Diagnostic of the pathogenic bacterium Pseudomonas aeruginosa by bacteriophages

Pseudomonas aeruginosa ist ein opportunistisches Humanpathogen, das Infektionen in Atemwegen, Harnwegen und Wunden verursachen kann. Es stellt ein Gesundheitsproblem bei der Produktion von Nahrungsmitteln und Kosmetikprodukten dar. Das Ziel dieser Arbeit war die Entwicklung eines Diagnostik-Systems zum Nachweis lebender P. aeruginosa Bakterien durch Bakteriophagen. Prinzipiell sollten P. aeruginosa-spezifische Bakteriophagen immobilisiert auf Nanopartikeln auf mit P. aeruginosa kontaminierten Proben appliziert werden. Die dabei in den P. aeruginosa Zellen entstehende Phagen-DNA sollte amplifiziert und nachgewiesen werden. Zuerst konnte gezeigt werden, dass die Phagen JG012, JG054 und JG057 ihren P. aeruginosa Wirt über Pili-Strukturen erkennen und eine Vielzahl an P. aeruginosa-Isolaten aus CF-Patienten und Harnwegsinfektionen infizieren können. Zudem konnte dsDNA identifiziert und die Phagen JG012 und JG054 der Familie Siphoviridae und JG057 der Familie Podoviridae zugeordnet werden. Von vier zur Immobilisierung getesteten Nanopartikeln mit verschiedenen Oberflächen-funktionen zeigten die Dynabeads® M-270 Epoxy für den Phagen JG012 und die Dynabeads® M-280 Streptavidin für den Phagen JG024 die besten Ergebnisse. Zudem konnte ein im Vergleich zum freien Phagen unverändertes Infektionsverhalten bestätigt werden. Die Phagen-DNA Präparation und Amplifizierung wurde erfolgreich etabliert. Der Nachweis von P. aeruginosa gebildeter Phagen-DNA gelang mittels Lateral Flow Dipstick. Die Verifizierung des neuen Diagnostik-Systems mit drei realen Proben aus der Kosmetikindustrie zeigte eine hohe Sensitivität und Reproduzierbarkeit. Zusammengefasst wurden die kompletten Grundlagen eines Phagen-basierten Nachweises von lebenden P. aeruginosa-Zellen in klinisch, aber auch sonstigen kontaminationsgefährdeten Produkten, wie Kosmetika gelegt.Pseudomonas aeruginosa is an opportunistic human pathogen, which can cause infections of the respiratory tract, urinary tract and burn wound infections. It is a major health concern for the production of food and cosmetics. The aim of this work was the development of a fast and reliable diagnostic system based on bacteriophages for the detection of living P. aeruginosa cells. In principle, P. aeruginosa-specific bacteriophages immobilized on nanoparticles should be applied on with P. aeruginosa contaminated samples. The phage-DNA produced in the P. aeruginosa cells should be amplified and detected. At first, it could be shown that the screened phages JG012, JG054 and JG057 recognized their P. aeruginosa host via pili-structures and infected a variety of clinical isolates of CF-patients and patients with urinary tract infections. Furthermore, the contained dsDNA and JG012 and JG054 could be grouped to the family Siphoviridae and JG057 to the family Podoviridae. Four different nanoparticles with various surface functions were tested, where the Dynabeads® M-270 Epoxy showed the best binding results for the used phage JG012 and the Dynabeads® M-280 Streptavidin for the phage JG024. Furthermore, an unchanged infectivity could be confirmed compared to the free phages. The phage-DNA preparation and amplification was successfully established. The detection of P. aeruginosa produced phage-DNA succeeded by lateral flow dipstick. The verification of the new diagnostic system with three samples of the cosmetic industry showed a high sensitivity and reproducibility. In summary, the complete foundations were laid for a phage-based diagnosis of living P. aeruginosa cells in clinical, but also in other contamination endangered products, like cosmetics

Skill and added value of the MiKlip regional decadal prediction system for temperature over Europe

