The role of vitamins A, C, E and selenium as antioxidants against genotoxicity and cytotoxicity of cadmium, copper, lead and zinc on erythrocytes of Nile tilapia, Oreochromis niloticus.

This study was carried out to investigate the genotoxic and cytotoxic potentials of sublethal concentration (5mg L(-1)) of combined metals including Cd, Cu, Pb and Zn (1.25mg L(-1) of each) on erythrocytes of Nile tilapia, Oreochromis niloticus after exposure for five and seven days; and to evaluate the protective role of vitamin E alone and a combination of selenium (Se) with vitamins A, C and E which was added to the diet as antioxidants against the genotoxicity and cytotoxicity of these metals. This was accomplished by application of micronuclei (MN), binuclei (BN), nuclear abnormalities (NAs) assays in addition to morphological erythrocyte alteration (MAEs) assay. The results revealed that, exposure of O. niloticus to Cd, Cu, Pb and Zn induced the formation of nine genotoxic endpoints including MN, BN and seven patterns of NAs, kidney-shaped nuclei, blebbed nuclei, lobed nuclei, bilobed nuclei, notched nuclei, hook-shaped nuclei and vacuolated nuclei; and five patterns of morphological malformations were recorded as cytotoxic endpoints including echinocytes, acanthocytes, teardrop-like erythrocytes, microcytes and fused erythrocytes. Frequencies of these abnormalities were significantly different (p<0.05) in comparison to control group. The maximum number of MN, BN and most of NAs and MAEs were recorded in the 5th day of exposure and then start to decrease as recorded in the 7th day. Addition of the vitamin E alone to the diet significantly (p<0.05) decreased the frequencies of MN, BN, and most of NAs and MAEs to become less than those recorded in metals-treated fish. But, addition of a combination of Se with vitamins A, C and E in the diet resulted in more significant decrease (p<0.05) in frequencies of MN, BN, NAs and most MAEs to become less than those recorded in both, fish treated with metals only and fish treated with metals and supplied with vitamin E alone in the diet. Therefore, this study confirms the powerful protective potential of the vitamin E alone and a combination of SE with vitamins A, C and E as antioxidants against the genotoxicity and cytotoxicity of Cd, Cu, Pb and Zn in erythrocytes of O. niloticus. Also, confirmed on the validity of MN test and NAs in addition to MAEs as effective indicators and valuable sensitive monitoring tools for detecting genotoxic and cytotoxic agents in the aquatic environment.

Toxic responses and antioxidative enzymes activity of Scenedesmus obliquus exposed to fenhexamid and atrazine, alone and in mixture.

Laboratory studies were conducted to determine the effects of different concentrations of fenhexamid and atrazine (25, 50 and 100 µg L(-1)) on growth and oxidative stress on Scenedesmus obliquus (microalgae) after exposure for 24, 48, and 96 h. In addition, residues of fenhexamid and atrazine were determined in the culture medium after 96 h; 52%, 44% and 43% of fenhexamid remained in the medium for the lowest, middle and highest concentrations, respectively. Atrazine concentration decreased significantly in the medium with time. The reduction was faster with the lowest concentration (-53%), than in the highest concentration (-46%), while it was intermediate with 50 µg L(-1) (-47%). The antioxidative enzyme activities were used as biomarkers to evaluate the toxic effects of fenhexamid and atrazine on the microalgae. Enzymatic activities were measured in the presence of each compound alone after 24, 48 and 96 h and also in mixture after 24h exposure. The results showed that fenhexamid and atrazine induced antioxidative enzyme activities (GST, CAT and GR) at different concentrations. Catalase activities (CAT) in both pesticides treated-algae were significantly increased. Additionally, an increase in gulathione-S-transferase (GST) was observed in algae after 24, 48 and 96 h of exposure to both fenhexamid and atrazine. Antioxidative enzymes in fenhexamid and atrazine mixture treatment showed an antagonistic interaction after 24h of exposure in algae.

Toxic effects and bioaccumulation of the herbicide isoproturon in Tubifex tubifex (Oligocheate, Tubificidae): a study of significance of autotomy and its utility as a biomarker.

