Nucleotide Research Articles

Molecular Characterization of the Chicken Parvovirus Based on VP1 Gene Circulating in Brazilian Chicken Flocks.

Parvovirus infection affects several animal species, especially young animals. In birds, parvovirus infection has been described in Muscovy ducks, turkeys, and chickens, all of which had enteric diseases characterized by diarrhea. Chicken parvovirus (ChPV) has been detected in poultry around the world in animals affected by enteric problems, showing dwarfism, cloacal pasting, and diarrhea. In Brazil, ChPV was detected in chickens affected by diarrhea fifteen years ago. However, the genetic characteristics of ChPV circulating in chicken flocks were not determined. Therefore, the aim of the present investigation was to determine the genetic characteristics of the VP1 gene from ChPV detected in chickens affected by enteric diseases in Brazil. For this purpose, a molecular approach was used. Specific primers were designed to flank the complete VP1 gene of ChPV and amplify it using PCR. The amplified products from samples of chickens with enteric diseases were sequenced, and 22 complete CDs of the VP1 gene were obtained. These samples, compared to the ABU-P1 sequence, showed 17 sequences with high nucleotide (NT) similarity of 92.7-97.4% and amino acid (AA) similarity of 94.8-99.5% associated with Runting and Stunting syndrome (RSS); there were also five samples associated with hens with diarrhea with unusual jejunal dilatation (JD) that had less similarity than the RSS sequences (NT of 86.5% and AA of 93-93.1%). The phylogenetic analysis determined four groups. Group I had sequences from Korea. The second group included sequences from Korea, China, and Brazil (not included in this work). The third group had studied RSS sequences grouped with the ABU-P1 strain and sequences from China and the United States. Finally, the sequences from JD were clustered in a separate group with a bootstrap of 100%, a group that was denoted as group IV, and included sequences from China. RDP4 and SimPlot analysis showed one point of recombination with the sequences of group III ChPV in the JD sequences. Herein, we show that circulating strains of ChPV exhibit genetic differences in the VP1 gene in Brazilian chicken flocks. Nevertheless, more studies are needed to determine the probability of a new genetic group of ChPV based on the analysis of the complete genome.

Comparative Metabolome and Transcriptome Analyses Reveal Differential Enrichment of Metabolites with Age in Panax notoginseng Roots.

Panax notoginseng is a perennial plant well known for its versatile medicinal properties, including hepatoprotective, antioxidant, anti-inflammatory, anti-tumor, estrogen-like, and antidepressant characteristics. It has been reported that plant age affects the quality of P. notoginseng. This study aimed to explore the differential metabolome and transcriptome of 2-year (PN2) and 3-year-old (PN3) P. notoginseng plant root samples. Principal component analysis of metabolome and transcriptome data revealed major differences between the two groups (PN2 vs. PN3). A total of 1813 metabolites and 28,587 genes were detected in this study, of which 255 metabolites and 3141 genes were found to be differential (p < 0.05) between PN2 vs. PN3, respectively. Among differential metabolites and genes, 155 metabolites and 1217 genes were up-regulated, while 100 metabolites and 1924 genes were down-regulated. The KEGG pathway analysis revealed differentially enriched metabolites belonging to class lipids ("13S-hydroperoxy-9Z, 11E-octadecadionic acid", "9S-hydroxy-10E, 12Z-octadecadionic acid", "9S-oxo-10E, 12Z-octadecadionic acid", and "9,10,13-trihydroxy-11-octadecadionic acid"), nucleotides and derivatives (guanine and cytidine), and phenolic acids (chlorogenic acid) were found to be enriched (p < 0.05) in PN3 compared to PN2. Further, these differentially enriched metabolites were found to be significantly (p < 0.05) regulated via linoleic acid metabolism, nucleotide metabolism, plant hormone signal transduction, and arachidonic acid metabolism pathways. Furthermore, the transcriptome analysis showed the up-regulation of key genes MAT, DMAS, SDH, gallate 1-beta-glucosyltransferase, and beta-D-glucosidase in various plants' secondary metabolic pathways and SAUR, GID1, PP2C, ETR, CTR1, EBF1/2, and ERF1/2 genes observed in phytohormone signal transduction pathway that is involved in plant growth and development, and protection against the various stressors. This study concluded that the roots of a 3-year-old P. notoginseng plant have better metabolome and transcriptome profiles compared to a 2-year-old plant with importantly enriched metabolites and genes in pathways related to metabolism, plant hormone signal transduction, and various biological processes. These findings provide insights into the plant's dynamic biochemical and molecular changes during its growth that have several implications regarding its therapeutic use.

