Toxicological and ecotoxic impact of secondary and tertiary treated sewage effluents

Secondary sewage effluents are discharged in significant quantities in aquatic environments delivering pollutants that were not removed during treatment; yet advanced treated effluents are not lacking of contaminants. In this study, biochemical biomarkers were measured in liver and kidney of rainbow trout (Oncorynchus mykiss) exposed to unchlorinated, chlorinated and tertiary treated secondary sewage effluents. In addition, organic matter, nitrogen and suspended solids were assayed, while a common bioassay, Daphnia magna 21d reproduction test was also applied in order to examine potential relation between the performed bioassay and the biomarkers. Processes using oxidative conditions, such as ozonation and chlorination, resulted in significantly increased breeding rate (up to 74%) of the organism. Biomarkers measurements incorporated the determination of total glutathione (GSH), glutathione S-transferases (GST), glutathione peroxidase (GPX), lipid peroxidation (LPO) and an innovative biomarker in such applications, haem peroxidase. In general, the response of biomarkers was dependent upon the treatment method and it was tissue specific. Secondary effluents inhibited liver GST and haem peroxidase, while GSH levels and LPO were significantly provoked in liver. Ozonation provoked hepatic peroxidation, in terms of haem peroxidase and LPO, and GST; while the protective (to Reactive Oxidant Species – ROS) GSH was depleted, suggesting extended ROS attack to the organism. Similar response of biomarkers (but to a lesser extend) was observed after exposure of trout to effluents submitted to both coagulation and ozonation, emphasizing the significance of removing the residual organic matter by other methods than oxidative ones. Ozonation also enhanced renal LPO and GPX; however the former employment of coagulation limited the peroxidation phenomena. Chlorination mainly affected the levels of total GSH in both tissues.     

Antioxidant enzymes in brackishwater oyster, Saccostrea cucullata as potential biomarkers of polyaromatic hydrocarbon pollution in Hooghly Estuary (India): seasonality and its consequences.

Use of antioxidant enzymes as biomarkers often becomes a complicated process at application level because they show considerable seasonal fluctuation due to both natural and biological factors. In this study, we studied the consequences of seasonal variation of antioxidant enzymes [catalase (EC 1.11.1.6), superoxide dismutase (SOD, EC 1.15.1.1), glutathione peroxidase (GPX, EC 1.11.1.9) and microsomal NADPH-DT diaphorase (EC 1.6.99.2)] in the digestive gland of wild brackishwatcr oysters, Saccostrea cucullata for biomonitoring against polyaromatic hydrocarbon (PAH) contamination in Hooghly Estuary, north-eastern coast of India. As a general trend, maximum antioxidant enzyme activities were detected in pre-monsoon period or summer (March-June) followed by a gradual decrease during monsoon (July-October) with a minimum in post-monsoon period or winter (November-February) and this pattern was similar to tissue concentrations of PAHs also. The physiological fluctuations of the antioxidant defense systems were inversely-related to the lipid peroxidation indicating an enhanced susceptibility of oyster tissues to oxidative stress during post-monsoon or winter period. However, the oysters from polluted populations exhibited consistent very high PAHs load in their tissues as well as significant increases in the activities of antioxidant enzymes than in non-polluted populations in all three seasons. The results indicated that the antioxidant enzymes, catalase, SOD and microsomal NADPH-DT diaphorase in digestive gland of S. cucullata could be useful biomarkers of PAHs contamination. It also emphasized that seasonal variation of potential biomarkers like such enzymes should be incorporated into interpretation of biomonitoring studies by the use of appropriate controls and identical treatment in analysis of polluted and non-polluted samples.     

