Lipid peroxidation and antioxidant defense enzymes in Clarias gariepinus as useful biomarkers for monitoring exposure to polycyclic aromatic hydrocarbons

The toxicological evaluations of crude oil, xylene, toluene and benzene were carried out against juvenile stages of Clarias gariepinus. On the basis of LC(50) value, benzene (0.017 ml/l) was found to be the most toxic followed by xylene (0.086 ml/l), toluene (0.398 ml/l) and crude oil (2.219 ml/l) was the least toxic. The results of the lipid peroxidation assay showed that the level of malonaldehyde (MDA) in liver and gills of fish exposed to all the test chemicals increased significantly (P < 0.05) when compared to control. Measurement of activities of antioxidant enzymes, superoxide dismutase (SOD), catalase (CAT) and glutathione-S-transferase (GST) in liver and gill of fishes exposed to sublethal concentration of the test chemicals over a 28-day period was found to decrease significantly (P < 0.05) when compared to control. The observed reduction in the activities of antioxidant defense enzymes such as SOD, CAT and GST in conjunction with an increase in MDA levels in the gill and liver tissues of test animals exposed to aromatic hydrocarbon compounds were recommended as a good battery of biomarkers for early detection of pollution during biomonitoring programmes.     

Oxidative stress in Nile tilapia (Oreochromis niloticus) and armored catfish (Pterygoplichthys anisitsi) exposed to diesel oil

Considering that diesel oil is one of the most common aquatic contaminants, we compare the oxidative stress between two species of fish with different habitats (Pterygoplichthys anisitsi, benthic and Oreochromis niloticus, nektonic) exposed to diesel oil. Malondialdehyde concentrations (MDA) and the activities of ethoxyresorufin-O-deethylase (EROD), glutathione S-transferase (GST), catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase were all analyzed in the fishes' livers and gills after 2 and 7 days of exposure to different concentrations of diesel (0.1 and 0.5 mL/L). In the tilapia, MDA levels and the activities of EROD and GST activity in the liver, as well as MDA levels and the activities of GST and SOD in the gill had statistically significant differences between the treatments and between the times of exposure. For the catfish, the same occurred in the case of MDA, EROD, and SOD in the liver and in CAT and SOD in the gills. There were significant differences in the enzyme activity and lipid peroxidation between the species. Although the activity of most enzymes seemed to be more expressive and responsive to diesel in O. niloticus, diesel oil also caused significant effects on oxidative stress parameters in P. anisitsi, even though this species is benthic and thus has less access to insoluble fractions of diesel oil. Therefore, both species can be used as sentinel organisms in environmental biomonitoring of diesel contamination.     

水体突发性重金属污染胁迫下斑马鱼的行为反应分析

研究了斑马鱼在不同程度突发性的Zn2＋和Cr6＋胁迫下的行为反应,采用计算机视觉技术量化研究斑马鱼游动行为数据。结果表明,在Zn2＋和Cr6＋的突发性胁迫中,斑马鱼的行为反应快速且敏感,游动行为与污染物种类、质量浓度和暴露时间直接相关,并表现出比较相似的行为变化规律;随着胁迫程度的增加,斑马鱼游动速度的变化幅度增大,反应时间缩短,且其变化曲线符合生物行为的环境压力模型。斑马鱼的行为变化远远提前于生物的病理损伤或死亡,可利用斑马鱼暴露于污染物时的行为变化实现在线水体突发性重金属污染事故的监测预警。     

Integrated survey of water pollution in the Suquía River basin (Córdoba, Argentina)

We report a combined two-year seasonal monitoring of Suquía River basin using both chemical parameters and biomarkers measured in Jenynsia multidentata, aiming to correlate external levels of contaminants with the response of oxidative stress biomarkers in this fish. Identified pollution sources correspond to city sewage as well as agricultural and small industry activities downstream from Córdoba city. Physicochemical parameters integrated into a water quality index (WQI) were measured in Suquía River during dry and wet seasons. Ag, Mn, Cu, Cr, Ni, Fe, Pb and Zn were also monitored in water and sediment samples. Biomarkers include detoxication and antioxidant enzymes: catalase (CAT), glutathione peroxidase (GPX), glutathione S-transferase (GST) and glutathione reductase (GR). Enzymes showed a pollution dependent response, with increased activities in fish collected close to the sewage exit and progressive drop further downstream, matching changes in the Water Quality index. The combined use of biomarkers with water quality parameters allowed both the identification of pollution sources and the evaluation of effects of contaminants on the aquatic biota.     