Transcriptional changes during crown-root development and emergence in barley (Hordeum vulgare L.)

BackgroundRoots play an important role during plant growth and development, ensuring water and nutrient uptake. Understanding the mechanisms regulating their initiation and development opens doors towards root system architecture engineering.ResultsHere, we investigated by RNA-seq analysis the changes in gene expression in the barley stem base of 1 day-after-germination (DAG) and 10DAG seedlings when crown roots are formed. We identified 2,333 genes whose expression was lower in the stem base of 10DAG seedlings compared to 1DAG seedlings. Those genes were mostly related to basal cellular activity such as cell cycle organization, protein biosynthesis, chromatin organization, cytoskeleton organization or nucleotide metabolism. In opposite, 2,932 genes showed up-regulation in the stem base of 10DAG seedlings compared to 1DAG seedlings, and their function was related to phytohormone action, solute transport, redox homeostasis, protein modification, secondary metabolism. Our results highlighted genes that are likely involved in the different steps of crown root formation from initiation to primordia differentiation and emergence, and revealed the activation of different hormonal pathways during this process.ConclusionsThis whole transcriptomic study is the first study aiming at understanding the molecular mechanisms controlling crown root development in barley. The results shed light on crown root emergence that is likely associated with a strong cell wall modification, death of the cells covering the crown root primordium, and the production of defense molecules that might prevent pathogen infection at the site of root emergence.

Identification of blood exosomal metabolomic profiling for high-altitude cerebral edema

High-altitude cerebral edema (HACE) is a severe neurological condition that can occur at high altitudes. It is characterized by the accumulation of fluid in the brain, leading to a range of symptoms, including severe headache, confusion, loss of coordination, and even coma and death. Exosomes play a crucial role in intercellular communication, and their contents have been found to change in various diseases. This study analyzed the metabolomic characteristics of blood exosomes from HACE patients compared to those from healthy controls (HCs) with the aim of identifying specific metabolites or metabolic pathways associated with the development of HACE conditions. A total of 21 HACE patients and 21 healthy controls were recruited for this study. Comprehensive metabolomic profiling of the serum exosome samples was conducted using ultraperformance liquid chromatography–tandem mass spectrometry (UPLC‒MS/MS). Additionally, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was performed to identify the metabolic pathways affected in HACE patients. Twenty-six metabolites, including ( +)-camphoric acid, choline, adenosine, adenosine 5′-monophosphate, deoxyguanosine 5′-monophosphate, guanosine, and hypoxanthine-9-β-D-arabinofuranoside, among others, exhibited significant changes in expression in HACE patients compared to HCs. Additionally, these differentially abundant metabolites were confirmed to be potential biomarkers for HACE. KEGG pathway enrichment analysis revealed several pathways that significantly affect energy metabolism regulation (such as purine metabolism, thermogenesis, and nucleotide metabolism), estrogen-related pathways (the estrogen signaling pathway, GnRH signaling pathway, and GnRH pathway), cyclic nucleotide signaling pathways (the cGMP-PKG signaling pathway and cAMP signaling pathway), and hormone synthesis and secretion pathways (renin secretion, parathyroid hormone synthesis, secretion and action, and aldosterone synthesis and secretion). In patients with HACE, adenosine, guanosine, and hypoxanthine-9-β-D-arabinofuranoside were negatively correlated with height. Deoxyguanosine 5′-monophosphate is negatively correlated with weight and BMI. Additionally, LPE (18:2/0:0) and pregnanetriol were positively correlated with age. This study identified potential biomarkers for HACE and provided valuable insights into the underlying metabolic mechanisms of this disease. These findings may lead to potential targets for early diagnosis and therapeutic intervention in HACE patients.

Toxic mechanisms of the antiviral drug arbidol on microalgae in algal bloom water at transcriptomic level

Increasing antivirals in surface water caused by their excessive consumption pose serious threats to aquatic organisms. Our recent research found that the input of antiviral drug arbidol to algal bloom water can induce acute toxicity to the growth and metabolism of Microcystis aeruginosa, resulting in growth inhibition, as well as decrease in chlorophyll and ATP contents. However, the toxic mechanisms involved remained obscure, which were further investigated through transcriptomic analysis in this study. The results indicated that 885–1248 genes in algae were differentially expressed after exposure to 0.01–10.0 mg/L of arbidol, with the majority being down-regulated. Analysis of commonly down-regulated genes found that the cellular response to oxidative stress and damaged DNA bonding were affected, implying that the stress defense system and DNA repair function of algae might be damaged. The down-regulation of genes in porphyrin metabolism, photosynthesis, carbon fixation, glycolysis, tricarboxylic acid cycle, and oxidative phosphorylation might inhibit chlorophyll synthesis, photosynthesis, and ATP supply, thereby hindering the growth and metabolism of algae. Moreover, the down-regulation of genes related to nucleotide metabolism and DNA replication might influence the reproduction of algae. These findings provided effective strategies to elucidate toxic mechanisms of contaminants on algae in algal bloom water.

