Functional characterization of the Ustilago maydis effector protein Ten1

Ustilago maydis, the causal agent of corn smut disease, is a pathogen that establishes a biotrophic interaction with Zea mays. The interaction with the host plant is largely governed by a plethora of secreted effector proteins, many of which are encoded in gene clusters. The deletion of cluster 10A consisting of 10 effector-encoding genes results in strongly reduced virulence after maize seedling infection. In the present study, the gene UMAG_03744 (termed ten1) could be identified as a major virulence factor of gene cluster 10A. Via quantitative reverse transcription PCR an induction of ten1 during the biotrophic development of the fungus was detected. ten1 deletion strains showed a virulence phenotype mainly reflected by a reduced tumor size on seedling leaves. Moreover, by complementing the cluster 10A deletion strain for ten1, the strong virulence defect of the cluster mutant was partially rescued. After overexpression in U. maydis hyphae, secreted Ten1 protein could be detected in axenic culture supernatant. Furthermore, using immunoelectron microscopy, the translocation of secreted Ten1 to plant cells could be shown after maize seedling infection, manifested by a significant accumulation of the protein in the plant cytoplasm and especially in plant nuclei. Through a yeast two-hybrid screen ZmPP26, a type 2C maize protein phosphatase (PP2C) could be identified as interaction partner of Ten1. This interaction was supported by coimmunoprecipitation experiments after transient co-expression of Ten1 and ZmPP26 in Nicotiana benthamiana. Moreover, ZmPP26 could be detected by mass spectrometry after immunoprecipitation of Ten1 from U. maydis-infected leaf tissue. Via yeast two-hybrid assays the ZmPP26-interacting domain of Ten1 was mapped. The engineered protein Ten1m, harboring amino acid substitutions in the interacting domain, showed no interaction with ZmPP26 in yeast two-hybrid assays. By complementing the cluster 10A deletion strain with Ten1m, the virulence defect of the cluster mutant could not be rescued, suggesting that the interaction of Ten1 and ZmPP26 may be biologically relevant

A short endoscopic Secretin test for the diagnosis of chronic pancreatitis

Background: The diagnosis of CP is not yet clearly defined. Many national guidelines exist, but - as in many other not clearly defined diseases - there is no worldwide consensus. In CP, evaluation of exocrine pancreatic function is crucial because symptoms are often diffuse and overlooked by the doctors. Additionally, early diagnosis of exocrine pancreatic failure is important as its consequence, malnutrition and commonly abdominal pain, leads to serious complications and reduced life expectancy. Direct pancreas function testing with analyses of enzymes in duodenal juice may give this information. Aims: The main aim was to develop and establish a multimodal algorithm for the diagnoses of CP, accurate and easy to handle in clinical practice. Secondary, we wanted to simplify direct pancreas function testing including a): the performance of a short endoscopic test (article I) and b): the analyses of ingredients in duodenal juice by automation: bicarbonate (article II), Amylase (article III) and Lipase (article IV). Materials and Methods: We examined consecutively healthy controls and patients referred to our outpatient clinic due to symptoms suspicious of CP. We assessed patients with a modified Layer (Mayo) score, which includes imaging, pancreas function testing and medical history. We established a short endoscopic secretin test and analysed bicarbonate, Amylase and Lipase in duodenal juice as markers for ductal and acinar exocrine pancreatic function. In article I, we determined sensitivity, specificity and accuracy of bicarbonate and faecal-elastase, using our modified secretin-stimulated upper endoscopy (short endoscopic secretin test, or EST). In article II, III, IV, we describe correlation between automation of analyses of Bicarbonate, Amylase and Lipase in duodenal juice to labour-intensive manual methods. Results: I. Short endoscopic secretin test: Fifty-two patients aged 19 to 67 years and 25 healthy controls aged 19 to 64 years were included. Twenty-four patients fulfilled the modified Layer score for CP or non-CP. The overall accuracy of the EST versus FE1 test was 85%/71%, with positive and negative predictive values of 100%/79% and 80%/69%, respectively. II. Automation of bicarbonate measurement: 177 samples from 71 patients were analysed. Correlation coefficient of all measurements was r = 0.98 (p < 0.001). Correlation coefficient of fresh versus frozen samples conducted with automatic spectrophotometry (n = 25): r = 0.96 (p < 0.001). III. Automation of amylase measurement: We analysed 52 samples for assay of amylase in pairs. Correlation between measurements with the two methods was r = 0.99 (p < 0.001). IV. Automation of lipase measurement: We tested stability of 54 samples from 21 patients. Diluting samples with MOPS buffer, added BSA gave stable results, and was superior to diluting samples in saline. We compared the two assays in 50 samples from 20 patients and found a good correlation between the two assays (r=0.91, p < 0.001). Conclusions: I: Short EST is rapid and easy to perform and can be incorporated in daily routine in every clinical endoscopic unit. EST is superior to FE1 in the assessment of pancreatic insufficiency, leading to earlier diagnosis of moderate and early or mild CP. II: The measurement of bicarbonate in fresh and thawed samples by automatic spectrophotometric analysis correlates excellent with the back-titration gold standard. III and IV: Quantification of duodenal amylase and lipase activity with automated spectrophotometry has excellent correlation to measurements made by the manual methods. Overall, Endoscopic secretin test is easy to perform, and can be incorporated in a diagnostic endoscopic examination. Automated measurement of bicarbonate, lipase and amylase in duodenal juice simplifies the analytical methods and shortens time from test to result substantially. Standardized, centre-independent analyses of duodenal juice with quantification of ductal and acinar function in any unit with basic endoscopic and laboratory services is within reach

