoV-2 variants. Neutralizing antibody titers against Omicron-BA.1 and especially BA.5 were strongly reduced. Double-vaccinated patients with Delta breakthrough infection showed a significantly increased neutralizing antibody response against Delta compared to double-vaccinated uninfected controls (median complete neutralization titer (NT100) 640 versus 80, p

The pandemic caused by SARS-CoV-2 is still a major health problem. Newly emerging variants and long-COVID-19 represent a challenge for the global health system. In particular, individuals in developing countries with insufficient health care need easily accessible, affordable and effective treatments of COVID-19. Previous studies have demonstrated the efficacy of functional inhibitors of acid sphingomyelinase against infections with various viruses, including early variants of SARS-CoV-2. This work investigated whether the acid sphingomyelinase inhibitors fluoxetine and sertraline, usually used as antidepressant molecules in clinical practice, can inhibit the replication of the former and recently emerged SARS-CoV-2 variants in vitro. Fluoxetine and sertraline potently inhibited the infection with pseudotyped virus-like particles and SARS-CoV-2 variants D614G, alpha, delta, omicron BA.1 and omicron BA.5. These results highlight fluoxetine and sertraline as priority candidates for large-scale phase 3 clinical trials at different stages of SARS-CoV-2 infections, either alone or in combination with other medications.

The role of adaptive SARS-CoV-2 specific immunity in post-acute sequelae of COVID-19 (PASC) is not well explored, although a growing population of convalescent COVID-19 patients with manifestation of PASC is observed. We analyzed the SARS-CoV-2-specific immune response, via pseudovirus neutralizing assay and multiparametric flow cytometry in 40 post-acute sequelae of COVID-19 patients with non-specific PASC manifestation and 15 COVID-19 convalescent healthy donors. Although frequencies of SARS-CoV-2-reactive CD4+ T cells were similar between the studied cohorts, a stronger SARS-CoV-2 reactive CD8+ T cell response, characterized by IFNγ production and predominant TEMRA phenotype but low functional TCR avidity was detected in PASC patients compared to controls. Of interest, high avidity SARS-CoV-2-reactive CD4+ and CD8+ T cells were comparable between the groups demonstrating sufficient cellular antiviral response in PASC. In line with the cellular immunity, neutralizing capacity in PASC patients was not inferior compared to controls. In conclusion, our data suggest that PASC may be driven by an inflammatory response triggered by an expanded population of low avidity SARS-CoV-2 reactive pro-inflammatory CD8+ T cells. These pro-inflammatory T cells with TEMRA phenotype are known to be activated by a low or even without TCR stimulation and lead to a tissue damage. Further studies including animal models are required for a better understanding of underlying immunopathogensis. Summary: A CD8+ driven persistent inflammatory response triggered by SARS-CoV-2 may be responsible for the observed sequelae in PASC patients.

Background: The COVID-19 pandemic led to an alteration of algorithms in emergency medicine, which may influence the management of patients with similar symptoms but underlying cardiovascular diseases. We evaluated key differential diagnoses to acute COVID-19 infection and the prevalence and the prognosis of myocardial injury in patients presenting for suspected COIVD-19 infection. Methods: This prospective observational study includes patients presenting with symptoms suggestive of COVID-19 infection during the pandemic. In patients without COVID-19, leading diagnoses was classified according to ICD-10. Myocardial injury was defined as elevated high-sensitivity Troponin I with at least one value above the 99th percentile upper reference limit and its prevalence together with 90-days mortality rate was compared in patients with vs without COVID-infection. Results: From 497 included patients (age 62.9 ± 17.2 years, 56 % male), 314 (63 %) were tested positive on COVID-19 based on PCR-testing, while another cause of symptom was detected in 183 patients (37 %). Cardiovascular diseases were the most frequent differential diagnoses (40 % of patients without COVID-19), followed by bacterial infection (24 %) and malignancies (16 %). Myocardial injury was present in 91 patients (COVID-19 positive: n = 34, COVID-19 negative: n = 57). 90-day mortality rate was higher in patients with myocardial injury (13.4 vs 4.6 %, p = 0.009). Conclusion: Cardiovascular diseases represent the most frequent differential diagnoses in patients presenting to a tertiary care emergency department with symptoms suggestive of an acute infection. Screening for cardiovascular disease is crucial in the initial evaluation of symptomatic patients during the COVID pandemic to identify patients at increased risk.Trial Registration: Clinicaltrials.gov Identifier: NCT04327479.

Background. Although prolonged-release tacrolimus (PR-T) is widely approved for posttransplantation immunosuppression in kidney recipients, large-scale studies are required to assess long-term outcomes. We present follow-up data from the Advagraf-based Immunosuppression Regimen Examining New Onset Diabetes Mellitus in Kidney Transplant Recipients (ADVANCE) trial, in which kidney transplant patients (KTPs) received corticosteroid minimization with PR-T. Methods. ADVANCE was a 24-wk, randomized, open-label, phase-4 study. De novo KTPs received PR-T with basiliximab and mycophenolate mofetil and were randomized to receive an intraoperative corticosteroid bolus plus tapered corticosteroids until day 10 (arm 1) or an intraoperative corticosteroid bolus (arm 2). In this 5-y, noninterventional follow-up, patients received maintenance immunosuppression according to standard practice. The primary endpoint was graft survival (Kaplan-Meier). Secondary endpoints included patient survival, biopsy-confirmed acute rejection-free survival, and estimated glomerular filtration rate (4-variable modification of diet in renal disease). Results. Follow-up study included 1125 patients. Overall graft survival at 1 and 5 y posttransplantation was 93.8% and 88.1%, respectively, and was similar between treatment arms. At 1 and 5 y, patient survival was 97.8% and 94.4%, respectively. Five-year graft and patient survival rates in KTPs who remained on PR-T were 91.5% and 98.2%, respectively. Cox proportional hazards analysis demonstrated similar risk of graft loss and death between treatment arms. Five-year biopsy-confirmed acute rejection-free survival was 84.1%. Mean ± standard deviation values of estimated glomerular filtration rate were 52.7 ± 19.5 and 51.1 ± 22.4 mL/min/1.73 m2 at 1 and 5 y, respectively. Fifty adverse drug reactions were recorded, probably tacrolimus-related in 12 patients (1.5%). Conclusions. Graft survival and patient survival (overall and for KTPs who remained on PR-T) were numerically high and similar between treatment arms at 5 y posttransplantation.

