Background:Disease progression from human immunodeficiency virus (HIV) infection to acquired immunodeficiency syndrome (AIDS) is marked by chronic immune activation, partly due to increased translocation of gut-derived microbial antigens. Elevated mucosal tumour necrosis factor-α (TNF-α) and resulting epithelial cell apoptosis may be the etiology. However, studies using carcinoma cell lines have failed to find a causal link, possibly due to cellular abnormalities related to the malignant transformation of these immortal cell lines. Methods:We established intestinal organoid monolayers from healthy controls and HIV-infected adults and characterized their growth dynamics and cellular composition. We then examined the effects of HIV-associated cytokines on transepithelial resistance (TER), apoptosis and macromolecule translocation. Results:Organoid monolayers from HIV-infected patients grew similarly to healthy controls, forming confluent monolayers within one to two weeks containing enterocytes, Paneth, goblet and stem cells. IFN-γ synergized with TNF-α, allowing TNF-α to cause caspase-mediated apoptosis and TER reduction within 5 ± 3 hours, reflecting patient sample heterogeneity. This led to paracellular passage of 4 kDa Dextran and transcytosis of 44 kDa horse radish peroxidase, both of which could be blocked by pan-caspase inhibitor, Q-VD-Oph. Conclusion:Our study confirms that intestinal organoid monolayers from biopsies of HIV-infected individuals can be used to study apoptosis-related epithelial barrier dysfunction and macromolecular translocation. We provide direct evidence that TNF-α-induced apoptosis triggered two pathways of macromolecular translocation: paracellular passage via apoptotic leaks and transcytosis. Therapies targeting apoptosis may be useful in preventing disease progression from HIV to AIDS.

BackgroundTraditionally, transformed cell line monolayers have been the standard model for studying epithelial barrier and transport function. Recently, intestinal organoids were proposed as superior in recapitulating the intestine. Typically, 3D organoids are digested and seeded as monolayers on gelatinous matrix pre-coated surfaces for anchorage. As this coat could potentially act as a diffusion barrier, we aimed to generate robust human duodenum-derived organoid monolayers that do not need a gelatinous matrix for anchorage to improve upon existing models to study epithelial transport and barrier function.ResultsWe characterized these monolayers phenotypically regarding polarization, tight junction formation and cellular composition, and functionally regarding uptake of nutrients, ion transport and cytokine-induced barrier dysfunction. The organoid monolayers recapitulated the duodenum phenotypically as well as functionally regarding glucose and short-chain fatty acid uptake. Tumour necrosis factor-alpha induced paracellular transport of 4-kDa Dextran and transcytosis of 44-kDa horseradish peroxidase. Notably, forskolin-stimulated chloride secretion was consistently lower when organoid monolayers were seeded on a layer of basement membrane extract (BME).ConclusionsBME-free organoid monolayers represent an improved model for studying transcytotic, paracellular but especially transcellular transport. As BME is extracted from mice, our model furthers efforts to make organoid culture more animal-free.

Das Ziel der interdisziplinären S2k-Leitlinie „Akute infektiöse Gastroenteritis im Säuglings-, Kindes- und Jugendalter“ ist es, den aktuellen Kenntnisstand zu klinischem Bild, Diagnostik, Therapie, Prävention und Hygiene der akuten infektiösen Gastroenteritis, einschließlich der nosokomialen gastrointestinalen Infektionen, bei Säuglingen, Kindern und Jugendlichen auf Basis der wissenschaftlichen Evidenz zusammenzufassen, im Expertenkonsens zu bewerten und daraus praxisrelevante Empfehlungen abzuleiten. Die Leitlinie liefert einen Handlungskorridor für häufige Entscheidungen. Sie dient zudem der evidenzbasierten Fort- und Weiterbildung und soll damit eine Verbesserung der medizinischen Versorgung von Kindern mit akuter Gastroenteritis erreichen. Insbesondere sollen durch die Leitlinie unnötige stationäre Aufnahmen von Kindern mit AGE vermieden und präventive Maßnahmen zur Infektionsvermeidung und -ausbreitung ergriffen werden.

