Assessment of genotoxic and mutagenic effects of chlorpyrifos in freshwater fish Channa punctatus (Bloch) using micronucleus assay and alkaline single-cell gel electrophoresis

Chlorpyrifos (CPF) is the single largest selling agrochemical that has been widely detected in surface waters in India. The studies on long-term genotoxic effects of CPF in different tissues of fish using genotoxic biomarkers are limited. Therefore, in the present study DNA damage by CPF in freshwater fish Channapunctatus using micronucleus (MN) and comet assays was investigated. The LC₅₀ – 96 h of CPF was estimated for the fish in a semi-static system. On this basis of LC₅₀ value sublethal and nonlethal concentrations were determined. The DNA damage was measured in lymphocytes and gill cells as the percentage of DNA in comet tails and micronuclei were scored in erythrocytes of fishes exposed to above concentrations of CPF. In general, significant effects for both the concentrations and time of exposure were observed in treated fish. It was found that MN induction in the blood was highest on day 14 at 203.0 μg/l of CPF. The highest DNA damage was observed on day 5, followed by a gradual non-linear decline in the lymphocytes and gill cells. The study indicated MN and comet assays to be sensitive and rapid methods to detect mutagenicity and genotoxicity of CPF and other pollutants in fishes.     

Influence of cypermethrin toxicity on ionic regulation and gill Na(+)/K(+)-ATPase activity of a freshwater teleost fish Cyprinus carpio

The effects of acute and sublethal toxicity of cypermethrin, a synthetic pyrethroid insecticide on plasma electrolytes (Na(+), K(+) and Cl(-)) levels and gill Na(+)/K(+)-ATPase activity in a common carp Cyprinus carpio were examined. The 24h LC(50) value of cypermethrin for C. carpio was 1.86ppm. During acute exposure, cypermethrin caused adverse effects in the form of hyponatreima, hypokalemia and hypochloremia and inhibition of gill Na(+)/K(+)-ATPase activity. In sublethal exposure to cypermethrin (0.186ppm for 35 days), plasma Na(+) was decreased throughout the exposure period except 7th day whereas plasma K(+) level was increased up to 28th day, then declined. However, plasma Cl(-) level was decreased. Likewise, gill Na(+)/K(+)-ATPase activity was decreased as the exposure period extended. The present study indicates that cypermethrin was highly toxic to freshwater fish and ion levels in blood as measured by specific ion concentrations (Na(+), K(+) and Cl(-)) and changes in gill Na(+)/K(+)-ATPase activity may represent a sensitive and useful nonspecific biomarkers of chemical exposure and effects.     

Effects of water quality and fish size on toxicity of methiocarb, a carbamate pesticide, to rainbow trout

The acute toxicity of methiocarb in juvenile rainbow trout (Oncorhynchus mykiss, 3.25±0.79g) was evaluated in glass aquaria under static conditions. Nominal concentrations of methiocarb in the toxicity test ranged from 1.25 to 7.50mgL(-1). The concentrations of methiocarb that killed 50% of the rainbow trout within 24-h (24-h LC(50)), 48-h LC(50), 72-h LC(50), and 96-h LC(50) were 5.43±0.19, 5.04±0.18, 4.95±0.19, and 4.82±0.21mgL(-1) (95% confidence limits), respectively. Mortality of fish increased with increasing water temperature. Increasing alkalinity from 19mgL(-1) as CaCO(3) to 40, 60, or 90mgL(-1) as CaCO(3) significantly decreased mortality of fish. Total hardness ranging from 50mgL(-1) as CaCO(3) to 147mgL(-1) as CaCO(3) did not affect mortality of fish exposed to methiocarb. Fish exposed to methiocarb had histological alterations such as lamellar edema, separation of epidermis from lamellae, and lamellar fusion. Methiocarb exposed fish had necrosis between molecular and granular layer of cerebellum where Purkinje cells present. Results indicate that alkalinity, temperature, and fish size affect methiocarb toxicity of rainbow trout.     

