Exposure to tebuconazol in rice field and laboratory conditions induces oxidative stress in carp (Cyprinus carpio)

Pesticides can have an effect on the biochemical and physiological functions of living organisms. The changes seen in fish and their response to pesticides can be used as an example for vertebrate toxicity. In this study, carp fish (Cyprinus carpio) were exposed to different concentrations of tebuconazol fungicide, by rice field (31.95 μg/L) and laboratory (33.47 and 36.23 μg/L) conditional testing, during a 7 day period. Parameters such thiobarbituric acid-reactive substance levels (TBARS), protein carbonyl, catalase, glutathione S-transferase and acetylcholinesterase activities were studied, using the liver, brain and white muscle of the fish. The field experiment showed that the TBARS levels were increased in all the analyzed tissues. Similarly, the protein carbonyl of the liver and the brain AChE activity increased after 7 days. The laboratory experiment demonstrated that the TBARS levels in the liver were increased in both of the concentration tests. TBARS levels in the muscle increased only by the lowest test concentration. On the other hand, the protein carbonyl was increased only by the highest concentration. The results indicate that the tebuconazol exposure from the field and laboratory conditions directly affected the health of the fish, showing the occurrence of oxidative stress.     

Combined toxicological effects of pesticides: A fish multi-biomarker approach

The combined effects of two synthetic insecticides on a wide array of biomarkers in the freshwater fish Piaractus mesopotamicus were studied. Fish were exposed to sublethal concentrations of endosulfan (ED), lambda-cyhalothrin (LC), and the combination of both pesticides for 96 h. The set of analyzed biomarkers included morphometric and hematological parameters, transaminases and alkaline phosphatase activities, antioxidant enzymes activities and oxidative damage biomarkers measured in gills, liver, kidney, brain and muscle. According to the principal component analysis, the most significant effects were produced by the ED–LC combination. So, the mixture of both insecticides produced an increase in the liver-somatic index, hematological changes related to immunological biomarkers (increased white blood cells count, and alterations in the differential leukocytes count), decreased liver transaminases activity, antioxidant enzymes induction in almost every tissue, and lipid peroxidation levels increases in liver, kidney and brain of exposed fish. Our results suggest deleterious effects of ED and LC insecticides in combination, and support the usefulness of the multi-biomarker approach for the characterization of toxicological mechanisms induced by pesticides.     

Alterations in the general condition,biochemical parameters and locomotor activity in Cnesterodon decemmaculatus exposed to commercial formulations of chlorpyrifos,glyphosate and their mixtures

The Pampean region, an extensive area of South America is continuously impacted by agricultural activities and the pesticides related to them like chlorpyrifos and glyphosate. Both pesticides have been registered in freshwater bodies of the region. One of the most abundant and widely distributed fish species in Pampean streams is Cnesterodon decemmaculatus, which have to cope with this altered scenario.In the present study the toxicity of Clorfox^® and Roundup Max^®, the commercial formulations of chlorpyrifos and glyphosate, respectively, and their mixture where evaluated using a set of biomarkers at different biological organization levels in fish exposed to relevant environmentally pesticides concentrations. Somatic indexes such as the condition factor (K), and the hepato-somatic index (HSI), the locomotor activity through the distance traveled and the average speed, the enzymatic activities of acetylcholinesterase (AChE) in brain and muscle, catalase (CAT) in muscle and liver, glutathione-S-transferase (GST) in brain, liver, muscle and gills, aspartate amino-transferase (AST), alanine amino-transferase (ALT), AST/ALT ratio and alkaline phosphatase (ALP) in liver were measured on C. decemmaculatus. Adult females were exposed during 6 weeks to the following concentrations: 0.0084 μl/l and 0.00084 μl/l of Clorfox (CF), 0.2 and 2 mg/l of Roundup Max (RM) and all the combinations of these concentrations. The CF exposure caused a decrease in the condition factor and in the locomotor activity parameters and induced an increase brain AChE, liver CAT activity and AST/ALT ratio. On the other hand, the exposure to RM produced a decrease in liver GST, AST/ALT ratio and ALP activity. Finally, some pesticide combinations decrease general condition and liver GST activities, and increase brain GST and liver ALP activities. Different responses in biomarkers were observed in mixtures treatments, reflecting the complex interactions between these toxics and suggesting a suppressive action of RM on CF effects.Since the concentrations we tested are environmentally relevant and the overall fish health condition was affected, the presence of these pesticides in freshwater systems could impose a risk for populations by causing deleterious effects on C. decemmaculatus in Pampean region.     