In recent years, several decadal prediction systems have been developed to provide multi-year predictions of the climate for the next 5–10 years. On the global scale, high decadal predictability has been identified for the North Atlantic sector, often extending over Europe. The first full regional hindcast ensemble, derived from dynamical downscaling, was produced within the German MiKlip project (‘decadal predictions’). The ensemble features annual starting dates from 1960 to 2017, with 10 decadal hindcasts per starting year. The global component of the prediction system uses the MPI-ESM-LR and the downscaling is performed with the regional climate model COSMO-CLM (CCLM). The present study focusses on a range of aspects dealing with the skill and added value of regional decadal temperature predictions over Europe. The results substantiate the added value of the regional hindcasts compared to the forcing global model as well as to un-initialized simulations. The results show that the hindcasts are skilful both for annual and seasonal means, and that the scores are comparable for different observational reference data sets. The predictive skill increases from earlier to more recent start-years. A recalibration of the simulation data generally improves the skill further, which can also be transferred to more user-relevant variables and extreme values like daily maximum temperatures and heating degree-days. These results provide evidence of the potential for the regional climate predictions to provide valuable climate information on the Abstract Formulae display:MathJax Logo? In recent years, several decadal prediction systems have been developed to provide multi-year predictions of the climate for the next 5–10 years. On the global scale, high decadal predictability has been identified for the North Atlantic sector, often extending over Europe. The first full regional hindcast ensemble, derived from dynamical downscaling, was produced within the German MiKlip project (‘decadal predictions’). The ensemble features annual starting dates from 1960 to 2017, with 10 decadal hindcasts per starting year. The global component of the prediction system uses the MPI-ESM-LR and the downscaling is performed with the regional climate model COSMO-CLM (CCLM). The present study focusses on a range of aspects dealing with the skill and added value of regional decadal temperature predictions over Europe. The results substantiate the added value of the regional hindcasts compared to the forcing global model as well as to un-initialized simulations. The results show that the hindcasts are skilful both for annual and seasonal means, and that the scores are comparable for different observational reference data sets. The predictive skill increases from earlier to more recent start-years. A recalibration of the simulation data generally improves the skill further, which can also be transferred to more user-relevant variables and extreme values like daily maximum temperatures and heating degree-days. These results provide evidence of the potential for the regional climate predictions to provide valuable climate information on the decadal time-scale to users

Pharmacokinetics of a long-acting subcutaneous eprinomectin injection in semi-domesticated reindeer (Rangifer tarandus tarandus) - a pilot study

Reindeer (Rangifer tarandus tarandus) are exposed to the pathogenic parasitic nematode Elaphostrongylus rangiferi during grazing. The severity of disease is dose-dependent. Prophylactic anthelmintic treatment is needed to improve animal health and reindeer herding sustainability. Herds are traditionally only gathered once during the summer, requiring a drug with a persistent effect. In this study we investigated the suitability of long-acting eprinomectin, given as a single subcutaneous injection at 1mg/kg bodyweight in adult reindeer and calves. Plasma and faeces concentrations were determined using ultra-high performance liquid chromatography high resolution mass spectrometry (UHPLC-HRMS). Plasma concentrations remained above the presumed effect level of 2ng/mL for 80 days, demonstrating the drug's potential. Pharmacokinetic parameters were compared to other species using allometric scaling. Calves and adults had slightly different profiles. No viable faecal nematode eggs were detected during treatment. Eprinomectin was measurable in the reindeer faeces up to 100 days, which is of environmental concern

Impaired antibacterial autophagy links granulomatous intestinal inflammation in Niemann-Pick disease type C1 and XIAP deficiency with NOD2 variants in Crohn's disease

OBJECTIVE: Patients with Niemann-Pick disease type C1 (NPC1), a lysosomal lipid storage disorder that causes neurodegeneration and liver damage, can present with IBD, but neither the significance nor the functional mechanism of this association is clear. We studied bacterial handling and antibacterial autophagy in patients with NPC1. DESIGN: We characterised intestinal inflammation in 14 patients with NPC1 who developed IBD. We investigated bacterial handling and cytokine production of NPC1 monocytes or macrophages in vitro and compared NPC1-associated functional defects to those caused by IBD-associated nucleotide-binding oligomerization domain-containing protein 2 (NOD2) variants or mutations in X-linked inhibitor of apoptosis (XIAP). RESULTS: Patients with the lysosomal lipid storage disorder NPC1 have increased susceptibility to early-onset fistulising colitis with granuloma formation, reminiscent of Crohn's disease (CD). Mutations in NPC1 cause impaired autophagy due to defective autophagosome function that abolishes NOD2-mediated bacterial handling in vitro similar to variants in NOD2 or XIAP deficiency. In contrast to genetic NOD2 and XIAP variants, NPC1 mutations do not impair NOD2-receptor-interacting kinase 2 (RIPK2)-XIAP-dependent cytokine production. Pharmacological activation of autophagy can rescue bacterial clearance in macrophages in vitro by increasing the autophagic flux and bypassing defects in NPC1. CONCLUSIONS: NPC1 confers increased risk of early-onset severe CD. Our data support the concept that genetic defects at different checkpoints of selective autophagy cause a shared outcome of CD-like immunopathology linking monogenic and polygenic forms of IBD. Muramyl dipeptide-driven cytokine responses and antibacterial autophagy induction are parallel and independent signalling cascades downstream of the NOD2-RIPK2-XIAP complex

Cystatin C predicts renal function impairment after partial or radical tumor nephrectomy

