Multi-Omics Profiling of Candida albicans Grown on Solid Versus Liquid Media

Funding Information: Special thanks to the University of Sharjah for the financial support. Competitive grant No: 2201110155 MHS; Seed grant No: 2001110138. This research is part of the Human Disease Biomarkers Discovery Research Group study. Publisher Copyright: © 2023 by the authors.Candida albicans is a common pathogenic fungus that presents a challenge to healthcare facilities. It can switch between a yeast cell form that diffuses through the bloodstream to colonize internal organs and a filamentous form that penetrates host mucosa. Understanding the pathogen’s strategies for environmental adaptation and, ultimately, survival, is crucial. As a complementary study, herein, a multi-omics analysis was performed using high-resolution timsTOF MS to compare the proteomes and metabolomes of Wild Type (WT) Candida albicans (strain DK318) grown on agar plates versus liquid media. Proteomic analysis revealed a total of 1793 proteins and 15,013 peptides. Out of the 1403 identified proteins, 313 proteins were significantly differentially abundant with a p-value < 0.05. Of these, 156 and 157 proteins were significantly increased in liquid and solid media, respectively. Metabolomics analysis identified 192 metabolites in total. The majority (42/48) of the significantly altered metabolites (p-value 0.05 FDR, FC 1.5), mainly amino acids, were significantly higher in solid media, while only 2 metabolites were significantly higher in liquid media. The combined multi-omics analysis provides insight into adaptative morphological changes supporting Candida albicans’ life cycle and identifies crucial virulence factors during biofilm formation and bloodstream infection.publishersversionpublishe

Proteomic and metabolomic profiling of methicillin resistant versus methicillin sensitive Staphylococcus aureus using a simultaneous extraction protocol

Background: Understanding the biology of methicillin resistant Staphylococcus aureus (MRSA) is crucial to unlocking insights for new targets in our fight against this antimicrobial resistant priority pathogen. Although proteomics and metabolomic profiling offer the potential to elucidating such biological markers, reports of methodological approaches for carrying this out in S. aureus isolates remain limited. We describe the use of a dual-functionality methanol extraction method for the concurrent extraction of protein and metabolites from S. aureus and report on the comparative analysis of the proteomic and metabolomic profiles of MRSA versus methicillin sensitive S. aureus (MSSA). Methods: Bacterial reference strains MRSA ATCC43300 and MSSA ATCC25923 were used. The conventional urea methodology was used for protein extraction and a methanol based method was used for concurrent proteins and metabolites extraction. Proteomic and metabolomic profiling was carried out using TimsTOF mass spectrometry. Data processing was carried out using the MaxQuant version 2.1.4.0. Results: This study represents the first report on the utilization of the methanol extraction method for concurrent protein and metabolite extraction in Gram positive bacteria. Our findings demonstrate good performance of the method for the dual extraction of proteins and metabolites from S. aureus with demonstration of reproducibility. Comparison of MRSA and MSSA strains revealed 407 proteins with significantly different expression levels. Enrichment analysis of those proteins revealed distinct pathways involved in fatty acid degradation, metabolism and beta-lactam resistance. Penicillin-binding protein PBP2a, the key determinant of MRSA resistance, exhibited distinct expression patterns in MRSA isolates. Metabolomic analysis identified 146 metabolites with only one exclusive to the MRSA. The enriched pathways identified were related to arginine metabolism and biosynthesis. Conclusion: Our findings demonstrate the effectiveness of the methanol-based dual-extraction method, providing simultaneous insights into the proteomic and metabolomic landscapes of S. aureus strains. These findings demonstrate the utility of proteomic and metabolomic profiling for elucidating the biological basis of antimicrobial resistance

Proteomic and metabolomic profiling of methicillin resistant versus methicillin sensitive Staphylococcus aureus using a simultaneous extraction protocol

