Suitability of Daphnia similis as an alternative organism in ecotoxicological tests: implications for metal toxicity

The acute toxicity of metals to Daphnia similis was determined and compared to other daphnid species to evaluate the suitability of this organism in ecotoxicology bioassays. To verify the performance D. similis in toxicity tests, we also investigated the effect of Pseudokirchneriella subcapitata at 1 x 10(5) and 1 x 10(6) cells ml(-1) on Cd and Cr acute toxicity to the cladoceran. Daphnid neonates were exposed to a range of chromium and cadmium concentrations in the absence and presence of the algal cells. Metal speciation calculations using MINEQL(+) showed that total dissolved metal concentrations in zooplankton culture corresponded to 96.2% free Cd and 100% free Cr concentrations. Initial total dissolved metal concentrations were used for 48 h-LC(50) determination. LC(50) for D. similis was 5.15 x 10(-7) mol l(-1) dissolved Cd without algal cells, whereas with 1 x 10(5) cells ml(-1), it was significantly higher (7.15 x 10(-7) mol l(-1) dissolved Cd). For Cr, the 48 h-LC(50) value of 9.17 x 10(-7) mol l(-1) obtained for the cladoceran in tests with 1 x 10(6) cells ml(-1) of P. subcapitata was also significantly higher than that obtained in tests without algal cells (5.28 x 10(-7) mol l(-1) dissolved Cr). The presence of algal cells reduced the toxicity of metals to D. similis, as observed in other studies that investigated the effects of food on metal toxicity to standard cladocerans. Comparing our results to those of literature, we observed that D. similis is as sensitive to metals as other standardized Daphnia species and may serve as a potential test species in ecotoxicological evaluations.

Copper-mediated amyloid-beta toxicity is associated with an intermolecular histidine bridge

Amyloid-beta peptide (A beta) is pivotal to the pathogenesis of Alzheimer disease. Here we report the formation of a toxic A beta-Cu2+ complex formed via a histidine-bridged dimer, as observed at Cu2+/ peptide ratios of > 0.6:1 by EPR spectroscopy. The toxicity of the A beta-Cu2+ complex to cultured primary cortical neurons was attenuated when either the pi- or tau-nitrogen of the imidazole side chains of His were methylated, thereby inhibiting formation of the His bridge. Toxicity did not correlate with the ability to form amyloid or perturb the acyl-chain region of a lipid membrane as measured by diphenyl- 1,3,5-hexatriene anisotropy, but did correlate with lipid peroxidation and dityrosine formation. P-31 magic angle spinning solid-state NMR showed that A beta and A beta-Cu2+ complexes interacted at the surface of a lipid membrane. These findings indicate that the generation of the A beta toxic species is modulated by the Cu2+ concentration and the ability to form an intermolecular His bridge.

Toxicity, accumulation and depuration of heavy metalsin the brown mussel perna indica

In this thesis certain important aspects of heavy metal toxicity have been worked out. Recent studies have clearly shown that when experimental media contained more than one heavy metals, such metals could conspicuously influence the toxic reaction of the animals both in terms of quantity and nature. The experimental results available on individual metal toxicity show that, in majority of such results, unrealistically high concentrations of dissolved metals are involved. A remarkable number of factors have been shown to influence metal toxicity such as various environmental factors particularly temperature and salinity, the condition of the organism and the ability of some of the marine organisms to adapt to metallic contamination. Further, some of the more sensitive functions like embryonic and larval development, growth and fecundity, oxygen utilization and the function of various enzymes are found to be demonstrably sensitive in the presence of heavy metals. However, some of the above functions could be compensated for by adaptive process. If it is assumed that the presence of a single metal in higher concentrations could affect the life function of marine animals, more than one metal in the experimental media should manifest such effects in a greater scale. Commonly known as synergism or more than additivity, majority of heavy metals bring about synergistic reaction .The work presented in this thesis comprises lethal and sublethal toxicities of different salt forms of copper and silver on the brown mussel Perna indica. during the present investigation sublethal concentrations of copper and silver in their dent effects on survival, oxygen consumption, filtration, accumulation and depuration on Perna indica. The results are presented under different sections to make the presentation meaningful .