Einführung Die Clostridioides-difficile-Infektion (CDI) ist mit einer hohen Morbidität und Mortalität assoziiert. Auch wenn ihre Inzidenz in den letzten Jahren in Deutschland rückläufig ist, kann die individuelle Infektion trotz therapeutischen Fortschritts eine medizinische Herausforderung darstellen. Hier soll geklärt werden, welche Lücken verantwortliche Behandler/-innen in der Versorgung und in der Evidenzlage als besonders gravierend ansehen. Methoden In einem moderierten Workshop von in Deutschland tätigen CDI-Experten/-innen, wurden als relevant eingeschätzte Fragestellungen ermittelt. Eine bereits in fünf anderen Ländern (Australien, Frankreich, Großbritannien, Kanada und Italien) durchgeführte Befragung wurde ergänzend angepasst und von 27 Behandlern/-innen bearbeitet. Bei der Auswertung wurden die als besonders wichtig empfundenen Themen herausgearbeitet, die Aussagen der Fachgruppen verglichen und Meinungsänderungen berücksichtigt. Ergebnisse 27 vollständig ausgefüllte Fragebögen wurden ausgewertet. Verbesserungsbedarf wurde vorrangig bei der Verhinderung von CDI-Rezidiven (74,1%), sowie Therapie der Rezidive (55,6%) gesehen. Evidenzdefizite wurden bei der Behandlung von Rezidiven (55,6%) und der Identifikation der Risikofaktoren für Rezidive konstatiert (48,1%). Die Verbesserung der Versorgung mittels fäkalem Mikrobiota-Transfer (FMT) nannten 70,4%. Bei Leitlinien wurde mehr Klarheit (48,1%) und regelmäßigere Aktualisierung (40,7%) gewünscht. Für Patienten/-innen wurden bessere Aufklärung der über angemessenen Antibiotikaeinsatz (52,0%) und Wahlmöglichkeiten der FMT angestrebt (48,1%). Zusammenfassung Bei der Frage nach Verbesserungsbedarf in der Versorgung und Evidenzlücken bei der Behandlung von Patienten/-innen mit CDI ist die nationale Experten/-innensicht in Deutschland, wie auch die internationale Einschätzung ähnlich: Im Zentrum steht die Prävention und Therapie der rezidivierenden CDI. Die Problematik des Zugangs zum FMT ist eine deutsche Besonderheit, die verbesserungspflichtig erscheint.

Introduction Clostridioides difficile infection (CDI), as a nosocomial disease, is associated with high morbidity and mortality. Even though the incidence of CDI has been declining in Germany in recent years, the individual infection may pose a medical challenge despite therapeutic advances. The aim here is to clarify which gaps practitioners consider to be particularly serious in care and in the existing evidence base. Methods In a moderated workshop of German CDI experts the topics considered as relevant were identified. A survey already conducted in five other countries (Australia, France, Great Britain, Canada, and Italy) was adapted and processed by 27 practitioners. During the evaluation, the topics perceived as particularly important were identified, the statements of the specialist groups were compared and changes in opinion were considered. Results 27 fully completed questionnaires were evaluated. The need for improvement was primarily seen in the prevention of CDI recurrences (74.1%) and the treatment of recurrences (55.6%). Evidence deficits were noted in the treatment of recurrences (55.6%) and identification of risk factors for recurrences (48.1%). Improving care via fecal microbiota transfer (FMT) was named by 70.4%. For guidelines, more clarity (48.1%) and more regular updates (40.7%) were desired. For patients, better education on appropriate antibiotic use (52.0%) and choice of FMT were desired (48.1%). Summary The German expert view and the international assessment is similar, when asked about the need for improvement in care and evidence gaps in the treatment of patients with CDI: The focus is on prevention and therapy of recurrent CDI. The problem of access to FMT is a German peculiarity that seems to need improvement.

