Copper monitoring in vineyard soils of central Italy subjected to three antifungal treatments, and effects of copper sub-lethal doses on the earthworm Eisenia fetida.

none6The extensive employment of copper-based fungicides has increased copper concentration in vineyard soils. The present study’s objectives were to monitor copper concentration in two vineyard soils during two cropping seasons and study the ecotoxicological effects on the earthworm Eisenia fetida. Total, soluble, and bioavailable copper fractions were measured at the end of two cropping seasons and different depths in two vineyards of central Italy, characterised by three anticryptogamic control methods: copper compounds, chitosan, and combined treatments of them. A laboratory experiment to assess the effects on Eisenia fetida was conducted with soil samples collected in the vineyards with a mean copper concentration of 60 mg/kg and two higher concentrations of 90 and 150 mg/kg. Results showed low levels of total copper concentration in the first 20 cm of soils, regardless of antifungal treatment, highlighting prudent management of the vineyards under study, but the soluble fractions showed a significant increase in all samples during the two cropping seasons. At the dose of 150 mg/kg, earthworms suffer during the first two days, showing weight loss and DNA damage, but they are able to recover until day 28, showing no permanent harm at this copper concentration in soil.openArianna De Bernardi, Enrica Marini, Cristiano Casucci, Luca Tiano, Fabio Marcheggiani, Costantino VischettiDE BERNARDI, Arianna; Marini, Enrica; Casucci, Cristiano; Tiano, Luca; Marcheggiani, Fabio; Vischetti, Costantin

DNA damage in different Eisenia andrei coelomocytes sub-populations after in vitro exposure to hydrogen peroxide

Earthworms play an essential role in providing soil fertility and may represent an important soil contamination bio-indicator. They are able to ingest soil particles, adsorb substances throughout the intestinal epithelium into the coelomic cavity, where chemicals can come in direct contact with coelomic fluid. Earthworm coelomic fluid shelters leucocytes (coelomocytes) that differ significantly both structurally and functionally. Cellular variability could lead to different susceptibility towards contaminants possibly present in soil ecosystem. In order to define population specific dose response to chemicals and to identify a homogeneous cell population to be used as a relevant biomarker, we investigated different coelomocytes subpopulation, obtained by Percoll density gradient centrifugation (5–35 %), exposed ex vivo to H2O2 in the range of concentration 15–120 µM. DNA damage levels were assessed by the comet assay on unseparated coelomocytes and on three enriched cellular fractions (light, medium and heavy density subpopulations). All tested samples showed a dose–response genotoxic effect following H2O2 exposure. Moreover, light density sub-population appeared more susceptible to oxidative insult highlighted by a significant increase in DNA damage indexes at lower concentrations of H2O2. Present data suggested that in these experimental condition coelomocytes light fraction may represent a more sensitive biomarker of genotoxic insult

Rheological properties of guar and its methyl, hydroxypropyl and hydroxypropyl-methyl derivatives in semidilute and concentrated aqueous solutions

We report on a comparative study of the theological properties of guar [GG], methyl guar [MG], hydroxypropyl guar [HPG] and hydroxypropyl-methyl guar [MHPG] polymers aqueous solutions in semidilute (both unentangled and entangled) and concentrated regimes, using oscillatory and steady-shear techniques. In the dilute regime, molecular weights and radii of gyration have been investigated by means of light scattering measurements. Data obtained from steady-shear rheology were satisfactorily analyzed according to Cross model and the effects of polymer concentration and temperature on the theological behaviour of guar and guar derivatives have been investigated and discussed in terms of theological parameters, such as the zero-shear viscosity eta(0), the characteristic time tau and critical coil-overlap concentration C*. The storage and loss moduli of guar and guar derivatives aqueous solutions have been measured using angular frequencies in the range between 10(-3) and 10 rad/s. The data have been analyzed using the "blob" model for semidilute solutions and the scaling approach proposed by Rubinstein, Dobrynin and Colby for concentrated solutions. These theological parameters obey a time-concentration superposition principle, so that master curves can be constructed over a wide frequency range. Moreover, we show that, at lower temperatures, these systems behave as thermo-rheological simple systems, in that the oscillatory shear response at different temperatures can be superimposed according to the empirical time- temperature superposition principle. Although these systems can be conveniently described within a unifying scaling model, the behaviour of guar derivatives are somewhat different. At higher temperatures, relatively small deviations from the scaling behaviour of the storage modulus of MG and MHPG polymers were observed. These findings can be justified by a structural re-organization of the macromolecular network, due to the hydrophobic interactions. (C) 2010 Elsevier Ltd. All rights reserved

Evaluation of gene expression of different molecular biomarkers of stress response as an effect of copper exposure on the earthworm eisenia andrei.