Effects of the organophosphate insecticide,monocrotophos, on acetylcholinesterase activity in the nile tilapia fish (Oreochromis niloticus) brain

The neurotoxic effects of monocrotophos on the brain of the nile tilapia fish (Oreochromis niloticus) were examined, using a static bioassay under laboratory conditions. By probit analysis the 96 h LC₅₀ value of monocrotophos was 4.9 mg/l. After 96 h exposure to acute levels of monocrotophos, the brain acetylcholinesterase (AChE) activity decreased progressively as the concentration of monocrotophos increased. In addition, four weeks following transfer to toxicant-free water after exposure to 1 mg monocrotophos, nile tilapia fish brain regained 95% of control AChE activity. The results indicate that inhibition of AChE activity in fish exposed to monocrotophos may serve as an indicator of hazard due to application of this chemical in the natural environment.Special issue dedicated to Dr. Robert Balazs.     

Toxicity and effects of a glyphosate-based herbicide on the Neotropical fish Prochilodus lineatus

The toxicity of Roundup, a glyphosate-based herbicide widely used in agriculture, was determined for the Neotropical fish Prochilodus lineatus. The 96 h-LC(50) of Roundup was 13.69 mg L(-1), indicating that this fish is more sensitive to Roundup than rainbow trout (Oncorhynchus mykiss) and Atlantic salmon (Salmo salar). These differences should be considered when establishing criteria for water quality and animal well-being in the Neotropical region. Short-term (6, 24 and 96 h) toxicity tests were then performed to evaluate the effects of sub-lethal concentrations of the herbicide (7.5 and 10 mg L(-1)) to P. lineatus. Roundup did not interfere with the maintenance of the ionic balance and there was no significant alteration in plasma cortisol levels in Roundup-exposed fish. However an increase in plasma glucose was noted in fish exposed to 10 mg L(-1) of the herbicide, indicating a typical stress response. Catalase liver activity also showed an increase in fish exposed to 10 mg L(-1) of the herbicide, suggesting the activation of antioxidant defenses after Roundup exposure. In addition, Roundup induced several liver histological alterations that might impair normal organ functioning. Therefore, short-term exposure to Roundup at subletal concentrations induced biochemical, physiological and histological alterations in P. lineatus.     

Temperature increase results in oxidative stress in goldfish tissues. 1. Indices of oxidative stress

Levels of lipid peroxides (LOOH), thiobarbituric-acid reactive substances (TBARS), protein carbonyls and low- and high-molecular weight thiols were measured in brain, liver, kidney, and white muscle of goldfish, Carassius auratus L., over 1-12 h of high temperature (35 degrees C) exposure followed by 4 or 24 h of lower (21 degrees C) temperature recovery. LOOH and TBARS contents increased during heat shock exposure with a maximal rise of 20-fold for liver TBARS, but both mainly reversed at recovery. Protein carbonyl content was unaffected by heat shock but rose in brain, liver, and kidney during recovery. Low-molecular weight thiol concentrations unexpectedly increased up to approximately 4-fold in brain, kidney and muscle under heat shock and remained high during recovery. Protein thiol contents also rose in liver and muscle during high temperature exposure by 2- and 3-fold, respectively, and decreased to control values or below in all tissues at late recovery. Low- and high-molecular weight thiol levels inversely correlated in liver (R2=0.87) suggesting that the former was used to reduce the latter over the experiment. It is concluded that the redox balance in goldfish tissues is strictly maintained probably contributing to the high tolerance of this species to heat shock.     