Effects of dimethoate on spiders from metal pollution gradient

In this study, an attempt has been made to assess whether a chronic exposure to metals in habitats under a strong industrial pressure might have equipped spiders with biochemical defensive mechanisms enabling them to survive an additional chemical stress. To check this, non-web-building wolf spiders Pardosa lugubris (Lycosidae) and funnel web Agelena labyrinthica (Agelenidae) were collected at five variously polluted meadows and, under laboratory conditions, intoxicated with either single or multiple dose of dimethoate (OP pesticide). Then the activities of detoxifying (carboxylesterase: CarE, glutathione S-transferase: GST), antioxidative (selene-dependent and selene-independent glutathione peroxidases: GPOX and GSTPx) enzymes as well as acetylcholinesterase as a biomarker of exposure to OP pesticides were measured. In web-building A. labyrinthica, even a single application of the pesticide caused the inhibition of CarE, GSTPx and GPOX in individuals from less polluted sites and AChE and GST in specimens pre-exposed to high metal concentrations. Multiple intoxication, irrespectively of the site, caused significant, in comparison to controls, decrease in CarE, AChE and GSTPx activities. Actively hunting P. lugubris seem more resistant to acute pesticide intoxication, since the spiders from each site had a constant level of GST and AChE. In individuals of this species from heavily polluted sites, the inhibition caused by multiple intoxication with dimethoate was stated only for glutathione peroxidases.     

Field validation of autometallographical black silver deposit (BSD) extent in three bivalve species from the Lagoon of Venice, Italy (Mytilus galloprovincialis, Tapes philippinarum, Scapharca inaequivalvis) for metal bioavailability assessment

The autometallographic approach was applied to three filter-feeder bivalves from differently polluted sites of the Lagoon of Venice. The area density (AD) values of black silver deposits (BSDs) were therefore seasonally quantified by image analysis in digestive cell lysosomes. Sediments were characterised in terms of grain size, organic content and metal concentrations (Cd, Cu, Ni, Pb, Zn). The physiological status of the bivalves was also evaluated by applying the "survival in air" test as a generic stress index. Chemical results showed that Zn contributed to the total metal load for more than 60% in sediments and for 85%-94% in mussel and clam digestive glands, respectively (data processed from published results obtained on the same samples). Regression analyses between extent of BSDs and both total metal and Zn concentrations in digestive gland followed a linear pattern in mussels and clams, but not in arks. Similarly, in mussels and clams only, BSD amount was significantly correlated with total metal concentrations in sediments according to a logarithmic model. Consequently, results obtained from autometallography can discriminate sites on the basis of their metal pollution levels, like chemical analyses on both sediments and digestive glands, evaluations being supported by the results of the physiological stress index. As a conclusion, it is suggested that autometallography in bivalves may provide overall estimates of metal fractions moving from sediments towards organisms through the ingestion of metal-enriched particles. Future work is needed to define the bioaccumulation model in arks and to clarify their possible use as metal biomonitors.     

Clams as pollution bioindicators in Kuwait''s marine environment: metal accumulation and depuration

This study has developed and verified clam transplantation for marine pollution monitoring in Kuwait and has assessed its reliability for monitoring pollution by heavy metals through laboratory and field transplantation experiments. In field experiments, live specimens of the clam Circenita callipyga were transplanted, either suspended in seawater or buried in sand, to 11 coastal sites in Kuwait; subsamples of transplanted clams were recovered at intervals over a 6-month period. In laboratory experiments, heavy metal accumulation for 36 days and depuration (body's release of pollutants) for 60 days in small and big clams were investigated. Clams subsampled in laboratory and field experiments were analyzed for Hg, Cu, V, Cd and Pb. Results showed the ability of clams to survive under the transplantation conditions. Accumulation of biofouling materials was a problem at only two transplantation sites and was overcome by periodic cleaning. Statistical analysis of laboratory experimental data showed significant accumulation of all test metals and significant depuration of Hg, Cu, V and Cd. Rates of metal accumulation and depuration differed in relation to clam size class. Field experiments indicated statistically significant increases in Hg and Cu concentrations in the transplanted clams at most stations, no change in Cd concentrations and an increase or, occasionally, a decrease in V and Pb concentrations.     

