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The aims of the present study were to compare the toxic effects of benzo[ a]pyrene (B aP) and to screen for rapid and sensitive biomarkers that can be used to assess the environmental risks of B aP in earthworms in different natural soil types. The authors exposed Eisenia fetida to 2 types of soil (red soil and fluvo-aquic soil) spiked with different concentrations (0 mg kg−1, 1 mg kg−1, 10 mg kg−1, 100 mg kg−1, and 500 mg kg−1) of B aP for 7 d or 14 d. Benzo[ a]pyrene-induced weight variation altered the activities of antioxidant enzymes (superoxide dismutase [SOD]; catalase [CAT]; and guaiacol peroxidase [POD]) and changed the content of malondialdehyde (MDA). In addition, using the comet assay, the authors determined the DNA damage in earthworms. The results revealed that the comet assay was suitable for evaluating the genotoxicity of B aP in the soil, even at the lowest examined concentration. The MDA content was the least sensitive indicator of B aP toxicity. A 3-way analysis of variance (ANOVA) was used to determine whether the soil type, exposure concentration, and duration affected the B aP toxicity. The antioxidant enzyme activities and the MDA content were shown to be significantly correlated with the exposure concentration. The percentage of weight variation ( p < 0.001), CAT activity ( p < 0.05), and SOD activity ( p < 0.01) were significantly affected by the soil type, and the POD activity ( p < 0.01), CAT activity ( p < 0.001), and SOD activity ( p < 0.001) were significantly affected by the exposure duration. Therefore, measuring DNA damage in earthworms is a simple and efficient means of assessing B aP genotoxicity in a terrestrial environment, and the effects of the soil type and exposure time on the other parameters that were investigated in E. fetida, which were used as responsive biomarkers, should be considered. Environ Toxicol Chem 2014;9999:1-8. © 2014 SETAC [ABSTRACT FROM AUTHOR]