Oxidative stress in freshwater fish, Labeo rohita as a biomarker of malathion exposure

This study examined the effect of lethal (4.5 μg/l) and sublethal (0.45 μg/l) malathion levels on oxidative stress responses of the freshwater edible fish, Labeo rohita. Fish were exposed to lethal (1–4 days) and sublethal (1, 5, 10, and 15 days) periods. In the present study, catalase and protease activity, hydrogen peroxide, malondialdehyde, protein carbonyls, and free amino acids levels increased in the gill, liver, and kidney tissues of fish exposed to lethal and sublethal concentrations of malathion except protein content. Time- and concentration-dependent induction/reduction of the above parameters by lethal and sublethal concentrations of malathion was observed in the tissues (the gill, liver, and kidney) of L. rohita. Thus, the results clearly infer oxidative damage and decline in antioxidant defense due to malathion-induced oxidative stress.     

Using condition factor and blood variable biomarkers in fish to assess water quality

The condition factor and blood variables, including erythrocyte lipid peroxidation (LPO) and the activity of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx), in two ecologically distinct fish species (Astyanax fasciatus and Pimelodus maculatus) were evaluated at five sites in the Furnas Hydroelectric Power Station reservoir (Brazil) to assess water quality. Aldrin/dieldrin, endosulfan, heptachlor epoxide, and metolachlor were detected at different concentrations in four of the sites. Condition factor was not directly affected by such contaminants. A negative correlation between hematocrit and heptachlor was detected in P. maculatus. Positive correlations between red blood cells and heptachlor as well as an interactive effect of metolachlor and aldrin/dieldrin were detected in A. fasciatus. The erythrocytes of both species collected from the contaminated sites showed high levels of LPO, an increase in SOD and GPx activities and a decrease in CAT activity. Although the leukocyte number and the differential percentage of leukocytes varied among the sites, the hematological variables, the LPO levels, and the antioxidant enzyme activities could be used to assess water quality, regardless of the differences in the responses of the fish species.     

Comparative toxicity of nano-ZnO and bulk ZnO suspensions to zebrafish and the effects of sedimentation, ˙OH production and particle dissolution in distilled water

With the common application of nanoscale zinc oxide (nZnO) and significant potential for its release directly into aquatic environments, it is urgent to carry out research on ecotoxicological impact of nZnO. The characterization of nZnO, the amount of ˙OH in suspensions in the presence of light and the acute toxicity of nZnO and its bulk counterpart suspensions, as well as the acute toxicity of Zn(2+) solution to zebrafish (Danio rerio) at 96 h were studied. It was found that nZnO aggregated into irregular shapes in suspensions, and showed a relationship between its size distribution and concentration. In the presence of light, nZnO suspensions could generate ˙OH, the concentration of which increased with time. Although it was generally thought that ˙OH played a role in the biotoxicity to zebrafish, similar toxicity was observed for the nZnO and bulk ZnO suspensions (96 h LC(50) 3.969 mg L(-1), 2.525 mg L(-1), respectively). Furthermore, the sedimentation of nZnO and bulk ZnO in suspensions, and the accumulation of Zn in zebrafish were studied. The results showed that dissolved Zn(2+), from nZnO and bulk ZnO in suspensions, were toxic to zebrafish, while the aggregation and sedimentation of nZnO suspensions reduced the toxicity of nZnO. However, Zn(2+) may not be the main source of acute toxicity of nZnO and bulk ZnO to zebrafish. The experimental results highlight the importance of a systematic assessment of toxicity mechanisms of metal oxide nanoparticles (NPs) to determine definitively whether their toxicity is caused by nano-effects.     

