Abstract:

In this experiment, fish (southern flounder) were exposed to oiled (Louisiana crude oil) sediments in a mesocosm experiment for 30 days. At the end of this period, the fish were examined for oxidative stress and DNA damage. The dataset contains measurements of glutathione (GSH), malondialdehye assay (MDA), AP (apurinic or apyrimidinic) sites, percent cell viability as well as basic details of fish such as gender, length and weight.

Suggested Citation:

Dana Wetzel, Tracy Sherwood, Christelle Miller and Rebecca Medvecky. 2019. Sub-adult Southern flounder exposure to crude oil-contaminated sediment: Oxidative Stress Assays & DNA Damage Assays. Distributed by: GRIIDC, Harte Research Institute, Texas A&M University–Corpus Christi. doi:10.7266/N7WH2NGJ

Publications:

Sherwood, T. A., Medvecky, R. L., Miller, C. A., Tarnecki, A. M., Schloesser, R. W., Main, K. L., … Wetzel, D. L. (2019). Nonlethal Biomarkers of Oxidative Stress in Oiled Sediment Exposed Southern Flounder (Paralichthys lethostigma): Utility for Field-Base Monitoring Exposure and Potential Recovery. Environmental Science & Technology, 53(24), 14734–14743. doi:10.1021/acs.est.9b05930

Purpose:

Controlled oil exposure study to evaluate the effects of south Louisiana crude oil, via contaminated sediment, on fish health.

Data Parameters and Units:

Worksheets "Test 211 Sample Key", "Oxidative Stress", "DNA Damage", and "Comet Cell Viability": gender, weight (g), length (cm) and treatment type, GSH (uM), GSSH (uM), trolox concentration (uM), MDA concentration (uM), AP Sites per 100,000bp, % comet cell viability. TAP = Oxford Biomedical Research Colorimetric Microplate Assay for Total Antioxidant Power Kit (product number TA02); MDA = malondialdehye assay; 8-OHdG= 8-hydroxydeoxyguanosine assay; and AP Sites = apurinic or apyrimidinic sites. Note: The dashes or empty cells signify that no data was generated for the sample ID listed.

Methods:

5 fish were put in each tank that contained between 246-250 g of oil, and 81.6-84 Kg of sediment. The amount of water in tanks were Test 211 - 552L tanks and Test 208 - 533L tanks. Negative controls had no oil. There were 3 replicates. Fish were fed twice a day. Temperature, pH, salinity, and dissolved oxygen were measured daily. At the end of the 30-day exposure to sediment, fish were anaesthetized with MS‌-222 prior to full euthanization, fish. Blood was drawn from each fish with a heparinized syringe and then divided into a sodium citrate vacutainer and an EDTA vacutainer. Samples of whole blood were collected from the EDTA vacutainer for comet assay and oxidative stress assessments. Remaining whole blood was centrifuged, and plasma collected for further oxidative assessments and ELISA. Plasma samples were stored at -80°C until analysis. Whole blood was collected for GSH/GSSG analysis via Oxford Biomedical Research Microplate Assay for GSH/GSSG (product number GT40). For each fish, 50ul whole blood was frozen at -80°C until analysis and 100ul whole blood was added to 30 ul Scavenger and frozen at -80°C until analysis. Blood was analyzed according to manufacturer protocols. DNA damage was assessed with OxiSelect™ Oxidative DNA Damage Quantitation Kit (Cell Biolabs, STA‌-324) and Comet Assay: Mitchelmore, et al (1998). Comet cell viability was determined by staining cells with Trypan Blue solution - CAS No.: 72-57-1 and counting viable cells with an optical microscope.

Provenance and Historical References:

Mitchelmore, C. & Chipman, J. (1998). Detection of DNA strand breaks in brown trout (Salmo trutta) hepatocytes and blood cells using the single cell gel electrophoresis (comet) assay. Aquatic Toxicology, 41(1-2), 161–182. doi:10.1016/s0166-445x(97)00064-7

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