Metabolomics Reveals Target and Off-Target Toxicities of a Model Organophosphate Pesticide to Roach (Rutilus rutilus): Implications for Biomonitoring

The ability of targeted and nontargeted metabolomics to discover chronic ecotoxicological effects is largely unexplored. Fenitrothion, an organophosphate pesticide, is categorized as a “red list” pollutant, being particularly hazardous to aquatic life. It acts primarily as a cholinesterase inhibitor, but evidence suggests it can also act as an androgen receptor antagonist. Whole-organism fenitrothion-induced toxicity is well-established, but information regarding target and off-target molecular toxicities is limited. Here we study the molecular responses of male roach (Rutilus rutilus) exposed to fenitrothion, including environmentally realistic concentrations, for 28 days. Acetylcholine was assessed in brain; steroid metabolism was measured in testes and plasma; and NMR and mass spectrometry-based metabolomics were conducted on testes and liver to discover off-target toxicity. O-demethylation was confirmed as a major route of pesticide degradation. Fenitrothion significantly depleted acetylcholine, confirming its primary mode of action, and 11-ketotestosterone in plasma and cortisone in testes, showing disruption of steroid metabolism. Metabolomics revealed significant perturbations to the hepatic phosphagen system and previously undocumented effects on phenylalanine metabolism in liver and testes. On the basis of several unexpected molecular responses that were opposite to the anticipated acute toxicity, we propose that chronic pesticide exposure induces an adapting phenotype in roach, which may have considerable implications for interpreting molecular biomarker responses in field-sampled fish.

[1]  S. Bessman,et al.  The creatine-creatine phosphate energy shuttle. , 1985, Annual review of biochemistry.

[2]  S. Morris,et al.  Detection of the anti-androgenic effect of endocrine disrupting environmental contaminants using in vivo and in vitro assays in the three-spined stickleback. , 2009, Aquatic toxicology.

[3]  Mark R. Viant,et al.  MI-Pack: Increased confidence of metabolite identification in mass spectra by integrating accurate masses and metabolic pathways , 2010 .

[4]  Ralf J M Weber,et al.  Discriminating between different acute chemical toxicities via changes in the daphnid metabolome. , 2010, Toxicological sciences : an official journal of the Society of Toxicology.

[5]  E. Hill,et al.  Methodology for profiling the steroid metabolome in animal tissues using ultraperformance liquid chromatography-electrospray-time-of-flight mass spectrometry. , 2008, Analytical chemistry.

[6]  Francesco Falciani,et al.  Discovery of metabolic signatures for predicting whole organism toxicology. , 2010, Toxicological sciences : an official journal of the Society of Toxicology.

[7]  R. Baudinette,et al.  Fenitrothion, an organophosphate, affects running endurance but not aerobic capacity in fat-tailed dunnarts (Sminthopsis crassicaudata). , 2008, Chemosphere.

[8]  B. Borg.  Androgens in teleost fishes , 1994 .

[9]  C. Tyler,et al.  Altered sexual development in roach (Rutilus rutilus) exposed to environmental concentrations of the pharmaceutical 17alpha-ethinylestradiol and associated expression dynamics of aromatases and estrogen receptors. , 2008, Toxicological sciences : an official journal of the Society of Toxicology.

[10]  Edward J Calabrese,et al.  The hormetic dose-response model is more common than the threshold model in toxicology. , 2003, Toxicological sciences : an official journal of the Society of Toxicology.

[11]  M. Brosnan,et al.  Creatine synthesis: hepatic metabolism of guanidinoacetate and creatine in the rat in vitro and in vivo. , 2009, American journal of physiology. Endocrinology and metabolism.

[12]  C. Tyler,et al.  The roach (Rutilus rutilus) as a sentinel for assessing endocrine disruption. , 2007, Environmental sciences : an international journal of environmental physiology and toxicology.

[13]  S. Peng,et al.  Chronic organophosphate (OP)-induced neuropsychiatric disorder is a withdrawal syndrome. , 2009, Medical hypotheses.

[14]  M. Viant,et al.  High-throughput tissue extraction protocol for NMR- and MS-based metabolomics. , 2008, Analytical biochemistry.

[15]  M. Jokanović.  Biotransformation of organophosphorus compounds. , 2001, Toxicology.

[16]  I. Soojhawon,et al.  Biotransformation of nitroaromatics and their effects on mixed function oxidase system , 2005 .

[17]  A. Scott,et al.  Endocrine changes during natural spawning in the white sucker, Catostomus commersoni. II. Steroid hormones. , 1984, General and comparative endocrinology.

[18]  T. Marchant,et al.  Endocrine changes during natural spawning in the white sucker, Catostomus commersoni. I. Gonadotropin, growth hormone, and thyroid hormones. , 1984, General and comparative endocrinology.

