Ecotoxicological status of two bird species (Falco tinnunculus and Parus major) and the influence of changing environtments: a multi-biomarker and complementary approach

Birds are crucial for ecosystem balance, but urbanization, agricultural intensification, and industrialization impact wild birds by modifying natural environments and releasing a wide range of contaminants. Multiple stressors can cause severe physiological, behavioral, and toxicological responses. A multi-tiered approach is necessary to monitor these effects, integrating biological information with data on chemical contamination and on the environment in which the different populations live. This thesis aimed to: a) conduct a broad spectrum assessment of the toxicological status of two bird species (Falco tinnunculus and Parus major) sampled in the wild using a multi-biomarker and complementary approach based on contaminant analysis, biomarkers responses and behavioural and breeding performances; b) investigate the influence of environmental changes and multi-stressors on the physiological state/adaptation ability of the two bird species. The thesis permitted to develop and apply genotoxicity and immunotoxicity biomarkers aiming to extend the biomarkers battery that can be used in vivo and in vitro studies on Common Kestrel, and in other bird species, enabling the measurement of fundamental endpoints for evaluating the health status of the avian populations. In vitro laboratory studies on Common Kestrel blood aiming to evaluate cytotoxicity and genotoxicity of two commercial pesticides, an herbicide (glyphosate) and a fungicide, using the comet assay were performed. The comet assay, used to evaluate potential DNA damage, showed substantial differences in the genotoxic potential of the two compounds, glyphosate was found to be more toxic compared to fungicide. Glyphosate was strongly toxic to cells even at concentrations well below usage doses and for very short exposure time. The developed in vitro approach showed interesting toxicological effects of two pesticides widely used and less investigated and the effectiveness in testing the toxicity of commercial pesticide formulations instead of the active principles. A two-year monitoring study on Common Kestrel aiming to evaluate the ecotoxicological status and the physiological responses of populations from areas with different anthropic pressures by applying a multi-biomarker approach combined with contaminant analysis was conducted. A wide set of biomarkers was applied on blood of nestlings sampled in urban, rural and natural areas of Lazio Region. Some of the biomarkers used, such as respiratory burst and comet assay, were developed for the first time in this species in the laboratory studies and successfully applied in this investigation. All the results suggest that the urban environment is not necessarily more risky for birds than other types of habitats. Our multi-marker approach suggests that kestrels probably face with environmental stressors that differ in typology or intensity across habitats, requiring adequate physiological responses. Indeed, important differences of contaminants concentrations among the studied environments were also found. A three-year monitoring study on Great tit aiming to evaluate the ecotoxicological status of populations from areas with different anthropic impacts using a multi-biomarker and complementary approach was conducted. The study was carried out in wood, urban and agricultural areas of the Veneto Region. The integrated monitoring approach allowed to get a complete picture of the toxicological status of great tit populations by integrating biomarkers responses with contaminant analysis data and information on behavioral traits and reproduction success parameters. We found that the great tit populations of urban and agricultural (agri 1) areas were those with the most significant impacts resulting from environmental pressures. In addition, thanks to this holistic approach, it was also possible to identify sub-lethal effects in individuals from the other areas, showing the importance of evaluating different endpoints during a monitoring study. Although biomarker alterations emerged, no evident effects at the population level were found. However, it must be taken into account that biomarker alterations represent early warning signals of possible long-term effects for individuals and populations, therefore a longer-term monitoring is needed. Overall, the multi-biomarker and complementary approach effectively assessed the impact of different environments on bird populations, providing valuable insights for land use and urban planning to better protect wildlife. This method can also be applied to other bird species and for regulatory purposes to conserve endangered birds.