Linking mechanistic and behavioral responses to sublethal esfenvalerate exposure in the endangered delta smelt; Hypomesus transpacificus (Fam. Osmeridae)

BackgroundThe delta smelt (Hypomesus transpacificus) is a pelagic fish species listed as endangered under both the USA Federal and Californian State Endangered Species Acts and considered an indicator of ecosystem health in its habitat range, which is limited to the Sacramento-San Joaquin estuary in California, USA. Anthropogenic contaminants are one of multiple stressors affecting this system, and among them, current-use insecticides are of major concern. Interrogative tools are required to successfully monitor effects of contaminants on the delta smelt, and to research potential causes of population decline in this species. We have created a microarray to investigate genome-wide effects of potentially causative stressors, and applied this tool to assess effects of the pyrethroid insecticide esfenvalerate on larval delta smelt. Selected genes were further investigated as molecular biomarkers using quantitative PCR analyses.ResultsExposure to esfenvalerate affected swimming behavior of larval delta smelt at concentrations as low as 0.0625 μg.L-1, and significant differences in expression were measured in genes involved in neuromuscular activity. Alterations in the expression of genes associated with immune responses, along with apoptosis, redox, osmotic stress, detoxification, and growth and development appear to have been invoked by esfenvalerate exposure. Swimming impairment correlated significantly with expression of aspartoacylase (ASPA), an enzyme involved in brain cell function and associated with numerous human diseases. Selected genes were investigated for their use as molecular biomarkers, and strong links were determined between measured downregulation in ASPA and observed behavioral responses in fish exposed to environmentally relevant pyrethroid concentrations.ConclusionsThe results of this study show that microarray technology is a useful approach in screening for, and generation of molecular biomarkers in endangered, non-model organisms, identifying specific genes that can be directly linked with sublethal toxicological endpoints; such as changes in expression levels of neuromuscular genes resulting in measurable swimming impairments. The developed microarrays were successfully applied on larval fish exposed to esfenvalerate, a known contaminant of the Sacramento-San Joaquin estuary, and has permitted the identification of specific biomarkers which could provide insight into the factors contributing to delta smelt population decline.

[1]  Inge Werner,et al.  Acute, sublethal exposure to a pyrethroid insecticide alters behavior, growth, and predation risk in larvae of the fathead minnow (Pimephales promelas) , 2008, Environmental toxicology and chemistry.

[2]  William H. Benson,et al.  Genomic approaches for cross-species extrapolation in toxicology : proceedings from the Workshop on Emerging Molecular and Computational Approaches for Cross-Species Extrapolations, 18-22 July 2004, Portland, Oregon USA , 2006 .

[3]  M. Ro̸rth,et al.  Beta-2-microglobulin excretion: An indicator of long term nephrotoxicity during cis-platinum treatment? , 2004, Cancer Chemotherapy and Pharmacology.

[4]  H. Masuyama,et al.  Down-Regulation of Pregnane X Receptor Contributes to Cell Growth Inhibition and Apoptosis by Anticancer Agents in Endometrial Cancer Cells , 2007, Molecular Pharmacology.

[5]  George N. Phillips,et al.  Structure of aspartoacylase, the brain enzyme impaired in Canavan disease , 2007, Proceedings of the National Academy of Sciences.

[6]  K. Michals‐Matalon,et al.  Biochemistry and Molecular Biology of Canavan Disease , 1999, Neurochemical Research.

[7]  I. Fujita,et al.  The olfactory system, not the terminal nerve, functions as the primary chemosensory pathway mediating responses to sex pheromones in male goldfish. , 1991, Brain, behavior and evolution.

[8]  Elizabeth C. Theil,et al.  Daphnia magna ecotoxicogenomics provides mechanistic insights into metal toxicity. , 2007, Environmental science & technology.

[9]  M. Baslow,et al.  Functions of N‐Acetyl‐l‐Aspartate and N‐Acetyl‐l‐Aspartylglutamate in the Vertebrate Brain , 2000, Journal of neurochemistry.

[10]  S. Stürzenbaum,et al.  Genes and environment - striking the fine balance between sophisticated biomonitoring and true functional environmental genomics. , 2008, The Science of the total environment.

[11]  S. Köprücü,et al.  Acute Toxicity of Synthetic Pyrethroid Cypermethrin on the Common Carp (Cyprinus carpio L.) Embryos and Larvae , 2005, Aquaculture International.

[12]  Gary M. Rand,et al.  Fundamentals of Aquatic Toxicology , 1985 .

[13]  N. Scholz,et al.  Comparative thresholds for acetylcholinesterase inhibition and behavioral impairment in coho salmon exposed to chlorpyrifos , 2005, Environmental toxicology and chemistry.