Evaluation of genotoxicity in Rhamdia quelen (Pisces, Siluriformes) after sub-chronic contamination with Fipronil

Diverse genetic biomarkers have been used to evaluate the effects of pollution by mutagenic agents such as metals and pesticides, as well as a large variety of chemical substances derived from human activities. This work researched the effects that an exposure of 60 days to the insecticide Fipronil (concentrations of 0.05, 0.10 and 0.23 μg/L) can cause in the fish Rhamdia quelen using Comet assay with gills, histopathological analysis of gills and the Piscine Micronucleus test and Nuclear Morphological Alterations. The results for the Comet assay and for gills histopathological injuries showed no difference between the control group and the contaminated groups. In the Piscine Micronucleus test, the smallest concentration of Fipronil (0.05 μg/L) was similar as the control group, while concentrations of 0.10 and 0.23 μg/L caused more damage to the DNA. These results suggested that only the highest concentrations of Fipronil tested cause damage in erythrocytes, but none of these concentrations was sufficient to alter the DNA in the gill cells. R. quelen may be a less sensitive bioindicator than other fish that have been tested. On the other hand, the concentrations used may not have been sufficient to detect alterations in the DNA of R. quelen with the chosen tests. Works like this take on great importance given the enormous quantity of substances that are thrown daily into the environment in an uncontrolled way, without evaluation of the consequences. The application of these tests with other concentrations, tissues and exposure times is suggested for future works.     

Assessing the effects of Cu, Cd, and exposure period on metallothionein production in gills of the Brazilian brown mussel Perna perna by using factorial design

Factorial design plan was experimented in Perna perna in order to find out the contribution of day exposure, Cd and Cu concentrations in water, and their interactions on metallothioneins (MT), Cd, and Cu (the dependent measured variables) in the gills. The picture obtained is more adequate than by studying the factor effect separately. Compared with the control group, the MT concentration after 22?days exposure period in the mixture of 100?μg/mL Cu and Cd is increased almost two times, showing that P. perna might be used as a biomonitor. Cd showed stronger effect than Cu on MT inducing.     

Histopathology alterations and lipid peroxidation as biomarkers of hydrocarbon-induced stress in earthworm, Eudrilus eugeniae

This study investigated the biomarker responses of the earthworm, Eudrilus eugeniae, exposed to sublethal concentrations of benzene, toluene and xylene (BTX) for a time span of 28 days. Lipid peroxidation (LPO) and histopathological alterations were examined. Toxicological evaluations of BTX were carried out against E. eugeniae. On the basis of the 96-h LC₅₀ value, xylene (1.212 mg/kg) was found to be the most toxic followed by toluene (1.335 mg/kg) and benzene (1.896 mg/kg) was the least toxic. The exposure of earthworms to sublethal concentrations (1/10th and 1/100th of 96 h LC₅₀) of BTX premixed with the substrate (loamy and humus soil) induced pathological changes in the clitella such as severe lesion, necrosis and dark brown pigments. The result of the lipid peroxidation assay showed a significant increase in oxidative damage with LPO values ranging from 2.58–7.8 nM/g in exposed animals when compared to 0.07 nM/g in the control group. The findings from this study suggest the use of LPO and histopathology as useful biomarkers of exposure for early detection of petroleum related stress in terrestrial ecosystems     

Alteration of antioxidant enzymes and impairment of DNA in the SiO2 nanoparticles exposed zebra fish (Danio rerio)

The incorporation of nanoparticles in industrial and biomedical applications has increased significantly in recent years, yet their hazardous and toxic effects have not been studied extensively. While standard toxicological test methods are generally capable of detecting the toxic effects, the choice of relevant methods for nanomaterials is still discussed. Among the various oxide nanomaterials, silica nanoparticles are widely used in biological applications that include nano-medicine. But studies on adverse effects of silica nanoparticle exposure to fish remain unclear. Therefore, the present study was designed to investigate the oxidative toxic effects of silicon dioxide nanoparticles using fish model. The size of the SiO₂ nanoparticles was between 68 and 100 nm which was confirmed by X-ray diffractometer, dynamic light scattering, scanning electron microscope and transmission electron microscope. The zebra fish were exposed to sub-lethal concentrations (5 and 2.5 mg/L) of characterized SiO₂ nanoparticles for a period of 7 days. After 7 days, SiO₂ nanoparticle-treated fishes were sacrificed, and tissues such as liver, muscle and gill were dissected out for the analysis of antioxidant enzymes and DNA fragmentation. The DNA profiles were analysed in the tissues of zebra fish that treated with SiO₂ nanoparticles. Tissues of fish from clean water were used as control, and DNA profiles were analysed. It is found that DNA from control tissues was intact, whereas the tissues treated with SiO₂ were all fragmented. SiO₂ nanoparticle-mediated antioxidant enzymes activities, such as catalase, superoxide dismutase, glutathione (GSH)-S-transferase, glutathione reductase and GSH, in the tissues of zebra fish were measured. The results revealed that alteration of antioxidant enzymes due to SiO₂ nanoparticle can be considered as a biomarker to SiO₂-mediated oxidative stress in biological samples.     

