Clinical and immunological outcomes of SARS-CoV-2-infected vaccine responders, vaccine non-responders, and unvaccinated patients evaluated for neutralizing monoclonal antibody treatment at a single German tertiary care center: a retrospective cohort study with prospective follow-up.

PURPOSE: This study assessed the clinical and immunological outcomes of SARS-CoV-2-infected patients with risk factors for severe disease depending on their immunological status. METHODS: In this retrospective study with single follow-up visit, clinical outcome and humoral immunity was monitored in SARS-CoV-2 infected patients at risk. The results were compared based on the patients' initial immunological status: unvaccinated (UV), patients who did not develop neutralizing antibodies after vaccination (vaccine non-responders, VNR), and patients who expressed neutralizing antibodies after vaccination (vaccine responders, VR). Patients who lacked neutralizing antibodies (VNR and UV) were treated with nMABs. RESULTS: In total, 113 patients at risk of severe COVID-19 consented to participate in the study. VR and UV were not admitted to the hospital. During the observation period, UVs had the highest rate of SARS-CoV-2 re-infections. Three of 41 VNRs (7.3%) were hospitalized due to severe COVID-19, with two of them having undergone iatrogenic B-cell depletion. The humoral immune response after infection was significantly lower in the VNR group than in the VR group in terms of anti-N, anti-receptor-binding domain (RBD), anti-S antibody titers, and anti-S antibody avidity. In a sub-analysis of VNR, B cell-deficient non-responders had significantly lower levels of anti-N antibodies and anti-S avidity after infection than other VNRs. CONCLUSION: VNR, particularly B-cell-depleted VNR, remained at risk of hospitalization due to COVID-19. In the VR group, however, no clinical complications or severe disease were observed, despite not receiving nMAbs. Tailoring the administration of nMABs according to patient vaccination and immunological status may be advisable.

Lentiviral hepatitis B pseudotype entry requires sodium taurocholate co-transporting polypeptide and additional hepatocyte-specific factors.

Hepatitis B virus (HBV) is one of the world's major unconquered infections, resulting in progressive liver disease, and current treatments rarely cure infection. A limitation to discovering new therapies is our limited knowledge of HBV entry and dissemination pathways that hinders the development of in vitro culture systems. To address this gap in our understanding we optimized the genesis of infectious lentiviral pseudoparticles (HBVpps). The recent discovery that the bile salt transporter sodium taurocholate co-transporting polypeptide (NTCP) acts as a receptor for HBV enabled us to assess the receptor dependency of HBVpp infection. HBVpps preferentially infect hepatoma cells expressing NTCP, whereas other non-liver cells engineered to express NTCP do not support infection, suggesting that additional hepatocyte-specific factors are required for HBVpp internalization. These results highlight the value of the HBVpp system to dissect the pathways of HBV entry and dissemination.

Schistosome infection aggravates HCV-related liver disease and induces changes in the regulatory T-cell phenotype.

Schistosome infections are renowned for their ability to induce regulatory networks such as regulatory T cells (Treg) that control immune responses against homologous and heterologous antigens such as allergies. However, in the case of co-infections with hepatitis C virus (HCV), schistosomes accentuate disease progression and we hypothesized that expanding schistosome-induced Treg populations change their phenotype and could thereby suppress beneficial anti-HCV responses. We therefore analysed effector T cells and n/iTreg subsets applying the markers Granzyme B (GrzB) and Helios in Egyptian cohorts of HCV mono-infected (HCV), schistosome-co-infected (Sm/HCV) and infection-free individuals. Interestingly, viral load and liver transaminases were significantly elevated in Sm/HCV individuals when compared to HCV patients. Moreover, overall Treg frequencies and Helios(pos) Treg were not elevated in Sm/HCV individuals, but frequencies of GrzB(+) Treg were significantly increased. Simultaneously, GrzB(+) CD8(+) T cells were not suppressed in co-infected individuals. This study demonstrates that in Sm/HCV co-infected cohorts, liver disease is aggravated with enhanced virus replication and Treg do not expand but rather change their phenotype with GrzB possibly being a more reliable marker than Helios for iTreg. Therefore, curing concurrent schistosome disease could be an important prerequisite for successful HCV treatment as co-infected individuals respond poorly to interferon therapy.

