Cluster analysis and profiling of airway fluid metabolites in pediatric acute hypoxemic respiratory failure

Hierarchal clustering of amino acid metabolites may identify a metabolic signature in children with pediatric acute hypoxemic respiratory failure. Seventy-four immunocompetent children, 41 (55.4%) with pediatric acute respiratory distress syndrome (PARDS), who were between 2 days to 18 years of age and within 72 h of intubation for acute hypoxemic respiratory failure, were enrolled. We used hierarchal clustering and partial least squares-discriminant analysis to profile the tracheal aspirate airway fluid using quantitative LC–MS/MS to explore clusters of metabolites that correlated with acute hypoxemia severity and ventilator-free days. Three clusters of children that differed by severity of hypoxemia and ventilator-free days were identified. Quantitative pathway enrichment analysis showed that cysteine and methionine metabolism, selenocompound metabolism, glycine, serine and threonine metabolism, arginine biosynthesis, and valine, leucine, and isoleucine biosynthesis were the top five enriched, impactful pathways. We identified three clusters of amino acid metabolites found in the airway fluid of intubated children important to acute hypoxemia severity that correlated with ventilator-free days < 21 days. Further studies are needed to validate our findings and to test our models.

Transcriptome Analysis of Neuroendocrine Regulation of Ovine Hypothalamus-Pituitary-Ovary Axis during Ovine Anestrus and the Breeding Season

Most sheep are seasonal estrus, and they breed in autumn when the days get shorter. Seasonal estrus is an important factor that affects the productivity and fertility of sheep. The key point to solve this problem is to explore the regulation mechanism of estrus in sheep. Therefore, in this study, transcriptomic sequencing technology was used to identify differentially expressed mRNAs in the hypothalamus, pituitary and ovary of Small Tail Han sheep (year-round estrus) and tan sheep (seasonal estrus) among luteal, proestrus and estrus stages. There were 256,923,304,156 mRNAs being identified in the hypothalamus, pituitary and ovary, respectively. Functional analysis showed that the photosensor, leucine and isoleucine biosynthesis pathways were enriched significantly. It is speculated that photoperiod may initiate estrus by stimulating the corresponding pathways in hypothalamus. ODC1, PRLH, CRYBB2, SMAD5, OPN1SW, TPH1 are believed to be key genes involved in the estrogen process. In conclusion, this study expanded the database of indigenous sheep breeds, and also provided new candidate genes for future genetic and molecular studies on the seasonal estrus trait in sheep.

Screening of Altered Metabolites and Metabolic Pathways in Celiac Disease Using NMR Spectroscopy

Background. Celiac disease (CeD) is an autoimmune intestinal disorder caused by gluten protein consumption in genetically predisposed individuals. As biopsy sampling is an invasive procedure, finding novel noninvasive serological markers for screening of at-risk CeD population is a priority. Metabolomics is helpful in monitoring metabolite changes in body fluids and tissues. In the present study, we evaluated serum metabolite levels of CeD patients relative to healthy controls with the aim of introducing new biomarkers for population screening. Method. We compared the serum metabolic profile of CeD patients ( n = 42 ) and healthy controls ( n = 22 ) using NMR spectroscopy and multivariate analysis. Result. 25 metabolites were identified by serum metabolic profiling. Levels of 3-hydroxyisobutyric acid and isobutyrate showed significant differences in CeD patients’ samples compared with healthy controls ( p < 0.05 ). According to pathway analysis, our data demonstrated that changes in nine metabolic pathways were significantly disrupted/affected in patients with CeD. These enriched pathways are involved in aminoacyl-tRNA biosynthesis; primary bile acid biosynthesis; nitrogen metabolism; glutamine and glutamate metabolism; valine, leucine, and isoleucine biosynthesis and degradation; taurine and hypotaurine metabolism; glyoxylate and dicarboxylate metabolism; glycine, serine, and threonine metabolism; and arginine biosynthesis. Conclusion. In summary, our results demonstrated that changes in the serum level of 25 metabolites may be useful in distinguishing CeD patients from healthy controls, which have the potential to be considered candidate biomarkers of CeD.

