Genomic investigation of year-long and multigenerational exposures of fathead minnow to the munitions compound RDX
ABSTRACT We assessed the impacts of exposure to an environmentally representative concentration (0.83 mg/L) of the explosive cyclotrimethylenetrinitramine (RDX) on fathead minnows (Pimephales promelas) in one-year and multigenerational bioassays. In the one-year bioassay, impacts were assessed by statistical comparisons of females from breeding groups reared in control or RDX-exposure conditions. The RDX had no significant effect on gonadosomatic index or condition factor assayed at 1 d and at one, three, six, nine, and 12 months. The liver-somatic index was significantly increased versus controls only at the 12-month timepoint. RDX had no significant effect on live-prey capture rates, egg production, or fertilization. RDX caused minimal differential-transcript expression with no consistent discernable effect on gene-functional categories for either brain or liver tissues in the one-year exposure. In the multigenerational assay, the effects of acute (96 h) exposure to RDX were compared in fish reared to the F(2) generation in either control or RDX-exposure conditions. Enrichment of gene functions including neuroexcitatory glutamate metabolism, sensory signaling, and neurological development were observed comparing control-reared and RDX-reared fish. Our results indicated that exposure to RDX at a concentration representing the highest levels observed in the environment (0.83 mg/L) had limited impacts on genomic, individual, and population-level endpoints in fathead minnows in a one-year exposure. However, multigenerational exposures altered transcript expression related to neural development and function. Environ.
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ABSTRACT: The earthworm Eisenia fetida is one of the most used species in standardized soil ecotoxicity tests. End points such as survival, growth, and reproduction are eco-toxicologically relevant but provide little mechanistic insight into toxicity pathways, especially at the molecular level. Here we apply a toxicogenomic approach to investigate the mode of action underlying the reversible neurotoxicity of hexanitrohexaazaisowurtzitane (CL-20), a cyclic nitroamine explosives compound. We developed an E. fetida-specific shotgun microarray targeting 15119 unique E. fetida transcripts. Using this array we profiled gene expression in E. fetida in response to exposure to CL-20. Eighteen earthworms were exposed for 6 days to 0.2 μg/cm(2) of CL-20 on filter paper, half of which were allowed to recover in a clean environment for 7 days. Nine vehicle control earthworms were sacrificed at days 6 and 13, separately. Electrophysiological measurements indicated that the conduction velocity of earthworm medial giant nerve fiber decreased significantly after 6-day exposure to CL-20, but was restored after 7 days of recovery. Total RNA was isolated from the four treatment groups including 6-day control, 6-day exposed, 13-day control, and 13-day exposed (i.e., 6-day exposure followed by 7-day recovery), and was hybridized to the 15K shotgun oligo array. Statistical and bioinformatic analyses suggest that CL-20 initiated neurotoxicity by noncompetitively blocking the ligand-gated GABA(A) receptor ion channel, leading to altered expression of genes involved in GABAergic, cholinergic, and Agrin-MuSK pathways. In the recovery phase, expression of affected genes returned to normality, possibly as a result of autophagy and CL-20 dissociation/metabolism. This study provides significant insights into potential mechanisms of CL-20-induced neurotoxicity and the recovery of earthworms from transient neurotoxicity stress.Environmental Science & Technology 02/2012; 46(2):1223-32. DOI:10.1021/es203642e
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ABSTRACT: Genomics-enabled technologies have transformed ecological, evolutionary and environmental sciences. The nature of this transformation became obvious at the 2003 Gordon Research Conference on EEFG: Evolutionary and Ecological Functional Genomics, after which high quality papers have influenced the environmental science literature. More recently next-generation DNA sequencing has brought about a new revolution, as the non-model genomes of environmental science have become as tractable as genomes of classical genetic models. In this paper we aim to critically review the possible contributions of environmental genomics to risk assessment of chemicals. Environmental (ecological) genomics can be defined as: "A scientific discipline that studies the structure and functioning of a genome with the aim of understanding the relationship between the organism and its biotic