Microbial Diversity Impacts Non-Protein Amino Acid Production in Cyanobacterial Bloom Cultures Collected from Lake Winnipeg

Lake Winnipeg in Manitoba, Canada is heavily impacted by harmful algal blooms that contain non-protein amino acids (NPAAs) produced by cyanobacteria: N-(2-aminoethyl)glycine (AEG), β-aminomethyl-L-alanine (BAMA), β-N-methylamino-L-alanine (BMAA), and 2,4-diaminobutyric acid (DAB). Our objective was to investigate the impact of microbial diversity on NPAA production by cyanobacteria using semi-purified crude cyanobacterial cultures established from field samples collected by the Lake Winnipeg Research Consortium between 2016 and 2021. NPAAs were detected and quantified by ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) using validated analytical methods, while Shannon and Simpson alpha diversity scores were determined from 16S rRNA metagenomic sequences. Alpha diversity in isolate cultures was significantly decreased compared to crude cyanobacterial cultures (p < 0.001), indicating successful semi-purification. BMAA and AEG concentrations were higher in crude compared to isolate cultures (p < 0.0001), and AEG concentrations were correlated to the alpha diversity in cultures (r = 0.554; p < 0.0001). BAMA concentrations were increased in isolate cultures (p < 0.05), while DAB concentrations were similar in crude and isolate cultures. These results demonstrate that microbial community complexity impacts NPAA production by cyanobacteria and related organisms.Science, Faculty ofNon UBCChemistry, Department of (Okanagan)ReviewedFacultyResearche

The Morphoregulatory Role of Thidiazuron: Metabolomics-Guided Hypothesis Generation for Mechanisms of Activity

Thidiazuron (TDZ) is a diphenylurea synthetic herbicide and plant growth regulator used to defoliate cotton crops and to induce regeneration of recalcitrant species in plant tissue culture. In vitro cultures of African violet thin petiole sections are an ideal model system for studies of TDZ-induced morphogenesis. TDZ induces de novo shoot organogenesis at low concentrations and somatic embryogenesis at higher concentrations of exposure. We used an untargeted metabolomics approach to identify metabolites in control and TDZ-treated tissues. Statistical analysis including metabolite clustering, pattern and pathway tools, logical algorithms, synthetic biotransformations and hormonomics identified TDZ-induced changes in metabolism. A total of 18,602 putative metabolites with extracted masses and predicted formulae were identified with 1412 features that were found only in TDZ-treated tissues and 312 that increased in response to TDZ. The monomer of TDZ was not detected intact in the tissues but putative oligomers were found in the database and we hypothesize that these may form by a Diels–Alder reaction. Accumulation oligomers in the tissue may act as a reservoir, slowly releasing the active TDZ monomer over time. Cleavage of the amide bridge released TDZ-metabolites into the tissues including organic nitrogen and sulfur containing compounds. Metabolomics data analysis generated six novel hypotheses that can be summarized as an overall increase in uptake of sugars from the culture media, increase in primary metabolism, redirection of terpene metabolism and mediation of stress metabolism via indoleamine and phenylpropanoid metabolism. Further research into the specific mechanisms hypothesized is likely to unravel the mode of action of TDZ and to provide new insights into the control of plant morphogenesis.Science, Faculty ofNon UBCChemistry, Department ofReviewedFacult

Antibacterial Activity and Untargeted Metabolomics Profiling of <i>Acalypha arvensis</i> Poepp

