Final Activity Report Summary - BIOECOTOX (Biomarkers: the early warning sentinel of chemical pollution risk assessment)
Water quality is impacted by several anthropogenic pressures, which impair the value of aquatic ecosystem and sustainable use of water resources. The EU policy (e.g. Water Framework Directive) requires development of bioindicators in order to monitor and assess water quality and aquatic ecosystem health. Biomarkers have long been recognised as useful tools for the environmental risk assessment in aquatic ecosystems. We define biomarkers as rapid, measurable changes occurring at lower levels of biological organisation (biochemical, molecular and cellular) as a consequence of exposure to pollutants. Ecotoxicogenomic, defined as the study of gene and protein expression in responses to environmental toxicant exposures, is an emerging field which intended to develop new molecular biomarkers to assess ecotoxicological stress at an early stage and to explain its mode of action. Moreover, some biomarker responses might be transient and adaptive, whereas other might be followed by later changes at tissue or organism level which ultimately affect populations and whole ecosystems.
In this scenario, the BIOECOTOX project proposed to use a recent and very sensitive technique, the Real Time PCR (RT-PCR) to assess the effects of pollutants at the expression level of genes of the model organism Saccaromyces cerevisiae which is a very common yeast (e.g. ingredient in beer and bread). Two aquatic pollutants of concern, belonging to two different chemical families, Benzo(a)pyrene (B(a)P) and Cadmium (Cd) were tested focusing on two crucial points(1) non toxic concentrations assessed by whole-organism responses in standardised methods (No Observed Effect Concentration or NOEC and Predicted No Effect Concentration or PNEC) and (2) full life cycle of the organism to predict the influence of life cycle on genes responses. Considering our objective to develop those molecular biomarkers for EU legislation a particular attention was brought to the quality control and the validation of our results.
None of the six tested genes were modified by the B(a)P but their responses varied with life cycle. Because we didn't observed any toxicity on the cell growth even at highest concentrations, we hypothesised that S. cerevisiae may not to be the relevant organism to detect B(a)P contamination. On the other hand, among the ten selected genes to detect cadmium, nine of them were induced after 4h exposure. Such results showed that the RT-PCR approach allowed us to investigate genes profiles in yeast at different life stage (B(a)P and Cd) and discriminate this natural variation from the deleterious effect of pollutants (Cd). This technique also allowed detecting very low concentrations of cadmium which are not detected by traditional toxicological methods based on whole-organism responses and confirmed mechanisms of action for cadmium detoxification at environmental level. Finally, the results of the BIOECOTOX project identified nine genes as promising early warning indicators of exposure to and effects of cadmium on the eukaryotic organisms in the environment.
In this scenario, the BIOECOTOX project proposed to use a recent and very sensitive technique, the Real Time PCR (RT-PCR) to assess the effects of pollutants at the expression level of genes of the model organism Saccaromyces cerevisiae which is a very common yeast (e.g. ingredient in beer and bread). Two aquatic pollutants of concern, belonging to two different chemical families, Benzo(a)pyrene (B(a)P) and Cadmium (Cd) were tested focusing on two crucial points(1) non toxic concentrations assessed by whole-organism responses in standardised methods (No Observed Effect Concentration or NOEC and Predicted No Effect Concentration or PNEC) and (2) full life cycle of the organism to predict the influence of life cycle on genes responses. Considering our objective to develop those molecular biomarkers for EU legislation a particular attention was brought to the quality control and the validation of our results.
None of the six tested genes were modified by the B(a)P but their responses varied with life cycle. Because we didn't observed any toxicity on the cell growth even at highest concentrations, we hypothesised that S. cerevisiae may not to be the relevant organism to detect B(a)P contamination. On the other hand, among the ten selected genes to detect cadmium, nine of them were induced after 4h exposure. Such results showed that the RT-PCR approach allowed us to investigate genes profiles in yeast at different life stage (B(a)P and Cd) and discriminate this natural variation from the deleterious effect of pollutants (Cd). This technique also allowed detecting very low concentrations of cadmium which are not detected by traditional toxicological methods based on whole-organism responses and confirmed mechanisms of action for cadmium detoxification at environmental level. Finally, the results of the BIOECOTOX project identified nine genes as promising early warning indicators of exposure to and effects of cadmium on the eukaryotic organisms in the environment.