Investigation of antimicrobial potential of nanocomposites based on functionalization of graphene oxide with zinc porphyrin complexes

Herein, zinc porphyrin complexes were synthesised by passing through the formation of free base porphyrin ligands and metalation of free base porphyrins with zinc metal ions. The synthesised zinc porphyrin complexes were functionalised with graphene oxide leading to the formation of zinc porphyrin anchored graphene oxide hybrid nanocomposites. The zinc porphyrins complexes were anchored with graphene oxide through the particular functional group present at the periphery of zinc porphyrins (hydroxyl and carboxyl functional groups in Pr1 and Pr2 respectively). 1H NMR and UV–visible spectroscopic techniques have monitored the successful formation of free base porphyrins and their matching zinc-integrated metalloporphyrin complexes. Various spectroscopic techniques like, photophysical properties (UV–Visible spectroscopy and Fluorescence spectroscopy), Fourier transform infrared (FT-IR), powdered X-ray diffraction (P-XRD) patterns, FE-SEM and EDAX analysis were used to examine the intrinsic characteristics of the prepared nanocomposites. The in-vitro antimicrobial activity of the synthesized complexes and nanocomposites was tested against Staphylococcus aureus gram-positive strain, Klebsiella pneumonia, and Escherichia coli. By working on the antimicrobial behaviour, it has been observed that the compound containing –COOH substitute has shown enhanced antimicrobial activity. The in silico molecular docking studies of the synthesized compounds with the S. aureus nucleoside diphosphate kinase receptor showed strong interactions via hydrogen bonding, π − π interactions and hydrophobic interactions. Molecular dynamics (MDs) simulations were used to study the dynamic behaviour of the complexes as well as nanocomposites which revealed the stability of docked structures with S. aureus nucleoside diphosphate kinase receptor. The in-silico pharmacokinetics studies showed the drug-likeness, non-toxic nature and safe oral administration of the synthesized compounds.

Decitabine cytotoxicity is promoted by dCMP deaminase DCTD and mitigated by SUMO-dependent E3 ligase TOPORS

The nucleoside analogue decitabine (or 5-aza-dC) is used to treat several haematological cancers. Upon its triphosphorylation and incorporation into DNA, 5-aza-dC induces covalent DNA methyltransferase 1 DNA–protein crosslinks (DNMT1-DPCs), leading to DNA hypomethylation. However, 5-aza-dC’s clinical outcomes vary, and relapse is common. Using genome-scale CRISPR/Cas9 screens, we map factors determining 5-aza-dC sensitivity. Unexpectedly, we find that loss of the dCMP deaminase DCTD causes 5-aza-dC resistance, suggesting that 5-aza-dUMP generation is cytotoxic. Combining results from a subsequent genetic screen in DCTD-deficient cells with the identification of the DNMT1-DPC-proximal proteome, we uncover the ubiquitin and SUMO1 E3 ligase, TOPORS, as a new DPC repair factor. TOPORS is recruited to SUMOylated DNMT1-DPCs and promotes their degradation. Our study suggests that 5-aza-dC-induced DPCs cause cytotoxicity when DPC repair is compromised, while cytotoxicity in wild-type cells arises from perturbed nucleotide metabolism, potentially laying the foundations for future identification of predictive biomarkers for decitabine treatment.

Risk assessment model based on nucleotide metabolism-related genes highlights SLC27A2 as a potential therapeutic target in breast cancer