Analysis of the Involvement of the Isoleucine Biosynthesis Pathway in Photoheterotrophic Metabolism of Rhodospirillum rubrum

Purple non-sulfur bacteria (PNSB) are recognized as a highly versatile group of bacteria that assimilate a broad range of carbon sources. Growing heterotrophically, PNSB such as Rhodospirillum rubrum (Rs. rubrum) generate reduced equivalents that are used for biomass production. However, under photoheterotrophic conditions, more reduced electron carriers than required to produce biomass are generated. The excess of reduced equivalents still needs to be oxidized for the metabolism to optimally operate. These metabolic reactions are known as electron sinks. Most PNSB rely on the CO2-fixing Calvin cycle and H2 production to oxidize these reduced equivalents. In addition to these well-described electron sinks, the involvement of some pathways, such as polyhydroxyalkanoate (PHA) biosynthesis, in redox poise is still controversial and requires further studies. Among them, isoleucine biosynthesis has been recently highlighted as one of these potential pathways. Here, we explore the role of isoleucine biosynthesis in Rs. rubrum. Our results demonstrate that the isoleucine content is higher under illuminated conditions and that submitting Rs. rubrum to light stress further increases this phenomenon. Moreover, we explore the production of (p)ppGpp in Rs. rubrum and its potential link with light stress. We further demonstrate that a fully functional isoleucine biosynthesis pathway could be an important feature for the onset of Rs. rubrum growth under photoheterotrophic conditions even in the presence of an exogenous isoleucine source. Altogether, our data suggest that isoleucine biosynthesis could play a key role in redox homeostasis.

Metabolic profiling of liver and faeces in mice infected with echinococcosis

Background Echinococcosis is a severe zoonotic parasitic disease which severely affects the health of the hosts. The diagnosis of echinococcosis depends mainly on imaging examination. However, the patient is often in the late stage of the disease when the symptoms appear, thus limiting the early diagnosis of echinococcosis. The treatment and prognosis of the patients are hampered because of long-term asymptomatic latency. Metabolomics is a new discipline developed in the late 1990s. It reflects a series of biological responses in pathophysiological processes by demonstrating the changes in metabolism under the influence of internal and external factors. When the organism is invaded by pathogens, the alteration in the characteristics of metabolites in cells becomes extremely sensitive. Here, we used a metabolomics approach involving liquid chromatography coupled with tandem mass spectrometry (LC–MS/MS) to determine the molecular mechanism of cystic echinococcosis (CE) and to develop an effective method for CE diagnosis. Methods Twenty 8-week-old female BALB/c mice were divided into normal and Echinococcus granulosus infection groups. To develop the E. granulosus infection model, mice were infected with protoscoleces. Six weeks later, the abdomens of the mice showed significant bulging. An LC–MS/MS system-based metabolomics approach was used to analyse the liver and faeces to reveal the metabolic profiles of mice with echinococcosis. Results We found that the metabolism of nucleotides, alkaloids, amino acids, amides, and organic acids in mice is closely interrelated with E. granulosus infection. In the liver, the metabolic pathways of tyrosine and tryptophan biosynthesis; phenylalanine, valine, leucine and isoleucine biosynthesis; and phenylalanine metabolism were notably associated with the occurrence and development of hydatid disease, and in the faeces, pantothenate and CoA biosynthesis are thought to be closely associated with the development of CE. Conclusion The metabolomics approach used in this study provides a reference for a highly sensitive and specific diagnostic and screening method for echinococcosis. Graphic Abstract

Substitution of histone H3 arginine 72 to alanine leads to deregulation of isoleucine biosynthesis in budding yeast Saccharomyces cerevisiae

Histone residues play an essential role in the regulation of various biological processes. In the present study, we have utilized the H3/H4 histone mutant library to probe functional aspects of histone residues in amino acid biosynthesis. We found that histone residue H3R72 plays a crucial role in the regulation of isoleucine biosynthesis. Substitution of arginine residue (H3R72) of histone H3 to alanine (H3R72A) renders yeast cells unable to grow in the minimal media. Histone mutant H3R72A requires the external supplementation of either isoleucine, serine, or threonine for the growth in minimal media. We also observed that H3R72 residue and leucine amino acid in synthetic complete media might play a crucial role in determining the intake of isoleucine and threonine in yeast. Further, gene deletion analysis of ILV1 and CHA1 in H3R72A mutant confirmed that isoleucine is the sole requirement for growth in minimal medium. Altogether, we have identified that histone H3R72 residue may be crucial for yeast growth in the minimal medium by regulating isoleucine biosynthesis through the Ilv1 enzyme in budding yeast Saccharomyces cerevisiae.

