Detection of 8-oxodG in Dreissena polymorpha gill cells exposed to model contaminants

Genotoxic end-points are routinely measured in various sentinel organisms in aquatic environments in order to monitor the impact of water pollution on organisms. As a first step towards the evaluation of oxidative DNA damage (8-oxodG) in organisms exposed to chemical water pollution, we have optimized the association between the comet assay and the hOGG1 enzyme for use on zebra mussel (Dreissena polymorpha) gill cells by in vitro exposure to H₂O₂. Firstly, we observed that in vitro exposure of D. polymorpha gill cells to benzo[a]pyrene (B[a]P, 98.4nM) induced an increase of the Olive Tail Moment (OTM) in both the comet-hOGG1 and comet-Fpg assays, indicating that B[a]P causes oxidative DNA damage. By contrast, methylmethane sulfonate (MMS, 33μM) only induced an increase of the Fpg-sensitive sites, indicating that MMS caused alkylating DNA damage and confirming that hOGG1 does not detect alkylating damage. Thus, the hOGG1 enzyme seems to be more specific towards oxidative DNA damage, such as 8-oxodG than Fpg. Secondly, as was observed in vitro, the in vivo exposure of D. polymorpha to B[a]P (24.6 and 98.4nM) increased oxidative DNA damage in gill cells, whereas only Fpg-sensitive sites were detected in mussels exposed to MMS (240μM). These results show that the comet-hOGG1 assay detects oxidative DNA lesions induced in vitro by H₂O₂ and in vivo with BaP. The comet-hOGG1 assay will be used to detect oxidative DNA lesions (8-oxodG) in mussels exposed in situ.