Tubifex is the only animal reported to respond with autotomy to contamination. This response of contaminated worm is understood as a mode of metal excretion. Few data concern the potential of organic compounds to induce tubifex autotomy. The objective of this study was to investigate if autotomy can be induced by a herbicide isoproturon (IP) and be related to the way of excretion. Isoproturon accumulation in worm tissues and its effect on tubifex mortality, autotomy and regeneration rates were analysed after 4 and 7 days of exposure to the herbicide and also when worms were replaced for 10 days in clean water. IP accumulated in the same way in all parts of the worm body but IP metabolite rates were significantly higher in the posterior part of the worm. Thus the loss of the posterior part allows the worm to eliminate an important amount of pesticide. Autotomy has a population importance and is related to the degree of worm contamination so it may become an interesting biomarker.

Effects of chitosan on oxidative stress and metallothioneins in aquatic worm Tubifex tubifex (Oligochaeta, Tubificidae).

Chitosan is a natural polymer which has the property to elicit the natural defenses mechanism in plant and which can be an interesting biopesticides. It is then necessary to investigate the potential toxicity of chitosan for aquatic animal health. Metallothioneins (MTs) are low molecular weight proteins, mainly implicated in metal ion detoxification. Increase in MTs contents had been considered as a specific biomarker of metal exposure. However recently it has been demonstrated that MTs participate in several cellular functions such as regulation of growth and anti-oxidative defenses. Therefore, the induction of MTs has been investigated in the aquatic worms Tubifex tubifex exposed to chitosan. MTs levels in exposed worm increased significantly (p > 0.05) after 2, 4, and 7 days of exposure to different concentrations of chitosan (maximum + 158.19 +/- 10.2% after 2 days of exposure to 125 mgl(-1) of chitosan). Several antioxidant parameters including glutathione (GSH), glutathione-S-transferase (GST), glutathione reductase (GR), and catalase (CAT) were quantified in T. tubifex after 2, 4, and 7 days of exposure to chitosan. Exposure to chitosan had a negative effect on T. tubifex growth (maximum effect -6.11 +/- 1.6% after 7 days with 125 mgl(-1)) demonstrating the toxic effect of the pesticide. This growth rate decrease was accompanied by a reduction in protein contents. The activity of catalase (CAT), glutathione-S-transferase (GST), and glutathione reductase (GR) increased in response to the chitosan demonstrating an oxidative stress in the worms.

Metallothioneins induction and antioxidative response in aquatic worms Tubifex tubifex (Oligochaeta, Tubificidae) exposed to copper.

Metallothioneins (MTs), are low molecular weight proteins, mainly implicated in metal ion detoxification. Increase in MT contents is considered as a specific biomarker of metal exposure. Recently it has been demonstrated that MTs participate in several cellular functions such as regulation of growth, and antioxidative defences. Tubifex tubifex were exposed to different copper concentrations (50, 100, and 200 microgl(-1)) for 7 and 15 days. MT levels in exposed worms increased significantly (p<0.05) after 7 and 15 days of exposure to different concentrations of copper (maximum +208% for 100 microgl(-1) after 7 days of exposure). Also important perturbation in metal-metallothionein content occurred, along with an increase in total soluble protein content in all treated worms after 7 and 15 days (max. +88.49%). Catalase activities (CAT) in Cu treated-worms were significantly increased, and demonstrated a development of antioxidative defenses. Additionally a reduction of gulathione-S-transferase (GST) was observed in all treated worms after 7 days of exposure to Cu (max. -44.42%). The high induction of MTs observed during T. tubifex exposure to Cu make them potentially useful biomarkers to monitor metal pollution.

Effects of the herbicide isoproturon on metallothioneins, growth, and antioxidative defenses in the aquatic worm Tubifex tubifex (Oligochaeta, Tubificidae).

Metallothioneins (MTs) are low molecular weight proteins, mainly implicated in metal ion detoxification. Increase in MT contents is considered to be a specific biomarker of metal exposure. Recently it has been demonstrated that MTs participate in several cellular functions such as regulation of growth, and antioxidative defenses. Therefore, the induction of MTs as biomarkers of exposure to the pesticide isoproturon has been investigated in the aquatic worms Tubifex tubifex. MT levels in exposed worms increased significantly (p < 0.05) after 2, 4, and 7 days of exposure to different concentrations of isoproturon (maximum increase compared to unexposed controls: +148.56% for 10 mg l(-1) after 4 days of exposure). In response to isoproturon, the activity of glutathione-S-transferase (max. +52%), glutathione-reductase (max. +100%), and catalase (max. +117%) increased, demonstrating the occurrence of an oxidative stress response to the herbicide. Thus, the increase in MT contents caused by isoproturon was interpreted as a defense response towards increased oxidative stress generated by the herbicide. Residues of isoproturon and its metabolites, 1-(4-isopropylphenyl)-3-methylurea, 1-(4-isopropylphenyl) urea, and 4-isopropylanilin were detected in the worm growth medium. Half-life of the herbicide was shorter at a low (0.1 mg l(-1)) initial concentration. The herbicide accumulated in T. tubifex but no metabolite could be detected.