BackgroundHerpes simplex viruses (HSV) cause ubiquitous human infections. For vaccine development, knowledge concerning correlates of protection is essential. Therefore, we investigated (I) if humans are in principle capable producing cell-to-cell spread inhibiting antibodies against HSV and (II) whether this capacity is associated with a reduced HSV-1 reactivation risk.MethodsWe established a high-throughput HSV-1-ΔgE-GFP reporter virus-based assay and evaluated 2,496 human plasma samples for HSV-1 glycoprotein E (gE) independent cell-to-cell spread inhibiting antibodies. Subsequently, we conducted a retrospective survey among the blood donors to analyze the correlation between the presence of cell-to-cell spread inhibiting antibodies in plasma and the frequency of HSV reactivations.ResultsIn total, 128 of the 2,496 blood donors (5.1%) exhibited high levels of HSV-1 gE independent cell-to-cell spread inhibiting antibodies in the plasma. None of the 147 HSV-1 seronegative plasmas exhibited partial or complete cell-to-cell spread inhibition, demonstrating the specificity of our assay. Individuals with cell-to-cell spread inhibiting antibodies showed a significantly lower frequency of HSV reactivations compared to subjects without sufficient levels of such antibodies.ConclusionThis study contains two important findings: (I) upon natural HSV infection, some humans produce cell-to-cell spread inhibiting antibodies and (II) such antibodies correlate with protection against recurrent HSV-1. Moreover, these elite neutralizers may provide promising material for immunoglobulin therapy and information for the design of a protective vaccine against HSV-1.

Abstract Allograft-specific regulatory T cells (Treg cells) are crucial for long-term graft acceptance after transplantation. Although adoptive Treg cell transfer has been proposed, major challenges include graft-specificity and stability. Thus, there is an unmet need for the direct induction of graft-specific Treg cells. We hypothesized a synergism of the immunotolerogenic effects of rapamycin (mTOR inhibition) and plerixafor (CXCR4 antagonist) for Treg cell induction. Thus, we performed fully-mismatched heart transplantations and found combination treatment to result in prolonged allograft survival. Moreover, fibrosis and myocyte lesions were reduced. Although less CD3+ T cell infiltrated, higher Treg cell numbers were observed. Noteworthy, this was accompanied by a plerixafor-dependent plasmacytoid dendritic cells-(pDCs)-mobilization. Furthermore, in vivo pDC-depletion abrogated the plerixafor-mediated Treg cell number increase and reduced allograft survival. Our pharmacological approach allowed to increase Treg cell numbers due to pDC-mediated immune regulation. Therefore pDCs can be an attractive immunotherapeutic target in addition to plerixafor treatment.

Abstract Background Sepsis is triggered by an infection and represents one of the greatest challenges of modern intensive care medicine. With regard to a targeted antimicrobial treatment strategy, the earliest possible pathogen detection is of crucial importance. Until now, culture-based detection methods represent the diagnostic gold standard, although they are characterized by numerous limitations. Culture-independent molecular diagnostic procedures represent a promising alternative. In particular, the plasmatic detection of circulating, cell-free DNA by next-generation sequencing (NGS) has shown to be suitable for identifying disease-causing pathogens in patients with bloodstream infections. Methods The DigiSep-Trial is a randomized, controlled, interventional, open-label, multicenter trial characterizing the effect of the combination of NGS-based digital precision diagnostics with standard-of-care microbiological analyses compared to solely standard-of-care microbiological analyses in the clinical picture of sepsis/septic shock. Additional anti-infective expert consultations are provided for both study groups. In 410 patients (n = 205 per arm) with sepsis/septic shock, the study examines whether the so-called DOOR-RADAR (Desirability of Outcome Ranking/Response Adjusted for Duration of Antibiotic Risk) score (representing a combined endpoint including the criteria (1) intensive/intermediate care unit length of stay, (2) consumption of antibiotics, (3) mortality, and (4) acute kidney injury (AKI)) can be improved by an additional NGS-based diagnostic concept. We also aim to investigate the cost-effectiveness of this new diagnostic procedure. It is postulated that intensive/intermediate care unit length of stay, mortality rate, incidence of AKI, the duration of antimicrobial therapy as well as the costs caused by complications and outpatient aftercare can be reduced. Moreover, a significant improvement in patient’s quality of life is expected. Discussion The authors´ previous work suggests that NGS-based diagnostics have a higher specificity and sensitivity compared to standard-of-care microbiological analyses for detecting bloodstream infections. In combination with the here presented DigiSep-Trial, this work provides the optimal basis to establish a new NGS-driven concept as part of the national standard based on the best possible evidence. Trial registrations DRKS-ID DRKS00022782 . Registered on August 25, 2020 ClinicalTrials.gov NCT04571801 . Registered October 1, 2020

This study compared the immunogenicity and safety of a booster dose of HepB-CpG (HEPLISAV-B® vaccine) with HepB-Eng (Engerix-B®) and HepB-AS04 (Fendrix®) in patients receiving chronic hemodialysis. This was a multicenter, randomized, open-label, phase 3 study of adults receiving hemodialysis with antibodies to HBsAg (anti-HBs)

Immunocompromised patients are at increased risk for a severe course of COVID-19. Treatment of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection with anti-SARS-CoV-2 monoclonal antibodies (mAbs) has become widely accepted. However, the effects of mAb treatment on the long-term primary cellular response to SARS-CoV-2 are unknown. In the following study, we investigated the long-term cellular immune responses to SARS-CoV-2 Spike S1, Membrane (M) and Nucleocapsid (N) antigens using the ELISpot assay in unvaccinated, mAb-treated immunocompromised high-risk patients. Anti-SARS-CoV-2 mAb untreated though vaccinated COVID-19 immunocompromised patients, vaccinated SARS-CoV-2 immunocompromised patients without COVID-19 and vaccinated healthy control subjects served as control groups. The cellular immune response was determined at a median of 5 months after SARS-CoV-2 infection. Our data suggest that immunocompromised patients develop an endogenous long-term cellular immune response after COVID-19, although at low levels. A better understanding of the cellular immune response will help guide clinical decision making for these vulnerable patient cohorts.

