Girdling interruption between source and sink in Quercus pubescens does not trigger leaf senescence

Metabolite changes and senescence behaviour after mechanical phloem girdling were studied in leaf tissue of Quercus pubescens. Sugar accumulation is not only considered to be an important part of several developmental signalling pathways, but is also seen as one of the basic triggers for senescence induction, or at least an obligatory accessory phenomenon. Our survey showed that an accumulation of the soluble sugars, glucose and fructose, was not on its own obligatorily connected with the induction of leaf senescence, since no indication or even an onset of senescence could be observed during the course of the experiment. Instead, we observed an inhibition of leaf development with a decrease of photosynthesis and a slow-down of development in nearly all chlorophyll a fluorescence analysis parameters using the JIP-test. We detected a change of metabolites linked to oxidative stress, possibly due to an overexcitation of the developmentally inhibited photosynthetic apparatus.     

Solving the Differential Biochemical Jacobian from Metabolomics Covariance Data

High-throughput molecular analysis has become an integral part in organismal systems biology. In contrast, due to a missing systematic linkage of the data with functional and predictive theoretical models of the underlying metabolic network the understanding of the resulting complex data sets is lacking far behind. Here, we present a biomathematical method addressing this problem by using metabolomics data for the inverse calculation of a biochemical Jacobian matrix, thereby linking computer-based genome-scale metabolic reconstruction and in vivo metabolic dynamics. The incongruity of metabolome coverage by typical metabolite profiling approaches and genome-scale metabolic reconstruction was solved by the design of superpathways to define a metabolic interaction matrix. A differential biochemical Jacobian was calculated using an approach which links this metabolic interaction matrix and the covariance of metabolomics data satisfying a Lyapunov equation. The predictions of the differential Jacobian from real metabolomic data were found to be correct by testing the corresponding enzymatic activities. Moreover it is demonstrated that the predictions of the biochemical Jacobian matrix allow for the design of parameter optimization strategies for ODE-based kinetic models of the system. The presented concept combines dynamic modelling strategies with large-scale steady state profiling approaches without the explicit knowledge of individual kinetic parameters. In summary, the presented strategy allows for the identification of regulatory key processes in the biochemical network directly from metabolomics data and is a fundamental achievement for the functional interpretation of metabolomics data.     

Phytochemical composition of Potentilla anserina L. analyzed by an integrative GC-MS and LC-MS metabolomics platform

Potentilla anserina L. (Rosaceae) is known for its beneficial effects of prevention of pre-menstrual syndrome (PMS). For this reason P. anserina is processed into many food supplements and pharmaceutical preparations. Here we analyzed hydroalcoholic reference extracts and compared them with various extracts of different pharmacies using an integrative metabolomics platform comprising GC-MS and LC-MS analysis and software toolboxes for data alignment (MetMAX Beta 1.0) and multivariate statistical analysis (COVAIN 1.0). Multivariate statistics of the integrated GC-MS and LC-MS data showed strong differences between the different plant extract formulations. Different groups of compounds such as chlorogenic acid, kaempferol 3-O-rutinoside, acacetin 7-O-rutinoside, and genistein were reported for the first time in this species. The typical fragmentation pathway of the isoflavone genistein confirmed the identification of this active compound that was present with different abundances in all the extracts analyzed. As a result we have revealed that different extraction procedures from different vendors produce different chemical compositions, e.g. different genistein concentrations. Consequently, the treatment may have different effects. The integrative metabolomics platform provides the highest resolution of the phytochemical composition and a mean to define subtle differences in plant extract formulations.     

Primary Metabolism,Phenylpropanoids and Antioxidant Pathways Are Regulated in Potato as a Response to Potato virus Y Infection

Potato production is one of the most important agricultural sectors, and it is challenged by various detrimental factors, including virus infections. To control losses in potato production, knowledge about the virus—plant interactions is crucial. Here, we investigated the molecular processes in potato plants as a result of Potato virus Y (PVY) infection, the most economically important potato viral pathogen. We performed an integrative study that links changes in the metabolome and gene expression in potato leaves inoculated with the mild PVY^N and aggressive PVY^NTN isolates, for different times through disease development. At the beginning of infection (1 day post-inoculation), virus-infected plants showed an initial decrease in the concentrations of metabolites connected to sugar and amino-acid metabolism, the TCA cycle, the GABA shunt, ROS scavangers, and phenylpropanoids, relative to the control plants. A pronounced increase in those metabolites was detected at the start of the strong viral multiplication in infected leaves. The alterations in these metabolic pathways were also seen at the gene expression level, as analysed by quantitative PCR. In addition, the systemic response in the metabolome to PVY infection was analysed. Systemic leaves showed a less-pronounced response with fewer metabolites altered, while phenylpropanoid-associated metabolites were strongly accumulated. There was a more rapid onset of accumulation of ROS scavengers in leaves inoculated with PVY^N than those inoculated with PVY^NTN. This appears to be related to the lower damage observed for leaves of potato infected with the milder PVY^N strain, and at least partially explains the differences between the phenotypes observed.     