Background: Tropheryma whipplei (TW) can cause different pathologies, e.g., Whipple’s disease and transient gastroenteritis. The mechanism by which the bacteria pass the intestinal epithelial barrier, and the mechanism of TW-induced gastroenteritis are currently unknown. Methods: Using ex vivo disease models comprising human duodenal mucosa exposed to TW in Ussing chambers, various intestinal epithelial cell (IEC) cultures exposed to TW and a macrophage/IEC coculture model served to characterize endocytic uptake mechanisms and barrier function. Results: TW exposed ex vivo to human small intestinal mucosae is capable of autonomously entering IECs, thereby invading the mucosa. Using dominant-negative mutants, TW uptake was shown to be dynamin- and caveolin-dependent but independent of clathrin-mediated endocytosis. Complementary inhibitor experiments suggested a role for the activation of the Ras/Rac1 pathway and actin polymerization. TW-invaded IECs underwent apoptosis, thereby causing an epithelial barrier defect, and were subsequently subject to phagocytosis by macrophages. Conclusions: TW enters epithelia via an actin-, dynamin-, caveolin-, and Ras-Rac1-dependent endocytosis mechanism and consecutively causes IEC apoptosis primarily in IECs invaded by multiple TW bacteria. This results in a barrier leak. Moreover, we propose that TW-packed IECs can be subject to phagocytic uptake by macrophages, thereby opening a potential entry point of TW into intestinal macrophages.

Based on indirect evidence, increased mucosal translocation of gut-derived microbial macromolecules has been proposed as an important pathomechanism in HIV infection. Here, we quantified macromolecule translocation across intestinal mucosa from treatment-naive HIV-infected patients, HIV-infected patients treated by combination antiretroviral therapy, and HIV-negative controls and analyzed the translocation pathways involved. Macromolecule permeability was quantified by FITC-Dextran 4000 (FD4) and horseradish peroxidase (HRP) flux measurements. Translocation pathways were addressed using cold inhibition experiments. Tight junction proteins were characterized by immunoblotting. Epithelial apoptosis was quantified and translocation pathways were further characterized by flux studies in T84 cell monolayers using inducers and inhibitors of apoptosis and endocytosis. In duodenal mucosa of untreated but not treated HIV-infected patients, FD4 and HRP permeabilities were more than a 4-fold increase compared to the HIV-negative controls. Duodenal macromolecule permeability was partially temperature-dependent and associated with epithelial apoptosis without altered expression of the analyzed tight junction proteins. In T84 monolayers, apoptosis induction increased, and both apoptosis and endocytosis inhibitors reduced macromolecule permeability. Using quantitative analysis, we demonstrate the increased macromolecule permeability of the intestinal mucosa in untreated HIV-infected patients. Combining structural and mechanistic studies, we identified two pathways of increased macromolecule translocation in HIV infection: transcytosis and passage through apoptotic leaks.

Despite reports of a declining incidence over the last decade, Clostridioides difficile infection (CDI) is still considered the most important healthcare-associated causes of diarrhea worldwide. In Germany, several measures have been taken to observe, report, and influence this development. This report aims to analyze the development of hospital coding for CDI in Germany over the last decade and to use it to estimate the public health burden caused by CDI. Reports from the Institute for Hospital Remuneration Systems, German Federal Statistical Office (DESTATIS), the Robert-Koch-Institute (RKI), Saxonian authorities and hospital quality reports during 2010–2021 were examined for CDI coding and assessed in a structured expert consultation. Analysis was performed using 2019 versions of Microsoft Excel® and Microsoft Access®. Peaks of 32,203 cases with a primary diagnosis (PD) of CDI and 78,648 cases with a secondary diagnosis (SD) of CDI were observed in 2015. The number of cases had decreased to 15,412 PD cases (− 52.1

A recently developed haemostatic peptide gel for endoscopic application has been introduced to improve the management of gastrointestinal bleeding. The aim of this pilot study was to evaluate the feasibility, safety, efficacy and indication profiles of PuraStat in a clinical setting. In this prospective observational multicentre pilot study, patients with acute non-variceal gastrointestinal bleeding (upper and lower) were included. Primary and secondary application of PuraStat was evaluated. Haemoglobin, prothrombin time, platelets and transfusion behaviour were documented before and after haemostasis. The efficacy of PuraStat was assessed during the procedure, at 3 days and 1 week after application. 111 patients with acute gastrointestinal bleeding were recruited into the study. 70 percent (78/111) of the patients had upper gastrointestinal bleeding and 30% (33/111) had lower gastrointestinal bleeding. After primary application of PuraStat, initial haemostatic success was achieved in 94% of patients (74/79, 95% CI 88–99%), and in 75% of the patients when used as a secondary haemostatic product, following failure of established techniques (24/32, 95% CI 59–91%). The therapeutic success rates (absence of rebleeding) after 3 and 7 days were 91% and 87% after primary use, and 87% and 81% in all study patients. Overall rebleeding rate at 30 day follow-up was 16% (18/111). In the 5 patients who finally required surgery (4.5%), PuraStat allowed temporary haemostasis and stabilisation. PuraStat expanded the therapeutic toolbox available for an effective treatment of gastrointestinal bleeding sources. It could be safely applied and administered without complications as a primary or secondary therapy. PuraStat may additionally serve as a bridge to surgery in order to achieve temporary haemostasis in case of refractory severe bleeding, possibly playing a role in preventing immediate emergency surgery.

