Vasiliki Tsarpali

Environmental and human risk assessment of landfill leachate: an integrated approach with the use of cytotoxic and genotoxic stress indices in mussel and human cells.

The present study investigates leachate hazardous effects on marine biota and human cells, with the use of a battery of assays, both under in vivo and in vitro conditions. According to the results, mussels exposed for 4 days to 0.01 and 0.1% (v/v) of leachate showed increased levels of DNA damage and micronuclei (MN) frequencies in their hemocytes. Similarly, enhanced levels of DNA damage were also observed in hemocytes treated in vitro with relevant concentrations of leachate, followed by a significant enhancement of both superoxide anions (O₂(-)) and lipid peroxidation products (malondialdehyde/MDA). On the other hand, human lymphocyte cultures treated with such a low concentrations of leachate (0.1, 0.2 and 1%, v/v), showed increased frequencies of MN formation and large MN size ratio, as well as decreased cell proliferation, as indicated by the use of the cytokinesis block micronucleus (CBMN) assay and Cytokinesis Block Proliferation Index (CBPI) respectively. These findings showed the clear-cut genotoxic and cytotoxic effects of leachate on both cellular types, as well as its potential aneugenic activity in human lymphocytes.

Seasonal alterations of landfill leachate composition and toxic potency in semi-arid regions

The present study investigates seasonal variations of leachate composition and its toxic potency on different species, such as the brine shrimp Artemia franciscana (formerly Artemia salina), the fairy shrimp Thamnocephalus platyurus, the estuarine rotifer Brachionus plicatilis and the microalgal flagellate Dunaliella tertiolecta. In specific, leachate regularly collected from the municipal landfill site of Aigeira (Peloponissos, Greece) during the year 2011, showed significant alterations of almost all its physicochemical parameters with time. Further analysis showed that seasonal alterations of leachate composition are related with the amount of rainfall obtained throughout the year. In fact, rainfall-related parameters, such as conductivity (Cond), nitrates (NO(3)(-)), total nitrogen (TN), ammonium (NH(4)-N), total dissolved solids (TDS) and the BOD(5)/NH(4)-N ratio could merely reflect the leachate strength and toxicity, as verified by the significant correlations occurred among each of them with the toxic endpoints, 24 h LC(50) and/or 72 h IC(50), obtained in all species tested. According to the result of the present study, it could be suggested that the aforementioned leachate parameters could be used independently, or in combination as a low-cost effective tools for estimating leachate strength and toxic potency, at least in the case of semi-arid areas such as the most of the Mediterranean countries.

Investigation of landfill leachate toxic potency: An integrated approach with the use of stress indices in tissues of mussels

The present study investigates the harmful impacts of landfill leachate release and/or disposal into the marine environment, as well as its ability to induce lethal and pre-pathological alterations in marine organisms, such as the mussel Mytilus galloprovincialis. In specific, mortality test (96 h), performed first in order to estimate leachate lethal endpoints, showed increased levels of mussel mortality after exposure to leachate higher than 0.5%, v/v (96 h LC(50)=0.526%, v/v), while the exposure to 0.01 and 0.1% (v/v) of leachate showed negligible levels of mortality (96 h LC(10)=0.167%, v/v). Furthermore, the estimation of lysosomal membrane integrity in hemocytes of exposed mussels (Neutral Red Retention Time assay) showed increased levels of lysosomal destabilization in cells of mussels exposed to sub-lethal concentrations of leachate (0.01, 0.1 and 0.5%, v/v) for 4 days. In order to exclude parameters, such as mussel mortality and cell death, which could interfere with the obtained results, leachate at final concentrations of 0.01 and 0.1% (v/v) were finally used for the estimation of a battery of stress indices in target tissues of mussels, such as hemolymph, gills and digestive gland. According to the results, leachate-exposed mussels showed a significant inhibition of acetylcholinesterase activity, increased levels of nuclear abnormalities, as well as increased levels of metallothionein, superoxide anion and lipid peroxides (in terms of malondialdehyde equivalents) in each tissue tested. The results of the present study clearly indicate leachate-induced lethal effects as well as the ability of leachate to induce disturbances on different levels of organism function before mortality occurs.