Emerging variants of concern (VOC) raise obstacles in shaping vaccination strategies and ending the pandemic. Vaccinated SARS-CoV-2 convalescence shapes the current immune dynamics. We analyzed the SARS-CoV-2 VOC-specific cellular and humoral response of 57 adults: 42 convalescent mRNA vaccinated patients (C+V+), 8 uninfected mRNA vaccinated (C-V+) and 7 unvaccinated convalescent individuals (C+V-). While C+V+ demonstrated a superior humoral SARS-CoV-2 response against all analyzed VOC (alpha, delta, omicron) compared to C-V+ and C+V-, SARS-CoV-2 reactive CD4+ and CD8+ T cells, which can cross-recognize the alpha, delta and omicron VOC after infection and/or vaccination were observed in all there groups without significant differences between the groups. We observed a preserved cross-reactive C+V+ and C-V+ T cell memory. An inferior humoral response but preserved cross-reactive T cell memory in C+V- compared to C+V+ was observed, as well as an inferior humoral response but preserved cross-reactive T cell memory in C+V- compared to C-V+. Adaptive immunity generated after SARS-CoV-2 infection and vaccination leads to superior humoral immune response against VOC compared to isolated infection or vaccination. Despite the apparent loss of neutralization potential caused by viral evolution, a preserved SARS-CoV-2 reactive T cell response with a robust potential for cross-recognition of the alpha, delta and omicron VOC was detected in all studied cohorts. Our results may have implications on current vaccination strategies.

We investigated immune responses against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) among a group of convalescent, potential blood donors in Germany who had PCR-confirmed SARS-CoV-2 infection. Sixty days after onset of symptoms, 13/78 (17%) study participants had borderline or negative results to an ELISA detecting IgG against the S1 protein of SARS-CoV-2. We analyzed participants with PCR-confirmed infection who had strong antibody responses (ratio >3) as positive controls and participants without symptoms of SARS-CoV-2 infection and without household contact with infected patients as negative controls. Using interferon-γ ELISpot, we observed that 78% of PCR-positive volunteers with undetectable antibodies showed T cell immunity against SARS-CoV-2. We observed a similar frequency (80%) of T-cell immunity in convalescent donors with strong antibody responses but did not detect immunity in negative controls. We concluded that, in convalescent patients with undetectable SARS-CoV-2 IgG, immunity may be mediated through T cells.

Abstract Background Orbital aspergillosis is a rare sight- and life-threatening fungal infection affecting immunocompromised or otherwise healthy patients. It is often misdiagnosed due to its unspecific clinical and radiologic appearance. Therapeutic delay can have dramatic consequences. However, progress in microbiological diagnostic techniques and therapeutic experience from case series help improve the management of this disease. Case presentation A 78-year-old immunocompetent woman presented at an eye clinic for subacute swelling, reddening, and ptosis of her left upper eyelid. Based on radiologic and histologic considerations, she was treated for idiopathic orbital inflammation, but her condition worsened. After a second biopsy of the orbital mass, aspergillosis was diagnosed. Her condition improved promptly after initiation of an oral voriconazole treatment. Additionally, using a polymerase chain reaction (PCR) assay, A. fumigatus was identified on tissue of both biopsies and its azole susceptibility was examined simultaneously. Conclusions In the case described here, oral antifungal treatment was sufficient for the therapy of invasive orbital aspergillosis. Performing fungal PCR on orbital tissue can accelerate the diagnostic process and should be performed in ambiguous cases of slowly growing orbital mass. Finally, interdisciplinary management is the key to optimal treatment of orbital tumours and infections.

Clinical and experimental studies indicate that the bacterial and fungal gut microbiota modulates immune responses in distant organs including the lungs. Immune dysregulation is associated with severe SARS-CoV-2 infection, and several groups have observed gut bacterial dysbiosis in SARS-CoV-2 infected patients, while the fungal gut microbiota remains poorly defined in these patients. We analyzed the fungal gut microbiome from rectal swabs taken prior to anti-infective treatment in 30 SARS-CoV-2 positive (21 non-severe COVID-19 and 9 developing severe/critical COVID-19 patients) and 23 SARS-CoV-2 negative patients by ITS2-sequencing. Pronounced but distinct interconnected fungal communities distinguished SARS-CoV-2 positive and negative patients. Fungal gut microbiota in severe/critical COVID-19 illness was characterized by a reduced diversity, richness and evenness and by an increase of the relative abundance of the Ascomycota phylum compared with non-severe COVID-19 illness. A dominance of a single fungal species with a relative abundance of >75% was a frequent feature in severe/critical COVID-19. The dominating fungal species were highly variable between patients even within the groups. Several fungal taxa were depleted in patients with severe/critical COVID-19.The distinct compositional changes of the fungal gut microbiome in SARS-CoV-2 infection, especially in severe COVID-19 illness, illuminate the necessity of a broader approach to investigate whether the differences in the fungal gut microbiome are consequences of SARS-CoV-2 infection or a predisposing factor for critical illness.

Background: In order to control their anti-infectives consumption, hospitals are required to provide multidisciplinary teams comprising among others an infectiologist, a microbiologist and a pharmacist. Small hospitals though often do not dispose of the defaulted personnel. This study illustrates a solution for an antimicrobial stewardship program (ASP) in small community hospitals in a rural area in Germany. Methods: Four hospitals of ca. 200 beds each, jointly hired an antimicrobial stewardship expert to start a common ASP. This expert did rounds on every ward once a week, mostly as chard reviews with the physician in charge. Outside the rounds, he could be consulted by mail. Working time and number of visited patients were documented. Anti-infectives consumption, incidence of Clostridioides difficile infections (CDI) and mortality rates were retrieved from routinely collected data. The intervention period (01/2018–12/2018) was compared to the preintervention period (01/2017–12/2017). Results: 3321 patients were visited in the intervention period. In average, 20 patients were seen per day and 20 min were needed per patient/ chard. About 65% of the expert’s working time was needed for rounds, 15% for driving between the hospitals. The anti-infectives consumption of the 4 hospitals in the preintervention period amounted to 50 defined daily doses per 100 occupied bed days. The total consumption was reduced by 10% and of quinolones by 36%. The incidence of hospital-acquired CDI receded from 0.14 to 0.07 cases per 100 patient days (−50%, p = 0.001). The overall in-hospital mortality did not change. Conclusions: A single expert was able to implement a successfull ASP in 4 hospitals. While multidisciplinary antimicrobial stewardship teams are ideal for tertiary care hospitals, small hospitals need a more practical solution. This survey shows that one expert can be sufficient for several small hospitals even with the distances in a rural setting.