The search for potent antimicrobial compounds is critical in the face of growing antibiotic resistance. This study explores Acalypha arvensis Poepp. (A. arvensis), a Caribbean plant traditionally used for disease treatment. The dried plant powder was subjected to successive extractions using different solvents: hexane (F1), dichloromethane (F2), methanol (F3), a 50:50 mixture of methanol and water (F4), and water (F5). Additionally, a parallel extraction was conducted using a 50:50 mixture of methanol and chloroform (F6). All the fractions were evaluated for their antimicrobial activity, and the F6 fraction was characterized using untargeted metabolomics using SPME-GC×GC-TOFMS. The extracts of A. arvensis F3, F4, and F5 showed antibacterial activity against Staphylococcus aureus ATCC 25923 (5 mg/mL), MRSA BA22038 (5 mg/mL), and Pseudomonas aeruginosa ATCC 27853 (10 mg/mL), and fraction F6 showed antibacterial activity against Staphylococcus aureus ATCC 29213 (2 mg/mL), Escherichia coli ATCC 25922 (20 mg/mL), Pseudomonas aeruginosa ATCC 27853 (10 mg/mL), Enterococcus faecalis ATCC 29212 (10 mg/mL), Staphylococcus aureus 024 (2 mg/mL), and Staphylococcus aureus 003 (2 mg/mL). Metabolomic analysis of F6 revealed 2861 peaks with 58 identified compounds through SPME and 3654 peaks with 29 identified compounds through derivatization. The compounds included methyl ester fatty acids, ethyl ester fatty acids, terpenes, ketones, sugars, amino acids, and fatty acids. This study represents the first exploration of A. arvensis metabolomics and its antimicrobial potential, providing valuable insights for plant classification, phytochemical research, and drug discovery

A Preclinical Model of Obesity-Independent Metabolic Syndrome for Studying the Effects of Novel Antidiabetic Therapy Beyond Glycemic Control

Accumulating data from several large, placebo-controlled studies suggests that sodium-glucose transporter 2 (SGLT-2) inhibitors and glucagon-like peptide 1 receptor (GLP-1) receptor agonists offer therapeutic benefits in the management of cardiovascular diseases, regardless of the patient's diabetic status. In addition to their effects on glucose excretion, SGLT2-inhibitors have a positive impact on systemic metabolism by reducing inflammation and oxidative stress, shifting metabolism towards ketone body production, and suppressing glycation end-product signaling. The aim of this study was to establish a non-invasive preclinical model of metabolic syndrome (MetS) to investigate the effects of novel antidiabetic therapies beyond glucose reduction, independent of obesity. Eighteen healthy adult Beagle dogs were fed isocalorically a Western diet (WD) adjusted from parameters of the National Health and Nutrition Examination Survey for ten weeks. Blood samples were collected at baseline (BAS1) when dogs were fed their regular diet, and then again after ten weeks of WD feeding (BAS2) for measurement of blood count and serum chemistry, lipoprotein profiling, fasting blood glucose, glucagon, insulin, NT-proBNP, BUN, creatinine, angiotensins and oxidative stress biomarkers. Blood pressure (BP) was measured at BAS1 and BAS2 using Doppler. Serum, urine and fecal metabolomics were derived by mass spectrometry to assess general metabolism, complex lipids and biogenic amines. Differences between BAS1 and BAS2 were analyzed using non-parametric Wilcoxon signed rank testing with continuity correction, as appropriate. Body weight changes did not exceed 13% after ten weeks of feeding with the WD. The isocaloric WD model induced significant variations in several markers of MetS, including (1) elevated BP, (2) increased fasting glucose levels, and (3) reduced HDL-cholesterol. It also triggered a significant decrease in circulating insulin, as well as an increase in circulating NT-proBNP levels and a decrease in serum bicarbonate levels. Marked and significant changes in overall metabolism, lipids, and biogenic amines were finally reported at BAS2. Short-term, isocaloric feeding with a WD in dogs replicates key biological features of MetS, while also causing low-grade metabolic acidosis and elevating natriuretic peptides. These findings support the use of the WD canine model for studying the metabolic effects of new antidiabetic therapies independent of obesity.This is a preprint from Mochel, Jonathan P., Jessica L. Ward, Thomas Blondel, Debosmita Kundu, Maria M. Merodio, Claudine Zemirline, Emilie Guillot et al. "A Preclinical Model of Obesity-Independent Metabolic Syndrome for Studying the Effects of Novel Antidiabetic Therapy Beyond Glycemic Control." (2023). doi: https://doi.org/10.21203/rs.3.rs-3569600/v1. Copyright The Authors 2023. This work is licensed under a Creative Commons Attribution 4.0 International License