Dietary copper absorption and excretion in three semi-terrestrial grapsoid crabs with different levels of terrestrial adaptation

Three species of phylogenetically related semi-terrestrial crabs (Superfamily Grapsoidea - Sesarma rectum, Goniopsis cruentata and Neohelice granulata (formerly: Chasmagnathus granulatus) with different degrees of terrestriality were studied to quantify the accumulation of copper (Cu) in hemolymph, gills, hepatopancreas and antennal gland, and its excretion through the faeces. These crabs were fed for 15 days practical diets containing 0 (A), 0.5 (B), 1.0 (C), and 1.5% (D) of added CuCl2 (corresponding to 0, 0.2, 0.5 and 0.7% of Cu2+, respectively). The amount of food ingested was directly proportional to the degree of terrestriality: S. rectum, the most terrestrial species, ate around 2-3 times more than the other crabs, whereas G. cruentata ate 1.5-2 times more than N. granulata, the least terrestrial. The amount of Cu excreted in the feces was proportional to Cu ingestion, and was 76.8% and 64.2% higher for Sesarma fed diet D compared to G. cruentata and N. granulata, respectively. Sesarma also displayed higher Cu concentration in the haemolymph, gills and antennal glands, but not in the hepatopancreas. A detoxifying mechanism followed by elimination was probably present at this last organ, preventing Cu accumulation. More terrestrial crabs, such as Sesarma, may accumulate more Cu in hemolymph and tissues, showing a correlation between metal accumulation and increased terrestriality. In this aspect, contaminated feed sources with Cu may have more impact in conservation of terrestrial crabs. (C) 2008 Elsevier Inc. All rights reserved.

EVALUATION OF METAL TOXICITY BY A MODIFIED METHOD BASED ON THE FUNGUS GERRONEMA VIRIDILUCENS BIOLUMINESCENCE IN AGAR MEDIUM

Metal cation toxicity to basidiomycete fungi is poorly understood, despite its well-known importance in terrestrial ecosystems. Moreover, there is no reported methodology for the routine evaluation of metal toxicity to basidiomycetes. In the present study, we describe the development of a procedure to assess the acute toxicity of metal cations (Na(+), K(+), Li(+), Ca(2+), Mg(2+), Co(2+), Zn(2+), Ni(2+), Mn(2+), Cd(2+), and Cu(2+)) to the bioluminescent basidiomycete fungus Gerronema viridilucens. The method is based on the decrease in the intensity of bioluminescence resulting from injuries sustained by the fungus mycelium exposed to either essential or nonessential metal toxicants. The assay described herein enables LIS to propose a metal toxicity series to Gerronenia viridilucens based on data obtained from the bioluminescence intensity (median effective concentration [EC50] values) versus metal concentration: Cd(2+) > Cu(2+) > Mn(2+) approximate to Ni(2+) approximate to Co(2+) > Zn(2+) > Mg(2+) > Li(+) > K(+) approximate to Na(+) > Ca(2+), and to shed some li-ht on the mechanism of toxic action of metal cations to basidiomycete fungi. Environ. Toxicol. Chem. 2010;29:320-326. (C) 2009 SETAC

Copper oxychloride fungicide and its effect on growth and oxidative stress of potato plants

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Response of gram-positive bacteria to copper stress

The Gram-positive bacteria Enterococcus hirae, Lactococcus lactis, and Bacillus subtilis have received wide attention in the study of copper homeostasis. Consequently, copper extrusion by ATPases, gene regulation by copper, and intracellular copper chaperoning are understood in some detail. This has provided profound insight into basic principles of how organisms handle copper. It also emerged that many bacterial species may not require copper for life, making copper homeostatic systems pure defense mechanisms. Structural work on copper homeostatic proteins has given insight into copper coordination and bonding and has started to give molecular insight into copper handling in biological systems. Finally, recent biochemical work has shed new light on the mechanism of copper toxicity, which may not primarily be mediated by reactive oxygen radicals.

Identification of a Functional Homolog of the Yeast Copper Homeostasis Gene ATX1 from Arabidopsis1

A cDNA clone encoding a homolog of the yeast (Saccharomyces cerevisiae) gene Anti-oxidant 1 (ATX1) has been identified from Arabidopsis. This gene, referred to as Copper CHaperone (CCH), encodes a protein that is 36% identical to the amino acid sequence of ATX1 and has a 48-amino acid extension at the C-terminal end, which is absent from ATX1 homologs identified in animals. ATX1-deficient yeast (atx1) displayed a loss of high-affinity iron uptake. Expression of CCH in the atx1 strain restored high-affinity iron uptake, demonstrating that CCH is a functional homolog of ATX1. When overexpressed in yeast lacking the superoxide dismutase gene SOD1, both ATX1 and CCH protected the cell from the reactive oxygen toxicity that results from superoxide dismutase deficiency. CCH was unable to rescue the sod1 phenotype in the absence of copper, indicating that CCH function is copper dependent. In Arabidopsis CCH mRNA is present in the root, leaf, and inflorescence and is up-regulated 7-fold in leaves undergoing senescence. In plants treated with 800 nL/L ozone for 30 min, CCH mRNA levels increased by 30%. In excised leaves and whole plants treated with high levels of exogenous CuSO4, CCH mRNA levels decreased, indicating that CCH is regulated differently than characterized metallothionein proteins in Arabidopsis.

