Placental Metabolomics of Fetal Growth Restriction.

Fetal growth restriction is an obstetrical pathological condition that causes high neonatal mortality and morbidity. The mechanisms of its onset are not completely understood. Metabolites were extracted from 493 placentas from non-complicated pregnancies in Hamilton Country, TN (USA), and analyzed by gas chromatography-mass spectrometry (GC-MS). Newborns were classified according to raw fetal weight (low birth weight (LBW; <2500 g) and non-low birth weight (Non-LBW; >2500 g)), and according to the calculated birth weight centile as it relates to gestational age (small for gestational age (SGA), large for gestational age (LGA), and adequate for gestational age (AGA)). Mothers of LBW infants had a lower pre-pregnancy weight (66.2 ± 17.9 kg vs. 73.4 ± 21.3 kg, p < 0.0001), a lower body mass index (BMI) (25.27 ± 6.58 vs. 27.73 ± 7.83, p < 0.001), and a shorter gestation age (246.4 ± 24.0 days vs. 267.2 ± 19.4 days p < 0.001) compared with non-LBW. Marital status, tobacco use, and fetus sex affected birth weight centile classification according to gestational age. Multivariate statistical comparisons of the extracted metabolomes revealed that asparagine, aspartic acid, deoxyribose, erythritol, glycerophosphocholine, tyrosine, isoleucine, serine, and lactic acid were higher in both SGA and LBW placentas, while taurine, ethanolamine, ß-hydroxybutyrate, and glycine were lower in both SGA and LBW. Several metabolic pathways are implicated in fetal growth restriction, including those related to the hypoxia response and amino-acid uptake and metabolism. Inflammatory pathways are also involved, suggesting that fetal growth restriction might share some mechanisms with preeclampsia.

A screening test proposal for congenital defects based on maternal serum metabolomics profile.

BACKGROUND: Historically, noninvasive techniques are only able to identify chromosomal anomalies that accounted for <50% of all congenital defects; the other congenital defects are diagnosed via ultrasound evaluations in the later stages of pregnancy. Metabolomic analysis may provide an important improvement, potentially addressing the need for novel noninvasive and multicomprehensive early prenatal screening tools. A growing body of evidence outlines notable metabolic alterations in different biofluids derived from pregnant women carrying fetuses with malformations, suggesting that such an approach may allow the discovery of biomarkers common to most fetal malformations. In addition, metabolomic investigations are inexpensive, fast, and risk-free and often generate high performance screening tests that may allow early detection of a given pathology. OBJECTIVE: This study aimed to evaluate the diagnostic accuracy of an ensemble machine learning model based on maternal serum metabolomic signatures for detecting fetal malformations, including both chromosomal anomalies and structural defects. STUDY DESIGN: This was a multicenter observational retrospective study that included 2 different arms. In the first arm, a total of 654 Italian pregnant women (334 cases with fetuses with malformations and 320 controls with normal developing fetuses) were enrolled and used to train an ensemble machine learning classification model based on serum metabolomics profiles. In the second arm, serum samples obtained from 1935 participants of the New Zealand Screening for Pregnancy Endpoints study were blindly analyzed and used as a validation cohort. Untargeted metabolomics analysis was performed via gas chromatography-mass spectrometry. Of note, 9 individual machine learning classification models were built and optimized via cross-validation (partial least squares-discriminant analysis, linear discriminant analysis, naïve Bayes, decision tree, random forest, k-nearest neighbor, artificial neural network, support vector machine, and logistic regression). An ensemble of the models was developed according to a voting scheme statistically weighted by the cross-validation accuracy and classification confidence of the individual models. This ensemble machine learning system was used to screen the validation cohort. RESULTS: Significant metabolic differences were detected in women carrying fetuses with malformations, who exhibited lower amounts of palmitic, myristic, and stearic acids; N-α-acetyllysine; glucose; L-acetylcarnitine; fructose; para-cresol; and xylose and higher levels of serine, alanine, urea, progesterone, and valine (P<.05), compared with controls. When applied to the validation cohort, the screening test showed a 99.4%±0.6% accuracy (specificity of 99.9%±0.1% [1892 of 1894 controls correctly identified] with a sensitivity of 78%±6% [32 of 41 fetal malformations correctly identified]). CONCLUSION: This study provided clinical validation of a metabolomics-based prenatal screening test to detect the presence of congenital defects. Further investigations are needed to enable the identification of the type of malformation and to confirm these findings on even larger study populations.