Altered hematological and immunological parameters in silver catfish (Rhamdia quelen) following short term exposure to sublethal concentration of glyphosate

Using agrichemicals to control unwanted species has become a necessary and common worldwide practice to improve crop production. Although most currently used agrichemicals are considered relatively safe, continuous usage contributes for soil and water contamination and collateral toxic effects on aquatic species. Few studies correlated the presence of agrichemicals on fish blood cells and natural immune system. Thus, in this study, silver catfish (Rhamdia quelen) were exposed to sublethal concentrations (10% of the LC(50-96 h)) of a glyphosate based herbicide and hematological and natural immune system parameters were evaluated. Silver catfish fingerlings exposed to glyphosate for 96 h had a significant reduction on blood erythrocytes, thrombocytes, lymphocytes and total leukocytes in contrast to a significant increase in the number of immature circulating cells. The effect of glyphosate on natural immune system was evaluated after 24h or 10 days exposure by measuring the phagocytic index of coelomic cells, and lysozyme, total peroxidase, bacteria agglutination, bactericidal activity and natural complement hemolytic activity in the serum of fingerlings. A significant reduction on phagocytic index, serum bacteria agglutination and total peroxidase was observed only after 24h exposure to glyphosate. In contrast, fingerlings exposed to glyphosate for 10 days had a significant lower serum bacteria agglutination and lysozyme activity. Glyphosate had no effect on serum bactericidal and complement natural hemolytic activity after 24h or 10 days exposure. Nonetheless, the information obtained in this study indicates that glyphosate contaminated water contributes to alter blood cells parameters and to reduce the activity of natural immune components important to mediate fish resistance to infecting microorganisms.     

Fish Cholinesterases as Biomarkers of Sublethal Effects of Organophosphorus and Carbamates in Tissues of Labeo Rohita

Organophosphates and carbamates are major agrochemicals that strongly affect different neuroenzymes and the growth of various fish species. Here, we study the effect of sublethal concentrations of profenofos and carbofuran on the activity of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) and the associated health risk in fish. Labeo rohita fingerlings were exposed to three sublethal concentrations of profenofos and carbofuran. The minimum cholinesterase activities in the brain, gills, muscle, kidney, liver, and blood were after exposure to profenofos (0.06 mg/L). The minimum AChE and BuChE activities in the brain, gills, muscle, kidney, liver, and blood were after exposure to carbofuran (0.28 and 0.198 mg/L). Exposure to both types of pesticides affected the functions of these organs, including metabolism and neurotransmission, to various extents at different exposure concentrations. These findings suggest that they are required to be properly monitored in the environment, to reduce their toxic effects on nontarget organisms     

Impairment of the stress response in matrinxã juveniles (Brycon amazonicus) exposed to low concentrations of phenol

In this study we measured plasma cortisol, plasma glucose, plasma sodium and potassium, and liver and gill hsp70 levels in juvenile matrinxã (Brycon amazonicus) subjected to a 96 h exposure to phenol (0, 0.2, and 2.0 ppm), and the effect of this exposure on their ability to respond to a subsequent handling stress. Fish were sampled prior to initiation of exposure and 96 h, and at 1, 6, 12, and 24 h post-handling stress. During the 96 h exposure, plasma cortisol and glucose levels remained unchanged in all treatments. While plasma sodium levels were significantly reduced in all groups, plasma potassium levels only decreased in fish exposed to 0 and 0.2 ppm of phenol. Liver hsp70 levels decreased significantly at 96 h in fish exposed to 2.0 ppm of phenol. All groups, except fish exposed to 0.2 ppm of phenol, were able to increase plasma cortisol and glucose levels after handling stress. Fish exposed to 2.0 ppm of phenol showed decreased gill and liver hsp70 levels after the handling stress. Our data suggest that exposure to phenol may compromise the ability of matrinxã to elicit physiological responses to a subsequent stressor.     

