Training grapevines generates a metabolomic signature of wine.

Training systems are an option to handle the pronounced apical dominance of grapevines and to influence diverse traits of the corresponding wine. However, it is still unclear if different training systems generate signatures in the metabolome of the wine. By an untargeted metabolomics approach using (SPME) GC-MS wine (volatiles) and leaves were evaluated. Different training directions such as vertical shoot positioning systems, hanging shoot systems, and minimal pruning systems were distinguishable in wine. It was shown, that different training systems generate a metabolomic signature in the wine which was more pronounced than in leaves. Moreover, the sensory analysis showed some changes in the aroma of the different training systems. Thus, the influence of the training system ranges from the leaf metabolome to the wine metabolome.

CARMEN - Comparative Analysis and in silico Reconstruction of organism-specific MEtabolic Networks.

New sequencing technologies provide ultra-fast access to novel microbial genome data. For their interpretation, an efficient bioinformatics pipeline that facilitates in silico reconstruction of metabolic networks is highly desirable. The software tool CARMEN performs in silico reconstruction of metabolic networks to interpret genome data in a functional context. CARMEN supports the visualization of automatically derived metabolic networks based on pathway information from the KEGG database or from user-defined SBML templates; this software also enables comparative genomics. The reconstructed networks are stored in standardized SBML format. We demonstrated the functionality of CARMEN with a major application example focusing on the reconstruction of glycolysis and related metabolic reactions of Xanthomonas campestris pv. campestris B100. The curation of such pathways facilitates enhanced visualization of experimental results, simulations and comparative genomics. A second application of this software was performed on a set of corynebacteria to compare and to visualize their carbohydrate metabolism. In conclusion, using CARMEN, we developed highly automated data analysis software that rapidly converts sequence data into new knowledge, replacing the time-consuming manual reconstruction of metabolic networks. This tool is particularly useful for obtaining an overview of newly sequenced genomes and their metabolic blueprints and for comparative genome analysis. The generated pathways provide automated access to modeling and simulation tools that are compliant with the SBML standard. A user-friendly web interface of CARMEN is available at http://carmen.cebitec.uni-bielefeld.de.

Heat shock protein 27 modification is increased in the human diabetic failing heart.

Chronic conditions like diabetes mellitus (DM) leading to altered metabolism might cause cardiac dysfunction. Hyperglycemia plays an important role in the pathogenesis of diabetic complications including accumulation of methylglyoxal (MG), a highly reactive alpha-dicarbonyl metabolite of glucose degradation pathways and increased generation of advanced glycation endproducts (AGEs). The aim of this investigation was to study the extent of the MG-modification argpyrimidine in human diabetic heart and in rat cardiomyoblasts grown under hyperglycemic conditions. Left ventricular myocardial samples from explanted hearts of patients with cardiomyopathy with (n=8) or without DM (n=8) as well as nonfailing donor organs (n=6), and rat cardiac myoblasts H9c2 treated with glucose were screened for the MG-modification argpyrimidine. The small heat shock protein 27 (Hsp27) revealed to be the major argpyrimidine containing protein in cardiac tissue. Additionally, the modification of arginine leading to argpyrimidine and the phosphorylation of Hsp27 are increased in the myocardium of patients with DM. In H9c2 cells hyperglycemia leads to a decrease of the Hsp27-expression and an increase in argpyrimidine content and phosphorylation of Hsp27, which was accompanied by the induction of oxidative stress and apoptosis. This study shows an association between diabetes and increased argpyrimidine-modification of myocardial Hsp27, a protein which is involved in apoptosis, oxidative stress, and cytoskeleton stabilization.

Identification and characterization of a nodH ortholog from the alfalfa-nodulating Or191-like rhizobia.

Nodulation of Medicago sativa (alfalfa) is known to be restricted to Sinorhizobium meliloti and a few other rhizobia that include the poorly characterized isolates related to Rhizobium sp. strain Or191. Distinctive features of the symbiosis between alfalfa and S. meliloti are the marked specificity from the plant to the bacteria and the strict requirement for the presence of sulfated lipochitooligosaccharides (Nod factors [NFs]) at its reducing end. Here, we present evidence of the presence of a functional nodH-encoded NF sulfotransferase in the Or191-like rhizobia. The nodH gene, present in single copy, maps to a high molecular weight megaplasmid. As in S. meliloti, a nodF homolog was identified immediately upstream of nodH that was transcribed in the opposite direction (local synteny). This novel nodH ortholog was cloned and shown to restore both NF sulfation and the Nif+Fix+ phenotypes when introduced into an S. meliloti nodH mutant. Unexpectedly, however, nodH disruption in the Or191-like bacteria did not abolish their ability to nodulate alfalfa, resulting instead in a severely delayed nodulation. In agreement with evidence from other authors, the nodH sequence analysis strongly supports the idea that the Or191-like rhizobia most likely represent a genetic mosaic resulting from the horizontal transfer of symbiotic genes from a sinorhizobial megaplasmid to a not yet clearly identified ancestor.