Investigation of toxic effects of imidazolium ionic liquids, [bmim][BF4] and [omim][BF4], on marine mussel Mytilus galloprovincialis with or without the presence of conventional solvents, such as acetone.

This study investigated the cytotoxic, oxidative and genotoxic effects of two commonly used imidazolium ionic liquids (ILs), [bmim][BF4] (1-butyl-3-methylimidazolium) and [omim][BF4] (1-methyl-3-octylimidazolium tetrafluoroborate), on the marine mussel Mytilus galloprovincialis, as well as whether acetone could mediate their toxic profile. In this context, mussels were firstly exposed to different concentrations of [bmim][BF4] or [omim][BF4], with or without the presence of acetone (at a final concentration of 0.06% v/v), for a period of 96h, in order to determine the concentration that causes 50% mussel mortality (LC50 values) in each case. Thereafter, mussels were exposed to sub- and non-lethal concentrations of ILs for investigating their ability to cause lysosomal membrane impairment (with the use of neutral red retention assay/NRRT), superoxide anion and lipid peroxidation byproduct (malondialdehyde/MDA) formation, as well as DNA damage and formation of nuclear abnormalities in hemocytes. The results showed that [omim][BF4] was more toxic than [bmim][BF4] in all cases, while the presence of acetone resulted in a slight attenuation of its toxicity. The different toxic behavior of ILs was further revealed by the significantly lower levels of NRRT values observed in [omim][BF4]-treated mussels, compared to those occurring in [bmim][BF4] in all cases. Similarly, [bmim][BF4]-mediated oxidative and genotoxic effects were observed only in the highest concentration tested (10mgL(-1)), while [omim][BF4]-mediated effects were enhanced at lower concentrations (0.01-0.05mgL(-1)). Overall, the present study showed that [bmim][BF4] and [omim][BF4] could induce not only lethal but also nonlethal effects on mussel M. galloprovincialis. The extent of [bmim][BF4] and/or [omim][BF4]-mediated effects could be ascribed to the length of each IL alkyl chain, as well as to their lipophilicity. Moreover, the role of acetone on the obtained toxic effects of the specific ILs was reported for the first time, giving evidence for its interaction with the ILs and the modulation of their toxicity.

Effect of cultivation media on the toxicity of ZnO nanoparticles to freshwater and marine microalgae.

The aim of this work was to investigate the effect of zinc oxide nanoparticles (ZnO NPs) on freshwater and marine microalgae cultivated in different media. Freshwater species Chlorococcum sp. and Scenedesmus rubescens were cultivated in modified Blue-Green medium (BG-11) and Bolds Basal Medium (BBM), and marine species Dunaliella tertiolecta, and Tetraselmis suesica, cultured in salt modified BG-11 and f/2 medium. The microalgae species were exposed for 96 h with a daily reading of algal growth rate, to different ZnO NPs concentrations (0.081-810 mg/L). Significant differences were observed on microalgae growth rates, with the marine being more sensitive than the freshwater species, as revealed by their half inhibitory concentration values (IC50). The IC50 values in freshwater species were affected by the culture medium. The lowest IC50 values (<2.57 mg/L) were observed in the marine species. S. rubescens showed the less toxic effect in cultures with modified BG-11, compared to BBM cultures, with IC50 values >810 mg/L and 14.27 mg/L after 96 h exposure time, respectively. ZnO nanoparticles appeared to have toxic effects in all species tested, depended on the species type, the exposure time, the NPs concentration, and mainly the culture medium.

A Multidisciplinary Assessment of River Surface Water Quality in Areas Heavily Influenced by Human Activities