ObjectivesChloroquine (CQ) is approved for treatment of B-cell mediated diseases such as rheumatoid arthritis and systemic lupus erythematosus. However, the exact mode of action in these diseases has not been studied and it remains unclear which effect CQ has on B-cells. Thus, it was the aim of this study to investigate to which extent CQ affects functionality of effector and regulatory B-cell.MethodsFor this purpose, B-cells were isolated from peripheral blood of healthy controls and renal transplant patients. B-cells were stimulated in presence or absence of CQ and Interleukin-10 (IL-10) and Granzyme B (GrB) secretion were assessed. In addition, effector functions such as plasma cell formation, and Immunoglobulin G (IgG) secretion were studied.ResultsCQ suppressed Toll-Like-Receptor (TLR)-9 induced B-cell proliferation in a dose-dependent manner. IL-10pos regulatory B-cells were suppressed by CQ already at low concentrations whereas anti-IgG/IgM-induced GrB secreting regulatory B-cells were less susceptible. Plasma blast formation and IgG secretion was potently suppressed by CQ. Moreover, purified B-cells from renal transplant patients were also susceptible to CQ-induced suppression of effector B-cell functions as observed by diminished IgG secretion.ConclusionIn conclusion, CQ had a suppressive effect on IL-10 regulatory B-cells whereas GrB secreting regulatory B-cells were less affected. Effector functions of B-cells such as plasma blast formation and IgG secretion were also inhibited by CQ. Effector B-cells derived from renal transplant patients already under immunosuppression could be suppressed by CQ. These findings may partly explain the clinical efficacy of CQ in B-cell mediated autoimmune diseases. The application of CQ in other disease contexts where suppression of effector B-cells could offer a benefit, such as renal transplantation, may hypothetically be advantageous.

SARS-CoV-2 variants of concern (VOCs) can trigger severe endemic waves and vaccine breakthrough infections (VBI). We analyzed the cellular and humoral immune response in 8 patients infected with the alpha variant, resulting in moderate to fatal COVID-19 disease manifestation, after double mRNA-based anti-SARS-CoV-2 vaccination. In contrast to the uninfected vaccinated control cohort, the diseased individuals had no detectable high-avidity spike (S)-reactive CD4+ and CD8+ T cells against the alpha variant and wild type (WT) at disease onset, whereas a robust CD4+ T-cell response against the N- and M-proteins was generated. Furthermore, a delayed alpha S-reactive high-avidity CD4+ T-cell response was mounted during disease progression. Compared to the vaccinated control donors, these patients also had lower neutralizing antibody titers against the alpha variant at disease onset. The delayed development of alpha S-specific cellular and humoral immunity upon VBI indicates reduced immunogenicity against the S-protein of the alpha VOC, while there was a higher and earlier N- and M-reactive T-cell response. Our findings do not undermine the current vaccination strategies but underline a potential need for the inclusion of VBI patients in alternative vaccination strategies and additional antigenic targets in next-generation SARS-CoV-2 vaccines.

The human leukocyte antigen G (HLA-G) is an immune checkpoint molecule with a complex network of interactions with several inhibitory receptors. Although the effect of HLA-G on T cells and NK cells is well studied, the effect of HLA-G on B cells is still largely elusive. B cells are of particular interest in the context of the HLA-G-ILT-2 interaction because the ILT-2 receptor is constitutively expressed on most B cells, whereas it is only present on some subsets of T and NK cells. To characterize the effect of HLA-G5 molecules on B cells, we studied splenic B cells derived from cytomegalovirus (CMV) sero-positive donors after CMV stimulation with antigens in the presence and absence of soluble HLA-G5. In the presence of HLA-G5, increased expression of the ITIM-bearing Ig-like transcript (ILT-2) was observed on B cells, but its expression was not affected by stimulation with CMV antigens. Moreover, it became evident that HLA-G5 exposure resulted in a decreased expression of CD27 and CD38 and, accordingly, in lower proportions of CD19+CD27+CD38+ and higher proportions of CD19+CD27-CD38- B cells. Taken together, our in vitro findings demonstrate that soluble HLA-G5 suppresses markers of B cell activation, suggesting that HLA-G5 has an impact on splenic B cell differentiation and activation. Based on these results, further investigation regarding the role of HLA-G as a prognostic factor and a potential therapeutic agent with respect to B cell function appears reasonable.

Background: It is widely accepted that SARS-CoV-2 causes a dysregulation of immune and coagulation processes. In severely affected patients, viral sepsis may result in life endangering multiple organ dysfunction. Furthermore, most therapies for COVID-19 patients target either the immune system or coagulation processes. As the exact mechanism causing SARS-CoV-2-induced morbidity and mortality was unknown, we started an in-depth analysis of immunologic and coagulation processes. Methods: 127 COVID-19 patients were treated at the University Hospital Essen, Germany, between May 2020 and February 2022. Patients were divided according to their maximum COVID-19 WHO ordinal severity score (WHO 0–10) into hospitalized patients with a non-severe course of disease (WHO 4–5, n = 52) and those with a severe course of disease (WHO 6–10, n = 75). Non-infected individuals served as healthy controls (WHO 0, n = 42). Blood was analyzed with respect to cell numbers, clotting factors, as well as pro- and anti-inflammatory mediators in plasma. As functional parameters, phagocytosis and inflammatory responses to LPS and antigen-specific stimulation were determined in monocytes, granulocytes, and T cells using flow cytometry. Findings: In the present study, immune and coagulation systems were analyzed simultaneously. Interestingly, many severe COVID-19 patients showed an upregulation of pro-inflammatory mediators and at the same time clear signs of immunosuppression. Furthermore, severe COVID-19 patients not only exhibited a disturbed immune system, but in addition showed a pronounced pro-coagulation phenotype with impaired fibrinolysis. Therefore, our study adds another puzzle piece to the already complex picture of COVID-19 pathology implying that therapies in COVID-19 must be individualized. Conclusion: Despite years of research, COVID-19 has not been understood completely and still no therapies exist, fitting all requirements and phases of COVID-19 disease. This observation is highly reminiscent to sepsis. Research in sepsis has been going on for decades, while the disease is still not completely understood and therapies fitting all patients are lacking as well. In both septic and COVID-19 patients, immune activation can be accompanied by immune paralysis, complicating therapeutic intervention. Accordingly, therapies that lower immune activation may cause detrimental effects in patients, who are immune paralyzed by viral infections or sepsis. We therefore suggest individualizing therapies and to broaden the spectrum of immunological parameters analyzed before therapy. Only if the immune status of a patient is understood, can a therapeutic intervention be successful.