PurposeBreast cancer (BC) is the most prevalent malignant tumor worldwide among women, with the highest incidence rate. The mechanisms underlying nucleotide metabolism on biological functions in BC remain incompletely elucidated.Materials and MethodsWe harnessed differentially expressed nucleotide metabolism-related genes from The Cancer Genome Atlas-BRCA, constructing a prognostic risk model through univariate Cox regression and LASSO regression analyses. A validation set and the GSE7390 dataset were used to validate the risk model. Clinical relevance, survival and prognosis, immune infiltration, functional enrichment, and drug sensitivity analyses were conducted.ResultsOur findings identified four signature genes (DCTPP1, IFNG, SLC27A2, and MYH3) as nucleotide metabolism-related prognostic genes. Subsequently, patients were stratified into high- and low-risk groups, revealing the risk model's independence as a prognostic factor. Nomogram calibration underscored superior prediction accuracy. Gene Set Variation Analysis (GSVA) uncovered activated pathways in low-risk cohorts and mobilized pathways in high-risk cohorts. Distinctions in immune cells were noted between risk cohorts. Subsequent experiments validated that reducing SLC27A2 expression in BC cell lines or using the SLC27A2 inhibitor, Lipofermata, effectively inhibited tumor growth.ConclusionsWe pinpointed four nucleotide metabolism-related prognostic genes, demonstrating promising accuracy as a risk prediction tool for patients with BC. SLC27A2 appears to be a potential therapeutic target for BC among these genes.

Disruption of NADPH homeostasis by total flavonoids from Adinandra nitida Merr. ex Li leaves triggers ROS-dependent p53 activation leading to apoptosis in non-small cell lung cancer cells

Ethnopharmacological relevanceAdinandra nitida Merr. ex Li leaves serve as a herbal tea and hold a significant role in traditional Chinese medicine, being applied to assist in tumor treatment. Flavonoids present the primary bioactive constituents in Adinandra nitida Merr. ex Li leaves. Aim of the studyTo explore the potential of total flavonoids from Adinandra nitida Merr. ex Li Leaves (TFAN) in inhibiting non-small cell lung cancer (NSCLC) and further elucidate the underlying mechanisms. Materials and methodsHuman NSCLC cell lines and normal lung cell line were employed to assess the impact of TFAN (0–160 μg/mL for 24, 28 and 72 h) on cell proliferation in vitro. Immunofluorescence (IF) staining gauged p53 expression changes in NSCLC cells under TFAN present condition (150 μg/mL for 24 h). In vivo study utilized NSCLC cell derived xenograft tumors in nude mice, administering TFAN orally (200 and 400 mg/kg) for 14 days. Immunohistochemistry assessed Cleaved Caspase 3 expression change in A549 xenograft tumors treated with TFAN (400 mg/kg for 14 days). RNA-seq and KEGG analysis identified gene expression changes and enriched processes in A549 xenograft tumors treated with TFAN. CM-H2DCFDA and metabolomics assessed ROS level and GSH/GSSG pool changes in A549 cells under TFAN present condition. Cell viability assay and IF staining assessed A549 cell proliferation and p53 expression changes under H2O2-induced oxidative stress (0–40 μM for 24 h) and TFAN present conditions. GSEA and N-Acetyl-L-cysteine (NAC) rescue (0–1 μM for 24 h) analyzed the impact of TFAN on GSH de novo synthesis. NADPH/NADP+ pool measurement and NADPH rescue (0–10 μM for 24 h) analyzed the impact of TFAN on GSH salvage synthesis. GC-FID and HPLC-MS were utilized to detect ethanol and ethyl acetate residues, and to characterize the chemical constituents in TFAN, respectively. The total flavonoid content of TFAN was determined using a 330 nm wavelength. ResultsTFAN significantly inhibited A549 cells (wild-type p53) but not NCI–H1299 cells (p53-deficient), NCI–H596 cells (p53-mutant) or BEAS-2B in vitro. IF staining validated p53 genotype for the cell lines and revealed an increase in p53 expression in A549 cells after TFAN treatment. In vivo, TFAN selectively inhibited A549 xenograft tumor growth without discernible toxicity, inducing apoptosis evidenced by Cleaved Caspase 3 upregulation. RNA-seq and KEGG analysis suggested ROS biosynthesis was involved in TFAN-induced p53 activation in A549 cells. Elevated ROS level in TFAN-treated A549 cells were observed. Moreover, TFAN sensitized A549 cells to H2O2-induced oxidative stress, with higher p53 expression. Additionally, A549 cells compensated with GSH de novo synthesis under TFAN present condition, confirmed by GSEA and NAC rescue experiment. TFAN disrupted NADPH homeostasis to impair GSH salvage biosynthesis, supported by NADPH/NADP+ change and NADPH rescue experiment. The chemical constituents of TFAN, with acceptable limits for ethanol and ethyl acetate residues and a total flavonoid content of 68.87%, included Catechin, Epicatechin, Quercitroside, Camellianin A, and Apigenin. ConclusionThe disruption of NADPH homeostasis by TFAN triggers ROS-dependent p53 activation that leads to apoptotic cell death, ultimately suppressing NSCLC growth. These findings offer potential therapeutic implications of Adinandra nitida Merr. ex Li leaves in combating NSCLC.

