Exposure to tobacco smoke during pregnancy has been associated with a series of adverse reproductive outcomes; however, the underlying molecular mechanisms are not well-established. We conducted an untargeted metabolome-wide association study to identify the metabolic perturbations and molecular mechanisms underlying the association between cotinine, a widely used biomarker of tobacco exposure, and adverse birth outcomes. We collected early and late pregnancy urine samples for cotinine measurement and serum samples for high-resolution metabolomics (HRM) profiling from 105 pregnant women from the Atlanta African American Maternal-Child cohort (2014–2016). Maternal metabolome perturbations mediating prenatal tobacco smoke exposure and adverse birth outcomes were assessed by an untargeted HRM workflow using generalized linear models, followed by pathway enrichment analysis and chemical annotation, with a meet-in-the-middle approach. The median maternal urinary cotinine concentrations were 5.93 μg/g creatinine and 3.69 μg/g creatinine in early and late pregnancy, respectively. In total, 16,481 and 13,043 metabolic features were identified in serum samples at each visit from positive and negative electrospray ionization modes, respectively. Twelve metabolic pathways were found to be associated with both cotinine concentrations and adverse birth outcomes during early and late pregnancy, including tryptophan, histidine, urea cycle, arginine, and proline metabolism. We confirmed 47 metabolites associated with cotinine levels, preterm birth, and shorter gestational age, including glutamate, serine, choline, and taurine, which are closely involved in endogenous inflammation, vascular reactivity, and lipid peroxidation processes. The metabolic perturbations associated with cotinine levels were related to inflammation, oxidative stress, placental vascularization, and insulin action, which could contribute to shorter gestations. The findings will support the further understanding of potential internal responses in association with tobacco smoke exposures, especially among African American women who are disproportionately exposed to high tobacco smoke and experience higher rates of adverse birth outcomes.

Biomonitoring of aquatic environments requires new tools to characterize the effects of pollutants on living organisms. Zebra mussels (Dreissena polymorpha) from the same site in north-eastern France were caged for two months, upstream and downstream of three wastewater treatment plants (WWTPs) in the international watershed of the Meuse (Charleville-Mézières “CM” in France, Namur “Nam” and Charleroi “Cr” in Belgium). The aim was to test ¹H-NMR metabolomics for the assessment of water bodies’ quality. The metabolomic approach was combined with a more “classical” one, i.e., the measurement of a range of energy biomarkers: lactate dehydrogenase (LDH), lipase, acid phosphatase (ACP) and amylase activities, condition index (CI), total reserves, electron transport system (ETS) activity and cellular energy allocation (CEA). Five of the eight energy biomarkers were significantly impacted (LDH, ACP, lipase, total reserves and ETS), without a clear pattern between sites (Up and Down) and stations (CM, Nam and Cr). The metabolomic approach revealed variations among the three stations, and also between the upstream and downstream of Nam and CM WWTPs. A total of 28 known metabolites was detected, among which four (lactate, glycine, maltose and glutamate) explained the observed metabolome variations between sites and stations, in accordance with chemical exposure levels. Metabolome changes suggest that zebra mussel exposure to field contamination could alter their osmoregulation and anaerobic metabolism capacities. This study reveals that lactate is a potential biomarker of interest, and ¹H-NMR metabolomics can be an efficient approach to assess the health status of zebra mussels in the biomonitoring of aquatic environments.

Plastic particles, which are formed from routinely used plastics and their fragments, have become a new pollutant raising widespread concern about their potential effects. Several studies have been conducted to examine their toxicity, but the effects of nano-sized plastic fragments on freshwater organisms remain largely unclear and need to be further investigated. In this study, larval tilapia were first exposed to 100 nm polystyrene nanoparticles (PS-NPs, 20 mg/L) for seven days and then returned to freshwater without PS-NPs for another seven days in order to determine the toxic effects of PS-NPs at both transcriptomic and metabolomic levels. A total of 203 significantly changed metabolites, and 2,152 differentially expressed unigenes were identified between control and PS-NP treatment groups, control and recovery groups, as well as treatment and recovery groups. Our data suggested that PS-NPs induced abnormal metabolism of glycolipids, energy, and amino acids in tilapia after short-term exposure. Additionally, PS-NPs caused disturbed signaling, as suggested by the transcriptomic results. Different transcriptomic and metabolomic levels between the treatment group and recovery group indicated a persistent impact of PS-NPs on tilapia. The presence of adhesion molecule-related differentially expressed genes (DEGs) suggested that PS-NPs might cause early inflammatory responses. Notably, the detection of chemical stimulus involved in the sensory perception of smell was the most severely impacted biological process. Our work systemically studied the ecotoxicity of nano-sized plastics in aquatic creatures at the molecular and genetic levels, serving as a basis for future investigations on the prevention and treatment of such pollutants.