Purpose: To test the value of preoperative and postoperative cystatin C (CysC) as a predictor on kidney function after partial (PN) or radical nephrectomy (RN) in renal cell carcinoma (RCC) patients with normal preoperative renal function. Methods: From 01/2011 to 12/2014, 195 consecutive RCC patients with a preoperative estimated glomerular filtration rate (eGFR) > 60 ml/min/1.73m2 underwent surgical RCC treatment with either PN or RN. Logistic and linear regression models tested for the effect of CysC as a predictor of new-onset chronic kidney disease in follow-up (eGFR < 60 ml/min/1.73m2). Moreover, postoperative CysC and creatinine values were compared for kidney function estimation. Results: Of 195 patients, 129 (66.2%) underwent PN. In postoperative and in follow-up setting (median 14 months, IQR 10–20), rates of eGFR < 60 ml/min/1.73m2 were 55.9 and 30.2%. In multivariable logistic regression models, preoperative CysC [odds ratio (OR): 18.3] and RN (OR: 13.5) were independent predictors for a reduced eGFR < 60 ml/min/1.73m2 in follow-up (both p < 0.01), while creatinine was not. In multivariable linear regression models, a difference of the preoperative CysC level of 0.1 mg/dl estimated an eGFR decline in follow-up of about 5.8 ml/min/1.73m2. Finally, we observed a plateau of postoperative creatinine values in the range of 1.2–1.3 mg/dl, when graphically depicted vs. postoperative CysC values (‘creatinine blind area’). Conclusion: Preoperative CysC predicts renal function impairment following RCC surgery. Furthermore, CysC might be superior to creatinine for renal function monitoring in the early postoperative setting

Insights into In Vivo Absolute Oral Bioavailability, Biotransformation, and Toxicokinetics of Zearalenone, α-Zearalenol, β-Zearalenol, Zearalenone-14-glucoside, and Zearalenone-14-sulfate in Pigs

The aim of this study was to determine the toxicokinetic characteristics of ZEN and its modified forms, α-zearalenol (α-ZEL), β-zearalenol (β-ZEL), zearalenone-14-glucoside (ZEN14G), and zearalenone-14-sulfate (ZEN14S), including presystemic and systemic hydrolysis in pigs. Crossover pig trials were performed by means of intravenous and oral administration of ZEN and its modified forms. Systemic plasma concentrations of the administered toxins and their metabolites were quantified and further processed via tailor-made compartmental toxicokinetic models. Furthermore, portal plasma was analyzed to unravel the site of hydrolysis, and urine samples were analyzed to determine urinary excretion. Results demonstrate complete presystemic hydrolysis of ZEN14G and ZEN14S to ZEN and high oral bioavailability for all administered compounds, with further extensive first-pass glucuronidation. Conclusively, the modified-ZEN forms α-ZEL, β-ZEL, ZEN14G, and ZEN14S contribute to overall ZEN systemic toxicity in pigs and should be taken into account for risk assessment

Impaired antibacterial autophagy links granulomatous intestinal inflammation in Niemann–Pick disease type C1 and XIAP deficiency with NOD2 variants in Crohn's disease

Objective Patients with Niemann–Pick disease type C1 (NPC1), a lysosomal lipid storage disorder that causes neurodegeneration and liver damage, can present with IBD, but neither the significance nor the functional mechanism of this association is clear. We studied bacterial handling and antibacterial autophagy in patients with NPC1. Design We characterised intestinal inflammation in 14 patients with NPC1 who developed IBD. We investigated bacterial handling and cytokine production of NPC1 monocytes or macrophages in vitro and compared NPC1-associated functional defects to those caused by IBD-associated nucleotide-binding oligomerization domain-containing protein 2 (NOD2) variants or mutations in X-linked inhibitor of apoptosis (XIAP). Results Patients with the lysosomal lipid storage disorder NPC1 have increased susceptibility to early onset fistulising colitis with granuloma formation, reminiscent of Crohn’s disease (CD). Mutations in NPC1 cause impaired autophagy due to defective autophagosome function that abolishes NOD2-mediated bacterial handling in vitro similar to variants in NOD2 or XIAP deficiency. In contrast to genetic NOD2 and XIAP variants, NPC1 mutations do not impair NOD2-receptorinteracting kinase 2 (RIPK2)-XIAP-dependent cytokine production. Pharmacological activation of autophagy can rescue bacterial clearance in macrophages in vitro by increasing the autophagic flux and bypassing defects in NPC1. Conclusions NPC1 confers increased risk of earlyonset severe CD. Our data support the concept that genetic defects at different checkpoints of selective autophagy cause a shared outcome of CD-like immunopathology linking monogenic and polygenic forms of IBD. Muramyl dipeptide-driven cytokine responses and antibacterial autophagy induction are parallel and independent signalling cascades downstream of the NOD2-RIPK2-XIAP complex