The gut microbiota contributes to maintaining human health and regulating immune responses. Severe COVID-19 illness is associated with a dysregulated pro-inflammatory immune response. The effect of SARS-CoV-2 on altering the gut microbiome and the relevance of the gut microbiome on COVID-19 severity needs to be clarified. In this prospective study, we analyzed the gut microbiome of 212 patients of a tertiary care hospital (117 patients infected with SARS-CoV-2 and 95 SARS-CoV-2 negative patients) using 16S rRNA gene sequencing of the V3-V4 region. Inflammatory markers and immune cells were quantified from blood. The gut microbiome in SARS-CoV-2 infected patients was characterized by a lower bacterial richness and distinct differences in the gut microbiome composition, including an enrichment of the phyla Proteobacteria and Bacteroidetes and a decrease of Actinobacteria compared to SARS-CoV-2 negative patients. The relative abundance of several genera including Bifidobacterium, Streptococcus and Collinsella was lower in SARS-CoV-2 positive patients while the abundance of Bacteroides and Enterobacteriaceae was increased. Higher pro-inflammatory blood markers and a lower CD8+ T cell number characterized patients with severe COVID-19 illness. The gut microbiome of patients with severe/critical COVID-19 exhibited a lower abundance of butyrate-producing genera Faecalibacterium and Roseburia and a reduction in the connectivity of a distinct network of anti-inflammatory genera that was observed in patients with mild COVID-19 illness and in SARS-CoV-2 negative patients. Dysbiosis of the gut microbiome associated with a pro-inflammatory signature may contribute to the hyperinflammatory immune response characterizing severe COVID-19 illness.

Vulnerable patients such as immunosuppressed or elderly patients are at high risk for a severe course of COVID-19 upon SARS-CoV-2 infection. Immunotherapy with SARS-CoV-2 specific monoclonal antibodies (mAb) or convalescent plasma represents a considerable treatment option to protect these patients from a severe or lethal course of infection. However, monoclonal antibodies are not always available or less effective against emerging SARS-CoV-2 variants. Convalescent plasma is more commonly available and may represent a good treatment alternative in low-income countries. We retrospectively evaluated outcomes in individuals treated with mAbs or convalescent plasma and compared the 30-day overall survival with a patient cohort that received supportive care due to a lack of SARS-CoV-2 specific therapies between March 2020 and April 2021. Our data demonstrate that mAb treatment is highly effective in preventing severe courses of SARS-CoV-2 infection. All patients treated with mAb survived. Treatment with convalescent plasma improved overall survival to 82% compared with 61% in patients without SARS-CoV-2 targeted therapy. Our data indicate that early convalescent plasma treatment may be an option to improve the overall survival of high-risk COVID-19 patients. This is especially true when other antiviral drugs are not available or their efficacy is significantly reduced, which may be the case with emerging SARS-CoV-2 variants.

Abstract Background Dysregulation of the B-cell activating factor (BAFF) system is involved in the pathogenesis of systemic lupus erythematosus (SLE). Increased serum concentrations of BAFF are related to lupus nephritis and disease activity among SLE patients. Recently, a variant of the BAFF-encoding gene, BAFF-var, was identified to be associated with autoimmune diseases, in particular SLE, and to promote the production of soluble BAFF. The present study aimed to assess the prevalence of BAFF-var in a cohort of 195 SLE patients and to analyze the association of the BAFF-var genotype (TNSF13B) with various manifestations of SLE. Methods A cohort of 195 SLE patients from Central Europe, including 153 patients from the Swiss SLE Cohort Study and 42 patients from the University Hospital Essen, Germany, underwent genotyping for detection of BAFF-var allele. Results Of the 195 patients, 18 (9.2%) tested positive for BAFF-var variant according to the minor allele frequency of 4.6%. The presence of BAFF-var was associated with the occurrence of lupus nephritis (p = 0.038) (p = 0.03 and p = 0.003). Among various organ manifestations of SLE, the presence of BAFF-var was associated with the occurrence of lupus nephritis (p = 0.038; odds ratio [OR], 2.4; 95% confidence interval [CI], 0.89–6.34) and renal activity markers such as proteinuria and hematuria (p = 0.03; OR, 2.4; 95% CI, 0.9–6.4 for proteinuria; p = 0.003; OR, 3.9; 95% CI, 1.43–10.76 for hematuria). SLE patients carrying the BAFF-var allele exhibited increased disease activity at study entry, as determined by the physician’s global assessment (PGA: p = 0.002; OR, 4.8; 95% CI, 1.54–14.93) and the SLE Disease Activity Index (p = 0.012; OR, 3.5; 95% CI, 1.12–11.18). Consistent with that, the percentage of patients treated with immunosuppressive agents at study entry was higher among those carrying the BAFF-var allele than among those tested negative for BAFF-var (p = 0.006; OR, 3.7; 95% CI, 1.27–10.84). Conclusions Our results indicate an association between the BAFF-var genotype and increased severity of SLE. Determining the BAFF-var status of SLE patients may improve the risk stratification of patients for whom the development of lupus nephritis is more likely and thus may be helpful in the follow-up care and treatment of SLE patients.

Abstract Background Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by T-cell-dependent B-cell activation and altered T-cell response. Co-stimulatory and co-inhibitory molecules regulate and exert T-cell differentiation, survival and cytokine production. CD134+ and PD-1+ T-cells in SLE patients are increased in SLE. The aim of this study was to characterize CD134+ and PD-1+CD4+ T-cells according to their ability to produce IFN-γ, IL-21 and IL-22 in SLE patients. Methods Peripheral blood of 39 SLE patients and 19 healthy controls (HC) was stimulated with phorbol myristate acetate (PMA) calcium ionophore (Ca-Io). The expression of IFN-γ, IL-21 and IL-22 T-cells within the CD134+ and PD-1+ T-cells was analyzed by flow cytometry. Disease activity was assessed by SLE Disease Activity Index. Results Peripheral unstimulated CD134+ and PD-1+ CD4+ T-cells were significantly increased in patients with lupus nephritis. Upon stimulation both, CD134+ and PD-1+ CD4+ T-cells, produced significantly less IFN-γ in SLE patients as compared to HC. The percentages of IL-22 within the CD134+CD4+ T-cells were also significantly decreased in SLE as compared to HC. Conclusion CD134+ and PD-1+CD4+ T-cells have mainly a Th1 effector T-cell signature. A lower proportion produces also IL-21 and IL-22. The impaired capacity to produce IFN-γ and IL-22 in SLE patients may contribute to the pathogenesis of the disease.