Exogenous polyunsaturated fatty acids (PUFAs) influence permeability, antimicrobial peptide resistance, biofilm formation and membrane phospholipid structure in an A-layer and non-A-layer strain of Aeromonas salmonicida.

Aeromonas salmonicida is a Gram-negative bacterium that can infect a wide host range of fish populations, including salmonids and non-salmonids as well as freshwater and marine life. Some strains of A. salmonicida cause the disease furunculosis, which can cause lethargy, intestinal inflammation, ulcers, haemorrhaging and death. The infection is spread through fish-to-fish contact, and the presence of infection can have devastating effects on cultivated fish populations. The purpose of this study was to explore the ability of non-A-layer and A-layer A. salmonicida strains to incorporate polyunsaturated fatty acids (PUFAs) into their lipid profile and test the phenotypic effects thereof. Lipids were extracted from PUFA-exposed cultures and analysed for lipid modification by thin-layer chromatography and ultraperformance liquid chromatography-mass spectrometry, showing A. salmonicida, regardless of A-layer, capable of incorporating all seven of the PUFAs studied. Phenotypic effects were determined through the use of assays that tested for biofilm formation, membrane permeability and cyclic peptide susceptibility. Temperature-dependent effects on biofilm formation were observed, and PUFA exposure showed significant (p < .001) increases in membrane permeability as tested by the uptake of the hydrophobic compounds crystal violet and ethidium bromide. Additionally, some PUFAs elicited modest protection and vulnerability against the membrane-targeting cyclic peptides polymyxin B (PMB) and colistin. The diverse, strain-specific responses to exogenous PUFAs may allude to evolved adaptive strategies that enhance survival, persistence and virulence of non-pathogenic and pathogenic members of bacteria that oscillate between environmental and fish host niches.

The Metabolomic Approach for the Screening of Endometrial Cancer: Validation from a Large Cohort of Women Scheduled for Gynecological Surgery.

Endometrial cancer (EC) is the most common gynecological neoplasm in high-income countries. Five-year survival rates are related to stage at diagnosis, but currently, no validated screening tests are available in clinical practice. The metabolome offers an unprecedented overview of the molecules underlying EC. In this study, we aimed to validate a metabolomics signature as a screening test for EC on a large study population of symptomatic women. Serum samples collected from women scheduled for gynecological surgery (n = 691) were separated into training (n = 90), test (n = 38), and validation (n = 563) sets. The training set was used to train seven classification models. The best classification performance during the training phase was the PLS-DA model (96% accuracy). The subsequent screening test was based on an ensemble machine learning algorithm that summed all the voting results of the seven classification models, statistically weighted by each models' classification accuracy and confidence. The efficiency and accuracy of these models were evaluated using serum samples taken from 871 women who underwent endometrial biopsies. The EC serum metabolomes were characterized by lower levels of serine, glutamic acid, phenylalanine, and glyceraldehyde 3-phosphate. Our results illustrate that the serum metabolome can be an inexpensive, non-invasive, and accurate EC screening test.

A Metabolomics-Based Screening Proposal for Colorectal Cancer.

Colorectal cancer (CRC) is a high incidence disease, characterized by high morbidity and mortality rates. Early diagnosis remains challenging because fecal occult blood screening tests have performed sub-optimally, especially due to hemorrhoidal, inflammatory, and vascular diseases, while colonoscopy is invasive and requires a medical setting to be performed. The objective of the present study was to determine if serum metabolomic profiles could be used to develop a novel screening approach for colorectal cancer. Furthermore, the study evaluated the metabolic alterations associated with the disease. Untargeted serum metabolomic profiles were collected from 100 CRC subjects, 50 healthy controls, and 50 individuals with benign colorectal disease. Different machine learning models, as well as an ensemble model based on a voting scheme, were built to discern CRC patients from CTRLs. The ensemble model correctly classified all CRC and CTRL subjects (accuracy = 100%) using a random subset of the cohort as a test set. Relevant metabolites were examined in a metabolite-set enrichment analysis, revealing differences in patients and controls primarily associated with cell glucose metabolism. These results support a potential use of the metabolomic signature as a non-invasive screening tool for CRC. Moreover, metabolic pathway analysis can provide valuable information to enhance understanding of the pathophysiological mechanisms underlying cancer. Further studies with larger cohorts, including blind trials, could potentially validate the reported results.