Binary mixtures of diclofenac with paracetamol,ibuprofen, naproxen,and acetylsalicylic acid and these pharmaceuticals in isolated form induce oxidative stress on Hyalella azteca

Toxicity in natural ecosystems is usually not due to exposure to a single substance, but is rather the result of exposure to mixtures of toxic substances. Knowing the effects of contaminants as a mixture compared to their effects in isolated form is therefore important. This study aimed to evaluate the oxidative stress induced by binary mixtures of diclofenac with paracetamol, ibuprofen, naproxen, and acetylsalicylic acid and by these nonsteroidal anti-inflammatory drugs (NSAIDs) in isolated form, using Hyalella azteca as a bioindicator. The median lethal concentration (LC₅₀) and the lowest observed adverse effect level (LOAEL) of each NSAID were obtained. Amphipods were exposed for 72 h to the latter value in isolated form and as binary mixtures. The following biomarkers were evaluated: lipid peroxidation (LPX), protein carbonyl content (PCC), and activity of the antioxidant enzymes: superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx). Significant increases in LPX and PCC with respect to the control group (p?≤?0.05) were induced by NSAIDs both in isolated form and as binary mixtures. Changes in SOD, CAT, and GPx activity likewise occurred with NSAIDs in isolated form and as binary mixtures. In conclusion, NSAIDs used in this study induce oxidative stress on H. azteca both in isolated form and as binary mixtures, and the interactions occurring between these pharmaceuticals are probably antagonistic in type.     

Seasonal variation of oxidative stress biomarkers in clams Ruditapes decussatus sampled from Tunisian coastal areas

Seven (7) sites from to the Tunisian coastal area were monitored through four seasons using several oxidative stress biomarkers including lipid peroxidation. Catalase (CAT) specific activity was determined in clam Ruditapes decussatus digestive gland. Lipid peroxidation was assessed in the same tissues using malondialdehyde (MDA) concentration, neutral lipid (NL) and lipofuscin accumulation. Results show that catalase specific activity varies between sites and fluctuates in time. The highest CAT activities were recorded in sites from Bizerta Lagoon and sites neighbouring Sfax city. Malondialdehyde accumulation in digestive gland cells seems to be very pronounced in animals sampled in summer and autumn. Digestive gland sections of clams sampled from reported heavy metal polluted sites exhibited higher volume density of residual bodies. Our results showed that the neutral lipids and lipofuscin content in clam's digestive gland are more sensitive to general stress as compared to other biomarkers, and could be used successfully in clams as early warning tools in field biomonitoring programs.     

Response mechanisms of antioxidants in bryophyte (Hypnum plumaeforme) under the stress of single or combined Pb and/or Ni

The short-term responses and mechanisms of antioxidants in moss Hypnum plumaeforme subjected to single or combined Pb and/or Ni stress has been revealed in this study, in order to clarify (1) the relationship between the stress intensity and antioxidant fluctuation, (2) the difference between single and combined stress, and (3) the possibility of biomonitoring by the application of antioxidant fluctuation under stress. The results showed that the stress induced dose dependent formation of reactive oxygen species (ROS) and subsequent lipid peroxidation. Total chlorophyll (Chl) content and superoxide dismutase (SOD) activity were initiated under lower stress but were inhibited under higher stress. Both single and combined stress decreased catalase (CAT) activity but increased peroxidase (POD) activity, indicating POD in the moss played an important role in resisting the oxidative stress induced by Pb and Ni. The accumulation of (.)O2(-) and H2O2 in H. plumaeforme was respectively related to the low activity of SOD and the decreased activity of CAT. The study indicated that Pb and Ni had synergistic effect in inducing the oxidative stress in moss H. plumaeforme, especially under the combination of high concentration of Ni (0.1, 1.0 mM) and Pb. POD and CAT activity, as well as H2O2 and MDA content, which increased or decreased regularly with a dose dependent under Pb and Ni stress, could be used as an effective indicator in moss biomonitoring, especially in the case of light pollution caused by heavy metals without the changes in the appearance of mosses.     

