Comparative Study of the Comet Assay and the Micronucleus Test in Amphibian Larvae (Xenopus laevis) Using Benzo(a)pyrene, Ethyl Methanesulfonate, and Methyl Methanesulfonate: Establishment of a Positive Control in the Amphibian Comet Assay

Abstract

The present investigation explored the potential use of the comet assay (CA) as a genotoxicity test in the amphibian Xenopus laevis and compared it with the French standard micronucleus test (MNT). Benzo[a]pyrene (B[a]P), methyl methanesulfonate (MMS), and ethyl methanesulfonate (EMS) were used as model compounds for assessing DNA damage. Damage levels were measured as DNA strand breaks after alkaline electrophoresis of nuclei isolated from larval amphibian erythrocytes using the CA in order to establish a positive control for further ecotoxicological investigations. The results led to the selection of MMS as a positive control on the basis of the higher sensitivity of Xenopus laevis to this compound. The CA and MNT were compared for their ability to detect DNA damage with the doses of chemical agents and exposure times applied. EMS and MMS were shown to increase micronucleus and DNA strand break formation in larval erythrocytes concurrently. However, B[a]P increased micronucleus formation but not that of DNA strand breaks. Time–dose experiments over 12 days of exposure suggest that the CA provides an earlier significant response to genotoxicants than does the MNT. In Xenopus the CA appears to be a sensitive and suitable method for detecting genotoxicity like that caused by EMS and MMS. It can be considered a genotoxicityscreening tool. The results for B[a]P show that both tests should be used in a complementary manner on Xenopus.