Cooperative Action of Rhizobium meliloti Nodulation and Infection Mutants during the Process of Forming Mixed Infected Alfalfa Nodules.

Alfalfa plants co-inoculated with Rhizobium meliloti nodulation (Nod-) and infection mutants deficient in exopolysaccharide production (Inf-EPS-) formed mixed infected nodules that were capable of fixing atmospheric nitrogen. The formation of infected nodules was dependent on close contact between the inoculation partners. When the partners were separated by a filter, empty Fix- nodules were formed, suggesting that infection thread formation in alfalfa is dependent on signals from the nodulation and infection genes. In mixed infected nodules, both nodulation and infection mutants colonized the plant cells and differentiated into bacteroids. The formation of bacteroids was not dependent on cell-to-cell contact between the mutants. Immunogold/silver staining revealed that the ratio of the two mutants varied considerably in colonized plant cells following mixed inoculation. The introduction of an additional nif/fix mutation into one of the inoculation partners did not abolish nitrogen fixation in mixed infected nodules. The expression of nif D::lacZ fusions additionally demonstrated that mutations in the nodulation and infection genes did not prevent the nif genes from being expressed in the mutant bacteroids.

Successful diagnosis of lymphangioleiomyomatosis with transbronchial lung cryobiopsy.

A 47-year old female was evaluated in our clinic for an incidental discovery of diffuse cystic lung disease on high-resolution computed tomography (CT) scan of the chest. There was no personal or family history of tuberous sclerosis complex (TSC), sicca symptoms, pneumothorax, or skin or renal tumors. Review of her chest CT scan showed bilateral, round, uniform, thin-walled cysts present in a diffuse distribution characteristic of lymphangioleiomyomatosis (LAM). CT scan of the abdomen and pelvis did not reveal angiomyolipomas, lymphangioleiomyomas, abnormal lymphadenopathy, or chylous fluid collections. Serum vascular endothelial growth factor-D was non-diagnostic. In order to achieve diagnostic confirmation, the patient underwent transbronchial cryobiopsy of the lung, revealing changes consistent with LAM. Our case highlights the utility of transbronchial lung cryobiopsy in the evaluation of patients with suspected LAM and suggests that further investigation of this diagnostic technique is warranted in patients presenting with diffuse cystic lung disease.

Mixture effects of azole fungicides on the adrenal gland in a broad dose range.

Consumers are exposed to low concentrations of a variety of pesticide residues in or on food. Some of them might interfere with the endocrine system. While each individual active substance has been extensively tested for toxicity and safety, potential combination effects possibly resulting from combined exposure to different pesticides have seldomly been tested so far, especially in vivo. Since the adrenal gland is a key endocrine organ, we investigated if and how substances of a group of fungicides presumed to interfere with the biosynthesis of steroid hormones affect this organ when applied individually and in combination in a broad dose range. A 28-day feeding study was conducted in Wistar rats by using three (tri)azole fungicides considered to potentially affect the endocrine system (cyproconazole, epoxiconazole and prochloraz) individually at five dose levels, ranging from 0.9ppm to 2400ppm, and in combination at three dose levels. The parameters analysed included classical toxicology (pathology, histopathology, clinical chemistry) and molecular toxicology endpoints (gene expression arrays and quantitative real time PCR e.g. of Star, HSD3ß, Cyp11a1, Cyp11b1, Cyp11b2, Cyp 21, ApoE), as well as hormone analysis. A dose-dependent decrease in the adrenal gland weight of rats treated with epoxiconazole alone, which was accompanied by an atrophy of the adrenal gland as well as by an increase in the serum cholesterol level and which only became statistically significant at the top dose levels, was observed. These effects were attenuated in the combination experiments, although the same epoxiconazole concentration was used.