Changes in tissue adenylates and water content of bluegill, Lepomis macrochirus, exposed to copper

Bluegill were exposed to copper (2–0 mg/1⁻¹ unfiltered) for 24, 48, 72 or 96 h at 24° C. This concentration approximated the 96 h LC20. Water content of liver increased 4–8% after 24 h exposure and muscle 4–6% by 48 h. These changes persisted throughout the 96 h experiments. Copper exposed fish exhibited decreases in muscle ATP, ADP, total adenylates, and energy charge by 48 h. There was a trend toward recovery of ATP with further exposure. Liver ATP of exposed fish was lower than controls at all intervals with the greatest difference evident at 48 h. No significant changes in brain adenylates were observed. Muscle and liver lactic acid was unchanged at 48 h exposure, therefore tissue hypoxia was not the cause of the adenylate changes. It was concluded that copper causes decreases in muscle and liver ATP several days before probable death of copper exposed fish and the changes seen are the result of dilution by increased tissue water, and the copper acting on certain cellular enzymes involved in detoxification and energy metabolism.     

Physiological stress response of Mountain Whitefish (<Emphasis Type="Italic">Prosopium williamsoni</Emphasis>) and White Sucker (<Emphasis Type="Italic">Catostomus commersoni)</Emphasis> sampled along a gradient of temperature and agrichemicals in the Oldman River,Alberta

Species differences in tolerance to environmental stressors can contribute to differences in species distribution and abundance along river gradients. Climate change and intensive agriculture are likely to have major effects on fish populations in temperate zones, yet understanding of the interactions between temperature and chemical stressors on fish physiology is limited. The objective of this study was to compare the stress responses of the Mountain Whitefish, (Prosopium williamsoni, a cold-water fish) and White Sucker (Catostomus commersoni, a cool-water fish), along a temperature and pesticide gradient in the Oldman River, Southern Alberta in spring and summer. Fish were seined, placed into an enclosure, and plasma cortisol, glucose, liver glycogen, and condition factor were measured. Plasma acetylcholinesterase (AChE) activity was used as an indicator of exposure to organophosphate and carbamates pesticides. Whitefish had lower plasma AChE activity and lower liver glycogen reserves compared to suckers at all sites and all sampling times but the differences in plasma cortisol were not species-specific and there were no differences in plasma glucose levels, except at one site. Plasma cortisol increased, and plasma glucose decreased along a downstream river gradient in whitefish in both spring and summer; in sucker only plasma cortisol fluctuated and only in the summer. Liver glycogen decreased along the river gradient in both species at both seasons. Our study detected important species-specific differences in AChE activities and responses of the physiological stress axis, suggesting that whitefish are more sensitive to temperature and pesticide stress than suckers.     

Toxicity assessment of insecticides commonly used in rice pest management to the fry of common carp, Cyprinus carpio, a food fish culturable in rice fields

Toxicity of four insecticides commonly used in rice pest management, chlorpyrifos, dimethoate, carbaryl and carbosulfan, to the fry of common carp was assessed through median lethal concentrations (LC₅₀) and in vivo inhibition of the brain acetylcholinesterase (AChE) enzyme at sublethal concentrations. The 96‐h LC₅₀ values for these four insecticides were determined to be 0.008, 26.11, 7.85 and 0.60 mg L^?1 respectively. Exposure of fish to a series of sublethal concentrations (0.5–5% LC₅₀) of each insecticide for 14 days resulted in concentration‐dependent inhibition in AChE activity in comparison with the controls. AChE activity was greatly inhibited in the fish exposed to sublethal concentrations of chlorpyrifos. Upon transfer to insecticide‐free water, AChE activities in fry exposed to 0.5 and 1% LC₅₀ concentrations of carbaryl and carbosulfan were restored to the control level within 7–21 days whereas the fish exposed to chlorpyrifos or dimethoate did not fully recover from the insecticide‐induced anticholinesterase action. Of the four insecticides tested, chlorpyrifos was the most toxic for the fry of common carp. Although dimethoate was least toxic for the fish under acute exposure, the restoration level of normal AChE activity was slower under chronic exposure in comparison with carbaryl and carbosulfan. Hence, the use of carbamates, especially carbaryl, to control insect pests of rice in rice‐cum‐carp culture systems is recommended when considering survival, restoration of the normal AChE activity and stamina of the cultured fish.     