BackgroundUnderstanding the biology of methicillin resistant Staphylococcus aureus (MRSA) is crucial to unlocking insights for new targets in our fight against this antimicrobial resistant priority pathogen. Although proteomics and metabolomic profiling offer the potential to elucidating such biological markers, reports of methodological approaches for carrying this out in S. aureus isolates remain limited. We describe the use of a dual-functionality methanol extraction method for the concurrent extraction of protein and metabolites from S. aureus and report on the comparative analysis of the proteomic and metabolomic profiles of MRSA versus methicillin sensitive S. aureus (MSSA).MethodsBacterial reference strains MRSA ATCC43300 and MSSA ATCC25923 were used. The conventional urea methodology was used for protein extraction and a methanol based method was used for concurrent proteins and metabolites extraction. Proteomic and metabolomic profiling was carried out using TimsTOF mass spectrometry. Data processing was carried out using the MaxQuant version 2.1.4.0.ResultsThis study represents the first report on the utilization of the methanol extraction method for concurrent protein and metabolite extraction in Gram positive bacteria. Our findings demonstrate good performance of the method for the dual extraction of proteins and metabolites from S. aureus with demonstration of reproducibility. Comparison of MRSA and MSSA strains revealed 407 proteins with significantly different expression levels. Enrichment analysis of those proteins revealed distinct pathways involved in fatty acid degradation, metabolism and beta-lactam resistance. Penicillin-binding protein PBP2a, the key determinant of MRSA resistance, exhibited distinct expression patterns in MRSA isolates. Metabolomic analysis identified 146 metabolites with only one exclusive to the MRSA. The enriched pathways identified were related to arginine metabolism and biosynthesis.ConclusionOur findings demonstrate the effectiveness of the methanol-based dual-extraction method, providing simultaneous insights into the proteomic and metabolomic landscapes of S. aureus strains. These findings demonstrate the utility of proteomic and metabolomic profiling for elucidating the biological basis of antimicrobial resistance

Effects of medwakh smoking on salivary metabolomics and its association with altered oral redox homeostasis among youth

Abstract The use of alternative tobacco products, particularly medwakh, has expanded among youth in the Middle East and around the world. The present study is conducted to investigate the biochemical and pathophysiological changes caused by medwakh smoking, and to examine the salivary metabolomics profile of medwakh smokers. Saliva samples were collected from 30 non-smokers and 30 medwakh smokers and subjected to metabolomic analysis by UHPLC-ESI-QTOF-MS. The CRP and Glutathione Peroxidase 1 activity levels in the study samples were quantified by ELISA and the total antioxidant capacity (TAC) by TAC assay kits. Statistical measurements and thorough validation of data obtained from untargeted metabolomics identified 37 uniquely and differentially abundant metabolites in saliva of medwakh smokers. The levels of phthalate, L-sorbose, cytosine, uridine, alpha-hydroxy hippurate, and L-nicotine were noticeably high in medwakh smokers. Likewise, 20 metabolic pathways were differentially altered in medwakh smokers. This study identified a distinctive saliva metabolomics profile in medwakh smokers associated with altered redox homeostasis, metabolic pathways, antioxidant system, and CRP levels. The impact of the altered metabolites in medwakh smokers and their diagnostic utility require further research in large cohorts

Mass spectrometric analysis of bioactive conditioned media of bacteria isolated from reptilian gut

AIM: To determine whether selected gut bacteria of crocodile exhibit antibacterial properties.MATERIALS &amp; METHODS: Two bacteria isolated from Crocodylus porosus gut were used, namely: Pseudomonas aeruginosa and Aeromonas dhakensis. Conditioned media were tested against pathogenic bacteria and metabolites were analyzed using liquid chromatography-mass spectrometry.RESULTS &amp; CONCLUSION: Antibacterial assays revealed that conditioned media showed potent effects against pathogenic Gram-positive and Gram-negative bacteria. LC-MS revealed identity of 210 metabolites. The abundant metabolites were, N-Acetyl-L-tyrosine, Acetaminophen, Trans-Ferulic acid, N, N-Dimethylformamide, Pyrocatechol, Cyclohexanone, Diphenhydramine, Melatonin, Gamma-terpinene, Cysteamine, 3-phenoxypropionic acid, Indole-3-carbinol, Benzaldehyde, Benzocaine, 2-Aminobenzoic acid, 3-Methylindole. These findings suggest that crocodile gut bacteria are potential source of novel bioactive molecules that can be utilized as pre/post/antibiotics for the benefit of human health.</p

