Ecotoxicological Investigation in Three Model Species Exposed to Elutriates of Marine Sediments Inoculated With Bioplastics

The aim of this study was to evaluate a set of ecotoxicity biotests on three marine model species exposed to elutriates of marine sediments inoculated with the biodegradable plastic Mater-Bi or with cellulose. The sediments were incubated at 28°C and tested after 6 months, when clear signs of degradation were visible in the exposed samples, and after 12 months, when the samples had completely disappeared. The model organisms selected for the study were the unicellular algae Dunaliella tertiolecta, the sea urchin (Paracentrotus lividus) and sea bass (Dicentrarchus labrax) juveniles. The unicellular algae and sea urchins were used to determine the toxicological endpoint of growth inhibition and embryotoxicity, respectively, and the sea bass juveniles were tested to evaluate sublethal effect endpoints using lipid peroxidation and genotoxicity biomarkers. Elutriates of sediment inoculated with Mater-Bi for 6 and 12 months showed an absence of toxic effects in all of the model organisms exposed in this study. The Mater-Bi degradation process did not generate or transfer into the elutriates toxic substances that could cause alterations in the growth of D. tertiolecta, in the P. lividus embryo-toxicity assay or in the sensitive biomarker responses of the fish D. labrax. The tested species are considered to be representatives of different levels of the marine trophic chain. The results obtained in this study suggest that the ecotoxicological approach applied may be suitable for investigating the environmental impact of the degradation of bioplastics in marine sediments

Biochemical responses, feeding and survival in the solitary bee Osmia bicornis following exposure to an insecticide and a fungicide alone and in combination

In agricultural ecosystems, bees are exposed to combinations of pesticides that may have been applied at different times. For example, bees visiting a flowering crop may be chronically exposed to low concentrations of systemic insecticides applied before bloom and then to a pulse of fungicide, considered safe for bees, applied during bloom. In this study, we simulate this scenario under laboratory conditions with females of the solitary bee, Osmia bicornis L. We studied the effects of chronic exposure to the neonicotinoid insecticide, Confidor (R) (imidacloprid) at a realistic concentration, and of a pulse (1 day) exposure of the fungicide Folicur (R) SE (tebuconazole) at field application rate. Syrup consumption, survival, and four biomarkers: acetylcholinesterase (AChE), carboxylesterase (CaE), glutathione S-transferase (GST), and alkaline phosphatase (ALP) were evaluated at two different time points. An integrated biological response (IBRv2) index was elaborated with the biomarker results. The fungicide pulse had no impact on survival but temporarily reduced syrup consumption and increased the IBRv2 index, indicating potential molecular alterations. The neonicotinoid significantly reduced syrup consumption, survival, and the neurological activity of the enzymes. The co-exposure neonicotinoid-fungicide did not increase toxicity at the tested concentrations. AChE proved to be an efficient biomarker for the detection of early effects for both the insecticide and the fungicide. Our results highlight the importance of assessing individual and sub-individual endpoints to better understand pesticide effects on bees

Biochemical responses, feeding and survival in the solitary bee Osmia bicornis following exposure to an insecticide and a fungicide alone and in combination

In agricultural ecosystems, bees are exposed to combinations of pesticides that may have been applied at different times. For example, bees visiting a flowering crop may be chronically exposed to low concentrations of systemic insecticides applied before bloom and then to a pulse of fungicide, considered safe for bees, applied during bloom. In this study, we simulate this scenario under laboratory conditions with females of the solitary bee, Osmia bicornis L. We studied the effects of chronic exposure to the neonicotinoid insecticide, Confidor® (imidacloprid) at a realistic concentration, and of a pulse (1 day) exposure of the fungicide Folicur® SE (tebuconazole) at field application rate. Syrup consumption, survival, and four biomarkers: acetylcholinesterase (AChE), carboxylesterase (CaE), glutathione S-transferase (GST), and alkaline phosphatase (ALP) were evaluated at two different time points. An integrated biological response (IBRv2) index was elaborated with the biomarker results. The fungicide pulse had no impact on survival but temporarily reduced syrup consumption and increased the IBRv2 index, indicating potential molecular alterations. The neonicotinoid significantly reduced syrup consumption, survival, and the neurological activity of the enzymes. The co-exposure neonicotinoid-fungicide did not increase toxicity at the tested concentrations. AChE proved to be an efficient biomarker for the detection of early effects for both the insecticide and the fungicide. Our results highlight the importance of assessing individual and sub-individual endpoints to better understand pesticide effects on bees

A Novel Ex Vivo Approach Based on Proteomics and Biomarkers to Evaluate the Effects of Chrysene, MEHP, and PBDE-47 on Loggerhead Sea Turtles (Caretta caretta)