Abstract Background The influence of pre- or postprandial administration on pharmacokinetics of cyclosporine is supposed to be less in gel-based formulations than in microemulsions. This study was designed to investigate the influence of a high-fat meal on the pharmacokinetic profile of the two cyclosporine containing formulations Ciclosporin Pro (gel-based emulsion) and Sandimmun®Optoral (microemulsion) in renal transplant recipients. Methods A randomized, open-label, repeated-measurement, comparative phase IV trial was conducted with two sequence groups for nutrition condition (fasting→fed, fed→fasting) and two treatment phases (Sandimmun® Optoral → Ciclosporin Pro), each covering both nutrition conditions. Primary pharmacokinetic variable of interest was the reduction of bioavailability due to high-fat food compared to fasting conditions measured by the difference D of ln-transformed bioavailability variables (AUCSS, τ, Css, max, und Css, min). Results A nutrition effect was found for both study medications with respect to the parameters AUCSS, τ and CSS, max, but not to CSS, min. The reduction of bioavailability caused by high-fat food was not significantly different for Sandimmun®Optoral and Ciclosporin Pro. Conclusions An effect of high-fat breakfast prior to the morning dose on AUCSS, τ and CSS, max was found for Sandimmun® Optoral and for Ciclosporin Pro. Trough level monitoring did not capture ingestion-related variability. Conversion to Ciclosporin Pro seems to be safe with regard to intra-individual pharmacokinetic variability. Trial registration EudraCT No. 2009–011354-18 (29th April 2019)

In kidney transplant (KTX) patients, immune responses after booster vaccination against SARS-CoV-2 are inadequately examined. We analyzed these patients a median of four months after a third/fourth vaccination and compared them to healthy controls. Cellular responses were analyzed by interferon-gamma (IFN-γ) and interleukin-2 (IL-2) ELISpot assays. Neutralizing antibody titers were assessed against SARS-CoV-2 D614G (wild type) and the variants alpha, delta, and omicron by a cell culture-based neutralization assay. Humoral immunity was also determined by a competitive fluorescence assay, using 11 different variants of SARS-CoV-2. Antibody ratios were measured by ELISA. KTX patients showed significantly lower SARS-CoV-2-specific IFN-γ responses after booster vaccination than healthy controls. However, SARS-CoV-2-specific IL-2 responses were comparable to the T cell responses of healthy controls. Cell culture-based neutralizing antibody titers were 1.3-fold higher in healthy controls for D614G, alpha, and delta, and 7.8-fold higher for omicron (p < 0.01). Healthy controls had approximately 2-fold higher concentrations of potential neutralizing antibodies against all 11 variants than KTX patients. However, more than 60% of the KTX patients displayed antibodies to variants of SARS-CoV-2. Thus, KTX patients should be partly protected, due to neutralizing antibodies to variants of SARS-CoV-2 or by cross-reactive T cells, especially those producing IL-2.

Cytotoxic CD8+ T-cells play a pivotal role in the pathogenesis of systemic lupus erythematosus (SLE). The aim of this study was to investigate the role of CD107a (LAMP-1) on cytotoxic CD8+ T-cells in SLE-patients in particular with lupus nephritis. Peripheral blood of SLE-patients (n = 31) and healthy controls (n = 21) was analyzed for the expression of CD314 and CD107a by flow cytometry. Kidney biopsies of lupus nephritis patients were investigated for the presence of CD8+ and C107a+ cells by immunohistochemistry and immunofluorescence staining. The percentages of CD107a+ on CD8+ T-cells were significantly decreased in SLE-patients as compared to healthy controls (40.2 ± 18.5% vs. 47.9 ± 15.0%, p = 0.02). This was even more significant in SLE-patients with inactive disease. There was a significant correlation between the percentages of CD107a+CD8+ T-cells and SLEDAI. The evaluation of lupus nephritis biopsies showed a significant number of CD107a+CD8+ T-cells mainly located in the peritubular infiltrates. The intrarenal expression of CD107a+ was significantly correlated with proteinuria. These results demonstrate that CD8+ T-cells of patients with systemic lupus erythematosus have an altered expression of CD107a which seems to be associated with disease activity. The proof of intrarenal CD107a+CD8+ suggests a role in the pathogenesis of lupus nephritis.

Background: A growing number of reports suggest that infection with SARS-CoV-2 often leads to neurological involvement; however, data on the incidence and severity are limited to mainly case reports and retrospective studies. Methods: This prospective, cross-sectional study of 102 SARS-CoV-2 PCR positive patients investigated the frequency, type, severity and risk factors as well as underlying pathophysiological mechanisms of neurological involvement (NIV) in COVID-19 patients. Results: Across the cohort, 59.8% of patients had NIV. Unspecific NIV was suffered by 24.5%, mainly general weakness and cognitive decline or delirium. Mild NIV was found in 9.8%; most commonly, impaired taste or smell. Severe NIV was present in 23.5%; half of these suffered cerebral ischaemia. Incidence of NIV increased with respiratory symptoms of COVID-19. Mortality was higher with increasing NIV severity. Notably, 83.3% with severe NIV had a pre-existing neurological co-morbidity. All cerebrospinal fluid (CSF) samples were negative for SARS-CoV-2 RNA, and SARS-CoV-2 antibody quotient did not suggest intrathecal antibody synthesis. Of the patients with severe NIV, 50% had blood–brain barrier (BBB) disruption and showed a trend of elevated interleukin levels in CSF. Antibodies against neuronal and glial epitopes were detected in 35% of the patients tested. Conclusion: Cerebrovascular events were the most frequent severe NIV and severe NIV was associated with high mortality. Incidence of NIV increased with respiratory symptoms and NIV and pre-existing neurological morbidities were independent risk factors for fatality. Inflammatory involvement due to BBB disruption and cytokine release drives NIV, rather than direct viral invasion. These findings might help physicians define a further patient group requiring particular attention during the pandemic.

BackgroundCytomegalovirus (CMV) infection is amongst the most important factors complicating solid organ transplantation. In a large prospective randomized clinical trial, valganciclovir prophylaxis reduced the occurrence of CMV infection and disease compared with preemptive therapy in CMV-positive renal allograft recipients (VIPP study; NCT00372229). Here, we present a subanalysis of the VIPP study, investigating single nucleotide polymorphisms (SNPs) in immune-response-related genes and their association with active CMV infection, CMV disease, graft loss or death, rejection, infections, and leukopenia.MethodsBased on literature research ten SNPs were analyzed for TLR4, three for IFN-γ, six for IL10, nine for IL37, and two for TNF-α. An asymptotic independence test (Cochran-Armitage trend test) was used to examine associations between SNPs and the occurrence of CMV infection or other negative outcomes. Statistical significance was defined as pResultsSNPs were analyzed on 116 blood samples. No associations were found between the analyzed SNPs and the occurrence of CMV infection, rejection and leukopenia in all patients. For IL37 rs2723186, an association with CMV disease (p = 0.0499), for IL10 rs1800872, with graft loss or death (p = 0.0207) and for IL10 rs3024496, with infections (p = 0.0258) was observed in all patients, however did not hold true after correction for multiple testing.ConclusionThe study did not reveal significant associations between the analyzed SNPs and the occurrence of negative outcomes in CMV-positive renal transplant recipients after correction for multiple testing. The results of this association analysis may be of use in guiding future research efforts.

