A multibiomarker approach to evaluate the impact of anthropogenic contaminants on the ecotoxicological status of honey bees, Apis mellifera

A rapid decline of Apis mellifera, a keystone pollinator of wild plant species and agricultural crops, was recorded worldwide in recent years. The massive use of insecticides and fungicides in agriculture associated with pollution generated by other human activities and presence of parasites can cause toxicological effects in bees including a decrease of the immune defences, leading to the collapse of the colonies. Effective assessment of the ecotoxicological impacts of anthropogenic contaminants requires an approach that combines different biomarkers that enable a more precise diagnosis of exposure to environmental stressors through a combination of different biological responses. The aim of this study was to develop and apply a set of biomarkers to study the ecotoxicological status of honey bees. In the first phase, we investigated in the laboratory the effects of EMS, cadmium and a commercial fungicide (azoxistrobin 18.2% and ciproconazole 7.3%) in adult honey bees, evaluating eventual variation in glutathione S-transferase (GST), carboxylesterase (CaE), acetylcholinesterase (AChE) activities, alkaline phosphatase (ALP), lysozyme, erythrocyte nuclear abnormalities (ENA) assay and differential haemocytes count (DHC). Genotoxic effects, as well as alteration of the immune system, were found in bees treated with EMS, cadmium or the fungicide. Cadmium and the fungicide also inhibited AChE and CaE activities, GST was induced by all the compounds investigated. In the second phase, adult honey bees were collected from apiaries located in four environments characterized by different chemical input: a wooded environment (low input), an urban site, an orchard and a cultivated countryside site. Honey bees from the urban site were also collected and analyzed before and after treatment for parasites. ENA assay showed that bees taken from the countryside and the orchard had a greater number of abnormalities compared to the forest, confirming the presence of genotoxic substances in agricultural environments compared to control environments. GST activity was induced in bees from the urban environment, AChE was inhibited in the countryside compared to the forest, suggesting the presence of substances with neurotoxic effect in this environment. ALP activity was induced in all sites in comparison to wooded one. The bees collected after the parasites treatment showed an increase for GST activity as well as AChE inhibition.