Organic Pollutants in Environment - Markers and Biomarkers of Toxicity


Genetic Toxicology of Pesticides


The studies will be focused on:

  • evaluating the genotoxic effects of pesticide substances at exposure levels relevant for real-life scenarios (residential exposure level, occupational exposure limit, acceptable daily intake or if not estimated yet reference dose)
  • in vitro cell culture models will be used (primary lymphocyte cultures and HepG2 cell lines)
  • acute treatments (4h and 24h) will be performed
  • subacute effects will be evaluated on extended lymphocyte cultures
  • primary DNA will be assessed by applying the alkaline comet assay and further oxidative damage by hOGG1 modified comet assay
  • secondary DNA damage will be assessed by cytome assay (micronuclei, nuclear buds, nucleoplasmic bridges)
  • ploidy status due to malsegregation or loss of certain chromosomes will be assessed by coupling cytome assay with fluorescent in situ hybridization (FISH)
  • structural/functional integrity of gatekeeper genes TP 53 and c-Myc as biomarkers will be analyzed
  • in vivo evaluation will be performed on adult male Wistar rats, treated per os consecutively for 14 days
  • effects will be evaluated by alkaline comet assay on different tissue samples (liver, kidney, bone marrow, blood)
  • Possible transplacental genotoxicity will be estimated on pregnant Wistar rats
  • primary DNA damage in fetal blood and liver samples will be evaluated using alkaline comet assay


Picture 1. Fluorescence in situ hybridization (FISH) characterization applying pancentromeric probes in binucleated human lymphocytes. Centromeric DNA is red (Texas Red), and nuclei blue (DAPI). A represents binucleated cell containing micronucleus with two centromeric signals; B represents centromere-positive nucleoplasmic bridge



Picture 2. Comet-FISH images of a lymphocytes treated with pesticides. Total DNA is stained with DAPI. A represent comets hybridised with TP53 (TP53-red signal, cen 17 green signal); B, C  represent comets hybridised with c-Myc (c-Myc-red signal, cen 8 green signal)