The principal aim of the present study was to develop and apply novel ex vivo tests as an alternative to cell cultures able to evaluate the possible effects of emerging and legacy contaminants in Caretta caretta. To this end, we performed ex vivo experiments on non-invasively collected whole-blood and skin-biopsy slices treated with chrysene, MEHP, or PBDE-47. Blood samples were tested by oxidative stress (TAS), immune system (respiratory burst, lysozyme, and complement system), and genotoxicity (ENA assay) biomarkers, and genotoxic and immune system effects were observed. Skin slices were analyzed by applying a 2D-PAGE/MS proteomic approach, and specific contaminant signatures were delineated on the skin proteomic profile. These reflect biochemical effects induced by each treatment and allowed to identify glutathione S-transferase P, peptidyl-prolyl cis-trans isomerase A, mimecan, and protein S100-A6 as potential biomarkers of the health-threatening impact the texted toxicants have on C. caretta. Obtained results confirm the suitability of the ex vivo system and indicate the potential risk the loggerhead sea turtle is undergoing in the natural environment. In conclusion, this work proved the relevance that the applied ex vivo models may have in testing the toxicity of other compounds and mixtures and in biomarker discovery

“Test Tube Cetaceans”: From the Evaluation of Susceptibility to the Study of Genotoxic Effects of Different Environmental Contaminants Using Cetacean Fibroblast Cell Cultures

Population estimates of various species of cetaceans indicate that certain species have declined dramatically this century. Most studies of contamination and biomarker responses in marine mammals have been conducted using animals killed by hunting, tacitly approving this activity. The development of a series of nondestructive techniques to evaluate biomarker responses and residue levels is strongly recommended for the hazard assessment, protection and conservation of endangered species of marine mammals. A non-invasive sampling method, represented by skin biopsy or integument biopsy (epidermis, dermis and blubber), have been developed and validated in cetaceans. In this paper we present the cetacean fibroblast cell cultures obtained from the skin biopsies as the “test tube cetacean” for evaluate both the susceptibility that the genotoxicity of different environmental contaminants. Fibroblast cell cultures were obtained from many species of cetaceans sampled in Mediterranean Sea (Italy) and in the Mar de Cortez (Mexico). Using test tube cetaceans we can study the relationships between contamination and biochemical responses. One of the principal applications of this developed in vitro system was the assessment of interspecies differences in the mixed function oxidase activity (Cyp1A1 and Cyp2B) induced by in vitro treatment of various contaminants, such as some Persistent Organic Pollutants (POPs) but also emerging contaminants (such as bisphenol A (BpA)) and nanoparticles, added at different concentrations. The induction of Cyp1A1 and Cyp2B was evaluated with the indirect immunofluorescence technique. Therefore, another purpose of this work was to evaluate the qualitative and quantitative MICA protein expression in fibroblast cell cultures with immunofluorescence technique as toxicological stress marker of the immune system of different species of cetaceans. Finally, In this paper we will present how to evaluate the presence of DNA damage by comet assay in test tube cetaceans, after treatment with different genotoxic compounds (for example PCBs, DDTs, PAHs, BPA)

Brodifacoum Levels and Biomarkers in Coastal Fish Species following a Rodent Eradication in an Italian Marine Protected Area: Preliminary Results

Brodifacoum is the most common rodenticide used for the eradication of invasive rodents from islands. It blocks the vitamin K cycle, resulting in hemorrhages in target mammals. Non-target species may be incidentally exposed to brodifacoum, including marine species. A case study conducted on the Italian Marine Protected Area of Tavolara Island was reported after a rodent eradication using the aerial broadcast of a brodifacoum pellet. Brodifacoum presence and effects on non-target marine organisms were investigated. Different fish species were sampled, and a set of analyses was conducted to determine vitamin K and vitamin K epoxide reductase concentrations, prothrombin time, and erythrocytic nuclear abnormalities (ENA) assay. In all the examined organisms, brodifacoum was not detected. The results obtained showed differences in vitamin K and vitamin K epoxide concentrations among the samples studied, with a positive correlation for three species between vitamin K, vitamin K epoxide, and fish weight. The prothrombin time assay showed a good blood clotting capacity in the fish. Higher abnormality values were recorded for four species. The results of this study suggest that it is possible to hypothesize that the sampled fish were not likely to have been exposed to brodifacoum and that consequently there are no negative issues concerning human consumption

Could persistent organic pollutants affect future generations of sea turtles by maternal transfer? First results for Caretta caretta nests along the North-Western coast of Italy

Since 2013, loggerhead sea turtle (Caretta caretta, Linnaeus 1758) nesting has been observed further north along the Italian coast, reaching the Tuscan coast (NW Mediterranean Sea). The four nesting events that occurred in Tuscany in the summer of 2019 spurred the scientific community to monitor these occurrences more carefully, following them from egg deposition to hatching. This provided an opportunity to collect samples for conducting multidisciplinary investigations, including the toxicological investigations of the biological material collected from the four nests. The aim of this study was to conduct an initial assessment of persistent organic pollutants in the eggs laid in Tuscany, aiming to establish a baseline on this topic for subsequent nesting events that have occurred until today. Organochlorine compounds (OCs), specifically polychlorinated biphenyls (PCBs), dichlorodiphenyltrichloroethane (DDT) and its metabolites, and hexachlorobenzene (HCB), were analyzed and detected in unhatched eggs, embryos, and chorio-allantoic membranes (CAMs). OCs were detected in all samples, with PCBs > DDTs ≫ HCB. A significant spatial variation in pollutant levels and profiles among sea turtle nesting locations was found. Embryos showed higher levels of contamination than egg contents regardless of the developmental stages. Depth of the laying chamber and egg mass were not significant factors in OC bioaccumulation. For the first time in the Mediterranean Sea, this study assessed the role of CAM in the transfer of contaminants to the embryo. Overall, the OC levels found were lower compared to the results from other studies conducted worldwide on loggerhead sea turtle biological material. This was the first assessment of nest biological material for the North-Western coast of Italy