Maleic anhydride proton sponge as a novel MALDI matrix for the visualization of small molecules (<250 m/z) in brain tumors by routine MALDI ToF imaging mass spectrometry.

A novel vacuum stable proton sponge, 4-maleicanhydridoproton sponge (MAPS), was prepared and applied as the matrix in Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging (MALDI-MSI) of an aggressive brain tumor tissue (glioblastoma multiforme). Ionic maps of lactate, 2-hydroxyglutarate and chloride anions (m/z 89, 147, 35, respectively) were obtained using a routine MALDI ToF mass spectrometer.

A safe and feasible "clock-face" duct-to-mucosa pancreaticojejunostomy with a very low incidence of anastomotic failure: A single center experience of 248 patients.

INTRODUCTION: Postoperative pancreatic fistula (POPF) is one of the most frequent and serious postoperative complications of pancreatoduodenectomy (PD). We sought to assess the impact of a novel pancreaticojejunostomy (PJ) on the rates of POPF and overall postoperative complications. METHODS: Between 01/2010 and 12/2013, a total of 248 consecutive patients who underwent PD with a modified PJ were identified from our database and retrospectively analyzed. POPF cases were divided into three categories (ISGPF-international study group-guidelines): biochemical fistula without clinical sequelae (grade A), fistula requiring any therapeutic intervention (grade B), and fistula with severe clinical sequelae (grade C). Perioperative outcomes were recorded and analyzed. RESULTS: The overwhelming majority of patients had no evidence of fistula. Grade A POPF was observed in 9 (3.62%), grade B in 1 (0.40%), and grade C in 0 patients. There were no postoperative deaths. Overall complications occurred in 61 patients (24.59%) of patients after PD. CONCLUSIONS: This modified pancreaticojejunostomy is widely applicable and is associated with very low rates of POPF, low postoperative morbidity and mortality. Overall, it is a feasible and safe novel approach with excellent short-term outcomes.

A consolidated analysis of the physiologic and molecular responses induced under acid stress in the legume-symbiont model-soil bacterium Sinorhizobium meliloti.

Abiotic stresses in general and extracellular acidity in particular disturb and limit nitrogen-fixing symbioses between rhizobia and their host legumes. Except for valuable molecular-biological studies on different rhizobia, no consolidated models have been formulated to describe the central physiologic changes that occur in acid-stressed bacteria. We present here an integrated analysis entailing the main cultural, metabolic, and molecular responses of the model bacterium Sinorhizobium meliloti growing under controlled acid stress in a chemostat. A stepwise extracellular acidification of the culture medium had indicated that S. meliloti stopped growing at ca. pH 6.0-6.1. Under such stress the rhizobia increased the O2 consumption per cell by more than 5-fold. This phenotype, together with an increase in the transcripts for several membrane cytochromes, entails a higher aerobic-respiration rate in the acid-stressed rhizobia. Multivariate analysis of global metabolome data served to unequivocally correlate specific-metabolite profiles with the extracellular pH, showing that at low pH the pentose-phosphate pathway exhibited increases in several transcripts, enzymes, and metabolites. Further analyses should be focused on the time course of the observed changes, its associated intracellular signaling, and on the comparison with the changes that operate during the sub lethal acid-adaptive response (ATR) in rhizobia.

Health Informatics via Machine Learning for the Clinical Management of Patients.

OBJECTIVES: To review how health informatics systems based on machine learning methods have impacted the clinical management of patients, by affecting clinical practice. METHODS: We reviewed literature from 2010-2015 from databases such as Pubmed, IEEE xplore, and INSPEC, in which methods based on machine learning are likely to be reported. We bring together a broad body of literature, aiming to identify those leading examples of health informatics that have advanced the methodology of machine learning. While individual methods may have further examples that might be added, we have chosen some of the most representative, informative exemplars in each case. RESULTS: Our survey highlights that, while much research is taking place in this high-profile field, examples of those that affect the clinical management of patients are seldom found. We show that substantial progress is being made in terms of methodology, often by data scientists working in close collaboration with clinical groups. CONCLUSIONS: Health informatics systems based on machine learning are in their infancy and the translation of such systems into clinical management has yet to be performed at scale.

Molecular analysis of the Rhizobium meliloti mucR gene regulating the biosynthesis of the exopolysaccharides succinoglycan and galactoglucan.