The effects of selenium on oxidative stress biomarkers in the freshwater characid fish matrinxã, Brycon cephalus (Günther, 1869) exposed to organophosphate insecticide Folisuper 600 BR (methyl parathion)

Methyl parathion (MP), an organophosphate widely applied in agriculture and aquaculture, induces oxidative stress due to free radical generation and changes in the antioxidant defense system. The antioxidant roles of selenium (Se) were evaluated in Brycon cephalus exposed to 2 mg L(-1) of Folisuper 600 BR (MP commercial formulation - MPc, 600 g L(-1)) for 96 h. Catalase (CAT), glutathione peroxidase (GPx), superoxide dismutase (SOD), glutathione S-transferase (GST), reduced glutathione (GSH) and lipid peroxidation (LPO) levels in the gills, white muscle and liver were evaluated in fish fed on diets containing 0 or 1.5 mg Se kg(-1) for 8 weeks. In fish treated with a Se-free diet, the MPc exposure increased SOD and CAT activities in all tissues. However, the GPx activity decreased in white muscle and gills whereas no alterations were observed in the liver. MPc also increased GST activity in all tissues with a concurrent decrease in GSH levels. LPO values increased in white muscle and gills and did not change in liver after MPc exposure. A Se-supplemented diet reversed these findings, preventing increases in LPO levels and concurrent decreases in GPx activity in gills and white muscle. Similarly, GSH levels were maintained in all tissue after MPc exposure. These results suggest that dietary Se supplementation protects cells against MPc-induced oxidative stress.     

Correlation between metallothionein and energy metabolism in sea bass,Dicentrarchus labrax,exposed to cadmium

Specimens of sea bass (Dicentrarchus labrax) were exposed to two different cadmium concentrations (0.5 and 5 μg Cd²⁺/ml seawater) for a period of 7 days. Cadmium accumulated in the tissues of D. labrax in the following order: kidney > liver > gills at both concentrations. Accumulation patterns in fish exposed to 0.5 μg Cd²⁺/ml seawater were different with respect to 5.0 μg Cd²⁺/ml seawater. At both Cd concentrations a similar stress situation occurred during the first 4 hr as noted by the depletion of glycogen stores and the increase in free glucose in the muscle; metallothionein was induced in the liver, but failed to bind all the cytosolic Cd, which was in part bound to high-molecular-weight ligands. Fish recovered from this initial stress situation within 24 hr as indicated by the increase in glycogen and the decrease of glucose. Long-term effects were clearly dependent upon metal concentration: at lower Cd exposure, metallothionein induction increased linearly with time and counteracted the toxic effect of the metal; on the other hand, when fish were exposed to 5.0 μg Cd²⁺/ml seawater a clear stress occurred at the end of the exposure, as indicated by the notable decrease of glycogen stores, the increase of free glucose, the decrease of AEC in the muscle and the increase of Cd bound to high-molecular-weight ligands in the liver.     

Increased acetylcholinesterase activities in specimens of Sparus auratus exposed to sublethal copper concentrations

The present study looks at possible changes in the activity of acetylcholinesterase (AChE) in tissues (brain and white muscle) of the Mediterranean bony fish Sparus auratus after a 20 days exposure to sublethal concentrations (0.1 or 0.5 ppm) of copper in the marine water and on control untreated animals. The trials also included measurements of Cu concentration in the tissues to evaluate possible metal accumulation. Moreover, sedimentation analysis as well as V(max) and K(m) determination were carried out in tissue extracts of Cu-exposed or control animals. V(max) and K(m) were also determined with or without addition of Cu(2+) in the assay. No Cu accumulation occurred in brain and muscle after Cu exposure. AChE showed in both tissues a molecular polymorphism with putative globular (G) and asymmetric (A) forms. Cu exposition led to an increased specific activity and improved catalytic efficiency of AChE in brain and muscle, seemingly regarding G forms. The increase in catalytic efficiency also resulted from the in vitro assay with tissue extracts and Cu(2+) addition. The higher AChE activity and catalytic efficiency in both tissues after Cu exposition and without metal accumulation, suggests an increase of free Cu aliquot into the cells, likely due to mechanisms of metal homeostasis.     

