Ana M. Cameán

Time-dependent oxidative stress responses after acute exposure to toxic cyanobacterial cells containing microcystins in tilapia fish (Oreochromis niloticus) under laboratory conditions.

Microcystins (MCs) have been reported to induce oxidative stress in aquatic organisms including fish. The effect of acute exposure to toxic cyanobacterial material containing MCs on antioxidant enzymes and lipid peroxidation has been studied in liver, kidney and gills of tilapia fish (Oreochromis niloticus). Fish were orally exposed to a single dose of cyanobacterial cells containing 120 microg/fish MC-LR and sacrificed at 24 and 72 h. The activities of glutathione peroxidase (GPx), glutathione reductase (GR), superoxide dismutase (SOD) and catalase (CAT) enzymes in the studied organs decreased in general 24 and 72 h after the dose application, although elevation of CAT and GR was found in liver at 72 h post exposure in comparison to 24h values. In contrast, the lipid peroxidation level increased significantly in all the studied organs with the liver (3.6-fold) proving to be the most affected. Protein oxidation was also increased 1.5-fold in the liver. However, recovery in these parameters was observed in liver 72 h after exposure. The results show that an acute dose of MCs does not induce an adaptative response of the antioxidant enzymes, as a sub-chronic exposure to MCs in tilapia fish does, but a general decrease in them with an initial recovery of the oxidative damage after 72 h, expressed as enhancement of CAT and GR activities and a reduction of LPO and protein oxidation in comparison to 24h values.

Dose-dependent antioxidant responses and pathological changes in tenca (Tinca tinca) after acute oral exposure to Microcystis under laboratory conditions

The effects of cyanobacterial cells containing microcystins (MCs), toxins from cyanobacteria, on oxidative stress biomarkers from liver and kidney of Tenca fish (Tinca tinca) were investigated under laboratory conditions. Moreover, a histopathological study of liver, kidney, heart and intestine tissues was performed. Fish were orally exposed to cyanobacterial cells dosing 0, 5, 11, 25 and 55 microg MC-LR/fish mixed with the food. Results showed a dose-dependent decrease of superoxide dismutase (SOD) activity, and also of catalase (CAT) in the liver. Glutathione levels and protein oxidation, however, were not altered by the exposure to the cyanobacterial material. The microscopic study revealed tissue alterations even at the lower cyanobacterial cells doses. Onion-like hepatocytes in the liver, glomerulopathy in the kidney, loss of myofibrils in the heart and vacuolated enterocytes in the gastrointestinal tract were the main changes observed. These findings suggest that this fresh water fish can be adversely affected by cyanobacterial blooms in their natural habitats.

Cytotoxicity of carboxylic acid functionalized single wall carbon nanotubes on the human intestinal cell line Caco-2.

The unique properties of carbon nanotubes (CNTs) have been explored for their use in biomedical sciences and in biotechnological fields; however, their possible toxic effects are of concern. The sources of human exposure to nanomaterials include inhalation, ingestion, dermal contact and injection. The pulmonary and dermal effects of CNTs in vitro have been previously studied with contradictory results, but data on intestinal cells are scarce. The aim of this study was to evaluate the cytotoxicity of single wall CNTs functionalized with carboxylic acid on differentiated and non-differentiated Caco-2 cells, derived from a human intestinal adenocarcinoma. Biomarkers assessed were neutral red uptake (NR), protein content (PT), a tetrazolium salt (MTS) metabolization, LDH leakage (LDH) and cell viability by means of the trypan blue exclusion test (TBET). Moreover, a morphological study was performed. Cells were exposed to concentrations between 5 and 1,000 microg/ml CNTs and toxic effects were studied after 24h of exposure. NR and MTS results showed a concentration-dependent trend with an inhibitory response from 100 microg/ml CNT, together with an increase in LDH leakage. TBET resulted in an 80% reduction at higher concentrations, and finally PT was only modified at higher concentrations. Overall, results indicated cytotoxic effects on the Caco-2 cells with differentiated cultures showing a higher sensitivity. Thus, a hazard assessment of CNTs is necessary as the nanotechnology industry grows, and more nanoscale wastes are released into the environment.