The Rhizobium meliloti Tn5 mutant Rm3131, producing galactoglucan (EPS II) instead of succinoglycan (EPS I), was complemented by a 3.6-kb EcoRI-fragment of the Rhizobium meliloti genome. Sequencing of this fragment revealed six open reading frames (ORFs). The ORF found to be affected in the mutant Rm3131 codes for a putative protein of 15.7 kDa and forms a monocistronic transcriptional unit. Further genetic analysis revealed that the gene mutated in Rm3131 is identical to the previously described R. meliloti mucR gene (H. Zhan, S.B. Levery, C. C. Lee, and J.A. Leigh, 1989, Proc. Natl. Acad. Sci. USA 86:3055-3059). By hybridization it was shown that a mucR homologous gene is present in several rhizobacteria. The deduced amino acid sequence of MucR showed nearly 80% identity to the Agrobacterium tumefaciens Ros protein, a negative regulator of vir genes and necessary for succinoglycan production. MucR contains like Ros a putative zinc finger sequence of the C2H2 type. Transcriptional fusions of genes for EPS I and EPS II synthesis, the so-called exo and exp genes, with the marker gene lacZ were used to delineate the role of mucR for exo and exp gene expression. It was found that exp genes are negatively regulated by MucR on the transcriptional level, whereas a posttranscriptional regulation by MucR is assumed for exo genes. Furthermore, mucR is negatively regulating its own transcription.

Alfalfa and tobacco cells react differently to chitin oligosaccharides and sinorhizobium meliloti nodulation factors

Alfalfa (Medicago sativa L.) suspension cultures respond to yeast elicitors with a strong alkalinization of the culture medium, a transient synthesis of activated oxygen species, and typical late defence reactions such as phytoalexin accumulation and increased peroxidase activity. The alkalinization reaction as well as the oxidative burst were also observed when tobacco (Nicotiana tabacum L. ) cell-suspension cultures were treated with yeast elicitors. Depending on the degree of polymerization, N-acetyl chitin oligomers induced the alkalinization response in both plant cell-suspension cultures, while only tobacco cell cultures developed an oxidative burst. Suspension-cultured tobacco cells responded to Sinorhizobium meliloti nodulation factors with a maximal alkalinization of 0.25 pH units and a remarkable oxidative burst. In contrast, addition of Sinorhizobium meliloti nodulation factors to suspension-cultured alfalfa cells induced a slight acidification of the culture medium, instead of an alkalinization, but no oxidative burst.

Isolation and characterization of alfalfa-nodulating rhizobia present in acidic soils of central argentina and uruguay

We describe the isolation and characterization of alfalfa-nodulating rhizobia from acid soils of different locations in Central Argentina and Uruguay. A collection of 465 isolates was assembled, and the rhizobia were characterized for acid tolerance. Growth tests revealed the existence of 15 acid-tolerant (AT) isolates which were able to grow at pH 5.0 and formed nodules in alfalfa with a low rate of nitrogen fixation. Analysis of those isolates, including partial sequencing of the genes encoding 16S rRNA and genomic PCR-fingerprinting with MBOREP1 and BOXC1 primers, demonstrated that the new isolates share a genetic background closely related to that of the previously reported Rhizobium sp. Or191 recovered from an acid soil in Oregon (B. D. Eardly, J. P. Young, and R. K. Selander, Appl. Environ. Microbiol. 58:1809-1815, 1992). Growth curves, melanin production, temperature tolerance, and megaplasmid profiles of the AT isolates were all coincident with these characteristics in strain Or191. In addition to the ability of all of these strains to nodulate alfalfa (Medicago sativa) inefficiently, the AT isolates also nodulated the common bean and Leucaena leucocephala, showing an extended host range for nodulation of legumes. In alfalfa, the time course of nodule formation by the AT isolate LPU 83 showed a continued nodulation restricted to the emerging secondary roots, which was probably related to the low rate of nitrogen fixation by the largely ineffective nodules. Results demonstrate the complexity of the rhizobial populations present in the acidic soils represented by a main group of N2-fixing rhizobia and a second group of ineffective and less-predominant isolates related to the AT strain Or191.

Comparison of two Xanthomonas campestris pathovar campestris genomes revealed differences in their gene composition.