The typical alkyl organophosphorus flame retardant tributyl phosphate (TnBP) can leak from common products into the marine environment, with potential negative effects on marine organisms. However, risk assessments for TnBP regarding zooplankton are lacking. In this study, a marine rotifer, Brachionus plicatilis, was used to analyze the effect of TnBP (0.1 μg/L, environmental concentration; 1 and 6 mg/L) on reproduction, population growth, oxidative stress, mitochondrial function and metabolomics. Mortality increased as the TnBP concentration rose; the 24-h LC₅₀ value was 12.45 mg/L. All tested TnBP concentrations inhibited B. plicatilis population growth, with reproductive toxicity at the higher levels. Microstructural imaging showed ovary injury, the direct cause of reproductive toxicity. Despite elevated glutathione reductase activities, levels of reactive oxygen species and malonyldialdehyde increased under TnBP stress, indicating oxidative imbalance. TnBP induced mitochondrial malformation and activity suppression; the ROS scavenger N-acetylcysteine alleviated this inhibition, suggesting an internal connection. Nontargeted metabolomics revealed 398 and 583 differentially expressed metabolites in the 0.1 μg/L and 6 mg/L treatments relative to control, respectively, which were enriched in the pathways such as biosynthesis of amino acids, purine metabolism, aminoacyl-tRNA biosynthesis. According to metabolic pathway analysis, oxidative stress from purine degradation, mitochondrial dysfunction, disturbed lipid metabolism and elevated protein synthesis were jointly responsible for reproduction and population growth changes. This study echoes the results previously found in rotifer on trade-off among different life processes in response to environmental stress. Our systematic study uncovers the TnBP toxic mode of action.

Excess nitrate has been reported to be associated with many adverse effects in humans and experimental animals. However, there is a paucity of information of the effects of nitrate on intestinal microbial community. In this study, the effects of nitrate on development, intestinal microbial community, and metabolites of Bufo gargarizans tadpoles were investigated. B. gargarizans were exposed to control, 5, 20 and 100 mg/L nitrate-nitrogen (NO₃–N) from eggs to Gosner stage 38. Our data showed that the body size of tadpoles significantly decreased in the 20 and 100 mg/L NO₃–N treatment group when compared to control tadpoles. Exposure to 20 and 100 mg/L NO₃–N also caused indistinct cell boundaries and nuclear pyknosis of mucosal epithelial cells in intestine of tadpoles. In addition, exposure to NO₃–N significantly altered the intestinal microbiota diversity and structure. The facultative anaerobic Proteobacteria occupy the niche of the obligately anaerobic Bacteroidetes and Fusobacteria under the pressure of NO₃–N exposure. According to the results of functional prediction, NO₃–N exposure affected the fatty acid metabolism pathway and amino acid metabolism pathway. The whole-body fatty acid components were found to be changed after exposure to 100 mg/L NO₃–N. Therefore, we concluded that exposure to 20 and 100 mg/L NO₃–N could induce deficient nutrient absorption in intestine, resulting in malnutrition of B. gargarizans tadpoles. High levels of NO₃–N could also change the intestinal microbial communities, causing dysregulation of fatty acid metabolism and amino acid metabolism in B. gargarizans tadpoles.

Although bisphenol A (BPA) is commonly recognized as an endocrine disruptor, the metabolic consequences of its exposure are still poorly understood. In this study, we present a non-targeted LC-MS based metabolomic analysis in combination with a full-genome, high-throughput RNA sequencing (RNA-Seq) to reveal the metabolic effects and the subjacent regulatory pathways of exposing zebrafish embryos to BPA during the first 120 hours post-fertilization. We applied multivariate data analysis methods to extract biochemical information from the LC-MS and RNA-Seq complex datasets and to perform testable predictions of the phenotypic adverse effects. Metabolomic and transcriptomic data revealed a similar subset of altered pathways, despite the large difference in the number of identified biomarkers (around 50 metabolites and more than 1000 genes). These results suggest that even a moderate coverage of zebrafish metabolome may be representative of the global metabolic changes. These multi-omic responses indicate a specific metabolic disruption by BPA affecting different signaling pathways, such as retinoid and prostaglandin metabolism. The combination of transcriptomic and metabolomic data allowed a dynamic interpretation of the results that could not be drawn from either single dataset. These results illustrate the utility of -omic integrative analyses for characterizing the physiological effects of toxicants beyond the mere indication of the affected pathways.