Ecotoxicological evaluation of carbamazepine using six different model systems with eighteen endpoints

The occurrence of pharmaceutically active compounds in the aquatic environment has been recognized as one of the emerging issues in environmental chemistry. However, the ecotoxicological effects of pharmaceuticals have still not been researched adequately. Carbamazepine, an anticonvulsant commonly present in surface and groundwater, was studied, using six ecotoxicological model systems with eighteen endpoints evaluated at different exposure time periods. The battery included the immobilization of Daphnia magna, bioluminescence inhibition in the bacterium Vibrio fischeri, growth inhibition of the alga Chlorella vulgaris, and micronuclei induction and root growth inhibition in the plant Allium cepa. Cell morphology, neutral red uptake, total protein content, MTS metabolization, lactate dehydrogenase leakage and activity and glucose-6-phosphate dehydrogenase activity were studied in the salmonid fish cell line RTG-2. The total protein content, LDH activity, neutral red uptake and MTT metabolization in Vero monkey kidney cells were also investigated. The most sensitive system to carbamazepine was the Vero cell line, followed by Chlorella vulgaris, Vibrio fischeri, Daphnia magna, Allium cepa, and RTG-2 cells. EC50 values from 19 microM in Vero cells at 72 h to more than 1200 microM in other systems, were obtained. Comparing the concentrations in water and the toxicity quantified in our assay systems, carbamazepine is not expected to produce acute toxic effects in the aquatic biota under these circumstances, but chronic and synergistic effects with other chemicals cannot be excluded.

Multivariate characterization of wine vinegars from the south of Spain according to their metallic content.

Forty wine vinegar samples from the south of Spain were analyzed for mineral content. Nine metals (As, Ca, Cu, Fe, K, Mg, Mn, Na and Zn) were chosen as chemical features that may account for the rate of the vinegar fermentation process. Pattern recognition techniques were applied for distinguishing quick and slow processed vinegars. The results obtained indicated an excellent performance in both recalling and prediction ability.

Effects of dietary selenium on the oxidative stress and pathological changes in tilapia (Oreochromis niloticus) exposed to a microcystin-producing cyanobacterial water bloom

The present study investigates the role of selenium (Se) supplementation (as sodium selenite) on the oxidative stress and histopathological changes induced by cyanobacterial cells containing microcystins (MCs) in tilapia fish (Oreochromis niloticus). Variation in lipid peroxidation (LPO) levels and carbonyl groups content, reduced glutathione/oxidized glutathione (GSH/GSSG) ratio, and catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GR), glutathione peroxidase (GPx) and glutathione S-transferase (GST) activities in liver and kidney of tilapia fish exposed to a single oral dose of 120 microg MC-LR/fish and sacrificed in 24 h, were investigated in the absence and presence of 1.5, 3.0 and 6.0 microg Se/g diet. Results showed a protective role of Se depending on the dose and the biomarker considered. Thus, the lower Se dose made CAT, liver GR and kidney SOD converged to basal values, whereas LPO and liver SOD and GST needed the higher dose. Kidney GR, however, was not protected at any Se dose. Moreover, Se has also shown to have a pro-oxidant effect with increased kidney LPO values and liver and kidney GPx activities in MC-free fish. The microscopic study revealed tissue alterations induced by cyanobacterial cells in the liver, kidney, heart and gastrointestinal tract that were ameliorated by the highest Se dose assayed. The level of Se supplementation must be therefore carefully selected to provide beneficial effects and to avoid potential negative consequences.

Differential protein expression in two bivalve species; Mytilus galloprovincialis and Corbicula fluminea; exposed to Cylindrospermopsis raciborskii cells

The cyanobacteria Cylindrospermopsis raciborskii is considered a threat to aquatic organisms due to the production of the toxin cylindrospermopsin (CYN). Despite the numerous reports evidencing the toxic effects of C. raciborskii cells and CYN in different species, not much is known regarding the toxicity mechanisms associated with this toxin and the cyanobacteria. In this work, a proteomics approach based in the two-dimensional gel electrophoresis and mass spectrometry was used to study the effects of the exposure of two bivalve species, Mytilus galloprovincialis and Corbicula fluminea, to CYN producing (CYN+) and non-producing (CYN-) C. raciborskii cells. Additionally the activities of glutathione S-transferase (GST) and glutathione peroxidase (GPx) were determined. Alterations in actin and tubulin isoforms were detected in gills of both bivalve species and digestive gland of M. galloprovincialis when exposed to CYN- and CYN+ cells. Moreover, GST and GPx activities changed in gills and digestive tract of bivalves exposed to both C. raciborskii freeze dried cells, in comparison to control animals exposed to the green alga Chlorella vulgaris. These results suggest the induction of physiological stress and tissue injury in bivalves by C. raciborskii. This condition is supported by the changes observed in GPx and GST activities which indicate alterations in the oxidative stress defense mechanisms. The results also evidence the capacity of CYN non-producing C. raciborskii to induce biochemical responses and therefore its toxicity potential to bivalves. The heat shock protein 60 (HSP60), extrapallial (EP) fluid protein and triosephosphate isomerase homologous proteins from gills of M. galloprovincialis were down-regulated specifically with the presence of CYN+ C. raciborskii cells. The presence of CYN may lead to additional toxic effects in M. galloprovincialis. This work demonstrates that proteomics is a powerful approach to characterize the biochemical effects of C. raciborskii and to investigate the physiological condition of the exposed organisms.