Gli effetti dei contaminanti sulla vita delle api

L'agricoltura influisce in maniera significativa sul nostro benessere e su quello degli ecosistemi. Purtroppo, negli ultimi cinquant'anni il massiccio utilizzo della chimica ha favorito una produzione intensiva orientata più alla massimizzazione delle rese che alla qualità. Oggi, però, nel nostro paese è in corso una trasformazione radicale, guidata da una rete di produttori pionieri, esempio di sostenibilità ed eccellenza del made in Italy. Se infatti l'agroecologia è stata a lungo tenuta al margine delle politiche nazionali ed europee, grazie anche alle strategie Farm to Fork e Biodiversità 2030 della Commissione Europea negli ultimi anni si è imposta come una delle realtà più promettenti nel contrasto ai cambiamenti climatici e per la tutela della biodiversità. Attraverso i contributi di docenti universitari, professionisti del settore e le testimonianze di 30 tra le aziende italiane innovative, "Agroecologia circolare" analizza gli aspetti fondamentali di un settore agroalimentare rispettoso dell'ambiente e capace di produrre cibo sano ed equo. Inoltre, attraverso la valorizzazione degli scarti, l'utilizzo di materiali eco-compatibili, lo sviluppo delle rinnovabili e di nuove tecnologie, riesce a favorire la resilienza delle comunità e a trasferire l'amore per la terra alle nuove generazioni

Review on Sublethal Effects of Environmental Contaminants in Honey Bees (Apis mellifera), Knowledge Gaps and Future Perspectives

Honey bees and the pollination services they provide are fundamental for agriculture and biodiversity. Agrochemical products and other classes of contaminants, such as trace elements and polycyclic aromatic hydrocarbons, contribute to the general decline of bees’ populations. For this reason, effects, and particularly sublethal effects of contaminants need to be investigated. We conducted a review of the existing literature regarding the type of effects evaluated in Apis mellifera, collecting information about regions, methodological approaches, the type of contaminants, and honey bees’ life stages. Europe and North America are the regions in which A. mellifera biological responses were mostly studied and the most investigated compounds are insecticides. A. mellifera was studied more in the laboratory than in field conditions. Through the observation of the different responses examined, we found that there were several knowledge gaps that should be addressed, particularly within enzymatic and molecular responses, such as those regarding the immune system and genotoxicity. The importance of developing an integrated approach that combines responses at different levels, from molecular to organism and population, needs to be highlighted in order to evaluate the impact of anthropogenic contamination on this pollinator species

Biomarkers responses in the earthworm Eisenia fetida (Savigny, 1826) exposed to commercial fungicides: laboratory and field toxicity assessment

The use of pesticides in agriculture is a common practice to protect crops all over the world. However, pesticides potentially affect soil macro-and microorganism because only 0.1% of an applied pesticides reaches the target organism. Earthworm is a model organism in ecotoxicological research and it is a useful bioindicator of chemical toxicity in terrestrial ecosystem. The aim of this study was to evaluate the potential toxic effects of commercial fungicides (two foliar and two ear diseases with double and single active substance) on the earthworm Eisenia fetida (Savigny, 1826). E. fetida was exposed to sub lethal concentration of fungicides in laboratory and field condition and a set of biomarkers were investigated. In laboratory experiments, animals were exposed in a filter paper test to increasing concentration of the four fungicides. Field investigations were conducted transplanting E. fetida in cages in the soil of wheat and durum wheat fields during treatment with different combinations of the four fungicides. All the E. fetida were analysed to evaluate vitality, inhibition of acetylcholinesterase activity, glutathione S-transferase, lipid peroxidation, catalase activity, total glutathione, Comet assay and lysozyme activity. Laboratory studies with the fungicide with double active substances showed alterations in organism’s vitality which increased with increasing treatment doses. Significant alteration of GST enzymes with respect to controls were detected at environmentally relevant doses of ear deseases fungicide. The laboratory treatment with foliar fungicides caused an alteration of LPO which increased with increasing treatment doses, and an increase inhibition of lysozime activity. All the fungicides showed a total glutathione levels decrease probably due to a cellular degeneration caused by oxidative stress. The Comet assay showed the highest DNA fragmentation values in the experiment with fungicides with double active principle. In the field studies, the test proved that leaf fungicides have greater toxicity compared to those applied to the ears. Foliar fungicide with double active substances revealed to have the most toxic effects, inhibiting the lysozyme activity and determining high levels of GST and LPO. This study represents a first step towards a better understanding of commercial fungicides toxicological potential to non-target organisms and a wide set of biomarkers allowed to investigate the main toxicological effect on these organism