The ATX1 gene of Saccharomyces cerevisiae encodes a small metal homeostasis factor that protects cells against reactive oxygen toxicity.

In aerobic organisms, protection against oxidative damage involves the combined action of highly specialized antioxidant enzymes, such as superoxide dismutase (SOD) and catalase. Here we describe the isolation and characterization of another gene in the yeast Saccharomyces cerevisiae that plays a critical role in detoxification of reactive oxygen species. This gene, named ATX1, was originally isolated by its ability to suppress oxygen toxicity in yeast lacking SOD. ATX1 encodes a 8.2-kDa polypeptide exhibiting significant similarity and identity to various bacterial metal transporters. Potential ATX1 homologues were also identified in multicellular eukaryotes, including the plants Arabidopsis thaliana and Oryza sativa and the nematode Caenorhabditis elegans. In yeast cells, ATX1 evidently acts in the transport and/or partitioning of copper, and this role in copper homeostasis appears to be directly relevant to the ATX1 suppression of oxygen toxicity: ATX1 was incapable of compensating for SOD when cells were depleted of exogenous copper. Strains containing a deletion in the chromosomal ATX1 locus were generated. Loss of ATX1 function rendered both mutant and wild-type SOD strains hypersensitive toward paraquat (a generator of superoxide anion) and was also associated with an increased sensitivity toward hydrogen peroxide. Hence, ATX1 protects cells against the toxicity of both superoxide anion and hydrogen peroxide.

Copper sensitivity of cueO mutants of Escherichia coli K-12 and the biochemical suppression of this phenotype

The cueO gene of Escherichia coli encodes a multi-copper oxidase, which contributes to copper tolerance in this bacterium. It was observed that a cueO mutant was highly sensitive to killing by copper ions when cells were grown on defined minimal media. Copper sensitivity was correlated with accumulation of copper in the mutant strain. Growth of the cueO mutant in the presence of copper could be restored by addition of divalent zinc and manganese ions or ferrous iron but not by other first row transition metal ions or magnesium ions. Copper toxicity towards a cueO mutant Could also be suppressed by addition of the superoxide quencher 1,2-dihydroxybenzene-3,5-disulfonic acid (tiron), suggesting that a primary cause of copper toxicity is the copper-catalyzed production of superoxide anions in the cytoplasm. (C) 2005 Elsevier Inc. All rights reserved.

Chitosan Nanoparticles Loaded The Herbicide Paraquat: The Influence Of The Aquatic Humic Substances On The Colloidal Stability And Toxicity.

Polymeric nanoparticles have been developed for several applications, among them as carrier system of pesticides. However, few studies have investigated the fate of these materials in the environment in relation to colloidal stability and toxicity. In nature, humic substances are the main agents responsible for complexation with metals and organic compounds, as well as responsible for the dynamics of these nanoparticles in aquatic and terrestrial environments. In this context, the evaluation of the influence of aquatic humic substances (AHS) on the colloidal stability and toxicity of polymeric nanoparticles of chitosan/tripolyphosphate with or without paraquat was performed. In this study, the nanoparticles were prepared by the ionic gelation method and characterized by size distribution measurements (DLS and NTA), zeta potential, infrared and fluorescence spectroscopy. Allium cepa genotoxicity studies and ecotoxicity assays with the alga Pseudokirchneriella subcapitata were used to investigate the effect of aquatic humic substances (AHS) on the toxicity of this delivery system. No changes were observed in the physical-chemical stability of the nanoparticles due to the presence of AHS using DLS and NTA techniques. However some evidence of interaction between the nanoparticles and AHS was observed by infrared and fluorescence spectroscopies. The ecotoxicity and genotoxicity assays showed that humic substances can decrease the toxic effects of nanoparticles containing paraquat. These results are interesting because they are important for understanding the interaction of these nanostructured carrier systems with species present in aquatic ecosystems such as humic substances, and in this way, opening new perspectives for studies on the dynamics of these carrier systems in the ecosystem.