Hepatitis B virus-specific T-cell responses during IFN administration in a small cohort of chronic hepatitis B patients under nucleos(t)ide analogue treatment.

The effect of pegylated interferon-α (IFN) add-on therapy on HBV-specific T-cell responses was evaluated in 12 patients with stable, undetectable hepatitis B virus (HBV) load under nucleos(t)ide analogue therapy. Peripheral blood mononuclear cells were isolated at week 0, 4, 8, 12, 24 and 48 of IFN add-on therapy. Quantity and quality of circulating HBV S- and core-specific CD4 and CD8 T cells were analysed ex vivo by flow cytometry. HBV S- and core-specific CD4 T-cell numbers modestly increased within 8 weeks of IFN administration (P = 0.0391 and P = 0.0195), whereas HBV-specific CD8 T cells in general showed only minor changes under IFN add-on therapy. Functionality of HBV-specific CD4 but not CD8 T cells positively correlated with serum transaminase activity. In addition, we observed an increase in CD4 T cells producing tumour necrosis factor-α (TNFα) without antigen restimulation (P = 0.0039), which correlated with elevated transaminases. During IFN add-on therapy, two patients developed an anti-HBs seroconversion, only one of whom showed a relevant increase in HBV-specific T cells. In conclusion, IFN add-on therapy of chronic hepatitis B increased HBV-specific T-cell responses and affected a previously unrecognized TNFα-monofunctional CD4 T-cell population. Although the observed T-cell responses did not correlate with HBsAg seroconversion, we expect additional insights into the immunopathogenesis of hepatitis B, following the characterization of the newly identified TNF α-monofunctional T-cell population.

Rapid point-of-care detection of SARS-CoV-2 infection in exhaled breath using ion mobility spectrometry: a pilot study.

BACKGROUND: An effective testing strategy is essential for pandemic control of the novel Coronavirus disease 2019 (COVID-19) caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Breath gas analysis can expand the available toolbox for diagnostic tests by using a rapid, cost-beneficial, high-throughput point-of-care test. We conducted a bi-center clinical pilot study in Germany to evaluate breath gas analysis using multi-capillary column ion mobility spectrometry (MCC-IMS) to detect SARS-CoV-2 infection. METHODS: Between September 23, 2020, and June 11, 2021, breath gas measurements were performed on 380 patients (SARS-CoV-2 real-time polymerase chain reaction (PCR) positive: 186; PCR negative: 194) presenting to the emergency department (ED) with respiratory symptoms. RESULTS: Breath gas analysis using MCC-IMS identified 110 peaks; 54 showed statistically significant differences in peak intensity between the SARS-CoV-2 PCR-negative and PCR-positive groups. A decision tree analysis classification resulted in a sensitivity of 83% and specificity of 86%, but limited robustness to dataset changes. Modest values for the sensitivity (74%) and specificity (52%) were obtained using linear discriminant analysis. A systematic search for peaks led to a sensitivity of 77% and specificity of 67%; however, validation by transferability to other data is questionable. CONCLUSIONS: Despite identifying several peaks by MCC-IMS with significant differences in peak intensity between PCR-negative and PCR-positive samples, finding a classification system that allows reliable differentiation between the two groups proved to be difficult. However, with some modifications to the setup, breath gas analysis using MCC-IMS may be a useful diagnostic toolbox for SARS-CoV-2 infection. TRIAL REGISTRATION: This study was registered at ClinicalTrials.gov on September 21, 2020 (NCT04556318; Study-ID: HC-N-H-2004).

[Interdisciplinary ultrasound-guided, minimally invasive autopsy in COVID-19-deceased patients in the intensive care unit of a university hospital : A proof-of-concept study]. / Interdisziplinäre ultraschallgesteuerte minimal-invasive Autopsie bei COVID-19-Verstorbenen auf der Intensivstation einer Universitätsklinik : Eine Proof-of-Concept-Studie.

In this feasibility study, we carried out in an interdisciplinary team standardised, ultrasound-guided, minimally invasive autopsy (US-MIA) directly at the bedside of patients who died of COVID-19 in the intensive care unit of the Rechts der Isar Hospital of the Technical University Munich (TUM). The aim of the study was to verify the feasibility, time efficiency and infection hygiene aspects of the process, as well as the quality of the tissue samples. Our results show that bedside US-MIA is suitable for obtaining tissue samples before the onset of postmortem autolysis, and that it can also be carried out quickly and safely. The potential of US-MIA, which has gained little recognition so far, deserves special attention in the context of postmortem diagnosis, research and quality assurance. In the future, these strengths of US-MIA could help to lead postmortem diagnosis into the modern age of pathological deep analytics ("omics").