An exchange of single amino acid between the phosphohydrolase modules of Escherichia coli MutT and Mycobacterium smegmatis MutT1 switches their cleavage specificities

MutT proteins belong to the Nudix hydrolase superfamily that includes a diverse group of Mg2+ requiring enzymes. These proteins use a generalized substrate, nucleoside diphosphate linked to a chemical group X (NDP-X), to produce nucleoside monophosphate (NMP) and the moiety X linked with phosphate (XP). E. coli MutT (EcoMutT) and mycobacterial MutT1 (MsmMutT1) belong to the Nudix hydrolase superfamily that utilize 8-oxo-(d)GTP (referring to both 8-oxo-GTP or 8-oxo-dGTP). However, predominant products of their activities are different. While EcoMutT produces 8-oxo-(d)GMP, MsmMutT1 gives rise to 8-oxo-(d)GDP. Here, we show that the altered cleavage specificities of the two proteins are largely a consequence of the variation at the equivalent of Gly37 (G37) in EcoMutT to Lys (K65) in the MsmMutT1. Remarkably, mutations of G37K (EcoMutT) and K65G (MsmMutT1) switch their cleavage specificities to produce 8-oxo-(d)GDP, and 8-oxo-(d)GMP, respectively. Further, a time course analysis using 8-oxo-GTP suggests that MsmMutT1(K65G) hydrolyses 8-oxo-(d)GTP to 8-oxo-(d)GMP in a two-step reaction via 8-oxo-(d)GDP intermediate. Expectedly, unlike EcoMutT (G37K) and MsmMutT1, EcoMutT and MsmMutT1 (K65G) rescue an E. coli ΔmutT strain, better by decreasing A to C mutations.

Multi-omics analysis of kidney, bone and bone marrow explored potential mechanisms of Erzhi Wan against osteoporosis with kidney-Yin deficiency

Osteoporosis (OP) is a metabolic bone disease that can lead to major health challenges. The theory of Traditional Chinese medicine believes that kidney-Yin deficiency (KYD) is the main cause of postmenopausal osteoporosis. This study was aimed to investigate the effect of EZW on anti-osteoporosis with KYD, and explore potential mechanisms from the perspective of the kidney, bone and bone marrow through analysis of metabolomics and proteomics. The model of OP with KYD was established by rats treated with bilateral ovariectomy (OVX), and then given intragastric administration of thyroid and reserpine to induce. Micro-CT was applied to determine the microstructures of bone. Serum levels associated with bone turnover markers and kidney-Yin deficiency were detected by enzyme-linked immunosorbent (ELISA) assay. The differential metabolites in the kidney, bone and bone marrow were analyzed by metabolomics. The differentially expressed proteins in these three tissues were detected via proteomics. The findings suggested that EZW could alleviate a variety of metabolites and proteins among the kidney, bone and bone marrow, primarily in amino acid metabolism, carbohydrate metabolism, nucleotide metabolism and lipid metabolism, thus leading to improvements of OP with KYD, which provided theoretical basis for clinical treatment of EZW on OP with KYD.

Identification of metabolic components of carotid plaque in high-risk patients utilizing liquid chromatography-tandem mass spectrometry.

Carotid atherosclerosis is a chronic progressive vascular disease that can be complicated by stroke in severe cases. Prompt diagnosis and treatment of high-risk patients are quite difficult due to the lack of reliable clinical biomarkers. This study aimed to explore potential plaque metabolic markers of stroke-prone risk and relevant targets for pharmacological intervention. Carotid intima and plaque sample tissues were obtained from 20 patients with cerebrovascular symptoms of carotid origin. An untargeted metabolomics approach based on liquid chromatography-tandem mass spectrometry was utilized to characterize the metabolic profiles of the tissues. Multivariate and univariate analysis tools were used. A total of 154 metabolites were significantly altered in carotid plaque when compared with thickened intima. Of these, 62 metabolites were upregulated, whereas 92 metabolites were downregulated. Support vector machines identified the 15 most important metabolites, such as N-(cyclopropylmethyl)-N'-phenylurea, 9(S)-HOTrE, ACar 12:2, quinoxaline-2,3-dithiol, and l-thyroxine, as biomarkers for high-risk plaques. Metabolic pathway analysis showed that abnormal purine and nucleotide metabolism, amino acid metabolism, glutathione metabolism, and vitamin metabolism may contribute to the occurrence and progression of carotid atherosclerotic plaque. Our study identifies the biomarkers and related metabolic mechanisms of carotid plaque, which is stroke-prone, and provides insights and ideas for the precise prevention and targeted intervention of the disease.

