Toxicogenomic analysis provides new insights into molecular mechanisms of the sublethal toxicity of 2,4,6-trinitrotoluene in Eisenia fetida

Xenobiotics such as explosives and pesticides released into the environment can have lethal and sublethal impacts on soil organisms such as earthworms with potential subsequent impacts athighertrophic levels. To better understand the molecular toxicological mechanisms of 2,4,6-trinitrotoluene (TNT), a commonly used explosive, in Eisenia fetida, earthworms were exposed to a gradient of TNT-spiked soils for 28 days and impacts on gene expression were examined using a 4032 cDNA microarray. Reproduction was increased at low doses of TNT, whereas high doses of TNT reduced juvenile production. On the basis of reproduction responses to TNT, four treatments, that is, control, 2,10.6, and 38.7 mg/kg, were selected for gene expression studies in a balanced interwoven loop design microarray experiment in which the expression of 311 transcripts was significantly affected. Reverse-transcription quantitative polymerase chain reaction (RT-QPCR) data on 68 selected differentially and nondifferentially expressed transcripts showed a significant correlation with microarray results. The expression of genes involved in multiple biological processes was altered, including muscle contraction, neuronal signaling and growth, ubiquitinylation, fibrinolysis and coagulation, iron and calcium homeostasis, oxygen transport, and immunity. Chitinase activity assays confirmed down-regulation of chitinase genes as indicated by array and RT-QPCR data. An acute toxicity test provided evidence that dermal contact with TNT can cause bleeding, inflammation, and constriction, which may be explained by gene expression results. Sublethal doses of TNT affected the nervous system, caused blood disorders similar to methemoglobinemia, and weakened immunity in E. fetida.