Multidimensional Analysis of the Mother-to-child Transmission Risk Factors in Chronic Hepatitis B Virus Infection in Pregnant Women in Vietnam.

Background: Hepatitis B virus (HBV) infection remains a major public health problem. The interaction between HBV and the host inflammatory response is an important factor contributing to liver damage and disease development. We investigate of the correlation between peripheral blood cell levels, HBV DNA, and the risk of transmission to the baby in pregnant women infected with hepatitis B. Methods: A multidimensional analysis was performed on data collected from 60 Vietnamese pregnant women and their babies (cord blood). Results: Taking the risk ratio test results of cord blood HBsAg as a positive probability, the boundary of maternal PBMC concentration is 8.03x106 cells/ml (with negative correlation) and for CBMCs is 6.64x106 cells/ml (with positive correlation). That means that HBsAg positivity in the blood may be related to the increasing of CBMCs and the diminution of maternal PBMCs. When the maternal viral load is higher than 5x107 copies/ml, the risk of being HBsAg-positive in cord blood is 123% (RR=2.23 [1.48,3.36]); when the viral load is lower than this baseline, the risk is decreased by 55% (RR=0.45 [0.30,0.67]) (p<0.001). Conclusions: With several steps of the analysis, this study found maternal peripheral blood cell levels and cord blood positively cor-related in pregnant women with a load lower than 5x107 copies of HBV DNA/ml. The study's results suggest that the role of PBMCs and HBV DNA in vertical infection is essential.

Sequential control of hepatitis B virus in a mouse model of acute, self-resolving hepatitis B.

The determinants of an immune response to the human hepatitis B virus (HBV) are poorly understood. As studies in man and chimpanzees are limited, we aimed at developing a model of self-limiting hepatitis B in mice that helps to dissect the control of HBV by humoral and cellular immune responses. Adenoviral vectors containing 1.3-fold HBV genomes allowed an efficient and reproducible transfer of HBV genomes into mouse livers and initiated HBV replication in mice. HBV transcripts were detected in mouse livers for more than 3 months. HBsAg and HBeAg peaked around day 6 and slowly declined thereafter. A two-phase mild to moderate liver inflammation with elevated serum alanine transaminase activities was observed around day 7 and around day 70 when the vast majority of HBV-specific T cells were detected in the liver. HBV was initially controlled when specific and nonspecific T cells infiltrated the liver and intrahepatic interferon γ levels peaked around day 7, but replicated again from day 10 to day 24 and persisted at low levels thereafter despite the presence of HBV-specific T cells. Finally, HBV replication was terminated after a sufficient B-cell response had been mounted- indicated by anti-HBs seroconversion around day 35. HBV-specific T cells infiltrated the liver a second time around day 70 postinfection. This demonstrates that the established mouse model allows studying the onset and termination of HBV infection and will help to dissect the determinants of HBV control and clearance by the immune response.

A proof of concept study for the differentiation of SARS-CoV-2, hCoV-NL63, and IAV-H1N1 in vitro cultures using ion mobility spectrometry.

Rapid, high-throughput diagnostic tests are essential to decelerate the spread of the novel coronavirus disease 2019 (COVID-19) pandemic. While RT-PCR tests performed in centralized laboratories remain the gold standard, rapid point-of-care antigen tests might provide faster results. However, they are associated with markedly reduced sensitivity. Bedside breath gas analysis of volatile organic compounds detected by ion mobility spectrometry (IMS) may enable a quick and sensitive point-of-care testing alternative. In this proof-of-concept study, we investigated whether gas analysis by IMS can discriminate severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from other respiratory viruses in an experimental set-up. Repeated gas analyses of air samples collected from the headspace of virus-infected in vitro cultures were performed for 5 days. A three-step decision tree using the intensities of four spectrometry peaks correlating to unidentified volatile organic compounds allowed the correct classification of SARS-CoV-2, human coronavirus-NL63, and influenza A virus H1N1 without misassignment when the calculation was performed with data 3 days post infection. The forward selection assignment model allowed the identification of SARS-CoV-2 with high sensitivity and specificity, with only one of 231 measurements (0.43%) being misclassified. Thus, volatile organic compound analysis by IMS allows highly accurate differentiation of SARS-CoV-2 from other respiratory viruses in an experimental set-up, supporting further research and evaluation in clinical studies.