Metallothionein induction, antioxidative responses, glycogen and growth changes in Tubifex tubifex (Oligochaete) exposed to the fungicide, fenhexamid.

Laboratory studies were conducted to determine the effects of different concentrations of fenhexamid (0.1, 1, and 10 mg L(-1)) on growth, oxidative stress, protein, glycogen, and metallothionein (MT) contents in Tubifex tubifex after an exposure of 2, 4, and 7 days. In addition, residues of the fungicide were followed in water and in the worms. In water, fenhexamid concentration decreased slowly (maximum -2 +/- 0.03% after 2 days for 1 mg L(-1)). In the worms, it increased after 4 days and decreased thereafter, confirming that the worms were exposed to the fungicide and not to a degradation product. LC50 values were between 95.22 +/- 5.36 and 32.11 +/- 1.8 mg L(-1) depending on exposure time. Exposure to fenhexamid had a negative effect on T. tubifex growth (maximum effect -12.2 +/- 0.8% after 7 days with 10 mg L(-1)) demonstrating the toxic effect of the pesticide. This growth rate decrease was accompanied by a reduction in protein and glycogen contents. The activity of catalase (CAT), and glutathione reductase (GR) increased in response to the fungicide demonstrating an oxidative stress in the worms. In contrast glutathion-S-transferase activity (GST) decreased. Exposure to fenhexamid also induced synthesis of MT (maximum +78 +/- 8% after 2 days for 10 mg L(-1)). The specificity of MT concentration increase in response to metals is discussed.

Comparative toxicity and biochemical responses of certain pesticides to the mature earthworm Aporrectodea caliginosa under laboratory conditions.

This study was conducted to investigate the toxicity of aldicarb, cypermethrin, profenofos, chlorfluazuron, atrazine, and metalaxyl toward mature Aporrectodea caliginosa earthworms. The effects of the LC(25) values of these pesticides on the growth rate in relation to glucose, soluble protein, and activities of glutamic-oxaloacetic transaminase (GOT), glutamic-pyruvic transaminase (GPT), acid phosphatase (AcP), and alkaline phosphatase (AIP) were also studied. The results showed that aldicarb was the most toxic of the tested pesticides, followed in order by cypermethrin, profenofos, chlorfluazuron, atrazine, and metalaxyl. A reduction in growth rate was observed in all pesticide-treated worms, which was accompanied by a decrease in soluble protein and an increase in transaminases and phosphatases. Relationships between growth rate, protein content, transaminases, and phosphatases provided strong evidence for the involvement of pesticidal contamination in the biochemical changes in earthworms, which can be used as a bioindicator of soil contamination by pesticides.

Acute and sublethal effects of two insecticides on earthworms (Lumbricus terrestris L.) under laboratory conditions.

Earthworms (Lumbricus terrestris L.) were exposed to commercial formulations of endosulfan and aldicarb for 2, 7, and 15 days, and the LC(10), LC(25), and LC(50) were determined. Worms were then exposed to LC(10), LC(25), and LC(50) concentrations of endosulfan and LC(10) and LC(25) concentrations of aldicarb. The growth rate and total protein content were determined and related to endosulfan and aldicarb residues in soil and earthworms. Aldicarb was more toxic than endosulfan under the experimental conditions. The residues of endosulfan and aldicarb caused a significant reduction in the growth rate and total protein content of earthworms. The residues of endosulfan and aldicarb were monitored in soil and earthworms after 2, 7, and 15 days of exposure. The residues remaining in the soil after the experiments ranged between 37.75% and 68.54% of the applied concentration for endosulfan and between 10.13% and 67.71% of the applied concentration for aldicarb. Small amounts of both insecticides were detected in worms, and accumulation was more important for endosulfan. This study proposes the use of growth rate and total protein content as biomarkers for contamination by endosulfan and aldicarb.