Underground isoleucine biosynthesis pathways in E. coli

The promiscuous activities of enzymes provide fertile ground for the evolution of new metabolic pathways. Here, we systematically explore the ability of E. coli to harness underground metabolism to compensate for the deletion of an essential biosynthetic pathway. By deleting all threonine deaminases, we generated a strain in which isoleucine biosynthesis was interrupted at the level of 2-ketobutyrate. Incubation of this strain under aerobic conditions resulted in the emergence of a novel 2-ketobutyrate biosynthesis pathway based upon the promiscuous cleavage of O-succinyl-L-homoserine by cystathionine γ-synthase (MetB). Under anaerobic conditions, pyruvate formate-lyase enabled 2-ketobutyrate biosynthesis from propionyl-CoA and formate. Surprisingly, we found this anaerobic route to provide a substantial fraction of isoleucine in a wild-type strain when propionate is available in the medium. This study demonstrates the selective advantage underground metabolism offers, providing metabolic redundancy and flexibility which allow for the best use of environmental carbon sources.

Integrative Analysis of Metabolomic and Transcriptomic Profiles Uncovers Biological Pathways of Feed Efficiency in Pigs

Feed efficiency (FE) is an economically important trait. Thus, reliable predictors would help to reduce the production cost and provide sustainability to the pig industry. We carried out metabolome-transcriptome integration analysis on 40 purebred Duroc and Landrace uncastrated male pigs to identify potential gene-metabolite interactions and explore the molecular mechanisms underlying FE. To this end, we applied untargeted metabolomics and RNA-seq approaches to the same animals. After data quality control, we used a linear model approach to integrate the data and find significant differently correlated gene-metabolite pairs separately for the breeds (Duroc and Landrace) and FE groups (low and high FE) followed by a pathway over-representation analysis. We identified 21 and 12 significant gene-metabolite pairs for each group. The valine-leucine-isoleucine biosynthesis/degradation and arginine-proline metabolism pathways were associated with unique metabolites. The unique genes obtained from significant metabolite-gene pairs were associated with sphingolipid catabolism, multicellular organismal process, cGMP, and purine metabolic processes. While some of the genes and metabolites identified were known for their association with FE, others are novel and provide new avenues for further research. Further validation of genes, metabolites, and gene-metabolite interactions in larger cohorts will elucidate the regulatory mechanisms and pathways underlying FE.

Metagenomic analysis reveals significant differences in microbiome and metabolic profiles in the rumen of sheep fed low N diet with increased urea supplementation

ABSTRACT Urea is a cost-effective replacement for feed proteins in ruminant diets. However, its metabolism by the rumen microbiome is not fully understood. Here, rumen contents were collected from 18 male sheep fed one of the following three treatments: a low N basal diet with no urea (UC, 0 g/kg dry matter (DM)), low urea (LU, 10 g/kg DM) and high urea (HU, 30 g/kg DM). Principal coordinate analysis showed that the microbial composition and functional profiles of the LU treatment significantly differed from the UC and HU treatments. The genera Prevotella, Succinivibrio, Succinatimonas and Megasphaera were higher in the LU rumen, while the genera Clostridium, Ruminococcus and Butyrivibrio were enriched in the UC and HU rumen. The aspartate–glutamate and arginine–proline metabolic pathways and valine, leucine and isoleucine biosynthesis were higher in the LU rumen. The cysteine and methionine metabolism, lysine degradation and fructose and pentose phosphate metabolism pathways were higher in the UC and HU rumen. The protozoa population in the HU treatment was higher than in the UC and LU treatments. These findings suggest that the rumen microbiome of sheep fed low N diet with different urea supplementation are significantly different.