[14]  D. Wenger,et al.  Psychosine-induced apoptosis in a mouse oligodendrocyte progenitor cell line is mediated by caspase activation , 2004, Neuroscience Letters.

[15]  Elaine Holmes,et al.  Metabolic Profiling of CSF: Evidence That Early Intervention May Impact on Disease Progression and Outcome in Schizophrenia , 2006, PLoS medicine.

[16]  Prof. Dr. Peter J. Bentley.  Endocrines and Osmoregulation , 2002, Zoophysiology.

[17]  E. Taylor,et al.  Molecular resolution of the systematics of a problematic group of fishes (Teleostei: Osmeridae) and evidence for morphological homoplasy. , 2009, Molecular phylogenetics and evolution.

[18]  Timothy M Willson,et al.  The nuclear pregnane X receptor: a key regulator of xenobiotic metabolism. , 2002, Endocrine reviews.

[19]  D. Bhatnagar,et al.  Lipid peroxidative damage on pyrethroid exposure and alterations in antioxidant status in rat erythrocytes: a possible involvement of reactive oxygen species. , 1999, Toxicology letters.

[20]  A. Jarry,et al.  Interleukin 1 and interleukin 1β converting enzyme (caspase 1) expression in the human colonic epithelial barrier. Caspase 1 downregulation in colon cancer , 1999, Gut.

[21]  T. Sommer,et al.  Multidecadal trends for three declining fish species: habitat patterns and mechanisms in the San Francisco Estuary, California, USA , 2007 .

[22]  J. Connor,et al.  Iron acquisition and expression of iron regulatory proteins in the developing brain: manipulation by ethanol exposure, iron deprivation and cellular dysfunction. , 1994, Developmental neuroscience.

[23]  S. Madsen,et al.  Rapid modulation of Na+/K+-ATPase activity in osmoregulatory tissues of a salmonid fish. , 2001, The Journal of experimental biology.

[24]  R. Kamen,et al.  Caspase-1 processes IFN-γ-inducing factor and regulates LPS-induced IFN- γ production , 1997, Nature.

[25]  H. Köhler,et al.  Expression of immune-regulatory genes in juvenile Chinook salmon following exposure to pesticides and infectious hematopoietic necrosis virus (IHNV). , 2008, Fish & shellfish immunology.

[26]  P. Ferrara,et al.  Hemopexin is synthesized in peripheral nerves but not in central nervous system and accumulates after axotomy. , 1992, The Journal of biological chemistry.

[27]  K. Okumura,et al.  Beta2-microglobulin required for cell surface expression of blastocyst MHC. , 2005, Biochemical and biophysical research communications.

[28]  L. L. Porter,et al.  Effects of dopamine and estrogen upon cortical neurons that express parvalbumin in vitro. , 2002, Brain research. Developmental brain research.

[29]  H. Weiner,et al.  T-cell recognition of an immuno-dominant myelin basic protein epitope in multiple sclerosis , 1990, Nature.

[30]  R. Hunt,et al.  Hemopexin joins transferrin as representative members of a distinct class of receptor-mediated endocytic transport systems. , 1990, European journal of cell biology.

[31]  J. Gan,et al.  Synthetic Pyrethroids: Occurrence and Behavior in Aquatic Environments , 2008 .

[32]  L. Brown,et al.  Managing Water to Protect Fish: A Review of California’s Environmental Water Account, 2001–2005 , 2009, Environmental management.

[33]  B. Barlogie,et al.  Beta 2-microglobulin as a negative regulator of the immune system: high concentrations of the protein inhibit in vitro generation of functional dendritic cells. , 2003, Blood.

[34]  I. Ferrer,et al.  Parvalbumin immunocytochemistry and calcium deposition in muscle fiber necrosis and subsequent regeneration following intramuscular injection of metoclopramide , 1994, Muscle & nerve.

[35]  Nathan K. Truelove,et al.  Diazinon disrupts antipredator and homing behaviors in chinook salmon (Oncorhynchus tshawytscha) , 2000 .

[36]  N. Garcia-Reyero,et al.  Fish 'n' chips: the use of microarrays for aquatic toxicology. , 2007, Molecular bioSystems.

[37]  R. Welsh,et al.  NK cell response to viral infections in beta 2-microglobulin-deficient mice. , 1995, Journal of immunology.

[38]  J. Simpkins,et al.  Role of parvalbumin in estrogen protection from ethanol withdrawal syndrome. , 2005, Alcoholism, clinical and experimental research.