DNA damage and lipid peroxidation vs. protection responses in the gill of Dicentrarchus labrax L. from a contaminated coastal lagoon (Ria de Aveiro, Portugal)

The present research work aimed to investigate the damage vs. protection responses in gill of European sea bass (Dicentrarchus labrax) captured at a polluted coastal lagoon, Ria de Aveiro (Portugal), as a tool to evaluate the human impacts on environmental health. Damage was assessed as DNA strand breakage and lipid peroxidation (LPO) whereas protection was evaluated by measuring catalase (CAT), glutathione peroxidase (GPX), glutathione reductase (GR), glutathione S-transferase (GST), total glutathione (GSHt), thiols and metallothioneins (MT). Fish were caught at five locations: Gafanha (GAF), Rio Novo do Príncipe (RIO), Laranjo (LAR) and Vagos (VAG) presenting each a different recognized source of contamination, and Torreira (TOR), assumed as reference site. Among the surveyed sites, gill damage was observed only at GAF, as measured by DNA integrity loss and LPO increase. An overall induction in enzymatic antioxidant protection was perceptible in fish from GAF and VAG, expressed as higher GPX, GR and GST activities. In addition, LAR fish showed elevated GST and CAT activities. Thiols content was higher in all study sites irrespective to the pollution spectrum, whereas GSHt increase was only observed at GAF and VAG. The highest MT level was detected in fish from VAG and the lowest level from RIO suggesting a low contamination degree on this particular site. Globally, the results expressed site-specific response patterns, signalling two critical areas - GAF and VAG. Additionally, a joint analysis (damage vs. protection) allowed the following ordering of surveyed sites according to the contamination degree and risk to fish health; GAF>VAG>LAR>RIO>TOR. D. labrax gill responses demonstrated their efficacy as early warning signals of the contaminants presence. Moreover, the adopted approach, considering simultaneously protection responses and damaging effects, also revealed its usefulness on the pollution extent assessment.     

Natural organic matter (NOM) induces oxidative stress in freshwater amphipods Gammarus lacustris Sars and Gammarus tigrinus (Sexton)

Humic substances comprise the majority of natural organic matter (NOM) on Earth, including dissolved organic matter in freshwater systems. Recent studies show that these substances directly interact with aquatic organisms as chemical stressors. The aim of the present study was to investigate the mode of action of dissolved NOM on the freshwater amphipods Gammarus lacustris Sars and Gammarus tigrinus (Sexton), and in particular, to determine if NOM induces or promotes internal oxidative stress. NOM was isolated by reverse osmosis from a brown-water lake in Brandenburg State, Germany. Oxidative stress markers, such as lipid peroxidation, cell internal hydrogen peroxide concentration, as well as peroxidase, catalase and glutathione S-transferase activities, were quantified. Exposure of both amphipod species to NOM caused a significant increase in lipid peroxidation, hydrogen peroxide concentration, catalase, peroxidase and glutathione S-transferase activities. Both species showed a two-stage antioxidant response: the first stage allowed the organisms to effectively eliminate ROS and to protect cells from damage, whereas the second stage leads to H2O2 accumulation in combination with destruction of lipid structures in the cells and, finally, functional damage or even death of the organism.     

Hepatic elemental contents and antioxidant enzyme activities in Algerian mice (Mus spretus) inhabiting a mine area in central Portugal

In this study the effects of heavy metals (manganese, iron, copper, zinc) and selenium exposure on the hepatic activity of antioxidant enzymes, superoxide dismutase (SOD) and glutathione S-transferases (GST), were appraised on a seasonal basis in Algerian mice (Mus spretus) inhabiting an active copper mine area. A reference population of the same species was considered for comparative purposes. Different patterns of seasonal variation were found in both populations for the manganese, iron and selenium hepatic concentrations and SOD activity. When the two populations were compared, iron and selenium concentrations were enhanced in mice from the polluted area. In addition, SOD activity was significantly decreased in summer in exposed mice, but no other significant changes in SOD and GST activities between sites throughout the year were recorded. However, when seasonal data within each group of mice were pooled, significant differences were found between sites for the average concentrations of manganese, iron and selenium, which are higher in the polluted site. In addition, significant differences were obtained for the average values of SOD and of GST activities, due to simultaneously higher GST values and slightly lower SOD values in the polluted site. The population from the reference site was more homogeneous for all parameters measured than the population from the polluted area. These results, in particular the higher variability in data collected from mice exposed to heavy metals and selenium, combined with the negative associations between biochemical markers and heavy metals, may suggest, despite the good adaptability of the mice to their habitat, biochemical stress due to diminished environmental quality.     

