Compound-specific amino acid ¹⁵N stable isotope probing of nitrogen assimilation by the soil microbial biomass using gas chromatography/combustion/isotope ratio mass spectrometry

RATIONALE: Organic nitrogen (N) greatly exceeds inorganic N in soils, but the complexity and heterogeneity of this important soil N pool make investigations into the fate of N‐containing additions and soil organic N cycling challenging. This paper details a novel approach to investigate the fate of applied N in soils, generating quantitative measures of microbial assimilation and of newly synthesized soil protein. METHODS: Laboratory incubation experiments applying (15)N‐ammonium, (15)N‐nitrate and (15)N‐glutamate were carried out and the high sensitivity and selectivity of gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) exploited for compound‐specific (15)N stable isotope probing ((15)N‐SIP) of extracted incubation soil amino acids (AAs; as N‐acetyl, O‐isopropyl derivatives). We then describe the interpretation of these data to obtain a measure of the assimilation of the applied (15)N‐labelled substrate by the soil microbial biomass and an estimate of newly synthesised soil protein. RESULTS: The cycling of agriculturally relevant N additions is undetectable via bulk soil N content and δ (15)N values and AA concentrations. The assimilation pathways of the three substrates were revealed via patterns in AA δ (15)N values with time, reflecting known biosynthetic pathways (e.g. ammonium uptake occurs first via glutamate) and these data were used to expose differences in the rates and fluxes of the applied N substrates into the soil protein pool (glutamate > ammonium > nitrate). CONCLUSIONS: Our compound‐specific (15)N‐SIP approach using GC/C/IRMS offers a number of insights, inaccessible via existing techniques, into the fate of applied (15)N in soils and is potentially widely applicable to the study of N cycling in any soil, or indeed, in any complex ecosystem. © 2016 The Authors. Rapid Communications in Mass Spectrometry Published by John Wiley & Sons Ltd

Development of Alditol Acetate Derivatives for the Determination of 15N-Enriched Amino Sugars by Gas Chromatography-Combustion-Isotope Ratio Mass Spectrometry

Amino sugars can be used as indices to evaluate the role of soil microorganisms in active nitrogen (N) cycling in soil. This paper details the assessment of the suitability of gas chromatography–combustion–isotope ratio mass spectrometry (GC–C–IRMS) for the analysis of ¹⁵N-enriched amino sugars as alditol acetate derivatives prior to application of a novel ¹⁵N stable isotope probing (SIP) approach to amino sugars. The efficient derivatization and cleanup of alditol acetate derivatives for GC was achieved using commercially available amino sugars, including glucosamine, mannosamine, galactosamine, and muramic acid, as laboratory standards. A VF-23ms stationary phase was found to produce optimal separations of all four compounds. The structure of the alditol acetate derivatives was confirmed using gas chromatography/mass spectrometry (GC/MS). For GC–C–IRMS determinations, implementation of a two-point normalization confirmed the optimal carrier gas flow rate to be 1.7 mL min^–1. Linearity of δ¹⁵N value determinations up to δ¹⁵N_t of 469 ± 3.1‰ (where δ¹⁵N_t is the independently measured δ¹⁵N value) was confirmed when 30 nmol N was injected on-column, with the direction of deviation from δ¹⁵N_t at low sample amount dependent on the ¹⁵N abundance of the analyte. Observed between- and within-run memory effects were significant (<i>P</i> < 0.007) when a highly enriched standard (469 ± 3.1‰) was run; therefore, analytical run order and variation in ¹⁵N enrichment of analytes within the same sample must be considered. The investigated parameters have confirmed the isotopic robustness of alditol acetate derivatives of amino sugars for the GC–C–IRMS analysis of ¹⁵N-enriched amino sugars in terms of linearity over an enrichment range (natural abundance to 469 ± 3.1‰) with on-column analyte amount over 30 nmol N

Determination of the stretch tensor for structural transformations

The transformation stretch tensor plays an essential role in the evaluation of conditions of compatibility between phases and the use of the Cauchy-Born rule. This tensor is difficult to measure directly from experiment. We give an algorithm for the determination of the transformation stretch tensor from x-ray measurements of structure and lattice parameters. When evaluated on some traditional and emerging phase transformations the algorithm gives unexpected results.Comment: 3 figures, 1 tabl

The Milky Way:Mobility and Economy at the Turn of the 3rd Millennium in Southern Central Europe

