Imaging mass spectrometry for spatial metabolomics

Over the past decade, mass spectrometry (MS) has seen major technical advances that have increased the scope, applicability and adoption of the technology in a vast array of research areas (1)

Seismotectonic study of the Fergana region (Southern Kyrgyzstan): distribution and kinematics of local seismicity

We present new seismicity and focal-mechanism data for the Fergana basin and surrounding mountain belts in western Kyrgyzstan from a temporary local seismic network. A total of 210 crustal earthquakes with hypocentral depths shallower than 25 km were observed during a 12-month period in 2009/2010. The hypocenter distribution indicates a complex net of seismically active structures. The seismicity derived in this study is mainly concentrated at the edges of the Fergana basin, whereas the observed rate of seismicity within the basin is low. The seismicity at the dominant tectonic feature of the region, the Talas-Fergana fault, is likewise low, so the fault seems to be inactive or locked. To estimate the uncertainties of earthquake locations derived in this study, a strong explosion with known origin time and location is used as a ground truth calibration event which suggests a horizontal and vertical accuracy of about 1 km for our relocations. We derived 35 focal mechanisms using first motion polarities and retrieved a set of nine moment tensor solutions for earthquakes with moment magnitude (Mw) ranging from 3.3 to 4.9 by waveform inversion. The solutions reveal both thrust and strike-slip mechanisms compatible with a NW-SE direction of compression for the Fergana region. Two previously unknown tectonic structures in the Fergana region could be identified, both featuring strike-slip kinematics. The combined analysis of the results derived in this study allowed a detailed insight into the currently active tectonic structures and their kinematics where little information had previously been available

Mass spectrometry imaging for plant biology: A review

Mass spectrometry imaging (MSI) is a developing technique to measure the spatio-temporal distribution of many biomolecules in tissues. Over the preceding decade, MSI has been adopted by plant biologists and applied in a broad range of areas, including primary metabolism, natural products, plant defense, plant responses to abiotic and biotic stress, plant lipids and the developing field of spatial metabolomics. This review covers recent advances in plant-based MSI, general aspects of instrumentation, analytical approaches, sample preparation and the current trends in respective plant research

A new peak detection algorithm for MALDI mass spectrometry data based on a modified Asymmetric Pseudo-Voigt model

Background Mass Spectrometry (MS) is a ubiquitous analytical tool in biological research and is used to measure the mass-to-charge ratio of bio-molecules. Peak detection is the essential first step in MS data analysis. Precise estimation of peak parameters such as peak summit location and peak area are critical to identify underlying bio-molecules and to estimate their abundances accurately. We propose a new method to detect and quantify peaks in mass spectra. It uses dual-tree complex wavelet transformation along with Stein's unbiased risk estimator for spectra smoothing. Then, a new method, based on the modified Asymmetric Pseudo-Voigt (mAPV) model and hierarchical particle swarm optimization, is used for peak parameter estimation. Results Using simulated data, we demonstrated the benefit of using the mAPV model over Gaussian, Lorentz and Bi-Gaussian functions for MS peak modelling. The proposed mAPV model achieved the best fitting accuracy for asymmetric peaks, with lower percentage errors in peak summit location estimation, which were 0.17% to 4.46% less than that of the other models. It also outperformed the other models in peak area estimation, delivering lower percentage errors, which were about 0.7% less than its closest competitor - the Bi-Gaussian model. In addition, using data generated from a MALDI-TOF computer model, we showed that the proposed overall algorithm outperformed the existing methods mainly in terms of sensitivity. It achieved a sensitivity of 85%, compared to 77% and 71% of the two benchmark algorithms, continuous wavelet transformation based method and Cromwell respectively. Conclusions The proposed algorithm is particularly useful for peak detection and parameter estimation in MS data with overlapping peak distributions and asymmetric peaks. The algorithm is implemented using MATLAB and the source code is freely available at http://mapv.sourceforge.net

Insights into oxidized lipid modification in barley roots as an adaptation mechanism to salinity stress