Regionale Atmosphärenmodellierung über dem nördlichen Nordatlantik: das Letzte Glaziale Maximum

Regional atmospheric modelling results for the northern North Atlantic in the Last Glacial Maximum are presented. The forcing global modelling followed in most aspects the Paleoclimate Modelling Intercomparison Project boundary conditions. The regional modelling almost reproduce the forcing data, considering the monthly background circulation, the monthly variability of circulation and the surface-near wind. In addition, the regional model's higher resolution lead to the following improvements upon the forcing data: (1) in some cases precipitation over the Greenland-Iceland-Norway-Sea is more realistic; (2) more detailed katabatic winds were modelled and in the February experiments they were stronger; (3) partly orographic precipitation is stronger; (4) no spectral artefacts in the precipitation occur off the Norwegian coast

Catalytic Asymmetric Spirocyclizing Diels–Alder Reactions of Enones: Stereoselective Total and Formal Syntheses of α-Chamigrene, β-Chamigrene, Laurencenone C, Colletoic Acid, and Omphalic Acid

We disclose a general catalytic enantioselective Diels–Alder reaction of exo-enones with dienes to give spirocyclanes. The obtained products feature highly congested quaternary stereogenic spirocenters and are used in concise total and formal syntheses of several sesquiterpenes, including of α-chamigrene, β-chamigrene, laurencenone C, colletoic acid, and omphalic acid. The stereo- and regioselectivities of our spirocyclizing cycloaddition are effectively controlled by strongly acidic and confined imidodiphosphorimidate catalysts. Computational studies shed light on the origin of reactivity and selectivity

Measurement and Analysis of the Extreme Physical Shock Environment Experienced by Crane-Mounted Radiation Detection Systems

At ports of entry, radiation detectors could be mounted on container gantry crane spreaders to monitor cargo containers entering and leaving the country. These detectors would have to withstand the extreme physical conditions experienced by these spreaders during normal operations. Physical shock data from the gable ends of a spreader were recorded during the loading and unloading of a cargo ship by two hard mounted PCB Piezotronics model 340A50 accelerometers and two Lansmont SAVER 9X30 units (with padding). The majority of large shocks were observed in the vertical direction. The Lansmont units recorded mean shocks of 22.215 ± 1.174 and 23.776 ± 1.140 g, while the PCB accelerometers recorded mean shocks of 31.608 ± 1.798 and 37.072 ± 2.015 g in this direction. Maximum shocks were as high as 118.854 g. A scatter plot of observed peak acceleration versus velocity change is presented to allow comparison with the damage boundary curve for any planned instrumentation for future systems. It is hoped that the results of this research will aid in the design of future crane-mounted systems