MVA-nef induces HIV-1-specific polyfunctional and proliferative T-cell responses revealed by the combination of short- and long-term immune assays.

Several vaccination trials are evaluating the modified vaccinia virus Ankara (MVA) as a delivery vector in various clinical settings. In this paper, we present the reevaluation of a therapeutic vaccination trial in human immunodeficiency virus (HIV)-1-infected individuals treated with highly active antiretroviral therapy using MVA-expressing HIV-1 nef. Immunogenicity of MVA-nef was assessed using multicolor flow cytometry. Vaccine-induced polyfunctionality and proliferative capacity, which are associated with nonprogressive HIV-1 infection, were detectable by combining two immune assays. By means of short-term polychromatic intracellular cytokine staining, we observed a significant increase in polyfunctional Nef-specific CD4 T cells expressing interferon-γ, interleukin (IL)-2 and CD154 after vaccination, whereas changes in the quality of CD8 T-cell response could not be observed. Only the additional use of a long-term polychromatic Carboxyfluorescein succinimidyl ester (CFSE)-based proliferation assay revealed vaccine-induced Nef-specific CD8, as well as CD4 T cells with proliferative capacity. The correlation between vaccine-induced IL-2 production by CD4 T cells and the increase in proliferating Nef-specific CD8 T cells suggests a causal link between these two functions. These results highlight the importance of combining sophisticated immunomonitoring tools to unravel concealed effects of immunological interventions and support the use of the poxvirus-derived MVA vector to stimulate highly functional HIV-1-specific T-cell responses. However, the clinical benefit of these functional T cells remains to be determined.

Control of hepatitis B virus at the level of transcription.

Hepatitis B virus (HBV) is tightly controlled by a number of noncytotoxic mechanisms. This control occurs within the host hepatocyte at different steps of the HBV replication cycle. HBV persists by establishing a nuclear minichromosome, HBV cccDNA, serving as a transcription template for the viral pregenome and viral mRNAs. Nucleoside/nucleotide analogues widely used for antiviral therapy as well as most antiviral cytokines act at steps after transcription of HBV RNAs and thus can control virus replication but do not directly affect its gene expression. Control of HBV at the level of transcription in contrast is able to restrict both, HBV replication and gene expression. In the review, we focus on how HBV is controlled at the level of transcription. We discuss how the composition of transcription factors determines HBV gene expression and replication and how this may be influenced by antivirally active substances, e.g. the cytokine IL-6 or helioxanthin analogues, or by the differentiation state of the hepatocyte.

Antiviral gene therapy.

This chapter describes the major gene therapeutic approaches for viral infections. The vast majority of published approaches target severe chronic viral infections such as hepatitis B or C and HIV infection. Two basic gene therapy strategies are introduced here. The first involves the expression of a protein or an RNA that inhibits viral replication by targeting crucial steps of the viral life cycle or by interfering with a cellular factor required for virus replication. The major limitation of this approach is that primary levels of gene modification have generally not been sufficient to reduce the availability of target cells permissive for virus replication to a level that significantly decreases overall viral load. Thus, investigators have banked on the expectation that gene-protected cells have a sufficient selective advantage to accumulate and gain prevalence over time, a prediction that so far could not be confirmed in clinical trials. In vivo levels of gene modification can be improved, however, by introducing an additional selectable marker. In addition, a secreted antiviral gene product that exerts a bystander effect could significantly reduce overall virus replication despite relatively low levels of gene modification. In addition to these direct antiviral approaches, several strategies have been developed that employ or aim to enhance host immune responses. The innate immune response has been enhanced, for example, by the in vivo expression of interferons. Alternatively, T cells can be grafted with recombinant receptors to boost adaptive virus-specific immunity. These approaches are especially promising for chronic virus infection, where natural immune responses are evidently not sufficient to effectively control virus replication.

Liver sinusoidal endothelial cells are not permissive for adenovirus type 5.