In the light of discussions surrounding the social changes attributed to the arrival of the Corded Ware culture in central Europe, here we investigate the economic strategies of one of the cultural complexes of the immediately preceding Late Neolithic. The Cham culture of southern Bavaria is characterised by a variety of economic choices but problems remain in synthesising and combining archaeozoological and archaeobotanical evidence. Using lipid residue analysis from Cham culture pottery excavated at the unenclosed settlement of Riedling, Lower Bavaria, we succeed in identifying a dairying economy at this time. Compound-specific lipid radiocarbon dates are then combined with other samples to provide a formal estimate for the duration of activity at Riedling and the first Bayesian chronological model for the Cham culture as a whole. Although data are currently not fine-grained enough to distinguish between competing models for site permanence, we suggest that the Cham culture pattern fits into a wider central European trend of greater mobility and economic flexibility in the pre-Corded Ware horizon, concluding that key economic strategies previously associated with ‘steppe invasions’ were already present in the preceding centuries. Finally, the demonstrated use of cups for milk-based products, as opposed to alcoholic drinks as previously suggested, leads us to propose possible alternative uses and users for these items

Development of strategies for effective communication of food risks and benefits across Europe: Design and conceptual framework of the FoodRisC project

The FoodRisC project is funded under the Seventh Framework Programme (CORDIS FP7) of the European Commission; Grant agreement no.: 245124. Copyright @ 2011 Barnett et al.BACKGROUND: European consumers are faced with a myriad of food related risk and benefit information and it is regularly left up to the consumer to interpret these, often conflicting, pieces of information as a coherent message. This conflict is especially apparent in times of food crises and can have major public health implications. Scientific results and risk assessments cannot always be easily communicated into simple guidelines and advice that non-scientists like the public or the media can easily understand especially when there is conflicting, uncertain or complex information about a particular food or aspects thereof. The need for improved strategies and tools for communication about food risks and benefits is therefore paramount. The FoodRisC project ("Food Risk Communication - Perceptions and communication of food risks/benefits across Europe: development of effective communication strategies") aims to address this issue. The FoodRisC project will examine consumer perceptions and investigate how people acquire and use information in food domains in order to develop targeted strategies for food communication across Europe.METHODS/DESIGN: This project consists of 6 research work packages which, using qualitative and quantitative methodologies, are focused on development of a framework for investigating food risk/benefit issues across Europe, exploration of the role of new and traditional media in food communication and testing of the framework in order to develop evidence based communication strategies and tools. The main outcome of the FoodRisC project will be a toolkit to enable coherent communication of food risk/benefit messages in Europe. The toolkit will integrate theoretical models and new measurement paradigms as well as building on social marketing approaches around consumer segmentation. Use of the toolkit and guides will assist policy makers, food authorities and other end users in developing common approaches to communicating coherent messages to consumers in Europe.DISCUSSION: The FoodRisC project offers a unique approach to the investigation of food risk/benefit communication. The effective spread of food risk/benefit information will assist initiatives aimed at reducing the burden of food-related illness and disease, reducing the economic impact of food crises and ensuring that confidence in safe and nutritious food is fostered and maintained in Europe.This article is available through the Brunel Open Access Publishing Fund

Targeted Induction of Endoplasmic Reticulum Stress Induces Cartilage Pathology

Pathologies caused by mutations in extracellular matrix proteins are generally considered to result from the synthesis of extracellular matrices that are defective. Mutations in type X collagen cause metaphyseal chondrodysplasia type Schmid (MCDS), a disorder characterised by dwarfism and an expanded growth plate hypertrophic zone. We generated a knock-in mouse model of an MCDS–causing mutation (COL10A1 p.Asn617Lys) to investigate pathogenic mechanisms linking genotype and phenotype. Mice expressing the collagen X mutation had shortened limbs and an expanded hypertrophic zone. Chondrocytes in the hypertrophic zone exhibited endoplasmic reticulum (ER) stress and a robust unfolded protein response (UPR) due to intracellular retention of mutant protein. Hypertrophic chondrocyte differentiation and osteoclast recruitment were significantly reduced indicating that the hypertrophic zone was expanded due to a decreased rate of VEGF–mediated vascular invasion of the growth plate. To test directly the role of ER stress and UPR in generating the MCDS phenotype, we produced transgenic mouse lines that used the collagen X promoter to drive expression of an ER stress–inducing protein (the cog mutant of thyroglobulin) in hypertrophic chondrocytes. The hypertrophic chondrocytes in this mouse exhibited ER stress with a characteristic UPR response. In addition, the hypertrophic zone was expanded, gene expression patterns were disrupted, osteoclast recruitment to the vascular invasion front was reduced, and long bone growth decreased. Our data demonstrate that triggering ER stress per se in hypertrophic chondrocytes is sufficient to induce the essential features of the cartilage pathology associated with MCDS and confirm that ER stress is a central pathogenic factor in the disease mechanism. These findings support the contention that ER stress may play a direct role in the pathogenesis of many connective tissue disorders associated with the expression of mutant extracellular matrix proteins