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Applying Metabolomics to differentiate amphibian responses to multiple stressors
Citation:
Snyder, M., Tom Purucker, D. Glinski, AND Matt Henderson. Applying Metabolomics to differentiate amphibian responses to multiple stressors. Ecological Society of America, Ft. Lauderdale, FL, August 07 - 12, 2016.
Impact/Purpose:
This research explored the capability of metabolomics to distinguish and predict adverse outcomes by performing an experiment with adult leopard frogs exposed to multiple stressors (i.e. pesticide exposure and predation). One of the biggest challenges in ecological risk assessment is determining the impact of multiple stressors on individual organisms and populations in real world scenarios. Frequently, data derived from laboratory studies are used to estimate risk parameters and do not adequately address scenarios where other stressors exist. Emerging omic technologies, notably, metabolomics, provides us an opportunity to address the uncertainties surrounding ecological risk assessment of multiple stressors. Metabolomic profiling provides a fingerprint of potentially 1000s of endogenous metabolites. We found that metabolomics linked pesticide and predator stressed changes in biochemical fluxes to the same mechanism of action, however the four treatments’ were able to be classified based on differences in their metabolomic profiles. By demonstrating that metabolomics can be used to better characterize the endogenous impacts of multiple stressors on amphibian biochemical pathways we have demonstrated a methodology that can be used to decrease uncertainty in multiple stressor risk assessment in non-targeted amphibians. Ultimately, this methodology could be used to translate short-term toxicity assay results to long-term population effects on non-target organisms. The project was funded under a PIP year 2015 phase 1 grant.
Description:
Introduction/Objectives/Methods One of the biggest challenges in ecological risk assessment is determining the impact of multiple stressors on individual organisms and populations in ‘real world’ scenarios. Emerging ‘omic technologies, notably, metabolomics, provides an opportunity to address the uncertainties surrounding ecological risk assessment of multiple stressors. The objective of this study was to use a metabolomics biomarker approach to investigate the effect of multiple stressors on amphibian metamorphs. To this end, metamorphs of Rana pipiens (northern leopard frogs) were exposed to the insecticide Carbaryl (0.32 μg/L), a conspecific predator alarm call (Lithobates catesbeianus), Carbaryl and the predator alarm call, and a control with no stressor. In addition to metabolomic fingerprinting, we measured corticosterone levels in each treatment to assess general stress response. We analyzed relative abundances of endogenous metabolites collected in liver tissue with gas chromatography coupled with mass spectrometry. Support vector machine (SVM) methods with recursive feature elimination (RFE) were applied to rank the metabolomic profiles produced. Results/Conclusions SVM-RFE of the acquired metabolomic spectra demonstrated 85-96% classification accuracy among control and all treatment groups when using the top 75 ranked retention time bins. Biochemical fluxes observed in the groups exposed to carbaryl, predation threat, and the combined treatment include amino acids, sugar derivatives, and purine nucleotides. Biochemical pathways impacted include aminoacyl tRNA biosynthesis, nitrogen metabolism, and cyanoamino acid synthesis. Individual metabolite fluxes demonstrated 43% similarity among treatments. The extent of the overlap in endogenous metabolites impacted and pathway analysis suggests that pesticide and predation stressors act on similar modes of action.