Adenoviral vectors are known to transduce hepatocytes in normal liver tissue with high efficiency. The aim of this study was to investigate whether sinusoidal endothelial cells, which separate hepatocytes from the bloodstream in the sinusoidal lumen, are permissive for infection by adenoviruses. We show here that microvascular liver sinusoidal endothelial cells are not infected by adenovirus type 5 in vivo or in vitro unless high MOIs are used. In contrast, macrovascular endothelial cells from aorta are efficiently infected by adenovirus type 5. In addition, Kupffer cells, similar to sinusoidal endothelial cells, are not infected by adenovirus type 5. Liver sinusoidal endothelial cells do not express the integrin receptor alpha(v)beta3, which is required for efficient infection by adenoviruses. Our results demonstrate that hepatocytes are the main cell population of the liver that is infected by adenovirus type 5.

[Follow-up of Crohn's disease: can hydro-MRI replace fractionated gastrointestinal passage examination?]. / Verlaufskontrolle des Morbus Crohn: Kann die Hydro-MRT die fraktionierte Magen-Darm-Passage ersetzen?

PURPOSE: To compare the value of hydro-MRI with follow-through examinations in the follow-up of Crohn's disease. METHOD: 22 patients known to be suffering from Crohn's disease were examined via 1.5 T-MR system; an oral contrast examination using 1000 ml of a 2.5% mannitol solution was performed in all patients. T2-weighted TSE sequences and T1-weighted SE sequences were performed before and after the intravenous injection of Gd-DTPA. To reduce movement artifacts caused by peristalsis of the gut, intravenous injection of 40 mg Buscopan was given. The findings of hydro-MRI were compared with the follow-through examinations. RESULTS: In the upper gastrointestinal tract, the follow-through examination showed clear advantages compared with hydro-MRI for the demonstration of inflammatory changes in the gut; Hydro-MRI was, however, somewhat more reliable in the ileum and colon. It was also more sensitive than the follow-through for the demonstration of enteric fistulae (four as compared with two cases), and in demonstration extraluminal changes (free fluid in six against zero, and inflammatory adherent loops (four against zero)). Amongst the 22 patients, hydro-MRI was equal (in 10) or better (in 8) than the follow-through examination for demonstrating the intestinal manifestations of Crohn's disease, and follow-through was better in only four. CONCLUSION: For follow-up of Crohn's disease, hydro-MRI is at least as good as follow-through examination, and is even preferable, because of the absence of radiation exposure of the usually young patients.

[Enteropathic spondylarthritis in chronic inflammatory bowel diseases: prevalence, manifestation pattern and HLA association]. / Enteropathische Spondarthritiden bei chronisch entzündlichen Darmerkrankungen: Prävalenz, Befallsmuster und HLA-Assoziation.

BACKGROUND: Enteropathic spondylarthropathies (SpA) are the most frequent extraintestinal manifestation of the chronic inflammatory bowel disease (IBD), Crohn's disease (CD) and Ulcerative Colitis (UC). It was the aim of the present study, to analyze a large number of IBD patients for the prevalence and pattern of joint manifestation and the association of SpA with the extend of bowel involvement and HLA-haplotype. PATIENTS AND METHODS: 521 patients (409 CD and 112 UC) were prospectively analyzed over a period of one year. SpA was diagnosed on the basis of an appropriate patient history as well as clinical, radiological and immunoserological parameters. RESULTS: SpA was diagnosed in 10.7% of all CD and 14.4% of all UC patients. In 26.8% of all patients symptoms of SpA occurred prior to and in 14.4% simultaneously with IBD. 28.1% of all patients presented with isolated peripheral arthritis, 26.8% of all patients showed an isolated involvement of the spine or sacroiliic joints and 45.1% of all patients presented with combined involvement. 2/12 UC patients with SpA suffered from rectosigmoiditis, 5/12 from partial colitis and 5/12 had pancolitis. In CD patients with SpA, 8/59 had isolated colitis, 8/59 ileocolitis and 31/59 isolated small bowel involvement. There was a positive correlation between SpA and HLA-B27 (p < 0.01). CONCLUSION: Enteropathic spondylarthropathies are an important extraintestinal manifestation of IBD. Spondylarthropathies occur irrespective of the extend of IBD and frequently become symptomatic prior to IBD. These and recent data describing inflammatory bowel disease in patients with SpA of unknown etiology suggest that both diseases have a common pathogenetic background.