Alterations in Metabolome and Microbiome Associated with an Early Stress Stage in Male Wistar Rats: A Multi-Omics Approach

Stress disorders have dramatically increased in recent decades becoming the most prevalent psychiatric disorder in the United States and Europe. However, the diagnosis of stress disorders is currently based on symptom checklist and psychological questionnaires, thus making the identification of candidate biomarkers necessary to gain better insights into this pathology and its related metabolic alterations. Regarding the identification of potential biomarkers, omic profiling and metabolic footprint arise as promising approaches to recognize early biochemical changes in such disease and provide opportunities for the development of integrative candidate biomarkers. Here, we studied plasma and urine metabolites together with metagenomics in a 3 days Chronic Unpredictable Mild Stress (3d CUMS) animal approach that aims to focus on the early stress period of a well-established depression model. The multi-omics integration showed a profile composed by a signature of eight plasma metabolites, six urine metabolites and five microbes. Specifically, threonic acid, malic acid, alpha-ketoglutarate, succinic acid and cholesterol were proposed as key metabolites that could serve as key potential biomarkers in plasma metabolome of early stages of stress. Such findings targeted the threonic acid metabolism and the tricarboxylic acid (TCA) cycle as important pathways in early stress. Additionally, an increase in opportunistic microbes as virus of the Herpesvirales was observed in the microbiota as an effect of the primary stress stages. Our results provide an experimental biochemical characterization of the early stage of CUMS accompanied by a subsequent omic profiling and a metabolic footprinting that provide potential candidate biomarkers.

Metabolomics and lipid profile analysis of Coccomyxa melkonianii SCCA 048

With an unsupervised GC–MS metabolomics approach, polar metabolite changes of the microalgae Coccomyxa melkonianii SCCA 048 grown under standard conditions for seven weeks were studied. C. melkonianii was sampled at the Rio Irvi River, in the mining site of Montevecchio-Ingurtosu (Sardinia, Italy), which is severely contaminated by heavy metals and shows high concentrations of sulfates. The partial-least-square (PLS) analysis of the GC–MS data indicated that growth of C. melkonianii was characterized by an increase of the levels of threonic acid, myo-inositol, malic acid, and fumaric acid. Furthermore, at the sixth week of exponential phase the lipid fingerprint of C. melkonianii was studied by LC-QTOF-MS. C. melkonianii lipid extract characterized through an iterative MS/MS analysis showed the following percent levels: 61.34 ± 0.60% for triacylglycerols (TAG); 11.55 ± 0.09% for diacylglyceryltrimethyl homoserines (DGTS), 11.34 ± 0.10% for sulfoquinovosyldiacylglycerols (SQDG) and, 5.29 ± 0.04% for lysodiacylglyceryltrimethyl homoserines (LDGTS). Noteworthy, we were able to annotate different fatty acid ester of hydroxyl fatty acid, such as FAHFA (18:1_20:3), FAHFA (18:2_20:4), FAHFA (18:0_20:2), and FAHFA (18:1_18:0), with relevant biological activity. These approaches can be useful to study the biochemistry of this extremophile algae in the view of its potential exploitation in the phycoremediation of polluted mining areas.

AtFAHD1a: A New Player Influencing Seed Longevity and Dormancy in Arabidopsis?

Fumarylacetoacetate hydrolase (FAH) proteins form a superfamily found in Archaea, Bacteria, and Eukaryota. However, few fumarylacetoacetate hydrolase domain (FAHD)-containing proteins have been studied in Metazoa and their role in plants remains elusive. Sequence alignments revealed high homology between two Arabidopsis thaliana FAHD-containing proteins and human FAHD1 (hFAHD1) implicated in mitochondrial dysfunction-associated senescence. Transcripts of the closest hFAHD1 orthologue in Arabidopsis (AtFAHD1a) peak during seed maturation drying, which influences seed longevity and dormancy. Here, a homology study was conducted to assess if AtFAHD1a contributes to seed longevity and vigour. We found that an A. thaliana T-DNA insertional line (Atfahd1a-1) had extended seed longevity and shallower thermo-dormancy. Compared to the wild type, metabolite profiling of dry Atfahd1a-1 seeds showed that the concentrations of several amino acids, some reducing monosaccharides, and δ-tocopherol dropped, whereas the concentrations of dehydroascorbate, its catabolic intermediate threonic acid, and ascorbate accumulated. Furthermore, the redox state of the glutathione disulphide/glutathione couple shifted towards a more reducing state in dry mature Atfahd1a-1 seeds, suggesting that AtFAHD1a affects antioxidant redox poise during seed development. In summary, AtFAHD1a appears to be involved in seed redox regulation and to affect seed quality traits such as seed thermo-dormancy and longevity.