Toxic cyanobacteria strains isolated from blooms in the Guadiana river (southwestern Spain).

This paper describes the occurrence of toxic cyanobacteria along the Guadiana River over its course between Mérida and Badajoz (Extremadura, Spain). Water sampling for phytoplankton quantification and toxin analysis was carried out regularly between 1999 and 2001 in six different locations, including two shallow, slow-flowing river sites, two streamed river sites and two drinking water reservoirs. The cyanobacterial community differed significantly between these locations, especially during the summer. The predominant genera were Microcystis, Oscillatoria, Aphanizomenon and Anabaena. Using an ELISA assay the total microcystin contents of natural water samples from the most eutrophic locations ranged from 0.10 - 21.86 microg mcyst-LR equivalent x L(-1) in Valdelacalzada and 0.10-11.3 microg mcyst-LR equivalent x L(-1) in Vitonogales, and a seasonal variation of toxin content was observed. The amount of microcystins produced by each strain was determined by ELISA assay and the detection and identification of microcystin variants of three toxic strains of Microcystis aeruginosa was performed by high performance liquid chromatography (HPLC). The analysis of microcystins of the cultured strains revealed that toxin production was variable among different strains of M. aeruginosa isolated either from different blooms or from the same bloom.

Presence and bioaccumulation of microcystins and cylindrospermopsin in food and the effectiveness of some cooking techniques at decreasing their concentrations: a review.

Microcystins (MCs) and cylindrospermopsin (CYN) are among the cyanotoxins which occur naturally, produced by different cyanobacteria species when they grow or proliferate under favorable environmental conditions. From a toxicological point of view, their relevance is due to the deleterious effects that they have been reported to induce in a wide range of organisms, including humans. Cyanotoxins intake from contaminated water and food is an important source of human exposure. Various edible aquatic organisms, plants, and food supplements based on algae, can bioaccumulate these toxins. A thorough review of the scientific data available on this topic is provided, the studies on MCs being much more numerous than those focused on CYN. The scientific literature suggests that these cyanotoxins can be accumulated at concentrations higher than their respective recommended tolerable daily intake (TDI). Finally, the influence of different cooking procedures on their levels in food has been considered. In this regard, again studies on the matter dealing with CYN have been not yet raised. MCs contents have been reported to be reduced in muscle of fish after boiling, or cooking in a microwave-oven, although the effect of other traditional cooking processes such as frying, roasting or grilling have not been demonstrated.

Effects on growth and oxidative stress status of rice plants (Oryza sativa) exposed to two extracts of toxin-producing cyanobacteria (Aphanizomenon ovalisporum and Microcystis aeruginosa)

Toxic cyanobacteria are considered emerging world threats, being responsible for the degradation of the aquatic ecosystems. Aphanizomenon ovalisporum produces the toxin Cylindrospermopsin (CYN) being a concern in fresh water habitats. This work aims to increase our knowledge on the effects of this toxic cyanobacterium in plants by studying the alterations in growth parameters and oxidative stress status of rice (Oriza sativa) exposed to the cyanobacteria cell extracts containing CYN. Significant increases in glutathione S-transferase (GST) and glutathione peroxidase (GPx) activities were detected in the different experiments performed. The roots showed to be more sensitive than leaves regarding the enzyme activities. A reduction in the leaf tissue fresh weight was observed after 9 days of plant treatment suggesting a major physiological stress. The exposure of rice plants to a mixture of A. ovalisporum and Microcystis aeruginosa cell extracts containing CYN and microcystins including microcystin-LR, resulted in a significant increase in the GST and GPx activities, suggesting a synergistic effect of both extracts. Together these results point out the negative effects of cyanotoxins on plant growth and oxidative status, induced by A. ovalisporum cell extracts, raising also concerns in the accumulation of CYN.