Acute venous thromboembolism plasma and red blood cell metabolomic profiling reveals potential new early diagnostic biomarkers: observational clinical study

Abstract Background Venous thromboembolism (VTE) is a leading cause of cardiovascular mortality. The diagnosis of acute VTE is based on complex imaging exams due to the lack of biomarkers. Recent multi-omics based research has contributed to the development of novel biomarkers in cardiovascular diseases. Our aim was to determine whether patients with acute VTE have differences in the metabolomic profile compared to non-acute VTE. Methods This observational trial included 62 patients with clinical suspicion of acute deep vein thrombosis or pulmonary embolism, admitted to the emergency room. There were 50 patients diagnosed with acute VTE and 12 with non-acute VTE conditions and no significant differences were found between the two groups for clinical and demographic characteristics. Metabolomics assays identified and quantified a final number of 91 metabolites in plasma and 55 metabolites in red blood cells (RBCs). Plasma from acute VTE patients expressed tendency to a specific metabolomic signature, with univariate analyses revealing 23 significantly different molecules between acute VTE patients and controls (p < 0.05). The most relevant metabolic pathway with the strongest impact on the acute VTE phenotype was d-glutamine and d-glutamate (p = 0.001, false discovery rate = 0.06). RBCs revealed a specific metabolomic signature in patients with a confirmed diagnosis of DVT or PE that distinguished them from other acutely diseased patients, represented by 20 significantly higher metabolites and four lower metabolites. Three of those metabolites revealed high performant ROC curves, including adenosine 3′,5′-diphosphate (AUC 0.983), glutathione (AUC 0.923), and adenine (AUC 0.91). Overall, the metabolic pathway most impacting to the differences observed in the RBCs was the purine metabolism (p = 0.000354, false discovery rate = 0.68). Conclusions Our findings show that metabolite differences exist between acute VTE and nonacute VTE patients admitted to the ER in the early phases. Three potential biomarkers obtained from RBCs showed high performance for acute VTE diagnosis. Further studies should investigate accessible laboratory methods for the future daily practice usefulness of these metabolites for the early diagnosis of acute VTE in the ER

Serum multi-omics analysis in hindlimb unloading mice model:Insights into systemic molecular changes and potential diagnostic and therapeutic biomarkers

Microgravity, in space travel and prolonged bed rest conditions, induces cardiovascular deconditioning along with skeletal muscle mass loss and weakness. The findings of microgravity research may also aid in the understanding and treatment of human health conditions on Earth such as muscle atrophy, and cardiovascular diseases. Due to the paucity of biomarkers and the unknown underlying mechanisms of cardiovascular and skeletal muscle deconditioning in these environments, there are insufficient diagnostic and preventative measures. In this study, we employed hindlimb unloading (HU) mouse model, which mimics astronauts in space and bedridden patients, to first evaluate cardiovascular and skeletal muscle function, followed by proteomics and metabolomics LC-MS/MS-based analysis using serum samples. Three weeks of unloading caused changes in the function of the cardiovascular system in c57/Bl6 mice, as seen by a decrease in mean arterial pressure and heart weight. Unloading for three weeks also changed skeletal muscle function, causing a loss in grip strength in HU mice and atrophy of skeletal muscle indicated by a reduction in muscle mass. These modifications were partially reversed by a two-week recovery period of reloading condition, emphasizing the significance of the recovery process. Proteomics analysis revealed 12 dysregulated proteins among the groups, such as phospholipid transfer protein, Carbonic anhydrase 3, Parvalbumin alpha, Major urinary protein 20 (Mup20), Thrombospondin-1, and Apolipoprotein C-IV. On the other hand, metabolomics analysis showed altered metabolites among the groups such as inosine, hypoxanthine, xanthosine, sphinganine, L-valine, 3,4-Dihydroxyphenylglycol, and L-Glutamic acid. The joint data analysis revealed that HU conditions mainly impacted pathways such as ABC transporters, complement and coagulation cascades, nitrogen metabolism, and purine metabolism. Overall, our results indicate that microgravity environment induces significant alterations in the function, proteins, and metabolites of these mice. These observations suggest the potential utilization of these proteins and metabolites as novel biomarkers for assessing and mitigating cardiovascular and skeletal muscle deconditioning associated with such conditions.</p

