(P187) Differential gene expression of Caenorhabditis elegans when exposed to 2,4,6-trinitrotoluene (TNT) .
Davis, Emily*,1, Perkins, Edward2, 1 Analytical Services, Inc. (ASI), Vicksburg, MS, USA2 U.S. Army Engineer Research & Development Center, Vicksburg, MS, USA
ABSTRACT- Soil contaminated with 2,4,6-trinitrotoluene (TNT) and other explosive compounds has become a concern on many military sites, including firing ranges and ammunition plants. As it degrades in the environment, TNT and its breakdown products bind to soil organic matter. Currently, there is some controversy as to the bioavailability of TNT and its breakdown products to soil organisms. Using the soil nematode Caenorhabditis elegans, we are investigating how gene transcription changes in response to TNT exposure in liquid culture. Differential gene expression assessed at the transcriptional level in response to TNT exposure can be taken as an indication of bioavailability. Because the genome of C. elegans is completely sequenced, with most genes having known functions, this organism is an excellent indicator for gene-based assays of bioavailability. Our first step was to optimize conditions for whole organism bioavailability assays in liquid cultures. Synchronized liquid cultures were started using dauer stage larva. Once the organisms reached the larval 4 stage some were used as negative controls while others were exposed to concentrations of TNT between 5mg/L and 10 mg/L. Messenger RNA (mRNA) was extracted from controls and treatment populations after 4-hour exposures. Anchored differential display polymerase chain reaction assays were then used to identify genes affected by TNT exposure. Forty-five bands were isolated as differentially expressed gene candidates, ten of which have been sequenced for determination of gene identity. To date, we have designed primers to amplify five of the ten genes and are confirming their differential gene expression using real time polymerase chain reaction. Subsequent work will continue to focus on gene expression profiles. The ultimate goal of the work is to be able to utilize gene profiles from C. elegans to determine what explosive compounds are present in a soil sample and the approximate concentrations of those contaminates.
Key words: Bioavailability, Caenorhabditis elegans, 2,4, 6- Trinitrotoluene, Toxicogenomics