Background Ineffective antibiotic therapy increases mortality of acute cholangitis. The choice of antibiotics should reflect local resistance patterns and avoid the overuse of broad-spectrum agents. In this study, we analysed how results of bile and blood cultures and patient data can be used for selection of empirical antibiotic therapy in acute cholangits. Methods Pathogen frequencies and susceptibility rates were determined in 423 positive bile duct cultures and 197 corresponding blood cultures obtained from 348 consecutive patients with acute cholangitis. Patient data were retrieved from the medical records. Associations of patient and microbiological data were assessed using the Chi-2 test and multivariate binary logistic regression. Results In bile cultures, enterobacterales and enterococci were isolated with equal frequencies of approximately 30% whereas in blood cultures, enterobacterales predominated (56% compared to 21% enterococci). Antibiotic resistance rates of enterobacterales were > 20% for fluorochinolones, cephalosporines and acylureidopenicillins but not for carbapenems (< 2%). The efficacy of empirical therapy was poor with a coverage of bacterial bile and blood culture isolates in 51 and 69%, respectively. By multivariate analysis, predictors for pathogen species, antibiotic susceptibility and expected antibiotic coverage were identified. Conclusions In unselected patients treated for acute cholangitis in a large tertiary refferential center, use of carbapenems seems necessary to achieve a high antibiotic coverage. However, by analysis of patient and microbiological data, subgroups for highly effective carbapenem-sparing therapy can be defined. For patients with community-acquired cholangitis without biliary prosthesis who do not need intensive care, piperacillin/tazobactam represents a regimen with an expected excellent antibiotic coverage.

Abkürzungsverzeichnis AAD: antibiotikaassoziierte Diarrhoe AGE: akute Gastroenteritis AWMF: Arbeitsgemeinschaft der Wissenschaftlichen Medizinischen Fachgesellschaften e. V. BfR: Bundesinstitut für Risikobewertung BSG: Blutkörperchensenkungsgeschwindigkeit BVKJ: Berufsverband der Kinder- und Jugendärzte e. V. CDAD: Clostridoides-difficile-assoziierte Diarrhoe CDC: Centers for Disease Control and Prevention CDI: Clostridoides-difficile-Infektion CED: chronisch-entzündliche Darmerkrankung CRP: C-reaktives Protein DGHM: Deutsche Gesellschaft für Hygiene und Mikrobiologie DGKH: Deutsche Gesellschaft für Krankenhaushygiene e. V. DGKJ: Deutsche Gesellschaft für Kinder- und Jugendmedizin e. V. DGPI: Deutsche Gesellschaft für Pädiatrische Infektiologie e. V. DGPW: Deutsche Gesellschaft für Pflegewissenschaften e. V. DGVS: Deutsche Gesellschaft für Gastroenterologie, Verdauungs- und Stoffwechselerkrankungen e. V. EAEC: enteroaggregative Escherichia coli EHEC: enterohämorrhagische Escherichia coli EIA: enzymgekoppelter Immunadsorptionstest (ELISA): (engl. Enzyme-Linked Immunosorbent-Assay) EIEC: enteroinvasive Escherichia coli EPEC: enteropathogene Escherichia coli ETEC: enterotoxinbildende Escherichia coli GPGE: Gesellschaft für Pädiatrische Gastroenterologie und Ernährung e. V. HUS: hämolytisch-urämisches Syndrom IDSA: Infectious Diseases Society of America IfSG: Infektionsschutzgesetz KISS: Krankenhaus-Infektions-Surveillance-System des Nationalen Referenzzentrums für Surveillance von noskomialen Infektionen am RKI KRINKO: Kommission für Krankenhaushygiene und Infektionsprävention LGG: Lactobacillus GG NICE: National Institute for Health and Clinical Excellence (UK) MTS: Manchester-Triage-System ÖGKJ: Österreichische Gesellschaft für Kinder- und Jugendheilkunde ORL: orale Rehydrationslösung ORT: orale Rehydrationstherapie RCT: Randomized Controlled Trial RKI: Robert-Koch-Institut SBS: Säure-Basen-Status SIRS: systemisches inflammatorisches Response-Syndrom STEC: Shigatoxin-produzierende Escherichia coli STIKO: Ständige Impfkommission am RKI STX: Shigatoxin VRE: Vancomycin-resistente Enterokokken VTEC: Verotoxin-produzierende E. coli ZNS: Zentralnervensystem