The paper reports the results of a laboratory test on the bioaccumulation and toxicological effects of sub-lethal soil concentration of copper, a widely used fungicide in organic farming, on DNA damage, a critical marker increasingly used in ecotoxicology in the earthworm Eisenia andrei. In the same experimental setting we evaluated gene expression of classical biomarker of stress induced by xenobiotic. [Heat Shock Protein 70 (HSP70) and Metallothionein (MET)], as well as genes coding for enzymes involved in detoxification of reactive oxygen species [Superoxide dismutase (SOD) and catalase (CAT)]. Additionally, expression of genes involved in the immune response were investigated: a Toll-like receptor (TLR), a receptor with cytolytic activity named Cytolytic Factor (CCF) and two antimicrobial peptides, fetidin (FET) and lysenin (LYS). Results showed significant time-dependent bioaccumulation of Cu and DNA damage at concentrations remarkably lower than those found in most agricultural soils worldwide. MET was increased as was FET and TLR. The present work gives new insights into the mechanisms of sub-lethal toxicity of copper as an environmental pollutant and in the identification of novel sub-lethal biomarkers of cellular response to the stressor such as immune response genes

Soil Bacterial Community and Earthworms: Hand-in-Hand First Responders to Pesticides

Earthworms and microbial communities represent a large fraction of soil living biomass and play an essential role in soil functioning. They are also important non-target organisms in soil and sub-lethal dose toxicity assessment represents a decisive step for the regulatory authorities to evaluate the collateral impact of pesticides. Here we present a study where we evaluated through High Throughput Sequencing (HTS) of 16S rDNA amplicons the sub-lethal impact of spinosyn and organophosphate based pesticides on soil bacteria with or without earthworms during 28d long experiment. We found that community composition were changing over time rapidly starting from the first day with the most significant impact on the 7th and 14th days. Formation of hierarchical clusters mainly as a function of time and then treatments in the presence/absence of earthworms was observed. Multivariate canonical correspondence analysis that assessed the pesticides, earthworm's presence and their combined impact at the beginning (1d) and at the end of the experiments (28d) marked further differences caused by various treatments at the beginning of the experiments and fade-out phenomenon as time passed by. Specific OTUs with previously reported pesticide degradation potential and earthworm gut microbiota presence were also singled out as the main drivers of change in soil microbial community. Overall, these findings clearly indicated the buffering effect earthworms had on the bacterial communities starting from earliest sampling (1d) until the final sampling (28d) as well as bacterial community members’ degradation response to pesticides over time

Ecotoxicological effects of a synthetic and a natural insecticide on earthworms and soil bacterial community

Earthworms and microbial communities are essential non-target soil organisms that are useful to assess the collateral impact of pesticides. The present paper reports three laboratory experiments performed to investigate the effects of sub-lethal doses of two insecticides, a biologically-derived (spinosad) and a synthetic organophosphate (chlorpyrifos), on earthworm Eisenia foetida and microorganisms in organic soil. The effects were studied in terms of behaviour, reproduction, survival, and DNA damage (comet assay) in earthworms, and Next Generation Sequencing-Illumina was employed to detect the changes in the microbial community. In addition, the influence of earthworms on the degradation kinetics of insecticides and on microbial diversity was evaluated. The weights, reproductive activity and behaviour of earthworms were particularly compromised and followed a dose-dependent trend in chlorpyrifos trials, where the insecticide’s degradation wasn’t affected by the presence of Eisenia foetida. However, earthworms contributed to spinosad’s metabolisation without significantly impacting their health. Early DNA damage was estimated in earthworms exposed to chlorpyrifos, while the impact of spinosad was significant only at the end of the toxicity test. The analysis on the microbial community indicated the buffering effect earthworms had on the bacterial communities starting from earliest sampling until the end of the trial, as well as bacterial community members’ degradation response to pesticides over time