Solid organ transplant recipients have an up to ninefold higher risk of varicella–zoster virus (VZV) reactivation than the general population. Due to lifelong immunosuppressive therapy, vaccination against VZV may be less effective in kidney transplant (KTX) recipients. In the current study, twelve female and 17 male KTX recipients were vaccinated twice with the adjuvanted, recombinant zoster vaccine Shingrix™, which contains the VZV glycoprotein E (gE). Cellular immunity against various VZV antigens was analyzed with interferon-gamma ELISpot. We observed the strongest vaccination-induced changes after stimulation with a gE peptide pool. One month after the second vaccination, median responses were 8.0-fold higher than the responses prior to vaccination (p = 0.0006) and 4.8-fold higher than responses after the first vaccination (p = 0.0007). After the second vaccination, we observed an at least twofold increase in ELISpot responses towards gE peptides in 22 out of 29 patients (76%). Male sex, good kidney function, early time point after transplantation, and treatment with tacrolimus or mycophenolate were correlated significantly with higher VZV-specific cellular immunity, whereas diabetes mellitus was correlated with impaired responses. Thus, our data indicate that vaccination with Shingrix™ significantly augmented cellular, VZV gE-specific immunity in KTX recipients, which was dependent on several covariates.

Several polymorphisms within Fc receptors (FCR) have been described, some of which correlate with allograft function. In the current study, we determined three Fcγ receptor and five Fcα receptor dimorphisms in 47 kidney transplant recipients who had been vaccinated against Streptococcus pneumoniae. We analyzed if FCR genotypes correlated with pneumococcal antibodies and their serotype-specific opsonophagocytic function, tested prior to and at months 1 and 12 post-vaccination. In parallel, we assessed antibodies against HLA and MICA and determined kidney function. We observed that IgG2 antibodies against pneumococci at months 1 and 12 after vaccination and IgA antibodies at month 1 differed significantly between the carriers of the three genotypes of FCGR3A rs396991 (V158F, p = 0.02; 0.04 and 0.009, respectively). Moreover, the genotype of FCGR3A correlated with serotype-specific opsonophagocytic function, reaching statistical significance (p < 0.05) at month 1 for 9/13 serotypes and at month 12 for 6/13 serotypes. Heterozygotes for FCGR3A had the lowest antibody response after pneumococcal vaccination. On the contrary, heterozygotes tended to have more antibodies against HLA class I and impaired kidney function. Taken together, our current data indicate that heterozygosity for FCGR3A may be unfavorable in kidney transplant recipients.

Purpose: To evaluate the use of pulmonary computed tomography (CT) angiography during initial admission at an emergency department (ED), to identify COVID-19 patients with accompanying pulmonary embolism (PE) and its impact on clinical management. Methods: We performed a retrospective analysis of COVID-19 patients that underwent pulmonary CT angiography at the ED. CT scans were evaluated for the presence and extent of PE and for imaging changes suspicious of COVID-19. Patients were subdivided into two groups: (1) Group A consisted of patients with proven COVID-19 based on real-time polymerase chain reaction (RT-PCR), and (2) Group B of patients suspected for COVID-19, comprising patients positive on RT-PCR and/or COVID-19-suspicious CT findings. To assess the differences between patients with and without pulmonary embolism, Fisher’s exact test was used. Results: A total of 308 patients were admitted to the ED for diagnostic work-up of dyspnea and suspected COVID-19, and 95 patients underwent pulmonary CT angiography. PE was detected in 13.6% (3/22) of patients in Group A and 20.7% (6/29) in Group B. No significant differences were observed between patients with and without PE concerning hospitalization (Group B: 100% (6/6) vs. 91.3% (21/23)), the necessity of oxygen therapy (Group B: 66% (4/6) vs. 43.5% (10/23)), and death (Group B: 33% (2/6) vs. 4.3% (1/23) p > 0.05, respectively). Conclusions: In 20.7% of COVID-19 patients, PE was detected upon admission to the ED. Although the incorporation of early pulmonary CT angiography in patients suspicious of COVID-19 may be beneficial to identify concomitant PE, further studies are necessary to corroborate these findings.

The novel, highly transmissible severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has triggered a pandemic of acute respiratory illness worldwide and remains a huge threat to the healthcare system’s capacity to respond to COVID-19. Elderly and immunocompromised patients are at increased risk for a severe course of COVID-19. These high-risk groups have been identified as developing diminished humoral and cellular immune responses. Notably, SARS-CoV-2 RNA remains detectable in nasopharyngeal swabs of these patients for a prolonged period of time. These factors complicate the clinical management of these vulnerable patient groups. To date, there are no well-defined guidelines for an appropriate duration of isolation for elderly and immunocompromised patients, especially in hospitals or nursing homes. The aim of the present study was to characterize at-risk patient cohorts capable of producing a replication-competent virus over an extended period after symptomatic COVID-19, and to investigate the humoral and cellular immune responses and infectivity to provide a better basis for future clinical management. In our cohort, the rate of positive viral cultures and the sensitivity of SARS-CoV-2 antigen tests correlated with higher viral loads. Elderly patients and patients with diabetes mellitus had adequate cellular and humoral immune responses to SARS-CoV-2 infection, while immunocompromised patients had reduced humoral and cellular immune responses. Our patient cohort was hospitalized for longer compared with previously published cohorts. Longer hospitalization was associated with a high number of nosocomial infections, representing a potential hazard for additional complications to patients. Most importantly, regardless of positive SARS-CoV-2 RNA detection, no virus was culturable beyond a cycle threshold (ct) value of 33 in the majority of samples. Our data clearly indicate that elderly and diabetic patients develop a robust immune response to SARS-CoV-2 and may be safely de-isolated at a ct value of more than 35.