[39]  Juergen Geist,et al.  Effects of dietary exposure to the pyrethroid pesticide esfenvalerate on medaka (Oryzias latipes). , 2002, Marine environmental research.

[40]  H. Toyohara,et al.  A hyperosmotic stress-induced mRNA of carp cell encodes Na(+)- and Cl(-)-dependent high affinity taurine transporter. , 2000, Biochimica et biophysica acta.

[41]  A. Moore,et al.  The effects of a synthetic pyrethroid pesticide on some aspects of reproduction in Atlantic salmon (Salmo salar L.). , 2001, Aquatic toxicology.

[42]  Christopher R. Singh,et al.  Relating olfactory neurotoxicity to altered olfactory-mediated behaviors in rainbow trout exposed to three currently-used pesticides. , 2007, Aquatic toxicology.

[43]  J. Sumpter,et al.  Metabolism and environmental degradation of pyrethroid insecticides produce compounds with endocrine activities , 2000 .

[44]  Karl H. Clodfelter,et al.  Sexual dimorphism of rat liver gene expression: regulatory role of growth hormone revealed by deoxyribonucleic Acid microarray analysis. , 2004, Molecular endocrinology.

[45]  D. Dix,et al.  Selection of Surrogate Animal Species for Comparative Toxicogenomics , 2006 .

[46]  F. Altruda,et al.  Hemopexin: structure, function, and regulation. , 2002, DNA and cell biology.

[47]  G. C. Ferreira,et al.  Iron metabolism : inorganic biochemistry and regulatory mechanisms , 1999 .

[48]  F. J. Dwyer,et al.  Survival of bluegill and their behavioral responses during continuous and pulsed exposures to esfenvalerate, a pyrethroid insecticide , 1993 .

[49]  Simon C Watkins,et al.  Myozenin: an alpha-actinin- and gamma-filamin-binding protein of skeletal muscle Z lines. , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[50]  S. Hayashi,et al.  Identification of choriogenin cis-regulatory elements and production of estrogen-inducible, liver-specific transgenic Medaka , 2004, Mechanisms of Development.

[51]  N. Bertrand,et al.  Changes in expression and localization of hemopexin and its transcripts in injured nervous system: a comparison of central and peripheral tissues , 1997, Neuroscience.

[52]  K. Wallace,et al.  Mechanisms of pyrethroid neurotoxicity: implications for cumulative risk assessment. , 2002, Toxicology.

[53]  Xueping Chen,et al.  Choriogenin mRNA as a sensitive molecular biomarker for estrogenic chemicals in developing brackish medaka (Oryzias melastigma). , 2008, Ecotoxicology and environmental safety.

[54]  P. Bentley.  Endocrines and Osmoregulation: A Comparative Account in Vertebrates , 2002 .

[55]  R. Blust,et al.  Molecular responses during cadmium-induced stress in Daphnia magna: integration of differential gene expression with higher-level effects. , 2007, Aquatic toxicology.

[56]  H. Hartung,et al.  Apoptosis of myelin-reactive T cells induced by reactive oxygen and nitrogen intermediates in vitro. , 1997, Cellular immunology.

[57]  M. Hayden,et al.  Caspases and neurodegeneration: on the cutting edge of new therapeutic approaches , 2000, Clinical genetics.

[58]  Thomas H Hutchinson,et al.  Systems biology meets stress ecology: linking molecular and organismal stress responses in Daphnia magna , 2008, Genome Biology.

[59]  L. Brown,et al.  Pelagic Organism Decline Progress Report: 2007 Synthesis of Results , 2008 .

[60]  F. Spurlock,et al.  Residues and Toxicity of Esfenvalerate and Permethrin in Water and Sediment, in Tributaries of the Sacramento and San Joaquin Rivers, California, USA , 2005, Bulletin of environmental contamination and toxicology.

[61]  A. Porta,et al.  Expressed sequence tags (EST) identify genes preferentially expressed in catfish chemosensory tissues. , 1994, Brain research. Molecular brain research.

[62]  B. Pipy,et al.  Respective roles of inflammation and axonal breakdown in the regulation of peripheral nerve hemopexin: an analysis in rats and in C57BL/Wlds mice , 2000, Journal of Neuroimmunology.

[63]  Jullie W Pan,et al.  Interdependence of N‐acetyl aspartate and high‐energy phosphates in healthy human brain , 2005, Annals of neurology.

[64]  W. Horning,et al.  Short-term methods for estimating the chronic toxicity of effluents and receiving waters freshwater organisms , 1985 .

[65]  Thomas D. Schmittgen,et al.  Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. , 2001, Methods.