Assessment of contaminant impacts in a semi-enclosed estuary (Amvrakikos Gulf, NW Greece): Bioenergetics and biochemical biomarkers in mussels

A combination of bioenergetics and biochemical biomarkers in mussels was applied to assess possible pollution impacts in a protected semi-enclosed estuary (Amvrakikos Gulf, NW Greece) that receives pesticide discharges through riverine transport. Scope for growth, a physiological condition index representing the energy budget of the organism, was applied to detect general stress effects on the health status of mussels. The low energy budgets of mussels revealed stress conditions and provided early warning signals of possible consequences at higher levels of biological organization. Biochemical markers of exposure confirmed a risk of pesticide contamination. Decreased acetylcholinesterase activities indicated exposure to organophosphate and carbamate pesticides. Responses of the antioxidant enzyme glutathione peroxidase suggested the presence of contaminants capable of reactive oxygen species production that could be related to organochlorine pesticide contamination in the area. On the other hand, metallothionein levels implied low metal contamination.     

Alterations in protein metabolism in selected tissues of fish,Cyprinus carpio,during sublethal concentration of cypermethrin

Freshwater fish, Cyprinus carpio, were exposed to sublethal concentration of cypermethrin (20 µg/l) for 6, 12, 24 and 48 h to determine the protein fractions, amino acids, protease, alanine aminotransferase, aspartate aminotransferase, ammonia, urea and glutamine levels in gill, brain, liver and muscle tissues. Total, structural and soluble proteins showed decrement; whereas free amino acids and the activities of protease, aspartate aminotransferase and alanine aminotransferase significantly increased at all exposure periods in cypermethrin-exposed fish. Interestingly, ammonia content decreased but urea and glutamine levels increased in all the tissues during cypermethrin stress. It was observed that the changes steadily increased with an increase in the period of exposure and exhibited tissue specificity.     

The Use of Mytilus Galloprovincialis Acetylcholinesterase and Glutathione S-Transferases Activities as Biomarkers of Environmental Contamination Along the Northwest Portuguese Coast

With the aim of using Mytilus galloprovincialis acetylcholinesterase (AChE) and glutathione S-transferases (GST) activities as biomarkers of environmental contamination in risk assessment studies along the northwest Portuguese coast, the objective of this study was to provide background information related to: (1) baseline values of these enzymatic activities both in reference and contaminated areas and their responsiveness as indicators of exposure to environmental contaminants; (2) the importance of seasonal variations of such biomarkers in M. galloprovincialis along this area. For this, the activity of these enzymes was seasonally determined in mussels collected from four sites, including a reference and three stations with different contamination sources along the referred area. Statistically significant differences on both enzymatic activities were found among the four sampling stations, at the four sampling periods. In comparison to the reference station, lower AChE and higher GST activity values were found in mussels collected in stations potentially contaminated by pesticides and domestic/industrial effluents and in mussels collected in the vicinity of an oil refinery and an industrial/mercantile harbour, respectively. The results obtained in this work highlighted the potential suitability of these biomarkers to be used as components of environmental monitoring programs in risk assessment studies along the northwest Portuguese coast. Since a seasonal variation in both enzymatic activities was found, the possible implications of such variability in the use of these enzymes as environmental biomarkers are also discussed.     