Post-transplantation cytomegalovirus (CMV) syndrome can be prevented using the antiviral drug (val)ganciclovir. (Val)ganciclovir is typically administered following a prophylactic or a pre-emptive strategy. The prophylactic strategy entails early universal administration, the pre-emptive strategy, early treatment in case of infection. However, it is not clear which strategy is superior with respect to transplantation outcome; sex-specific effects of these prevention strategies are not known. We have retrospectively analyzed 540 patients from the multi-centre Harmony study along eight pre-defined visits: 308 were treated according to a prophylactic, 232 according to a pre-emptive strategy. As expected, we observed an association of prophylactic strategy with lower incidence of CMV syndrome, delayed onset and lower viral loads compared to the pre-emptive strategy. However, in female patients, the prophylactic strategy was associated with a strong impairment of glomerular filtration rate one year post-transplant (difference: −11.8 ± 4.3 ml min−1·1.73 m−2, p = 0.006). Additionally, we observed a tendency of higher incidence of acute rejection and severe BK virus reactivation in the prophylactic strategy group. While the prophylactic strategy was more effective for preventing CMV syndrome, our results suggest for the first time that the prophylactic strategy might lead to inferior transplantation outcomes in female patients, providing evidence for a strong association with sex. Further randomized controlled studies are necessary to confirm this potential negative effect.

ABSTRACT Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection induces a T cell response that most likely contributes to virus control in COVID-19 patients but may also induce immunopathology. Until now, the cytotoxic T cell response has not been very well characterized in COVID-19 patients. Here, we analyzed the differentiation and cytotoxic profile of T cells in 30 cases of mild COVID-19 during acute infection. SARS-CoV-2 infection induced a cytotoxic response of CD8+ T cells, but not CD4+ T cells, characterized by the simultaneous production of granzyme A and B as well as perforin within different effector CD8+ T cell subsets. PD-1-expressing CD8+ T cells also produced cytotoxic molecules during acute infection, indicating that they were not functionally exhausted. However, in COVID-19 patients over the age of 80 years, the cytotoxic T cell potential was diminished, especially in effector memory and terminally differentiated effector CD8+ cells, showing that elderly patients have impaired cellular immunity against SARS-CoV-2. Our data provide valuable information about T cell responses in COVID-19 patients that may also have important implications for vaccine development. IMPORTANCE Cytotoxic T cells are responsible for the elimination of infected cells and are key players in the control of viruses. CD8+ T cells with an effector phenotype express cytotoxic molecules and are able to perform target cell killing. COVID-19 patients with a mild disease course were analyzed for the differentiation status and cytotoxic profile of CD8+ T cells. SARS-CoV-2 infection induced a vigorous cytotoxic CD8+ T cell response. However, this cytotoxic profile of T cells was not detected in COVID-19 patients over the age of 80 years. Thus, the absence of a cytotoxic response in elderly patients might be a possible reason for the more frequent severity of COVID-19 in this age group than in younger patients.

Summary: T cell immunity toward SARS-CoV-2 spike (S-), membrane (M-), and nucleocapsid (N-) proteins may define COVID-19 severity. Therefore, we compare the SARS-CoV-2-reactive T cell responses in moderate, severe, and critical COVID-19 patients and unexposed donors. Overlapping peptide pools of all three proteins induce SARS-CoV-2-reactive T cell response with dominance of CD4+ over CD8+ T cells and demonstrate interindividual immunity against the three proteins. M-protein induces the highest frequencies of CD4+ T cells, suggesting its relevance for diagnosis and vaccination. The T cell response of critical COVID-19 patients is robust and comparable or even superior to non-critical patients. Virus clearance and COVID-19 survival are not associated with either SARS-CoV-2 T cell kinetics or magnitude of T cell responses, respectively. Thus, our data do not support the hypothesis of insufficient SARS-CoV-2-reactive immunity in critical COVID-19. Conversely, it indicates that activation of differentiated memory effector T cells could cause hyperreactivity and immunopathogenesis in critical patients.

This study analyzed binding and neutralizing antibody titers up to 6 months after standard vaccination with BNT162b2 (two doses of 30 µg each) in SARS-CoV-2 naïve patients (n = 59) on hemodialysis. Humoral vaccine responses were measured before and 6, 12, and 24 weeks after the first vaccination. A chemiluminescent immunoassay (CLIA) was used to quantify SARS-CoV-2 IgG against the spike glycoprotein. SARS-CoV-2 neutralizing activity was tested against the wild-type virus. A multivariable binary regression model was used to identify risk factors for the absence of humoral immune responses at 6 months. At week 6, vaccine-specific seroconversion was detected in 96.6% of all patients with median anti-SARS-CoV-2 IgGs of 918 BAU/mL. At weeks 12 and 24, seroconversion rates decreased to 91.5% and 79.7%, and corresponding median binding antibody titers declined to 298 BAU/mL and 89 BAU/mL, respectively. Neutralizing antibodies showed a decay from 79.6% at week 6 to 32.8% at week 24. The risk factor with the strongest association for vanishing immune responses was low serum albumin (p = 0.018). Regarding vaccine-specific humoral responses 6 months after the standard BNT162b2 vaccination schedule, SARS-CoV-2 naïve patients receiving hemodialysis must be considered at risk of becoming infected with SARS-CoV-2 and being infectious.

Abstract Background Conversion from calcineurin inhibitor (CNI) therapy to a mammalian target of rapamycin (mTOR) inhibitor following kidney transplantation may help to preserve graft function. Data are sparse, however, concerning the impact of conversion on posttransplant diabetes mellitus (PTDM) or the progression of pre-existing diabetes. Methods PTDM and other diabetes-related parameters were assessed post hoc in two large open-label multicenter trials. Kidney transplant recipients were randomized (i) at month 4.5 to switch to everolimus or remain on a standard cyclosporine (CsA)-based regimen (ZEUS, n = 300), or (ii) at month 3 to switch to everolimus, remain on standard CNI therapy or convert to everolimus with reduced-exposure CsA (HERAKLES, n = 497). Results There were no significant differences in the incidence of PTDM between treatment groups (log rank p = 0.97 [ZEUS], p = 0.90 [HERAKLES]). The mean change in random blood glucose from randomization to month 12 was also similar between treatment groups in both trials for patients with or without PTDM, and with or without pre-existing diabetes. The change in eGFR from randomization to month 12 showed a benefit for everolimus versus comparator groups in all subpopulations, but only reached significance in larger subgroups (no PTDM or no pre-existing diabetes). Conclusions Within the restrictions of this post hoc analysis, including non-standardized diagnostic criteria and limited glycemia laboratory parameters, these data do not indicate any difference in the incidence or severity of PTDM with early conversion from a CsA-based regimen to everolimus, or in the progression of pre-existing diabetes. Trial registration clinicaltrials.gov, NCT00154310 (registered September 2005) and NCT00514514 (registered August 2007); EudraCT (2006-007021-32 and 2004-004346-40).