Lipidomics is an emerging technology, which aims at the global characterization and quantification of lipids within biological matrices including biofluids, cells, whole organs and tissues. The changes in individual lipid molecular species in stress treated plant species and different cultivars can indicate the functions of genes affecting lipid metabolism or lipid signaling. Mass spectrometry–based lipid profiling has been used to track the changes of lipid levels and related metabolites in response to salinity stress. We have developed a comprehensive lipidomics platform for the identification and direct qualification and/or quantification of individual lipid species, including oxidized lipids, which enables a more systematic investigation of peroxidation of individual lipid species in barley roots under salinity stress. This new lipidomics approach has improved with an advantage of analyzing the composition of acyl chains at the molecular level, which facilitates to profile precisely the 18:3-containing diacyl-glycerophosphates and allowed individual comparison of lipids across varieties. Our findings revealed a general decrease in most of the galactolipids in plastid membranes, and an increase of glycerophospholipids and acylated steryl glycosides, which indicate that plastidial and extraplastidial membranes in barley roots ubiquitously tend to form a hexagonal II (HII) phase under salinity stress. In addition, salt-tolerant and salt-sensitive cultivars showed contrasting changes in the levels of oxidized membrane lipids. These results support the hypothesis that salt-induced oxidative damage to membrane lipids can be used as an indication of salt stress tolerance in barley

De novo transcriptome assembly and analysis of differentially expressed genes of two barley genotypes reveal root-zone-specific responses to salt exposure

Plant roots are the first organs sensing and responding to salinity stress, manifested differentially between different root types, and also at the individual tissue and cellular level. High genetic diversity and the current lack of an assembled map-based sequence of the barley genome severely limit barley research potential. We used over 580 and 600 million paired-end reads, respectively, to create two de novo assemblies of a barley landrace (Sahara) and a malting cultivar (Clipper) with known contrasting responses to salinity. Generalized linear models were used to statistically access spatial, treatment-related, and genotype-specific responses. This revealed a spatial gene expression gradient along the barley root, with more differentially expressed transcripts detected between different root zones than between treatments. The root transcriptome also showed a gradual transition from transcripts related to sugar-mediated signaling at the root meristematic zone to those involved in cell wall metabolism in the elongation zone, and defense response-related pathways toward the maturation zone, with significant differences between the two genotypes. The availability of these additional transcriptome reference sets will serve as a valuable resource to the cereal research community, and may identify valuable traits to assist in breeding programmes

A high-resolution HPLC-QqTOF platform using parallel reaction monitoring for in-depth lipid discovery and rapid profiling

Here, we developed a robust lipidomics workflow merging both targeted and untargeted approaches on a single liquid chromatography coupled to quadrupole-time of flight (LC-QqTOF) mass spectrometry platform with parallel reaction monitoring (PRM). PRM assays integrate both untargeted profiling from MS1 scans and targeted profiling obtained from MS/MS data. This workflow enabled the discovery of more than 2300 unidentified features and identification of more than 600 lipid species from 23 lipid classes at the level of fatty acid/long chain base/sterol composition in a barley root extracts. We detected the presence of 142 glycosyl inositol phosphorylceramides (GIPC) with HN(Ac)-HA as the core structure of the polar head, 12 cardiolipins and 17 glucuronosyl diacylglycerols (GlcADG) which have been rarely reported previously for cereal crops. Using a scheduled algorithm with up to 100 precursors multiplexed per duty cycle, the PRM assay was able to achieve a rapid profiling of 291 species based on MS/MS data by a single injection. We used this novel approach to demonstrate the applicability and efficiency of the workflow to study salt stress induced changes in the barley root lipidome. Results show that 221 targeted lipids and 888 unknown features were found to have changed significantly in response to salt stress. This combined targeted and untargeted single workflow approach provides novel applications of lipidomics addressing biological questions

The metabolic environment of the developing embryo: A multidisciplinary approach on oilseed rapeseed