Hepatic and extrahepatic expression of estrogen-responsive genes in male adult zebrafish (Danio rerio) as biomarkers of short-term exposure to 17β-estradiol

Growing concern over reproductive hormones in the environment demands sensitive and efficient methods by use of molecular biomarkers to detect these contaminants in oviparous vertebrates. In this study, a real-time quantitative RT-PCR was adopted to investigate the expressions of three estrogen-responsive genes, Vtg I, Vtg II and ERalpha, in hepatic and extrahepatic tissues of male adult zebrafish exposed to varying concentrations of 17beta-estradiol (E2) for selected periods. Without exposure to E2, all the genes were expressed in the tissues of male controls with their levels being much lower than those in the respective tissues of female controls. The expressions of hepatic Vtg I and II mRNAs were induced significantly after 1-day exposure to E2 at as low as 0.25 microg L(-1). Significant induction in the expression of hepatic ERalpha mRNA required a higher E2 concentration (> or = 0.5 microg L(-1)) and a longer exposure (> or = 2 days), suggesting that Vtg I and Vtg II are more sensitive to E2 exposure. The induction of Vtg mRNA in the skin of zebrafish was also significant following a short exposure (1 day) to low E2 concentration (0.25 microg L(-1)), with the levels of Vtg I and Vtg II mRNA being increased by 25 and 5 times, respectively. These results suggest that Vtg I mRNA is a highly sensitive biomarker for determining the estrogenic effects of E2 and that the skin of zebrafish may be an appropriate substitute for liver for such a determination.     

Contaminants-Induced Oxidative Damage on the Carp Cyprinus carpio Collected from the Upper Yellow River, China

The Yellow River, the second largest river in China, is the most important resource of water supply in North China. In the last 40 years, even in the upper Yellow River, with the development of industry and agriculture, more and more contaminants have been discharged into this river and greatly polluted the water. Although a routine chemical component analysis has been performed, little is known about the real toxic effects of the polluted water on organisms at environmental level. To explore whether the pollutants induced oxidative stress and damage to aquatic organisms, malondialdehyde (MDA) level and activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione S-transferase (GST) in hepatopancreas, kidney and intestine of the field-collected carp Cyprinus carpio from a mixed polluted (Lanzhou Region, LZR) and a relatively unpolluted (Liujiaxia Region, LJXR) sites of the upper Yellow River were measured. The results showed that when the values of LZR compared with those of LJXR, SOD and GST activities increased and GPx activity decreased significantly in all the three organs (P < 0.05–0.01); CAT activity decreased but MDA level increased significantly (P < 0.05–0.01) only in kidney and intestine. In conclusion, the results of this study suggest that the pollutants can induce obvious oxidative damage in the carp, and the SOD, GST and GPx might be better indicators for the oxidative damage in aquatic organisms.     

Use of biomarkers in Nile tilapia (Oreochromis niloticus) to assess the impacts of pollution in Bolgoda Lake, an urban water body in Sri Lanka

The present study reports the first analysis of water pollutants in Sri Lankan waters using a suite of biomarkers in Nile tilapia (Oreochromis niloticus) residing in Bolgoda Lake which receives urban, industrial and domestic wastes from multiple sources. The fish were collected from the lake in the dry period (April 2005) and wet periods (September 2005, October 2006) and the levels of biomarkers viz. hepatic ethoxyresorufin O-deethylase (EROD), glutathione S-transferase (GST), metallothioneins, biliary fluorescent aromatic compounds, brain and muscle cholinesterases (ChE) were compared with those of the laboratory reared control fish and the fish obtained from a less polluted water body, Bathalagoda reservoir (reference site). The results revealed that biomarker levels of the fish collected from the reference site were not significantly different from the controls. Hepatic EROD and GST activities in fish from Bolgoda Lake were induced 4.2-16.6 folds and 1.4-3.3 folds respectively compared with the control fish. Analysis of bile in the lake fish revealed recent uptake of naphthalene, pyrene and benzo(a)pyrene type polycyclic aromatic hydrocarbons (PAHs). The induction of EROD activities in feral fish reflects the exposure of fish to aryl hydrocarbon receptor agonists including PAHs present as pollutants in the Bolgoda Lake. Cholinesterase activity in the fish inhabiting one sampling site of Bolgoda Lake was lower (22-40% inhibition) than the activity measured in the control fish indicating the presence of anticholinesterase pollutants in the area. Hepatic metallothionein levels in the lake fish were higher (1.9-3.2 folds) in comparison to the controls indicating metal exposure. The results support the potential use of these biomarkers in Nile tilapia in assessing pollution in tropical water bodies.