Genotoxic and hematological effects of chlorpyrifos exposure on freshwater fish Labeo rohita

Chlorpyrifos is a commonly used organophosphate insecticide that causes toxicological effects in aquatic organisms especially in fish. This study determined the effects of chlorpyrifos on the genotoxic and hematological parameters of freshwater fish, Labeo rohita. The genotoxic effects of different sublethal concentrations of chlorpyrifos were investigated in the erythrocytes of Labeo rohita (commonly known as Rohu) using the Micronucleus test. Effects of chlorpyrifos on the hematological parameters of the fish were also observed. Fish specimens were exposed to three sublethal concentrations of chlorpyrifos viz., sublethal I (SL-I, 1/6th of LC₅₀?=?～73.8?μg/L), sublethal II (SL-II, 1/4th of LC₅₀?=?～110.7?μg/L) and sublethal III (SL-III, 1/2nd of LC₅₀?=?～221.4?μg/L) for 96?h. Blood samples were collected at every 24?h and were subjected to the Micronucleus assay. The observed micronucleus frequencies were concentration and time-dependent. The MN induction was significantly highest (p?相似文献   

Measurements of genotoxic potential of cadmium in different tissues of fresh water climbing perch Anabas testudineus (Bloch), using the comet assay

The present investigation was undertaken to study the induction of DNA damage by CdCl(2) in freshwater climbing perch Anabas testudineus (Bloch) using alkaline single cell gel electrophoresis (comet assay). The DNA damage was measured in the tissue of gill, kidney and liver as the percentage of DNA in comet tails and comet heads in the tissue of the fish specimens exposed to 0.1, 1.0, 2.0mgL(-1) concentrations of CdCl(2). It was found that at all the concentrations of CdCl(2), the liver tissue exhibited significantly (p<0.01) higher DNA damage, followed by kidney and gill tissue. The DNA damage was found to be concentration dependent, with the highest DNA damage at 2mgL(-1) concentration, followed by 1.0 and 0.1mgL(-1). At the concentration of 2mgL(-1) of CdCl(2), the tail and head DNA of liver tissue were 38.81% and 59.49%, in kidney tissue the values were 32.37% and 64.66% whereas in gill tissue the values were 31.30% and 66.40% respectively. This study conclude that comet assay can be used for in vivo laboratory experiment using fish as model for screening the genotoxic potential of cadmium.     

The acute toxicity of surfactants on fish cells, Daphnia magna and fish-a comparative study.

There is a need to replace acute toxicity tests on fish (LC(50)) with more cost effective assays. The main objective of this study was to explore whether gill epithelial cells, hepatocytes and Daphnia magna could be used to predict acute toxicity of surfactants on fish. The acute toxicity of 10 synthetic surfactants (anionic, cationic, nonionic and zwitterionic) and two resin acids were determined on hepatocytes and gill epithelial cells from rainbow trout (Oncorhynchus mykiss), on Daphnia magna and on fish. Cell viability was measured with the fluorescent viability probe calcein-AM, immobilization was determined for Daphnia and 24-hr LC(50) for rainbow trout. The EC(50) values for the cellular tests were clearly higher than the corresponding values for Daphnia and fish, indicating that the cellular tests with the endpoint used are less sensitive than whole organisms. A combination of the EC(50) values for Daphnia and freshly isolated gill epithelial cells in suspension showed, however, a good correlation with acute toxicity on fish (r(2)=0.91 and slope=1.09). The combination seems to be a promising in vitro alternative to acute toxicity tests on fish (LC(50)), but a more exhaustive comparison, including a broad spectrum of chemicals should be made before the predictive value of the combined in vitro test can be evaluated.     

Comparative study of toxic lead effect on gill and haemoglobin of tilapia fish

This study was designed to determine the 72-h LC50 of lead for tilapia fish (Oreochromis hornorum), as well as the effect of exposure to sublethal lead concentrations (15, 23, 31, 39 and 47% of the LC50) on gill tissue lysosomal membranes of the fish and thaemoglobin concentration in blood. The LC50 value was found to be 202 mg Pb2+ l-1. Exposure to sublethal lead concentrations for 72 h showed significant increases in the lability of gill lysosomal membranes, measured by the release of acid phosphatase. Changes in membrane lability and in haemoglobin concentration were dependent on the amount of lead used during the exposure. We considered that the membrane lability is an adequate parameter to assay for monitoring lead contamination in water, because it is more sensitive than the haemoglobin concentration in blood.     