New insights into the antibiofilm activity and mechanism of Mannosylerythritol Lipid-A against Listeria monocytogenes EGD-e

Listeria monocytogenes is one of the leading causative agents of foodborne disease outbreaks worldwide. Herein, the antibiofilm effect and mechanism of Mannosylerythritol Lipid-A against L. monocytogenes EGD-e is reported for the first time. MEL-A effectively attenuated biofilm formation while reducing the viability and motility of bacteria within the biofilm in the early stage, and influenced bacterial adhesion by affecting the secretion of extracellular polysaccharides and eDNA. RT-qPCR revealed that MEL-A significantly suppressed the expression of genes involved in flagellar movement and virulence. Untargeted LC-MS metabolomics indicated that MEL-A affected the fluidity and permeability of cell membranes by significantly upregulating unsaturated fatty acids, lipids and glycoside metabolites, and affected protein biosynthesis, nucleotide metabolism and DNA synthesis and repair by significantly downregulating amino acid metabolism and nucleic acid metabolism. These pathways may constitute the key targets of biofilm formation inhibition by MEL-A. Furthermore, MEL-A showed good removal effects on mature biofilms under different temperatures, different materials and milk. Our data indicated that MEL-A could be used as a novel antibiofilm agent to improve food safety. Our study provides new insights into the possible inhibitory mechanism of MEL-A and the response of L. monocytogenes EGD-e to MEL-A.

Comparative metabolomics study on the quality of Antarctic krill (Euphausia superba) stored at different temperatures

SummaryIn this study, the effect of short‐term storage at different temperatures (0, 4 and −8 °C) on the quality of Antarctic krill was investigated using the main quality indicators as a reference, combined with electronic nose and untargeted metabolomics. Antarctic krill stored at lower temperatures (−8 °C) were of better quality and had a flavour similar to that of the original Antarctic krill. Metabolomics analyses showed that differential metabolites (DMS) were more abundant in Antarctic krill at higher temperatures (4 °C), with sixty‐five metabolites having reduced expression and forty‐five metabolites having increased expression; Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that amino acid metabolism and nucleotide metabolism pathways were the Five DMS, including α‐ketoglutarate, glutamate, succinate, sucrose and glutarate, can be used as potential biomarkers to monitor quality changes during short‐term storage of Antarctic krill. In addition, the association between quality indicators and differential metabolites remains to be explored.

Metabolic perturbations in zebrafish (Danio rerio) larvae exposed to sulfentrazone and imidacloprid

The intensive and widespread application of pesticides in agroecosystems can lead to the simultaneous exposure of non-target aquatic organisms to insecticides and herbicides. However, the underlying mechanisms through which aquatic organisms undergo metabolic reprogramming to withstand the combined effects of the insecticide imidacloprid (IMI) and herbicide sulfentrazone (SUL) remain poorly elucidated. This study employs metabolomics to investigate the effects of individual and combined exposures to IMI and SUL on zebrafish (Danio rerio), aiming to simulate complex environmental conditions. Metabolomics analysis revealed extensive metabolic reprogramming in larvae induced by the selected agrochemicals. Both individual and combined exposures disrupted nucleotide metabolism, inhibited glycolysis, and led to the accumulation of acetylcholine through the shared modulation of differential metabolites. Notably, individual exposure exhibited a unique mode of action. Larvae exposed to IMI alone showed mitochondrial dysfunction, potentially stemming from interference with the electron transport chain, while SUL-induced disruptions were associated with glycerophospholipid accumulation, marking it as a critical target. Additionally, calculations of the metabolic effect level index indicated antagonistic interactions between SUL and IMI mixtures at an overall metabolic level. The results obtained through investigating the lethal and sub-lethal effects also revealed that the simultaneous application of SUL and IMI may have the potential to diminish acute and developmental toxicity in zebrafish. This study underscores the significance of metabolomics as a valuable and effective strategy for deciphering the toxicity and interactions of agrochemical mixtures.

Metabolomics unveil key pathways underlying the synergistic activities of aztreonam and avibactam against multidrug-resistant Escherichia coli.