Abstract Tu025: Exploring Metabolomic Profiles in Vitamin D Deficient and Obese Individuals: Relevance to Arterial Stiffness

Introduction: Cardiovascular diseases (CVDs) continue to pose a significant global health challenge, with vitamin D deficiency and obesity emerging as prominent risk factors. Arterial stiffness is recognized as a pivotal predictor and an independent risk element for CVDs. This study explores the metabolomic profiling of blood samples obtained from individuals afflicted with early arterial stiffness due to vitamin D deficiency and obesity, compared with a control group. Methods: The study comprised nine participants with vitamin D deficiency (vitamin D level of 20 ng/ml or lower), and obese participants (BMI of 30 or higher), along with eleven control participants (with vitamin D levels above 20 ng/ml and normal BMI). Pulse Wave Velocity (PWV) measurements were conducted using the SphygmoCor device. Whole blood was collected from each participant in EDTA tubes, followed by centrifugation at 4200 RPM for 10 minutes to obtain plasma samples, which were then stored in a -80°C freezer. Metabolomic analysis was carried out using Liquid Chromatography coupled with tandem mass spectrometry (LC-MS/MS). A two-tailed p-value &lt; 0.05 was considered statistically significant for all analyses. Results: The average PWV value relative to the participant's age was 19.4 ± 4.7 m/s in the group with both vitamin D deficiency and obesity, compared to 14.7 ± 2.1 m/s in the control group (p &lt; 0.05), indicating increased arterial stiffness. Furthermore, eleven metabolites exhibited statistically significant differences in patients with vitamin D deficiency and obesity. Of these, eight metabolites demonstrated increased expression levels, including Nutriacholic acid, N-Acetylputrescine, Succinylacetone, Adenosine monophosphate, Elaidic acid, Niacinamide, DUMP, and 2-Pyrrolidinone, while three metabolites exhibited decreased expression: PC (18:1(9Z)/18:1(9Z)), Trimethylamine, and Imidazole. Additionally, top enrichment analysis revealed metabolic pathways predicted to be altered in the context of dysfunctional enzymes, encompassing processes such as nicotinamide acid uptake, fatty-acyl-CoA synthase (n-C18:0CoA), and extracellular NADP nucleosidase, among others. Conclusion: The metabolomic alterations identified in our study offer insights into potential biomarkers for evaluating arterial stiffness and present novel targets for therapeutic interventions in vitamin D deficient and obese individuals

Evaluation of Two Simultaneous Metabolomic and Proteomic Extraction Protocols Assessed by Ultra-High-Performance Liquid Chromatography Tandem Mass Spectrometry

Untargeted multi-omics analysis of plasma is an emerging tool for the identification of novel biomarkers for evaluating disease prognosis, and for developing a better understanding of molecular mechanisms underlying human disease. The successful application of metabolomic and proteomic approaches relies on reproducibly quantifying a wide range of metabolites and proteins. Herein, we report the results of untargeted metabolomic and proteomic analyses from blood plasma samples following analyte extraction by two frequently-used solvent systems: chloroform/methanol and methanol-only. Whole blood samples were collected from participants (n = 6) at University Hospital Sharjah (UHS) hospital, then plasma was separated and extracted by two methods: (i) methanol precipitation and (ii) 4:3 methanol:chloroform extraction. The coverage and reproducibility of the two methods were assessed by ultra-high-performance liquid chromatography-electrospray ionization quadrupole time-of-flight mass spectrometry (UHPLC-ESI-QTOF-MS). The study revealed that metabolite extraction by methanol-only showed greater reproducibility for both metabolomic and proteomic quantifications than did methanol/chloroform, while yielding similar peptide coverage. However, coverage of extracted metabolites was higher with the methanol/chloroform precipitation