Polyunsaturated fatty acids cause physiological and behavioral changes in Vibrio alginolyticus and Vibrio fischeri.

Vibrio alginolyticus and Vibrio (Aliivibrio) fischeri are Gram-negative bacteria found globally in marine environments. During the past decade, studies have shown that certain Gram-negative bacteria, including Vibrio species (cholerae, parahaemolyticus, and vulnificus) are capable of using exogenous polyunsaturated fatty acids (PUFAs) to modify the phospholipids of their membrane. Moreover, exposure to exogenous PUFAs has been shown to affect certain phenotypes that are important factors of virulence. The purpose of this study was to investigate whether V. alginolyticus and V. fischeri are capable of responding to exogenous PUFAs by remodeling their membrane phospholipids and/or altering behaviors associated with virulence. Thin-layer chromatography (TLC) analyses and ultra-performance liquid chromatography-electrospray ionization mass spectrometry (UPLC/ESI-MS) confirmed incorporation of all PUFAs into membrane phosphatidylglycerol and phosphatidylethanolamine. Several growth phenotypes were identified when individual fatty acids were supplied in minimal media and as sole carbon sources. Interestingly, several PUFAs acids inhibited growth of V. fischeri. Significant alterations to membrane permeability were observed depending on fatty acid supplemented. Strikingly, arachidonic acid (20:4) reduced membrane permeability by approximately 35% in both V. alginolyticus and V. fischeri. Biofilm assays indicated that fatty acid influence was dependent on media composition and temperature. All fatty acids caused decreased swimming motility in V. alginolyticus, while only linoleic acid (18:2) significantly increased swimming motility in V. fischeri. In summary, exogenous fatty acids cause a variety of changes in V. alginolyticus and V. fischeri, thus adding these bacteria to a growing list of Gram-negatives that exhibit versatility in fatty acid utilization and highlighting the potential for environmental PUFAs to influence phenotypes associated with planktonic, beneficial, and pathogenic associations.

Exogenous fatty acids alter phospholipid composition, membrane permeability, capacity for biofilm formation, and antimicrobial peptide susceptibility in Klebsiella pneumoniae.

Klebsiella pneumoniae represents a major threat to human health due to a combination of its nosocomial emergence and a propensity for acquiring antibiotic resistance. Dissemination of the bacteria from its native intestinal location creates severe, complicated infections that are particularly problematic in healthcare settings. Thus, there is an urgency for identifying novel treatment regimens as the incidence of highly antibiotic-resistant bacteria rises. Recent findings have highlighted the ability of some Gram-negative bacteria to utilize exogenous fatty acids in ways that modify membrane phospholipids and influence virulence phenotypes, such as biofilm formation and antibiotic resistance. This study explores the ability of K. pneumoniae to assimilate and respond to exogenous fatty acids. The combination of thin-layer chromatography liquid chromatography-mass spectrometry confirmed adoption of numerous exogenous polyunsaturated fatty acids (PUFAs) into the phospholipid species of K. pneumoniae. Membrane permeability was variably affected as determined by two dye uptake assays. Furthermore, the availability of many PUFAs lowered the MICs to the antimicrobial peptides polymyxin B and colistin. Biofilm formation was significantly affected depending upon the supplemented fatty acid.

Placental concentrations of essential, toxic, and understudied metals and relationships with birth outcomes in Chattanooga, TN.