Aztreonam/avibactam is effective against serious infections caused by Gram-negative bacteria including Enterobacterales harboring metallo-β-lactamases. While the utility of this combination has been established in vitro and in clinical trials, the purpose of this study is to enhance our understanding of the underlying mechanism responsible for their activities through metabolomic profiling of a multidrug-resistant Escherichia coli clinical isolate. Metabolomic analyses of time-dependent changes in endogenous bacterial metabolites in a clinical isolate of a multidrug-resistant E. coli treated with aztreonam and avibactam were performed. E. coli metabolomes were compared at 15min, 1h and 24h following treatments with either avibactam (4mg/L), aztreonam (4mg/L), or aztreonam (4mg/L) + avibactam (4mg/L). Drug treatment affected 326 metabolites with magnitude changes of at least 2-fold, most of which are involved primarily in peptidoglycan biosynthesis, nucleotide metabolism, and lipid metabolism.The feedstocks for peptidoglycan synthesis were depleted by aztreonam/avibactam combination;a significant downstream increase in nucleotide metabolites and a release of lipids were observed at the three timepoints. The findings indicate that the aztreonam/avibactam combination accelerates structural damage to the bacterial membrane structure and their actions were immediate and sustained compared to aztreonam or avibactam alone. By inhibiting the production of crucial cell wall precursors, the combination may have inflicted damages on bacterial DNA.

Antibacterial activity of angelica essential oil, its mechanism against Pseudomonas fluorescens, and its application in the preservation of chilled fresh beef

The mechanism by which Angelica essential oil (AEO) can act as an antimicrobial agent at cellular and metabolic levels to inhibit Pseudomonas fluorescens (P. fluorescens), and its effectiveness in beef preservation, were investigated and evaluated in this study. Our results showed that the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of AEO to P. fluorescens were 12.5 μL/mL and 25 μL/mL, respectively, at which AEO could effectively inhibit P. fluorescens reproduction and retard its growth (p < 0.05). Severe morphological damage to the AEO-treated cells of the microorganism, including cell walls and membranes, were observed via scanning electron microscopy (SEM). AEO treatment was also seen to lead to an increase in alkaline phosphatase (AKPase) activity and conductivity, and a significant leakage of intracellular macromolecular components (such as nucleic acids and proteins), providing further evidence of damage to the integrity of cell wall and cell membrane. Metabolomics analysis revealed that the differential metabolites in the AEO hindered nucleic acid synthesis and expression in the P. fluorescens, leading to tricarboxylic acid (TCA) cycle blockage and amino acid restriction, energy metabolism disorders and membrane damage, mainly through nucleotide metabolism, amino acid metabolism, carbohydrate metabolism and membrane transport. Moreover, application testing demonstrated the efficacy of AEO in slowing the increase of beef pH and total volatile basic nitrogen (TVB-N), extending the shelf life of chilled beef by 5.39 d, and effectively maintaining the organoleptic quality of beef. These results lay a foundation for the development of AEO as a natural bacteriostatic agent.

Metabolomics revealed that toxic cyanobacteria stress reduced the flavor quality of Bellamya aeruginosa

The environmental impact of increased eutrophication and frequent cyanobacterial blooms on the growth and reproduction of aquatic organisms has become increasingly prominent in recent years. Bellamya aeruginosa is a nutrient-rich aquatic snail consumed by humans, and environmental changes may decrease its meat quality. However, the specific characteristics of muscle metabolism and flavor quality in freshwater snails and their response patterns to bloom-forming cyanobacterial stress have not been reported. Therefore, in this study, alterations in metabolites in the snail muscle after following feeding on toxic Microcystis aeruginosa were investigated using untargeted metabolomics. A total of 1,128 metabolites were identified, and the metabolic pathways of unsaturated fatty acid biosynthesis and purine, glutathione, and glycerophospholipid metabolism in snails fed toxic cyanobacteria differed from those in snails fed Chlorella vulgaris. Quantitative analysis showed increased levels of bitter-free amino acids, such as tyrosine, phenylalanine, and histidine, after consuming toxic cyanobacteria for 42 d, whereas a relative decrease was observed in the levels of umami- and sweet-free amino acids, such as glutamic acid, aspartic acid, serine, threonine, and glicine. The muscles of snails fed toxic cyanobacteria exhibited higher hypoxanthine and hypoxanthine nucleoside and lower adenosine triphosphate, adenine nucleotide, adenine nucleoside diphosphate, and hypoxanthine nucleotide contents than those of snails fed C. vulgaris. Furthermore, increased metabolites, such as stearic acid, palmitic acid, and cytidine diphosphate choline, and decreased metabolites, such as docosapentaenoic acid, docosahexaenoic acid, adrenoic acid, and L-glutamic acid, reflect the harmful effects of toxic cyanobacteria on the nutritional value and flesh quality of B. aeruginosa. This study comprehensively evaluated the effects of cyanobacterial blooms on freshwater gastropods, providing an important theoretical basis for the quality, safety, and sustainable development of snails as food and related processing industries.