Brassicaceae seeds consist of three genetically distinct structures: the embryo, endosperm and seed coat, all of which are involved in assimilate allocation during seed development. The complexity of their metabolic interrelations remains unresolved to date. In the present study, we apply state-of-the-art imaging and analytical approaches to assess the metabolic environment of the Brassica napus embryo. Nuclear magnetic resonance imaging (MRI) provided volumetric data on the living embryo and endosperm, revealing how the endosperm envelops the embryo, determining endosperm's priority in assimilate uptake from the seed coat during early development. MRI analysis showed higher levels of sugars in the peripheral endosperm facing the seed coat, but a lower sugar content within the central vacuole and the region surrounding the embryo. Feeding intact siliques with 13C-labeled sucrose allowed tracing of the post-phloem route of sucrose transfer within the seed at the heart stage of embryogenesis, by means of mass spectrometry imaging. Quantification of over 70 organic and inorganic compounds in the endosperm revealed shifts in their abundance over different stages of development, while sugars and potassium were the main determinants of osmolality throughout these stages. Our multidisciplinary approach allows access to the hidden aspects of endosperm metabolism, a task which remains unattainable for the small-seeded model plant Arabidopsis thaliana

High-mass-resolution MALDI mass spectrometry imaging reveals detailed spatial distribution of metabolites and lipids in roots of barley seedlings in response to salinity stress

Introduction Mass spectrometry imaging (MSI) is a technology that enables the visualization of the spatial distribution of hundreds to thousands of metabolites in the same tissue section simultaneously. Roots are below-ground plant organs that anchor plants to the soil, take up water and nutrients, and sense and respond to external stresses. Physiological responses to salinity are multifaceted and have predominantly been studied using whole plant tissues that cannot resolve plant salinity responses spatially. Objectives This study aimed to use a comprehensive approach to study the spatial distribution and profiles of metabolites, and to quantify the changes in the elemental content in young developing barley seminal roots before and after salinity stress. Methods Here, we used a combination of liquid chromatography–mass spectrometry (LC–MS), inductively coupled plasma mass spectrometry (ICP–MS), and matrix-assisted laser desorption/ionization (MALDI–MSI) platforms to profile and analyze the spatial distribution of ions, metabolites and lipids across three anatomically different barley root zones before and after a short-term salinity stress (150 mM NaCl). Results We localized, visualized and discriminated compounds in fine detail along longitudinal root sections and compared ion, metabolite, and lipid composition before and after salt stress. Large changes in the phosphatidylcholine (PC) profiles were observed as a response to salt stress with PC 34:n showing an overall reduction in salt treated roots. ICP–MS analysis quantified changes in the elemental content of roots with increases of Na+ and decreases of K+ content. Conclusion Our results established the suitability of combining three mass spectrometry platforms to analyze and map ionic and metabolic responses to salinity stress in plant roots and to elucidate tolerance mechanisms in response to abiotic stress, such as salinity stress

Metabolic profiling of diabetic cats in remission

Background: The majority of diabetic cats in remission have abnormal glucose tolerance, and approximately one third relapse within 1 year. Greater understanding of the metabolic characteristics of diabetic cats in remission, and predictors of relapse is required to effectively monitor and manage these cats. Objectives: To identify and compare differences in plasma metabolites between diabetic cats in remission and healthy control cats using a metabolomics approach. Secondly, to assess whether identified metabolites are predictors of diabetic relapse. Animals: Twenty cats in diabetic remission for a median of 101 days, and 22 healthy matched control cats. Methods: Cats were admitted to a clinic, and casual blood glucose was recorded. After a 24 h fast, blood glucose concentration was measured, then a blood sample was taken for metabolomic (GCMS and LCMS) analyses. Three hours later, a simplified intravenous glucose tolerance test (1 g glucose/kg) was performed. Cats were monitored for diabetes relapse for at least 9 months (270 days) after baseline testing. Results: Most cats in remission continued to display impaired glucose tolerance. Concentrations of 16 identified metabolites differed (P ≤ 0.05) between remission and control cats: 10 amino acids and stearic acid (all lower in remission cats), and glucose, glycine, xylitol, urea and carnitine (all higher in remission cats). Moderately close correlations were found between these 16 metabolites and variables assessing glycaemic responses (most |r| = 0.31 to 0.69). Five cats in remission relapsed during the study period. No metabolite was identified as a predictor of relapse. Conclusion and clinical importance: This study shows that cats in diabetic remission have abnormal metabolism