Whereas statistical association of hepatitis C virus (HCV) infection with cardiomyopathy is long known, establishment of a causal relationship has not been achieved so far. Patients with advanced heart failure (HF) are mostly unable to tolerate interferon (IFN)-based treatment, resulting in limited experience regarding the possible pathogenic role of HCV in this patient group. HCV infection often triggers disease in a broad spectrum of extrahepatic organs, with innate immune and autoimmune pathogenic processes involved. The fact that worldwide more than 70 million patients are chronically infected with HCV illustrates the possible clinical impact arising if cardiomyopathies were induced or aggravated by HCV, resulting in progressive HF or severe arrhythmias. A novel path has been opened to finally resolve the long-standing question of cause-effect relationship between HCV infection and cardiac dysfunction, by the recent development of IFN-free, highly efficient, and well tolerable anti-HCV regimens. The new direct-acting antiviral (DAA) agents are highly virus-specific and lack unspecific side-effects upon cardiac function which have always confounded the interpretation of IFN treatment data. The actual frequency of unexplained HF in chronic HCV infection will be determined from a planned large-scale study. Whereas such patients probably constitute a rather small fraction of all those harboring HCV, they have major clinical relevance. It is not yet known which fraction of these patients will significantly benefit from HCV eradication, but this issue will be addressed now in a prospective study.

Hepatitis C virus (HCV) has been associated with cardiomyopathies. Former anti-HCV therapies employing interferon could have serious side effects in patients with advanced heart failure since interferon may adversely impact upon cardiac function. We, therefore, examined whether the novel, interferon-free and highly virus-selective anti-HCV combination therapy might be applicable even in advanced or end-stage heart failure.

During six years as German National Consultant Laboratory for Spirochetes we investigated 149 intestinal biopsies from 91 patients, which were histopathologically diagnosed with human intestinal spirochetosis (HIS), using fluorescence in situ hybridization (FISH) combined with 16S rRNA gene PCR and sequencing. Aim of this study was to complement histopathological findings with FISH and PCR for definite diagnosis and species identification of the causative pathogens. HIS is characterized by colonization of the colonic mucosa of the human distal intestinal tract by Brachyspira spp. Microbiological diagnosis of HIS is not performed, because of the fastidious nature and slow growth of Brachyspira spp. in culture. In clinical practice, diagnosis of HIS relies solely on histopathology without differentiation of the spirochetes. We used a previously described FISH probe to detect and identify Brachyspira spp. in histological gut biopsies. FISH allowed rapid visualization and identification of Brachyspira spp. in 77 patients. In most cases, the bright FISH signal already allowed rapid localization of Brachyspira spp. at 400× magnification. By sequencing, 53 cases could be assigned to the B. aalborgi lineage including "B. ibaraki" and "B. hominis", and 23 cases to B. pilosicoli. One case showed mixed colonization. The cases reported here reaffirm all major HIS Brachyspira spp. clusters already described. However, the phylogenetic diversity seems to be even greater than previously reported. In 14 cases, we could not confirm HIS by either FISH or PCR, but found colonization of the epithelium by rods and cocci, indicating misdiagnosis by histopathology. FISH in combination with molecular identification by 16S rRNA gene sequencing has proved to be a valuable addition to histopathology. It provides definite diagnosis of HIS and allows insights into phylogeny and distribution of Brachyspira spp. HIS should be considered as a differential diagnosis in diarrhea of unknown origin, particularly in patients from risk groups (e.g. patients with colonic adenomas, inflammatory polyps, inflammatory bowel disease or HIV infection and in men who have sex with men).