How does the greater white-toothed shrew, Crocidura russula, responds to long-term heavy metal contamination? -- A case study

Heavy metals accumulation in parallel with the evaluation of physiological and biochemical effects resulting from continued metal exposure were considered here using for the first time the great white-toothed shrew Crocidura russula as an in vivo model. Shrews were originated from an abandoned lead/zinc mining area and from a reference area, both in Alentejo, southern Portugal. Hepatic contents of nickel, copper, zinc, cadmium, mercury and lead were quantified by Inductively Coupled Plasma Mass Spectrometer (ICP-MS). Haematological parameters (white blood cells, red blood cells, haemoglobin and haematocrit) were obtained in a Coulter Counter Analyser and biochemical markers of the redox balance (glutathione S-transferase, glutathione peroxidase, and glutathione reductase) activities were measured spectrophotometrically using a Duo-50 spectrophotometer. Compared with control animals, significantly higher concentration of hepatic cadmium (9.29 vs. 1.18 micorg/g dry weight) and nickel (1.56 vs. 0.343 microg/g dry weight) were detected in the shrews collected in the mining area. However, no significant changes were observed on haematological or enzymatic parameters in animals exposed to metal pollution. The obtained results show that shrews are good bioaccumulators of toxic heavy metals, but very tolerant to their effects, revealing an interesting long-term adaptation to polluted environments. In addition, this study provides reference values for haematological parameters and antioxidant enzymes levels in C. russula, which may be relevant for comparative purposes in further studies.     

Validation of two tropical marine bivalves as bioindicators of mining contamination in the New Caledonia lagoon: Field transplantation experiments

The bioaccumulation and retention capacities of some key local contaminants of the New Caledonia lagoon (Ag, As, Cd, Co, Cr, Cu, Mn, Ni and Zn) have been determined in the oyster Isognomon isognomon and the edible clam Gafrarium tumidum during transplantation experiments. In a first set of experiments, oysters and clams from a clean site were transplanted into contaminated sites. Uptake kinetics determined in the field indicated that for Cr and Cu in oysters and Co, Ni, and Zn in clams, concentrations in transplanted bivalves reached those of resident organisms after 100d, whereas for the other elements, it would require a longer time for transplanted bivalves to reach the same levels as in the resident populations (e.g., up to 3 years for Cd). However, the slow uptake rate for metals observed in the latter transplantation is rather related to low bioavailability of metals at the contaminated sites than to low bioaccumulation efficiency of the organisms. Indeed, results of a second transplantation experiment into two highly contaminated stations indicated a faster bioaccumulation of metals in both bivalves. Results of both transplantations point out that the clam G. tumidum is a more effective bioindicator of mining contamination than I. isognomon, since it is able to bioaccumulate the contaminants to a greater extent. However the very efficient metal retention capacity noted for most elements indicates that organisms originating from contaminated sites would not be suitable for monitoring areas of lower contamination. Hence, geographical origin of animals to be transplanted in a monitoring perspective should be carefully selected.     

Toxicity and physiological effects of brominated flame retardant PBDE-47 on two life stages of grass shrimp, Palaemonetes pugio

This study examined the effects of a polybrominated diphenyl ether (PBDE) compound, PBDE-47, on adult and larval stages of the estuarine grass shrimp (Palaemonetes pugio). The 96-h LC50 test resulted in an estimate of 23.60 microg/L (95% confidence interval=14.51-38.37 microg/L) for larval shrimp. Adult shrimp had a higher 96-h LC50 of 78.07 microg/L (95% CI=65.1-93.63 microg/L). Four physiological biomarkers glutathione (GSH), lipid peroxidation (LPx), cholesterol (CHL) and acetylcholinesterase (AChE) were then assessed to study the sublethal effects of PBDE-47 exposure. GSH, LPx and AChE levels in both adults and larvae were not affected by PBDE-47 at concentrations up to 50 microg/L for 96 h. CHL levels were elevated in adults and larvae at the lowest exposure concentrations tested, but significant differences were found only in adult exposures. Effects associated with PBDE-47 aqueous exposures were observed at levels well above those reported in the environment.     

