DNA damage detection by Comet Assay on Daphnia magna: Application in freshwater biomonitoring

Monitoring of water genotoxicity still remains underexploited in risk assessment. The present study aimed at standardizing and evaluating the sensitivity and applicability of the Comet Assay adapted for Daphnia magna in genotoxicological investigations in freshwater environments. Two sampling campaigns (2014–2015) were performed in the watercourses of a pilot basin located in the Parma district (Italy). Fourteen sampling stations with different Ecological Status and/or EBI values were selected, all with a good Chemical Status according to the EU–Water Framework Directive 2000/60. The Alkaline Comet Assay was performed on 48 h-aged daphnids exposed (24 h) to 23 water samples. In parallel, the acute toxicity test was carried out. Daphnids exposed to samples, collected upstream the main watercourses in non-impacted areas, showed low DNA migration (Tail Intensity percentage – TI% - in the range 2.97–13.21), similar to laboratory controls. An increase in genotoxicity (TI% in the range 20–40) proceeding from the mountain towards the plain area was observed, in agreement with the land uses and the ES/EBI values of the stations. The highest genotoxic damage was observed after exposure to samples from watercourses of the minor hydrographic network in the plain area and waterbodies receiving wastewater treatment plant outflows. A modified version of the Comet Assay able to identify the presence of genotoxins inducing DNA oxidative damage, after standardization, was applied to daphnids treated with waters from 4 selected monitoring stations. The presence of oxidative contaminants was detected downstream a wastewater treatment plant outflow.