The present study could serve as a multidisciplinary approach for the assessment of river surface water quality with the use of chemical and biological methods. Specifically, physicochemical parameters, heavy metals, and pesticides were measured in water samples from three different stations (sampling station S1, S2, and S3) along Asopos River (Greece). In parallel, algal species (primary producers)—such as Scenedesmus rubescens and Chlorococcum sp.; consumer invertebrate species, such as the fairy shrimp Thamnocephalus platyurus and the rotifer Brachionus calyciflorus; as well as human lymphocytes—were exposed to those samples for assessing their toxic and genotoxic/mutagenic effects. According to the results, although the values of almost all of the physicochemical parameters tested, heavy metals (zinc, cadmium, lead, and mercury) and pesticides were lower than or within the respective environmental quality standards, thus offering no clear evidence for their natural or anthropogenic origin. Values recorded for nickel, chromium, hexavalent chromium, and malathion represent a typical case of mixed influence from natural and anthropogenic enrichments. In contrast, the algal growth arrest, the acute toxic effects on the freshwater invertebrates, and the increased micronuclei frequencies observed in human lymphocytes showed the presence of human-derived hazardous substances, which were hardly determinable with the use of conventional chemical methods. Given that the presence of priority pollutants in river surface waters, heavily burdened by anthropogenic activities, could give no clear evidence for their biological risk, the results of the present study showed that chemical and biological assays should be applied in parallel, thus serving as a reliable tool for the assessment of river water quality.

The role of acetone in the [omim][BF4]-mediated adverse effects on tissues of mussels, human lymphocytes and the fruit fly Drosophila melanogaster

The present study investigated [omim][BF4]-mediated adverse effects on biological models widely used in toxicological studies. Specifically, mussels of the genus Mytilus, human lymphocytes and fruit flies of the species Drosophila melanogaster, were exposed to [omim][BF4] at concentrations ranging from micro- to milligrams per liter, with or without the presence of acetone as a carrier solvent and thereafter [omim][BF4]-mediated adverse effects were analyzed appropriately (stress indices, such as lipid peroxidation byproducts, acetylcholinesterase/AChE activity and micronucleus/MN formation frequency, in mussel gills, Cytokinesis Block Micronucleus/CBMN assay and SMART test in human lymphocytes and fruit flies respectively). LC-MS-TOF analysis was also performed for elucidating [omim][BF4] mode of action in the presence of the carrier solvent. The results showed the toxic potential of [omim][BF4], as well as acetones ability to attenuate [omim][BF4]-mediated toxicity in almost all cases, probably due to the significant effect of acetone on the hydrophilic-lipophilic character and the viscosity of [omim][BF4], as well as its interaction and permeability on the cell membranes. The slight involvement of acetone in the attenuation of [omim][BF4]-mediated genotoxic effects on D. melanogaster could be due to species feeding experimental conditions, thus favoring the induction of antioxidant defense system against the [omim][BF4]-mediated effects in all cases.

[omim][BF4]-mediated toxicity in mussel hemocytes includes its interaction with cellular membrane proteins

The current study is based on the increasing demand for the assessment of ionic liquid (IL)-mediated aquatic toxicity. Specifically, although a lot of studies have been performed so far, investigating IL-mediated adverse effects on numerous aquatic organisms, little is known about their mode of action. Given that the use of in vitro models is considered as a reliable tool for determining the mediated biological effects, the modulation of specific biochemical pathways and the onset of various forms of damage with great precision and reproducibility, mixed primary cultures of mussel Mytilus galloprovincialis hemocytes were used for investigating whether 1-octyl-3-methylimidazolium tetrafluoroborate ([omim][BF4]) mediated toxicity is related to its interaction with cellular membrane proteins. Specifically, [omim][BF4]-mediated cytotoxic, oxidative and genotoxic effects were investigated in mussel hemocytes before and after pre-treatment of cells with non-toxic concentration of guanidine hydrochloride (1 mM GndHCl). The results showed that [omim][BF4] at concentrations ranging from 0.7 to 1.75 μM can induce cytotoxic (almost <50% reduction of cell viability), oxidative (increased levels of O2•- production and lipid peroxidation by-products) and genotoxic (increased levels of DNA damage) effects, while cells pre-treated with 1 mM GndHCl showed a significant attenuation of ILs toxic potency in all cases. According to the latter, the current study showed that [omim][BF4]-mediated toxicity could be related not only to its well-known interaction with membrane lipid bilayers, but also to its interference with membrane proteins. Using GndHCl, a chaotropic agent that disrupts the hydrogen bonding network and the stability of membrane proteins via its interference with the intramolecular interactions mediated by non-covalent forces on cellular membranes, it was firstly shown that altering the membrane integrity as well as the native state of cellular membrane proteins, by weakening the hydrophobic effect, could attenuate the possible interaction of [omim][BF4] with cellular membranes and the concomitant induction of protein-based intracellular processes, commonly linked with the induction of severe cellular damage.