Acute effects of diflubenzuron on the freshwater fish Prochilodus lineatus

The effects of diflubenzuron (DFB), an insecticide to control ectoparasites in fish farms, on muscle acetylcholinesterase (AChE), detoxifying and antioxidant enzymes, hematological and physiological parameters, and liver histopathology were evaluated in Prochilodus lineatus after 6, 24 and 96 h of exposure to 25 mg L(-1) of DFB. The insecticide caused a reduction in the number of erythrocytes and hemoglobin content after 96 h exposure, probably due to hemolysis. Hyperglycemic response indicated energy mobilization, and may have contributed to the increase in osmolarity after 96 h exposure to DFB. The induction of glutathione-S-transferase (GST) and catalase activities in liver pointed to the activation of xenobiotic metabolic pathways and antioxidant defenses. The decrease in muscle AChE at all experimental times showed that DFB is an AChE inhibitor. In addition, DFB induced hepatic alterations that might impair normal liver functions. These results show that DFB can cause health disorders in fish and further studies are required to better define its safe use in aquaculture.     

Effect of hypoxia on carbohydrate metabolism in scorpion tissues

The levels of glycogen, glucose, lactate, as well the activities of ten enzymes of carbohydrates metabolism in brain, liver and white muscle of sea scorpion have been investigated. Metabolite concentrations didn't change in brain and the levels of glycogen and lactate were constant in the rest tissues investigated. Glucose concentration decreased in the liver and increased in muscle. In brain hypoxia decreased the activity of hexokinase and increased one of pyruvate kinase, phosphoglucoisomerase and fructoso-1,6-bisphosphatase. In liver most of the enzymes showed the tendency to decrease of their activities. In muscle the activities of phosphofructokinase and phosphoglucoisomerase decreased. Mechanisms of carbohydrates metabolism regulation under hypoxia are discussed.     

Genotoxicity and oxidative stress in fish after a short-term exposure to silver nanoparticles

This study examined the effects of waterborne silver nanoparticles (AgNPs) on juvenile fish Piaractus mesopotamicus (“pacú”), and analyzed toxicological endpoints such as metal burdens, oxidative stress and genotoxicity in a short-term assay. Fish were individually exposed to 0 (control), 2.5, 10, and 25 μg AgNPs/L. After 24 h, silver accumulation was greater in the brain than the liver and gills at all silver concentrations. Fish exposed to higher AgNPs concentrations showed major alterations in oxidative stress markers. An increase in lipid peroxidation (LPO) levels was observed in the liver of fish exposed to 10 μg AgNPs/L with no changes in the antioxidant enzymes activities. In the case of the 25 μg AgNPs/L treatment, a hepatic activation of the enzymatic antioxidant defense occurred, and LPO levels resulted unaltered. On the other hand, the brain presented the highest LPO levels at both 10 and 25 μg AgNPs/L exposures. The AgNPs toxicity was also evidenced by the DNA damage in fish erythrocytes at higher concentrations. Summarizing, a short exposure to sublethal concentrations of AgNPs is enough to generate deleterious effects on fish, including DNA damage.     

Sensitivity of brain cholinesterase to cypermethrin toxicity in freshwater teleost Tilapia mossambica

Cypermethrin at sublethal concentrations induced significant changes in acetylcholinesterase (AChE) activity and acetylcholine (ACh) content in the brain tissue of both juvenile and adult-fish. Maximum inhibition of AChE activity is noticed at 6h and 12h after exposure to cypermethrin in juvenile and adult fish respectively. In contrast, the ACh levels registered an elevation in both the cases. During subsequent periods the rate of recovery in AChE activity and ACh content is variable in both the groups.