Comparative Metabolomics Unravel the Effect of Magnesium Oversupply on Tomato Fruit Quality and Associated Plant Metabolism

In general, greenhouse cultivation involves the rampant application of chemical fertilizers, with the aim of achieving high yields. Oversaturation with mineral nutrients that aid plant growth, development, and yield may lead to abiotic stress conditions. We explore the effects of excess magnesium on tomato plant metabolism, as well as tomato fruit quality using non-targeted mass spectrometry (MS)-based metabolomic approaches. Tomato plants were subjected to three different experiments, including high magnesium stress (MgH), extremely high magnesium stress (MgEH), and a control with optimal nutrient levels. Leaves, roots, and fruits were harvested at 16 weeks following the treatment. A metabolic pathway analysis showed that the metabolism induced by Mg oversupply was remarkably different between the leaf and root. Tomato plants allocated more resources to roots by upregulating carbohydrate and polyamine metabolism, while these pathways were downregulated in leaves. Mg oversupply affects the fruit metabolome in plants. In particular, the relative abundance of threonic acid, xylose, fucose, glucose, fumaric acid, malic acid, citric acid, oxoglutaric acid, threonine, glutamic acid, phenylalanine, and asparagine responsible for the flavor of tomato fruits was significantly decreased in the presence of Mg oversupply. Altogether, we concluded that Mg oversupply leads to drastically higher metabolite transport from sources (fully expanded leaves) to sinks (young leaves and roots), and thus, produces unfavorable outcomes in fruit quality and development.

Aluminum toxicity could be mitigated with boron by altering the metabolic patterns of amino acids and carbohydrates rather than organic acids in trifoliate orange

Aluminum (Al) toxicity is the main constraint of root growth and productivity on arable acidic soil. Although boron (B) is used to ameliorate Al stress, the exact mechanisms underlying the effects of B on Al-induced alteration on root metabolites are poorly understood, especially in the trifoliate orange, which is an important rootstock in China. Therefore, a hydroponics experiment was conducted to explore the mechanisms of B mitigates Al toxicity in roots of citrus by metabolomics. A total of 60 metabolites were identified and analyzed in the present study. The 17 amino acids and 8 sugars were up-regulated in Al-treated roots, mainly histidine, cycloleucine, asparagine, citrulline, raffinose and trehalose, and increased by 38.5-, 8.7-, 6.0-, 6.0-, 7.5- and 6.6-fold, respectively. Meanwhile, significant down-regulation of aspartic acid, isoleucine, glutamic acid and six sugars were indicated under Al stress. Aluminum induced a decrease of nine organic acids, especially l-malic acid, citric acid and threonic acid, by 98.2, 93.6 and 95.1%, respectively. Interestingly, in the presence of Al, B application decreased the contents of asparagine, cycloleucine, citrulline and histidine as well as myo-inositol, raffinose, galactinol and 3,6-anhydro-d-galactose by 52.2, 57.4, 46.7, 63.0, 65.4, 74.3, 62.5 and 55.0%, respectively. However, there was no obvious difference in the organic acid contents in Al-stressed roots treated with B. Conclusively, our results show that B regulates the metabolic patterns of amino acids and carbohydrates and reduces Al toxicity. Nevertheless, B addition did not affect the Al-induced changes in the metabolic modes of organic acids.

METABOLOMIC PROFILE AT THE DIFFERENT STAGES OF PRUNELLA VULGARIS (LAMIACEAE) ONTOGENESIS AT GROWING IN THE CLIMATE CHAMBER

By gaz chromatography-mass spectrometry (GC-MS) technique based metabolite profiling of the methanol extract derived from leaves of Prunella vulgaris L. (Lamiaceae) was carried out. P. vulgaris was grown in climatic chamber. For every metabolomics profile, we estimated 102 target substances, and 41 of them were identified. Metabolite spectrum on vegetation stage is different than other stage by absent of groups: amino acids, terpenes and lipids. Comparison of the results of statistical analysis based on the data array, including all detected metabolites (metabolite profiling) show, that metabolomes of blossom and fructification stage of plants match often and only a small number of metabolites P. vulgaris at the this stage make it specific. It was studied the spatial heterogeneity in the structure of the metabolic network. The results of it are showed that metabolomes variety of the leaves from one plants much more than local variety in the range of parts of one leaf. Spatial distributions of metabolites of heterogeneity in a leaf have been illustrated on the example of threonic acid.

Systemic alterations in the metabolome of diabetic NOD mice delineate increased oxidative stress accompanied by reduced inflammation and hypertriglyceremia

Nonobese diabetic (NOD) mice are a commonly used model of type 1 diabetes (T1D). However, not all animals will develop overt diabetes despite undergoing similar autoimmune insult. In this study, a comprehensive metabolomic approach, consisting of gas chromatography time-of-flight (GC-TOF) mass spectrometry (MS), ultra-high-performance liquid chromatography-accurate mass quadruple time-of-flight (UHPLC-qTOF) MS and targeted UHPLC-tandem mass spectrometry-based methodologies, was used to capture metabolic alterations in the metabolome and lipidome of plasma from NOD mice progressing or not progressing to T1D. Using this multi-platform approach, we identified >1,000 circulating lipids and metabolites in male and female progressor and nonprogressor animals ( n = 71). Statistical and multivariate analyses were used to identify age- and sex-independent metabolic markers, which best differentiated metabolic profiles of progressors and nonprogressors. Key T1D-associated perturbations were related with 1) increases in oxidation products glucono-δ-lactone and galactonic acid and reductions in cysteine, methionine and threonic acid, suggesting increased oxidative stress; 2) reductions in circulating polyunsaturated fatty acids and lipid signaling mediators, most notably arachidonic acid (AA) and AA-derived eicosanoids, implying impaired states of systemic inflammation; 3) elevations in circulating triacylglyercides reflective of hypertriglyceridemia; and 4) reductions in major structural lipids, most notably lysophosphatidylcholines and phosphatidylcholines. Taken together, our results highlight the systemic perturbations that accompany a loss of glycemic control and development of overt T1D.