Vitamin B1 Supplementation Ameliorates Obesogenic Effects of a High Fat Diet in Male Mice

Dietary deficiencies in vitamins such as thiamine (Vitamin B1) are common even in developed countries and can have a range of deleterious health effects. This trend in Vitamin B1 deficiency coincides with increased fat consumption in the American diet, particularly of soybean oil, the most widely consumed cooking oil in the U.S. which is also present in many processed foods. Here, we hypothesized that the concurrent malnutrition caused by a soybean oil-based high fat diet (SO-HFD) and a physiological deficiency in thiamine can exacerbate conditions such as obesity, diabetes, and fatty liver that are associated with consumption of HFDs. To test this, C57BL/6N male mice (N=8-10 per group) were fed either a low-fat chow or an SO-HFD, with or without supplementation with 640 mg/kg of the thiamine analog thiamine tetrahydrofurfuryl disulfide (TTFD), for 18 weeks. Our results showed that the SO-HFD supplemented with TTFD leads to significantly decreased weight gain and delayed the onset of glucose intolerance compared to SO-HFD without added TTFD (p&lt;0.05). In contrast, TTFD supplementation did not have any effect on body weight or glucose intolerance in the context of a low-fat diet. Metabolomic analysis revealed that the SO-HFD decreased absolute thiamine levels both in the blood and liver tissue compared to the low fat control diet (p&lt;0.05); this decrease was reversed by TTFD supplementation suggesting that some of the negative effects of the SO-HFD may be due to its impact on the Vitamin B1 pathway. Untargeted metabolomic analysis identified several other metabolites — such as amino acids, organic acids, methylamine osmolytes and those involved in nucleotide metabolism — that were also affected by TTFD supplementation, but in a beneficial fashion. Transcriptomic analysis of the liver and intestines revealed alterations in thiamine absorption and metabolic pathways by the SO-HFD; effects of TTFD on hepatic gene expression and fatty liver will be presented. TTFD supplementation given after the onset of SO-HFD-induced obesity was not able to reverse the phenotype, suggesting that the deleterious effects of the SO-HFD-induced Vitamin B1 deficiency may be permanent. In conclusion, our findings indicate that consumption of an SO-enriched diet, similar in fat content to what is consumed by most Americans, can result in a Vitamin B1 deficiency. Although TTFD supplementation may mitigate some of the adverse metabolic effects of this diet, this intervention must be done at an early stage. Funding: The Thiamine Advocacy Foundation Research Grant (FMS, PHD, PD); NIH R01DK127082 (FMS)Acknowledgements: UCR Genomics Core Facility, UCR Metabolomics Core Facility, UCR High Performance Computing Center (HPCC). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract 2074: De Novo Nucleotide Synthesis Is Necessary For DNA Damage And Repair In Atheroprone Endothelial Cells

Aims: Endothelial cell (EC) metabolism is tightly regulated by hemodynamic changes to protect them from atherogenic injury induced by disturbed flow (d-flow) and other atherogenic risk factors. Therefore, characterizing the role of flow-induced metabolic reprogramming is important to develop new treatments for atherosclerosis and similar diseases. Currently, the effect of proatherogenic d-flow on EC nucleotide metabolism, including de novo purine and pyrimidine synthesis, remains under explored. ATIC and CAD are critical enzymes for de novo purine synthesis and pyrimidine synthesis, respectively. We investigated in the role of ATIC and CAD-mediated de novo nucleotide metabolism in atheroprone ECs. Methods and Results: (i) scRNA-Seq analysis results showed that DNA damage response was increased in ECs exposed to d-flow, this was accompanied by increased ATIC-mediated de novo purine synthesis and CAD-mediated de novo pyrimidine synthesis; (ii) Knockdown of ATIC or CAD with siRNA aggravated EC DNA damage, apoptosis and EC permeability induced by d-flow or other atherogenic factors; (iii) Supplement of nucleobases can rescue ATIC KD-mediated DNA damage and apoptosis in ECs; (iv) MYC, predicted high stringency binding with ATIC promoter, was highly expressed in d-flow induced ECs and MYC knockdown reduced the increased ATIC expression in d-flow induced ECs; (v) The atherosclerotic lesion size was markedly increased in EC-specific Atic deficient mice in atherosclerosis. Conclusion: ATIC and CAD-mediated de novo nucleotide synthesis in ECs supply nucleotides to repair DNA damage and preserve EC barrier integrity in vulnerable atheroprone regions and ultimately protect against the development and progression of atherosclerosis.