Response of antioxidant enzymes in freshwater fish populations (Leuciscus alburnoides complex) to inorganic pollutants exposure.

Evidence is accumulating indicating the importance of antioxidant enzyme activity measurements in eco-toxicological studies, as they may constitute markers for exposure to a large variety of pollutants. Variation of antioxidant enzymes, superoxide dismutase (SOD) and glutathione S-transferases (GST) and the effect of heavy metals and selenium exposure on these enzymes were investigated in the livers of Iberian endemic minnows (Leuciscus alburnoides complex) captured in a copper (Cu) mining area. Higher hepatic levels of copper and selenium were always observed in fish captured at the polluted site relative to the reference area population, reflecting the environmental monitoring results. A seasonal fluctuation in zinc and selenium levels for both populations was also observed which could be related to gonad maturation. The activity of SOD did not show significant regional alterations, but a seasonal variation occurred presumably associated with the Leuciscus life cycle. The GST activity was higher in the fish population from the polluted area (except in spring) and GST variability was associated with selenium and copper levels when both regions were compared. The increased GST activity was probably a metabolic adaptation to the continuous exposure to higher levels of those elements.     

Response of antioxidant enzymes in freshwater fish populations (Leuciscus alburnoides complex) to inorganic pollutants exposure

Evidence is accumulating indicating the importance of antioxidant enzyme activity measurements in eco-toxicological studies, as they may constitute markers for exposure to a large variety of pollutants. Variation of antioxidant enzymes, superoxide dismutase (SOD) and glutathione S-transferases (GST) and the effect of heavy metals and selenium exposure on these enzymes were investigated in the livers of Iberian endemic minnows (Leuciscus alburnoides complex) captured in a copper (Cu) mining area. Higher hepatic levels of copper and selenium were always observed in fish captured at the polluted site relative to the reference area population, reflecting the environmental monitoring results. A seasonal fluctuation in zinc and selenium levels for both populations was also observed which could be related to gonad maturation. The activity of SOD did not show significant regional alterations, but a seasonal variation occurred presumably associated with the Leuciscus life cycle. The GST activity was higher in the fish population from the polluted area (except in spring) and GST variability was associated with selenium and copper levels when both regions were compared. The increased GST activity was probably a metabolic adaptation to the continuous exposure to higher levels of those elements.     

Effect of lead concentration on the level of glutathione, glutathione S-transferase, reductase and peroxidase in human blood

Incubation of human whole blood for 24 h at 37 degrees C in the presence of 100-400 microg/dl lead chloride or lead acetate caused a concentration-dependent decrease in the level of reduced glutathione up to 40%. Similarly, the activities of glutathione reductase, glutathione peroxidase and glutathione S-transferase were decreased up to 25%, 50%, and 19%, respectively. Moreover, 100 microg/dl lead chloride or lead acetate slowed the process of glutathione regeneration, and delayed the time for complete regeneration from 20 to 40 min. When glutathione S-transferase was purified by affinity chromatography on Sepharose-linked glutathione, incubated with lead chloride or lead acetate, a concentration-dependent inhibition of the enzymatic activity was observed reaching 50% inhibition at a lead salt concentration of 6000 microg/dl.     