For the Xanthomonas campestris pathovar campestris wild-type strain B100 a plasmid-based clone library was constructed. The plasmids carried chromosomal fragments of 3-4 kb in size that were tagged in vitro with the artificial transposon KAN-2. More than 3000 of the transposon target sites were characterized by DNA sequencing. The sequences obtained were compared to the recently published genome of Xanthomonas campestris pathovar campestris strain ATCC 33913. Most of the sequenced clones derived from strain B100 matched the chromosomal sequence of strain ATCC 33913. An alignment to the circular map of this chromosome revealed that the similarities were statistically distributed over the entire genome of strain ATCC 33913. The similarity was obvious for protein coding sequences, as well as for mobile genetic elements. However, four regions in the genome of Xanthomonas campestris pathovar campestris strain ATCC 33913, ranging in size from 11 to 37 kb, were not represented in the sequenced clone library of Xanthomonas campestris pathovar campestris strain B100. On the other hand, 1.2% of the sequenced clones originating from Xanthomonas campestris pathovar campestris strain B100 showed no or insignificant similarities to the genome of strain ATCC 33913.

[Perihilar cholangiocarcinoma (Klatskin tumor)]. / Zentrales Gallengangskarzinom (Klatskin-Tumor).

Perihilar cholangiocarcinoma or Klatskin tumors are a rare entity arising from the extrahepatic bile duct bifurcation. Considering the close anatomical relationship of the bile duct bifurcation with the portal vein bifurcation and hepatic arteries, surgical treatment is demanding. With an incidence of only 2-4 cases/100,000 population/year patients should be referred to a specialized center. The tumors are usually poorly differentiated adenocarcinomas growing diffusely along the duct and also the perineural sheath. Only radical surgery offers a curative option and currently surgical strategy usually consists of en bloc resection of the bile duct, extended liver resection and portal vein resection. Proximal and lateral safety margin R0 resections are technically very demanding procedures because of the local anatomy.

Metabolite fingerprinting of barley whole seeds, endosperms, and embryos during industrial malting.

Samples of whole seeds, isolated endosperms including the aleurone layer and isolated embryos with attached scutellum from an industrial scale barley malting process (variety Braemar) were analysed for their water soluble metabolites by gas chromatography-mass spectrometry (GC-MS). 73 known metabolites and about 350 unknown signals were detected. Principal component analysis (PCA) showed a time dependent shift of sample profiles. Whole seeds and endosperm samples showed very similar patterns with nearly all compounds rising until the end of germination. In the embryos a maximum concentration of compounds was reached after 72-96 h of malting. Most concentrations decreased afterwards. The kilning step, namely the drying and roasting of germinated seeds, induced variable effects of increases, stability or decreases of metabolites and thereby separated kilned samples from germinated seeds in the PCA. A second barley cultivar (Quench) underwent the same malting and analysis procedures and gave nearly identical results. Fructose, malate, myo-inositol and raffinose exhibited the potential to serve as markers for specific developmental stages of seeds in both varieties. Biological markers represent targets for industrial process control. Their potential application would meet the maltsters' demand to flatten variances in germination properties and to produce equal composed malt by directed malting management.

Two differentially expressed MATE factor genes from apple complement the Arabidopsis transparent testa12 mutant.

Proanthocyanidins (PAs) are a class of flavonoids with numerous functions in plant ecology and development, including protection against microbial infection, animal foraging and damage by UV light. PAs are also beneficial in the human diet and livestock farming, preventing diseases of the cardiovascular system and lowering the risk of cancer, asthma and diabetes. Apples (Malus x domestica Borkh.) are naturally rich in flavonoids, but the flavonoid content and composition varies significantly between cultivars. In this work, we applied knowledge from the model plant Arabidopsis thaliana, for which the main features of flavonoid biosynthesis have been elucidated, to investigate PA accumulation in apple. We identified functional homologues of the Multidrug And Toxic compound Extrusion (MATE) gene TRANSPARENT TESTA12 from A. thaliana using a comparative genomics approach. MdMATE1 and MdMATE2 were differentially expressed, and the function of the encoded proteins was verified by complementation of the respective A. thaliana mutant. In addition, MdMATE genes have a different gene structure in comparison to homologues from other species. Based on our findings, we propose that MdMATE1 and MdMATE2 are vacuolar flavonoid/H(+) -antiporters, active in PA accumulating cells of apple fruit. The identification of these flavonoid transporter genes expands our understanding of secondary metabolite biosynthesis and transport in apple, and is a prerequisite to improve the nutritional value of apples and apple-derived beverages.