and abiotic environment". It can thus be viewed as a merger of genomics and ecology. The typical approach of genomics is to look upon the organism as a unitary whole, that is, to try and analyse as many genes as possible, their transcripts, the way they are regulated and interact, the proteins they encode, etc. Several activities together characterize the diverse field of environmental genomics: (1) Metagenomics, also called environmental community genomics, (2) Taxonomic and functional community genomics, (3) Genome scans and population genomics, (4) Gene expression profiling. The list does not exhaust the diversity of genomics approaches to environmental analysis, but we consider these four activities the most important. For environmental risk analysis of chemicals, the last mentioned activity, gene expression profiling, is most relevant. We discuss the limitations and possibilities of gene expression profiling with respect to four questions: (1) Biological limitations of transcription profiling. Is it possible at all to assess environmental quality based on transcriptomes (mRNA abundance)? (2) Classification of exposure or effect? Because of the genome-wide approach of gene expression profiling, the action spectrum of chemicals can be characterized on the basis of an extremely large number of endpoints. The question remains, what is measured by gene expression: exposure or effects? (3) Fishing in the quagmire of mixtures. The study of mixture toxicity is as old as toxicology itself, however, significant advances have been made recently regarding mathematical and statistical analysis of mixture toxicity data. The question is, how do these approaches perform on the level of gene expression? (4) Defining a range of fitness-neutral gene expression. Following from the classical paradigm of homeostasis, every animal will constantly adjust its gene expression as part of its "normal" operations. The collective set of fold-regulations associated with such normal functioning, which are neutral to fitness, may be called the "normal operating range". In conclusion, we identify a number of formidable problems for the use of transcription profiling in risk assessment. What is particularly needed is the generation of background data under normal, clean conditions. In addition, a close connection between traditional ecotoxicology and genomics seems to be particularly fruitful. Gene expression profiles must be generated for the same species and under the same conditions as are being deployed in internationally accepted ecotoxicity tests. Only in this way can transcriptome data be connected to endpoints accepted in risk assessment schemes.Environmental Science & Technology 11/2011; 46(1):3-9. DOI:10.1021/es2034153
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ABSTRACT: Interspecies uncertainty factors in ecological risk assessment provide conservative estimates of risk where limited or no toxicity data is available. We quantitatively examined the validity of interspecies uncertainty factors by comparing the responses of zebrafish (Danio rerio) and fathead minnow (Pimephales promelas) to the energetic compound 1,3,5-trinitroperhydro-1,3,5-triazine (RDX), a known neurotoxicant. Relative toxicity was measured through transcriptional, morphological, and behavioral end points in zebrafish and fathead minnow fry exposed for 96 h to RDX concentrations ranging from 0.9 to 27.7 mg/L. Spinal deformities and lethality occurred at 1.8 and 3.5 mg/L RDX respectively for fathead minnow and at 13.8 and 27.7 mg/L for zebrafish, indicating that zebrafish have an 8-fold greater tolerance for RDX than fathead minnow fry. The number and magnitude of differentially expressed transcripts increased with increasing RDX concentration for both species. Differentially expressed genes were enriched in functions related to neurological disease, oxidative-stress, acute-phase response, vitamin/mineral metabolism and skeletal/muscular disorders. Decreased expression of collagen-coding transcripts were associated with spinal deformity and likely involved in sensitivity to RDX. Our work provides a mechanistic explanation for species-specific sensitivity to RDX where zebrafish responded at lower concentrations with greater numbers of functions related to RDX tolerance than fathead minnow. While the 10-fold interspecies uncertainty factor does provide a reasonable cross-species estimate of toxicity in the present study, the observation that the responses between ZF and FHM are markedly different does initiate a call for concern regarding establishment of broad ecotoxicological conclusions based on model species such as zebrafish.Environmental Science & Technology 06/2012; 46(14):7790-8. DOI:10.1021/es300495c