CD8+ T cells in the intestinal mucosa influence the HIV-associated pathogenesis, but little is known about the dynamics of mucosal CD8+ T cell counts and activation of these cells during the course of infection. In this study, mucosal CD8+ T cells in the duodenum were studied at different stages of HIV infection, starting from the seronegative phase. In seronegative acute HIV infection, CD8+ T cell counts increased in the epithelium, but not in the lamina propria. Infiltration of the lamina propria by peripherally expanded CD8+ T cells was observed after seroconversion. Highest increase in the expression of perforin, the rate-limiting molecule for cytotoxic CD8+ T cell activity, was evident in the lamina propria of seronegative acutely HIV-infected patients. The number of perforin-expressing cells in the lamina propria of acutely HIV-infected patients was positively associated with biomarkers of enterocyte damage and microbial translocation. After seroconversion, perforin expression was downregulated in the lamina propria, but not in the epithelium. In conclusion, our findings demonstrate that intraepithelial and lamina propria CD8+ T cells exhibit different dynamics of numerical alteration and cytotoxic activity in HIV-infected patients. Moreover, our results suggest that perforin-dependent cytotoxic mechanisms by CD8+ T cells could impair the intestinal mucosal barrier already in the seronegative phase of acute HIV infection, thereby inducing microbial translocation as one of the earliest pathological events in HIV infection.

International guidelines recommend lifelong nucleos(t)ide analogue (NA) treatment in individuals with chronic hepatitis B (CHB) infection who are hepatitis B e antigen (HBeAg) seronegative, because hepatitis B surface antigen (HBsAg) seroconversion is rarely achieved. However, after terminating therapy, sustained responses and HBsAg loss have been observed. Clinical characteristics identifying persons with favorable outcomes after discontinuing NA therapy have not yet been defined. This case series describes outcomes of 6 individuals with HBeAg-seronegative CHB infection without cirrhosis and low plasma levels of HBsAg who discontinued long-term NA treatment. All individuals had a virologic relapse and 4 of 6 had a biochemical relapse; but 5 of 6 later developed a sustained virologic and biochemical response and a marked reduction of quantitative HBsAg (qHBsAg). Two of the 6 individuals experienced HBsAg loss. Only 1 patient was retreated, and none showed signs of hepatic decompensation. NA treatment can be safely stopped in selected HBeAg-seronegative patients. Sustained offtreatment responses seem to be frequently preceded by a virologic and biochemical flare. Loss of HBsAg possibly reflects restoration of antiviral immunity during prolonged NA treatment. Predictive factors, such as qHBsAg, may be valuable in selecting patients who could benefit from NA discontinuation.

BACKGROUND AND STUDY AIMS:Hemostatic powders have been introduced to improve the management of gastrointestinal (GI) bleeding and to extend the variety of tools available for emergency endoscopy. The aim of the present pilot study was to evaluate the indication profiles and the short-term outcome of EndoClot.PATIENTS, MATERIALS AND METHODS:In a prospective observational pilot study patients with acute nonvariceal GI bleeding were included. Primary or secondary application of EndoClot was assessed. Hemoglobin, prothrombine time and platelets were documented before and after hemostasis. The efficacy of EndoClot was assessed 72 hours and 1 week after application.RESULTS:Seventy patients with acute GI bleeding were recruited into the study. Eighty-three percent (58/70) of the patients had upper and 17% (12/70) had lower GI bleeding. In the upper GI tract treatment success was achieved in 64% (30/47, 95% confidence interval, 50%-76%) after primary use and in all patients, when used after established techniques had failed (95% confidence interval, 70%-100%). In lower GI bleeding hemostasis was achieved in 83% of cases (10/12, 95% confidence interval 54%-97%). Rebleeding occurred in 11% (8/70), in 10% EndoClot served as a bridge to surgery (7/70).CONCLUSIONS:EndoClot expanded the therapeutic options in the management of GI bleeding. It was applicable as a monotherapy or in combination with other techniques from oozing bleeding type or lower. It was most effective in diffuse or extensive bleeding activity or when access to the bleeding vessel was difficult. EndoClot can be applied as a bridge to surgery when classical methods of hemostasis have failed.