Assessment of Doñana National Park contamination in Procambarus clarkii: Integration of conventional biomarkers and proteomic approaches

Aquatic ecosystems of Doñana National Park (DNP) were monitored using Procambarus clarkii as bioindicator in four campaigns carried out between 2003 and 2004 to assess environmental quality possibly threatened by agrochemicals used in nearby areas. An integrated approach was carried out, by combining the responses of well-established biomarkers and the massive analysis of biological effects at the proteomic level. In sites potentially polluted, lower catalase, glucose-6-P dehydrogenase, and esterase activities, and higher malondialdehyde, metallothionein and glutathione levels were found. Two-dimensional gel electrophoresis resolved > 2500 gill spots, and image analysis detected that 35 showed significant intensity differences between the reference site and the other seven sites studied. The superiority of proteomic approaches was clearly recognized in our study since four different protein expression patterns were established based in the fold-number of up-/down-regulation of the 35 differentially expressed proteins. Sites located within Doñana Biological Reserve were essentially free of contaminants and those near the DNP limits were only slightly polluted. The higher proteomic responses found at the upper “Rocina” and “Partido” courses indicate that non-persistent agrochemicals are mainly used in Doñana surroundings. The highest responses corresponded to rice growing areas placed between the Guadiamar stream and the Guadalquivir River, according to the extended and intensive use of agrochemicals in such areas.     

Capacity of mercury volatilization by mer (from Escherichia coli) and glutathione S-transferase (from Schistosoma mansoni) genes cloned in Escherichia coli

A study was carried out to evaluate the capacity for mercury volatilization by genetically engineered strains that express the mer and glutathione S-transferase genes from Escherichia coli and Schistosoma mansoni, respectively. This method enabled strains containing simultaneously mer and glutathione S-transferase genes to grow in high concentrations of mercuric chloride (30 microg/ml) and to volatilize part of the mercury (248 microg/g cell dry wt.) present in the culture medium, while strains bearing only a single gene, did not have the same behavior. Up to 70% of the total mercury of bacterial volatilization occurred in the first 4 h. Although the findings were preliminary, the genetically engineered strain containing simultaneously the mer and glutathione S-transferase genes show a great potential for bioremediation. It may be used in a closed system to remove by volatilization, and recover mercury (Hg0) from contaminated effluents, such as industrial effluent, for instance.     

Acute effects of Benzo[a]pyrene, anthracene and a fuel oil on biomarkers of the common goby Pomatoschistus microps (Teleostei, Gobiidae)

The objective of this study was to investigate the effects of two different PAHs and a complex petrochemical mixture on the common goby, Pomatoschistus microps, using selected biomarkers as effect criteria. Benzo[a]pyrene (BaP) and anthracene were used as reference substances, while the water accommodated fraction of #4 fuel-oil (#4 WAF) was used as an example of a petrochemical mixture. P. microps was used since it is both a suitable bioindicator and a good test organism. Groups of fish were exposed to different concentrations of each of the test substances for 96 h and the activities of several enzymes commonly used as biomarkers were determined at the end of the bioassays. All the substances inhibited P. microps acetylcholinesterase (AChE) indicating that they have at least one mechanism of neurotoxicity in common: the disruption of cholinergic transmission by inhibition of AChE. An induction of lactate dehydrogenase (LDH) activity was found in fish exposed to BaP or to anthracene, suggesting an increase of the anaerobic pathway of energy production. On the contrary, inhibition of LDH was found in fish exposed to #4 WAF, suggesting a distinct effect of the mixture. An induction of P. microps glutathione S-transferase (GST) activity was found in fish exposed to BaP or to #4 WAF, while an inhibition was observed after exposure to anthracene. These results suggest that GST is involved in the detoxification of BaP and #4 WAF, but not of anthracene. All the substances increased catalase activity and isolated PAHs also increased superoxide dismutase, glutathione reductase and glutathione peroxidase activities, while #4 WAF did not cause significant alterations on these enzymes. These results suggest that all the substances may induce oxidative stress on P. microps, with BaP and anthracene apparently having more oxidative stress potential than #4 WAF.     