[66]  M. Tuszynski,et al.  Estrogen receptor‐beta colocalizes extensively with parvalbumin‐labeled inhibitory neurons in the cortex, amygdala, basal forebrain, and hippocampal formation of intact and ovariectomized adult rats , 2002, The Journal of comparative neurology.

[67]  Taisen Iguchi,et al.  Linking molecular and population stress responses in Daphnia magna exposed to cadmium. , 2008, Environmental science & technology.

[68]  R. Kong,et al.  Induction of hepatic choriogenin mRNA expression in male marine medaka: a highly sensitive biomarker for environmental estrogens. , 2006, Aquatic toxicology.

[69]  G. Evan,et al.  Caspase-mediated Cleavage of the Ca2+/Calmodulin-dependent Protein Kinase-like Kinase Facilitates Neuronal Apoptosis* , 2001, The Journal of Biological Chemistry.

[70]  M. Bernardi,et al.  Behavioral and endocrine changes induced by perinatal fenvalerate exposure in female rats. , 2005, Neurotoxicology and teratology.

[71]  Juergen Geist,et al.  Comparisons of tissue-specific transcription of stress response genes with whole animal endpoints of adverse effect in striped bass (Morone saxatilis) following treatment with copper and esfenvalerate. , 2007, Aquatic toxicology.

[72]  Tamás F. Freund,et al.  GABAergic septohippocampal neurons contain parvalbumin , 1989, Brain Research.

[73]  I. Mian,et al.  Exploratory differential gene expression analysis in microarray experiments with no or limited replication , 2004, Genome Biology.

[74]  Rafael A. Irizarry,et al.  Bioinformatics and Computational Biology Solutions using R and Bioconductor , 2005 .

[75]  Gordon K. Smyth,et al.  limma: Linear Models for Microarray Data , 2005 .

[76]  I. Werner,et al.  Synergistic effects of esfenvalerate and infectious hematopoietic necrosis virus on juvenile chinook salmon mortality , 2005, Environmental toxicology and chemistry.

[77]  G. Lees.  Effects of pyrethroid molecules on rat nerves in vitro: potential to reverse temperature‐sensitive conduction block of demyelinated peripheral axons , 1998, British journal of pharmacology.

[78]  Chulwoo Lee,et al.  Choriogenin mRNA induction in male medaka, Oryzias latipes as a biomarker of endocrine disruption. , 2002, Aquatic toxicology.

[79]  R. Kamen,et al.  Caspase-1 processes IFN-gamma-inducing factor and regulates LPS-induced IFN-gamma production. , 1997, Nature.

[80]  Å. Krogdahl,et al.  Carbohydrates in fish nutrition: digestion and absorption in postlarval stages , 2005 .

[81]  V. Shashoua,et al.  A peptide fragment of ependymin neurotrophic factor uses protein kinase C and the mitogen‐activated protein kinase pathway to activate c‐Jun N‐terminal kinase and a functional AP‐1 containing c‐Jun and c‐Fos proteins in mouse NB2a cells , 2003, Journal of neuroscience research.

[82]  R. M. Simmons,et al.  Elasticity and unfolding of single molecules of the giant muscle protein titin , 1997, Nature.

[83]  V. Dixit,et al.  Differential activation of the inflammasome by caspase-1 adaptors ASC and Ipaf , 2004, Nature.

[84]  J. Cech,et al.  Swimming performance of delta smelt: maximum performance, and behavioral and kinematic limitations on swimming at submaximal velocities. , 1998, The Journal of experimental biology.

[85]  D. Hinton,et al.  Application of whole effluent toxicity test procedures to ambient water quality assessment , 2000 .

[86]  H. Eppenberger,et al.  Muscle-type MM creatine kinase is specifically bound to sarcoplasmic reticulum and can support Ca2+ uptake and regulate local ATP/ADP ratios. , 1990, The Journal of biological chemistry.

[87]  I. Werner,et al.  Effects of Pyrethroid Insecticides on Aquatic Organisms , 2008 .

[88]  R. Rudick,et al.  Axonal transection in the lesions of multiple sclerosis. , 1998, The New England journal of medicine.

[89]  Bruce Herbold,et al.  The Collapse of Pelagic Fishes in the Upper San Francisco Estuary: El Colapso de los Peces Pelagicos en La Cabecera Del Estuario San Francisco , 2007 .

[90]  H. Jockusch,et al.  Serum parvalbumin, an indicator of muscle disease in murine dystrophy and myotonia , 1990, Muscle & nerve.

[91]  L. Mills,et al.  Individual effects of estrogens on a marine fish, Cunner (Tautogolabrus adspersus), extrapolated to the population level. , 2006, Ecotoxicology and environmental safety.