Altitudinal patterns of diversity and functional traits of metabolically active microorganisms in stream biofilms

Resources structure ecological communities and potentially link biodiversity to energy flow. It is commonly believed that functional traits (generalists versus specialists) involved in the exploitation of resources depend on resource availability and environmental fluctuations. The longitudinal nature of stream ecosystems provides changing resources to stream biota with yet unknown effects on microbial functional traits and community structure. We investigated the impact of autochthonous (algal extract) and allochthonous (spruce extract) resources, as they change along alpine streams from above to below the treeline, on microbial diversity, community composition and functions of benthic biofilms. Combining bromodeoxyuridine labelling and 454 pyrosequencing, we showed that diversity was lower upstream than downstream of the treeline and that community composition changed along the altitudinal gradient. We also found that, especially for allochthonous resources, specialisation by biofilm bacteria increased along that same gradient. Our results suggest that in streams below the treeline biofilm diversity, specialisation and functioning are associated with increasing niche differentiation as potentially modulated by divers allochthonous and autochthonous constituents contributing to resources. These findings expand our current understanding on biofilm structure and function in alpine streams.     

Die Mykostatische Wirkung von Trichophyton rubrum auf Kryptokokken

Zusammenfassung Schwarz pigmentierende Kulturformen desTrichophyton rubrum (Cast.)Sabouraud bilden auf Sabouraud's Agar ein Antimykotikum, das gegenCryptococcus neoformans, C. diffluens undTorulopsis stellata wirksam ist. Es ist uns bisher nicht gelungen, die wirksame Substanz zu isolieren.     

Liver disease in hog caused by Rhizopus cohnii

In a hog slaughtered for general deterioration of health and jaundice, tumoriform, nodular lesions were found in the liver. Histologically, there was coagulation necrosis sometimes liquefied in its central parts, lined with demarcation wall of neutrophilic granulocytes, or, with fibroblastic granulation tissue and infiltrates of histiocytes, eosinophilic granulocytes, lymphocytes and multinucleated foreign body giant cells. Within these lesions, there was mycelium of a Phycomycetes type. Cultures yielded Rhizopus cohnii.     

High-efficiency derivation of human embryonic stem cell lines following pre-implantation genetic diagnosis

Pre-implantation genetic diagnosis allows the characterisation of embryos that carry a gene responsible for a severe monogenic disease and to transfer to the mother’s uterus only the unaffected one(s). The genetically affected embryos can be used to establish human embryonic stem cell (hESC) lines. We are currently establishing a cell bank of ESC lines carrying specific disease-causing mutant genes. These cell lines are available to the scientific community. For this purpose, we have designed a technique that requires only minimal manipulation of the embryos. At the blastocyst stage, we just removed the zona pellucida before seeding the embryo as a whole on a layer of feeder cells. This approach gave a good success rate (>20%), whatever the quality of the embryos, and allowed us to derive 11 new hESC lines, representing seven different pathologies. Full phenotypic validation of the cell lines according to ISCI guidelines confirmed their pluripotent nature, as they were positive for hESC markers and able to differentiate in vitro in all three germ layers derivatives. Nine out of 11 stem cell lines had normal karyotypes. Our results indicate that inner cell mass isolation is not mandatory for hESC derivation and that minimal manipulation of embryos can lead to high success rate.     

Granger causality in integrated GC–MS and LC–MS metabolomics data reveals the interface of primary and secondary metabolism

Metabolomics has emerged as a key technique of modern life sciences in recent years. Two major techniques for metabolomics in the last 10 years are gas chromatography coupled to mass spectrometry (GC–MS) and liquid chromatography coupled to mass spectrometry (LC–MS). Each platform has a specific performance detecting subsets of metabolites. GC–MS in combination with derivatisation has a preference for small polar metabolites covering primary metabolism. In contrast, reversed phase LC–MS covers large hydrophobic metabolites predominant in secondary metabolism. Here, we present an integrative metabolomics platform providing a mean to reveal the interaction of primary and secondary metabolism in plants and other organisms. The strategy combines GC–MS and LC–MS analysis of the same sample, a novel alignment tool MetMAX and a statistical toolbox COVAIN for data integration and linkage of Granger Causality with metabolic modelling. For metabolic modelling we have implemented the combined GC–LC–MS metabolomics data covariance matrix and a stoichiometric matrix of the underlying biochemical reaction network. The changes in biochemical regulation are expressed as differential Jacobian matrices. Applying the Granger causality, a subset of secondary metabolites was detected with significant correlations to primary metabolites such as sugars and amino acids. These metabolic subsets were compiled into a stoichiometric matrix N. Using N the inverse calculation of a differential Jacobian J from metabolomics data was possible. Key points of regulation at the interface of primary and secondary metabolism were identified.