BACKGROUND: Comprehensive examinations of placental metal concentrations and correlations with infant parameters are under-investigated. Chattanooga, Tennessee's consistently high incidence of low birth weight and potential for metal exposure provides an ideal opportunity to investigate potential correlations. OBJECTIVES: To investigate the associations between a wide variety of metals in placental tissue and multiple infant parameters. METHODS: A total of 31 elements were screened via ICP-MS in 374 individual placental samples. Of those, 14 were quantifiable in > 86% of the samples. We examined correlations between metal concentrations and infant parameters (birth weight, gestational age, birth weight centile, placental weight, birth length and head circumference). We fit multivariable regression models to estimate the covariate-adjusted associations of birth weight with ln-transformed concentrations of each of the 14 metals and used generalized additive models to examine nonlinear relationships. RESULTS: Some of the strongest relationships with infant parameters came from several lesser-studied metals. Placental rhodium concentrations were negatively correlated with almost all infant parameters. In the fully adjusted regression model, birth weight was significantly associated with several metals. On an IQR (25th to the 75th percentile) basis, estimated changes in birthweight were: for cobalt (82.5 g, IQR=6.05 µg/kg, p = 0.006), iron (-51.5 g, IQR = 171800 µg/kg, p = 0.030), manganese (-27.2 g, IQR=152.1 µg/kg, p = 0.017), lead (-72.7 g, IQR=16.55 µg/kg, p = 0.004) and rhodium (-1365.5 g, IQR = 0.33 µg/kg, p < 0.001). Finally, a generalized additive model showed significant nonlinear relationships between birth weight and concentrations of Co and Rh. CONCLUSIONS: Our comprehensive examination of placental metals illustrate many strong associations between lesser-studied metals and infant parameters. These data, in combination with our correlations of well-studied metals, illustrate a need to consider in utero exposure to a broad array of metals when considering fetal development.

Exogenous polyunsaturated fatty acids (PUFAs) alter phospholipid composition, membrane permeability, biofilm formation and motility in Acinetobacter baumannii.

Acinetobacter baumannii is a ubiquitous multidrug-resistant bacteria that is found on a variety of surfaces, including skin, hair and soil. During the past decade, A. baumannii has emerged as a significant cause of nosocomial infections in the United States. Recent studies have highlighted the ability of some bacteria to utilize a wide variety of fatty acids as a membrane remodelling strategy. Considering this, we hypothesized that fatty acids may have an effect on the emerging pathogen A. baumannii. Thin-layer chromatography indicated structural alterations to major phospholipids. Liquid chromatography/mass spectrometry confirmed the assimilation of numerous exogenous polyunsaturated fatty acids (PUFAs) into the phospholipid species of A. baumannii. The incorporation of fatty acids affected several bacterial phenotypes, including membrane permeability, biofilm formation, surface motility and antimicrobial peptide resistance.

Exogenous Polyunsaturated Fatty Acids Impact Membrane Remodeling and Affect Virulence Phenotypes among Pathogenic Vibrio Species.

The pathogenic Vibrio species (V. cholerae, V. parahaemolyticus, and V. vulnificus) represent a constant threat to human health, causing foodborne and skin wound infections as a result of ingestion of or exposure to contaminated water and seafood. Recent studies have highlighted Vibrio's ability to acquire fatty acids from environmental sources and assimilate them into cell membranes. The possession and conservation of such machinery provokes consideration of fatty acids as important factors in the pathogenic lifestyle of Vibrio species. The findings here link exogenous fatty acid exposure to changes in bacterial membrane phospholipid structure, permeability, phenotypes associated with virulence, and consequent stress responses that may impact survival and persistence of pathogenic Vibrio species. Polyunsaturated fatty acids (PUFAs) (ranging in carbon length and unsaturation) supplied in growth medium were assimilated into bacterial phospholipids, as determined by thin-layer chromatography and liquid chromatography-mass spectrometry. The incorporation of fatty acids variably affected membrane permeability, as judged by uptake of the hydrophobic compound crystal violet. For each species, certain fatty acids were identified as affecting resistance to antimicrobial peptide treatment. Significant fluctuations were observed with regard to both motility and biofilm formation following growth in the presence of individual PUFAs. Our results illustrate the important and complex roles of exogenous fatty acids in the membrane physiology and virulence of a bacterial genus that inhabits aquatic and host environments containing an abundance of diverse fatty acids.IMPORTANCE Bacterial responses to fatty acids include, but are not limited to, degradation for metabolic gain, modification of membrane lipids, alteration of protein function, and regulation of gene expression. Vibrio species exhibit significant diversity with regard to the machinery known to participate in the uptake and incorporation of fatty acids into their membranes. Both aquatic and host niches occupied by Vibrio are rife with various free fatty acids and fatty acid-containing lipids. The roles of fatty acids in the environmental survival and pathogenesis of bacteria have begun to emerge and are expected to expand significantly. The current study demonstrates the responsiveness of V. cholerae, V. parahaemolyticus, and V. vulnificus to exogenous PUFAs. In addition to phospholipid remodeling, PUFA assimilation impacts membrane permeability, motility, biofilm formation, and resistance to polymyxin B.