Mercury (Hg) is recognized as a dangerous contaminant due to its bioaccumulation and biomagnification within trophic levels, leading to serious health risks to aquatic biota. Therefore, there is an urgent need to unravel the mechanisms underlying the toxicity of Hg. To this aim, a metabolomics approach based on protonic nuclear magnetic resonance (1H NMR), coupled with chemometrics, was performed on the gills of wild golden grey mullets L. aurata living in an Hg-polluted area in Ria de Aveiro (Portugal). Gills were selected as target organ due to their direct and continuous interaction with the surrounding environment. As a consequence of accumulated inorganic Hg and methylmercury, severe changes in the gill metabolome were observed, indicating a compromised health status of mullets. Numerous metabolites, i.e. amino acids, osmolytes, carbohydrates, and nucleotides, were identified as potential biomarkers of Hg toxicity in fish gills. Specifically, decrease of taurine and glycerophosphocholine, along with increased creatine level, suggested Hg interference with the ion-osmoregulatory processes. The rise of lactate indicated anaerobic metabolism enhancement. Moreover, the increased levels of amino acids suggested the occurrence of protein catabolism, further supported by the augmented alanine, involved in nitrogenous waste excretion. Increased level of isobutyrate, a marker of anoxia, was suggestive of onset of hypoxic stress at the Hg contaminated site. Moreover, the concomitant reduction in glycerophosphocholine and phosphocholine reflected the occurrence of membrane repair processes. Finally, perturbation in antioxidant defence system was revealed by the depletion in glutathione and its constituent amino acids. All these data were also compared to the differential Hg-induced metabolic responses previously observed in liver of the same mullets (Brandão et al., 2015). Overall, the environmental metabolomics approach demonstrated its effectiveness in the evaluation of Hg toxicity mechanisms in wild fish under realistic environmental conditions, uncovering tissue-specificities regarding Hg toxic effects namely in gills and liver.

We evaluated the correlation between soil organic carbon (OC) content and metabolic responses of Eisenia fetida earthworms after exposure to phenanthrene (58 ± 3 mg/kg) spiked into seven artificial soils with OC contents ranging from 1 to 27% OC. Principal component analysis of 1H nuclear magnetic resonance (NMR) spectra of aqueous extracts identified statistically significant differences in the metabolic profiles of control and phenanthrene-exposed E. fetida in the 1% OC soil only. Partial least squares analysis identified a metabolic response in the four soils with OC values ≤11% which was well correlated to estimated phenanthrene porewater concentrations. The results suggest that the higher sorption capability of high OC soils decreased the bioavailability of phenanthrene and the subsequent metabolic response of E. fetida.

One-dimensional (1-D) and two-dimensional (2-D) nuclear magnetic resonance (NMR)-based metabolomics was used to investigate the toxic mode of action (MOA) of endosulfan, an organochlorine pesticide, and its degradation product, endosulfan sulfate, to Eisenia fetida earthworms in soil. Three soil concentrations (0.1, 1.0 and 10.0 mg/kg) were used for both endosulfan and endosulfan sulfate. Both earthworm coelomic fluid (CF) and tissues were extracted and then analyzed using 1H and 1H–13C NMR techniques. A similar separation trajectory was observed for endosulfan and endosulfan sulfate-exposed earthworms in the mean principal component analysis (PCA) scores plot for both the earthworm CF and tissue extracts.A neurotoxic and apoptotic MOA was postulated for both endosulfan and endosulfan sulfate exposed earthworms as significant fluctuations in glutamine/GABA–glutamate cycle metabolites and spermidine were detected respectively. This study highlights the application of NMR-based metabolomics to understand molecular-level toxicity of persistent organochlorine pesticides and their degradation products directly in soil.

Metabolic profiling in plants can be used to differentiate between treatments and to search for biomarkers for exposure. A methodology for processing Ultra-High-Performance Liquid Chromatography–Diode-Array-Detection data is devised. This methodology includes a scheme for selecting informative wavelengths, baseline removal, retention time alignment, selection of relevant retention times, and principal component analysis (PCA). Plant crude extracts from rapeseed seedling exposed to sublethal concentrations of glyphosate are used as a study case. Through this approach, plants exposed to concentrations down to 5 μM could be distinguished from the controls. The compounds responsible for this differentiation were partially identified and were different from those specific for high exposure samples, which suggests that two different responses to glyphosate are elicited in rapeseed depending on the level of exposure. The PCA loadings indicate that a combination of other metabolites could be more sensitive than the response of shikimate to detect glyphosate exposure.