[19]  P. Kevan.  Forest application of the insecticide fenitrothion and its effect on wild bee pollinators (Hymenoptera: Apoidea) of lowbush blueberries (Vaccinium SPP.) in Southern New Brunswick, Canada , 1975 .

[20]  A. L. Filby,et al.  Molecular Characterization of Estrogen Receptors 1, 2a, and 2b and Their Tissue and Ontogenic Expression Profiles in Fathead Minnow (Pimephales promelas)1 , 2005, Biology of reproduction.

[21]  Theodoros N. Arvanitis,et al.  Dynamic range and mass accuracy of wide-scan direct infusion nanoelectrospray fourier transform ion cyclotron resonance mass spectrometry-based metabolomics increased by the spectral stitching method. , 2007, Analytical chemistry.

[22]  K. Kamaldeep,et al.  The effect of fenitrothion on reproduction of a teleost fish,Cyprinus carpio commuais linn: A biochemical study , 1978, Bulletin of environmental contamination and toxicology.

[23]  Mark R. Viant,et al.  Environmental metabolomics: a critical review and future perspectives , 2009, Metabolomics.

[24]  Mark R. Viant,et al.  Improved classification accuracy in 1- and 2-dimensional NMR metabolomics data using the variance stabilising generalised logarithm transformation , 2007, BMC Bioinformatics.

[25]  A. Seymour,et al.  High-throughput and multiplexed LC/MS/MRM method for targeted metabolomics. , 2010, Analytical chemistry.

[26]  R. P. Moody,et al.  The fate of fenitrothion in an aquatic ecosystem , 1978, Bulletin of environmental contamination and toxicology.

[27]  K. Kawata,et al.  Daily Variation of Pesticides in Surface Water of a Small River Flowing Through Paddy Field Area , 2009, Bulletin of environmental contamination and toxicology.

[28]  M. D. Ferrando,et al.  Liver energy metabolism of Anguilla anguilla after exposure to fenitrothion. , 1998, Ecotoxicology and environmental safety.

[29]  J. Caixach,et al.  Analysis of pesticides and metabolites in Spanish surface waters by isotope dilution gas chromatography/mass spectrometry with previous automated solid-phase extraction Estimation of the uncertainty of the analytical results. , 2006, Journal of chromatography. A.

[30]  M. Viant.  Improved methods for the acquisition and interpretation of NMR metabolomic data. , 2003, Biochemical and biophysical research communications.

[31]  Y. Benjamini,et al.  Controlling the false discovery rate: a practical and powerful approach to multiple testing , 1995 .

[32]  P. Kille,et al.  Metabolic profile biomarkers of metal contamination in a sentinel terrestrial species are applicable across multiple sites. , 2007, Environmental science & technology.

[33]  D. Hodgson,et al.  The organophosphorous pesticide, fenitrothion, acts as an anti-androgen and alters reproductive behavior of the male three-spined stickleback, Gasterosteus aculeatus , 2009, Ecotoxicology.

[34]  G. Ankley,et al.  Investigating compensation and recovery of fathead minnow (Pimephales promelas) exposed to 17alpha-ethynylestradiol with metabolite profiling. , 2008, Environmental science & technology.

[35]  P. Martin,et al.  Effects of fenitrothion on survival, behavior, and brain cholinesterase activity of white‐throated sparrows (Zonotrichia albicollis) , 1993 .

[36]  I. Katsiadaki,et al.  Non-invasive measurement of 11-ketotestosterone, cortisol and androstenedione in male three-spined stickleback (Gasterosteus aculeatus). , 2007, General and comparative endocrinology.

[37]  C. Tyler,et al.  Functional associations between two estrogen receptors, environmental estrogens, and sexual disruption in the roach (Rutilus rutilus). , 2007, Environmental science & technology.

[38]  L. Gray,et al.  Androgen receptor antagonism by the organophosphate insecticide fenitrothion. , 2001, Toxicological sciences : an official journal of the Society of Toxicology.

[39]  R. Harrison,et al.  Chemical pollution of the aquatic environment by priority pollutants and its control , 2001 .

[40]  H. Senn,et al.  Probabilistic quotient normalization as robust method to account for dilution of complex biological mixtures. Application in 1H NMR metabonomics. , 2006, Analytical chemistry.

[41]  M. Trevisan,et al.  Reduction of point contamination sources of pesticide from a vineyard farm. , 2007, Environmental science & technology.

[42]  K. Abu-Salah,et al.  Metabolism at a glance , 2001 .

[43]  Theodoros N. Arvanitis,et al.  A signal filtering method for improved quantification and noise discrimination in fourier transform ion cyclotron resonance mass spectrometry-based metabolomics data , 2009, Journal of the American Society for Mass Spectrometry.