Assessment of ocular surface toxicity after topical instillation of nitric oxide donors

PURPOSE: To evaluate the ocular surface toxicity of two nitric oxide donors in ex vivo and in vivo animal models: S-nitrosoglutathione (GSNO) and S-nitroso-N-acetylcysteine (SNAC) in a hydroxypropyl methylcellulose (HPMC) matrix at final concentrations 1.0 and 10.0 mM. METHODS: Ex vivo GSNO and SNAC toxicities were clinically and histologically analyzed using freshly excised pig eyeballs. In vivo experiments were performed with 20 albino rabbits which were randomized into 4 groups (5 animals each): Groups 1 and 2 received instillations of 150 µL of aqueous HPMC solution containing GSNO 1.0 and 10.0 mM, respectively, in one of the eyes; Groups 3 and 4 received instillations of 150 µL of aqueous HPMC solution-containing SNAC 1.0 and 10.0 mM, respectively, in one of the eyes. The contralateral eyes in each group received aqueous HPMC as a control. All animals underwent clinical evaluation on a slit lamp and the eyes were scored according to a modified Draize eye test and were histologically analyzed. RESULTS: Pig eyeballs showed no signs of perforation, erosion, corneal opacity or other gross damage. These findings were confirmed by histological analysis. There was no difference between control and treated rabbit eyes according to the Draize eye test score in all groups (p>0.05). All formulations showed a mean score under 1 and were classified as non-irritating. There was no evidence of tissue toxicity in the histological analysis in all animals. CONCLUSION: Aqueous HPMC solutions containing GSNO and SNAC at concentrations up to 10.0 mM do not induce ocular irritation.

Annual and polyetic progression of citrus canker on trees protected with copper sprays

The effects of copper sprays on annual and polyetic progress of citrus canker, caused by Xanthomonas citri subsp. citri, in the presence of the Asian citrus leafminer (Phyllocnistis citrella), were evaluated in a study conducted in a commercial orchard in northwest Parana state, Brazil, where citrus canker is endemic. Nonlinear monomolecular, logistic and Gompertz models were fitted to monthly disease incidence data (proportion of leaves with symptoms) for each treatment for three seasons. The logistic model provided the best estimate of disease progress for all years and treatments evaluated and logistic parameter estimates were used to describe polyetic disease dynamics. Although citrus canker incidence increased during each of the seasons studied, it decreased over the whole study period, more so in copper-treated trees than in water-sprayed controls. Copper treatment reduced disease incidence compared with controls in every year, especially 2004-2005, when incidence was ca. 10-fold higher in controls than in treated plots (estimated asymptote values 0 center dot 82 and 0 center dot 07, respectively). Copper treatment also reduced estimated initial disease incidence and epidemic growth rates every year.

Nutrition in bean plants and anthracnose intensity in function of silicon and copper application.

Nutrition in bean plants and anthracnose intensity in function of silicon and copper application. The objective of this work was to evaluate the effect of calcium silicate and copper sulfate on anthracnose intensity and nutrition of bean plants. The experiment was conducted using an experimental design in randomized blocks following a 4 x 4 factorial arrangement , (four levels of calcium silicate and four levels of copper sulfate) and two additional treatments (plants without inoculation and plants sprinkled with Benomyl). Four evaluations of the incidence and severity of anthracnose were done, in addition to measuring, total leaf area. At the end of the evaluations, incidence: and data were integrated over time, obtaining the area under disease progress curve (AUDPC). Contents of N, P, K, Ca, Mg, B, Cu, Fe, Mn, Zn, Si and lignin were determined in the aerial Part. A linear decrease of the intensity AUDPC was observed with the increase of the doses of calcium silicate. The severity AUDPC was influenced by the doses of copper, obtaining a reduction of 35% on the higher dosage. The supply of silicon and copper altered the content of the K, mg, S, Zn, Ca and Si in the aerial part of the bean plants.

Delivery of daunorubicin to cancer cells with decreased toxicity by association with a lipidic nanoemulsion that binds to LDL receptors

A lipidic nanoemulsion termed LDE concentrates in neoplastic cells after injection into the bloodstream and thus can be used as a drug carrier to tumour sites. The chemotherapeutic agent daunorubicin associates poorly with LDE; the aim of this study was to clarify whether the derivatization of daunorubicin by the attachment of an oleyl group increases the association with LDE, and to test the cytotoxicity and animal toxicity of the new preparation. The association of oleyl-daunorubicin (oDNR) to LDE showed high yield (93 +/- 2% and 84 +/- 4% at 1:10 and 1:5 drug:lipid mass, respectively) and was stable for at least 20 days. Association with oDNR increased the LDE particle diameter from 42 +/- 4 nm to 75 +/- 6 nm. Cytotoxicity of LDE-oDNR was reduced two-fold in HL-60 and K-562 cell lines, fourteen-fold in B16 cells and nine-fold in L1210 cells when compared with commercial daunorubicin. When tested in mice, LDE-oDNR showed remarkable reduced toxicity (maximum tolerated dose > 253 mu mol kg(-1), compared with <3 mu mol kg(-1) for commercial daunorubicin). At high doses, the cardiac tissue of LDE-oDNR-treated animals had much smaller structural lesions than with commercial daunorubicin. LDE-oDNR is therefore a promising new preparation that may offer superior tolerability compared with commercial daunorubicin.