Merging nano-genotoxicology with eco-genotoxicology: an integrated approach to determine interactive genotoxic and sub-lethal toxic effects of C(60) fullerenes and fluoranthene in marine mussels, Mytilus sp.

Whilst there is growing concern over the potential detrimental impact of engineered nanoparticles (ENPs) on the natural environment, little is known about their interactions with other contaminants. In the present study, marine mussels (Mytilus sp.) were exposed for 3 days to C(60) fullerenes (C(60); 0.10-1 mg l(-1)) and a model polycyclic aromatic hydrocarbon (PAH), fluoranthene (32-100 μg l(-1)), either alone or in combination. The first two experiments were conducted by exposing the organisms to different concentrations of C(60) and fluoranthene alone, in order to determine the effects on total glutathione levels (as a measure of generic oxidative stress), genotoxicity (DNA strand breaks using Comet assay in haemocytes), DNA adduct analyses (using (32)P-postlabelling method) in different organs, histopathological changes in different tissues (i.e. adductor muscle, digestive gland and gills) and physiological effects (feeding or clearance rate). Subsequently, in the third experiment, a combined exposure of C(60) plus fluoranthene (0.10 mg l(-1) and 32 μg l(-1), respectively) was carried out to evaluate all endpoints mentioned above. Both fluoranthene and C(60) on their own caused concentration-dependent increases in DNA strand breaks as determined by the Comet assay. Formation of DNA adducts however could not be detected for any exposure conditions. Combined exposure to C(60) and fluoranthene additively enhanced the levels of DNA strand breaks along with a 2-fold increase in the total glutathione content. In addition, significant accumulation of C(60) was observed in all organs, with highest levels in digestive gland (24.90 ± 4.91μg C(60) g(-1) ww). Interestingly, clear signs of abnormalities in adductor muscle, digestive gland and gills were observed by histopathology. Clearance rates indicated significant differences compared to the control with exposure to C(60), and C(60)/fluoranthene combined treatments, but not after fluoranthene exposure alone. This study demonstrated that at the selected concentrations, both C(60) and fluoranthene evoke toxic responses and genetic damage. The combined exposure produced enhanced damage with additive rather than synergistic effects.