Relationship between two oxidative stress biomarkers, malondialdehyde and 8-oxo-7,8-dihydro-2'-deoxyguanosine, in the freshwater bivalve Unio tumidus

This study addresses the question of the relation between cellular and genomic oxidative damages in freshwater bivalves in realistic conditions of exposure in the field. Membrane and genomic oxidative damages were studied by means of lipid peroxidation and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo), respectively. Malondialdehyde (MDA) was used as an indicator of lipid peroxidation. The biomarkers were measured in the digestive gland and the gills of mussels (Unio tumidus) after 7 and 21 days of exposure in four ponds of the Moselle Basin, during three field studies conducted in 1999 and 2000. Effects measured at three sites (1R, 3C and 4M) were moderate and lipid peroxidation was slightly enhanced in mussels transferred in these ponds. In contrast, an important degradation was observed at site 2V reflected by a sharp increase in MDA and 8-oxodGuo concentrations in 2000. The biomarker responses agreed with the results of physicochemical analyses that indicated a worsening of water quality at the same site. Globally, a relationship was found between MDA increase and 8-oxodGuo formation, in the digestive gland after 7 days of exposure, and later in the gills (21 days). Responses of the digestive glands and gills to the oxidative parameters appeared correlated only after 21 days of exposure. The biomarkers selected confirmed their sensitivity for appraising the water quality of hydrosystems.     

The influence of substrate, pH, diet and temperature upon cadmium accumulation in the asiatic clam (Corbicula fluminea) in laboratory artificial streams

The influence of substrate, pH, diet and temperature upon the accumulation of cadmium (0.05 mg l⁻¹ dose < 0.001 mg l⁻¹ control) in the visceral mass of the Asiatic clam Corbicula fluminea was studied in laboratory artificial stream systems at intervals of 0, 3, 5, 7, 10 and 14-day exposures. Four substrate conditions, sand; sand, silt and clay (SSC); sand, clay and organic matter (SCO); and no substrate (NoS), were considered. The greatest tissue accumulation of cadmium in C. fluminea occurred at 0.05 mg l⁻¹ Cd in NoS and the lowest in clams occupying SCO. Complexation of available metals, lower clam filtering rates and physical protection by the substrate were attributed to the depressed cadmium accumulation of clams exposed in the SCO substrate. Lower pH exposures (5.0 vs 7.8) significantly (P 0.05 level) reduced cadmium uptake at 21°C but had little effect at 9°C. Uptake was higher in clams fed with cadmium-exposed Chlamydomonas reinhardt at 21°C but not at 9°C. In temperature exposures alone at 0.05 mg l⁻¹ Cd, accumulation was significantly higher in C. fluminea exposed at 21°C than at 9°C. The results are discussed relative to the importance of standardized laboratory protocol and the use of C. fluminea as a bioindicator of heavy metal stress.     

Biomarkers of oxidative stress in Wallago attu (Bl. and Sch.) during and after a fish-kill episode at Panipat, India

The present investigation was carried out by sampling water, sediment and fish during a fish-kill episode at Panipat (Haryana, India), and again sampling at the same site was conducted after a gap of two months. During the second sampling no fish-kill was observed and the water was relatively less turbid and clear. Antioxidant profile and lipid peroxidation (LPO) in fish tissues were studied. Analysis was also carried out on the physico-chemical characteristics of water samples along with heavy metal and pesticide analysis in water and sediment samples during and after the episode. Dissolved oxygen level was substantially low during the fish-kill episode. Heavy metals (copper and chromium) and pesticides like BHC (Benzene hexachloride), DDT (1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane) were also detected during the episode in water and sediment samples. Various oxidative stress biomarkers in liver, kidney and gill tissues in the Indian freshwater fish Wallago attu (Bl. and Sch.) collected from the site were investigated. The levels of reduced glutathione and non-protein thiol were significantly (P<0.001) higher in the liver of Wallago attu collected from Panipat after the fish-kill episode. Ascorbic acid levels in all the tissues did not change significantly after the episode. The LPO in liver, kidney and gills was significantly low (P<0.01-0.001) in all tissues of fish collected after the fish-kill episode. The protein carbonyl levels were significantly low (P<0.05-0.01) in all the fish organs sampled after the fish-kill episode. The findings suggest that industrial effluent may result in the massive loss of a commercial commodity. The simultaneous measurement of the physicochemical parameters of the water samples showed a good correlation between the biomarkers responses and the environmental chemical stress conditions.