Toxic effects of profenofos on tissue acetylcholinesterase and gill morphology in a euryhaline fish,<Emphasis Type="Italic"> Oreochromis mossambicus</Emphasis>

Acute and sub-acute studies of profenofos [ O-(4-bromo-2-chlorophenyl)- O-ethyl- S-propyl phosphorothioate] on fish, Oreochromis (Tilapia) mossambicus, were carried out to assess the toxicity in relation to behaviour, morphology, and interactions with the targeted enzyme acetylcholinesterase (AChE, EC 3.1.1.7). Profenofos can be rated as highly toxic to O. mossambicus, with a median lethal concentration (LC(50)) of 0.272+/-0.0177 mg/l. The inhibitory and recovery pattern of brain and gill AChE was studied in vivo after exposure to a single LC(50) and multiple exposures to sub-lethal concentrations (0.108 mg/l) for 28 days, respectively. The LC(50)-exposed fish exhibited 90% inhibition of AChE activity in brain and gill in 24 h, and completely recovered within 23 days. Electron microscopy studies revealed an abnormal gill morphology, with distinct breakages in gill arches and rakers, along with deep lesions and erosions in the epithelium. Prolonged exposure at 0.108 mg/l also had similar effects, such as gill damage and AChE inhibition. The in vitro AChE study indicated that profenofos is neurotoxic and that it alters the apparent K(m) values widely in a concentration-dependent manner, resulting in a competitive type of inhibition. Based on the K(i) values, the sensitivity of AChE in brain was greater than that in gill tissue, at 2.38 x 10(-5) and 4.62 x 10(-5) M, respectively. The bioaccumulation values in head, body and viscera were estimated at regular intervals by gas chromatography method. The results indicated that the accumulation of profenofos was the highest in viscera followed by head and body, with depuration rates of 6.14, 0.16 and 0.12 micro g/h, respectively.     

2,4-Diaminotoluene (2,4-DAT)-induced DNA damage, DNA repair and micronucleus formation in the human hepatoma cell line HepG2

2,4-Diaminotoluene (2,4-DAT) is a widely used industrial intermediate and human exposure is possible in the dye and plastics industries. We investigated the genotoxicity of the environmental pollutant, 2,4-DAT, in human HepG2 cells using the unscheduled DNA synthesis (UDS) test, the micronucleus (MN) assay and single-cell gel electrophoresis (SCGE). 2,4-DAT was first tested by the RNA synthesis inhibition test as a cytotoxicity assay: the IC(50) of 2,4-DAT was 5.2 mM after 20 h of exposure. The compound had a genotoxic effect at concentrations from 1.45 to 6.80 mM in both micronucleus and comet assays. In the micronucleus assay, the number of MN/1000 BNC was 3.5 times higher at a concentration of 6.80 mM 2,4-DAT than in the negative control. At the same concentration, DNA migration (SCGE) showed an Olive tail moment (OTM) of 3.56+/-0.45, as compared to 0.19+/-0.02 for the negative control. The UDS test detected genotoxic effects at lower concentrations than did the other assays (0.01-5 mM). The percentage of cells in repair increased in a concentration-dependent manner to a maximum of 57% at 1mM. At the highest concentration tested (5 mM), the NNG/cell score was 13.6+/-0.5 whereas it was -2.7+/-0.5 for the negative control. These data, based on various endpoints, show a midly genotoxic effect of 2,4-DAT in the HepG2 cells and confirm that this cell line is a suitable model to study the toxic effects of aromatic amines.     

Effects of long term sublethal Cd exposure in rainbow trout during soft water exposure: implications for biotic ligand modelling

The objectives of the study were to determine the physiological and toxicological effects of chronic cadmium exposure on juvenile rainbow trout in soft water. Particular attention focused on acclimation, on comparison to an earlier hard water study, and on whether a gill surface binding model, originally developed in dilute soft water, could be applied in this water quality to fish chronically exposed to Cd. Juvenile rainbow trout, on 3% of body weight daily ration, were exposed to 0 (control), 0.07, and 0.11 microg l(-1) Cd [as Cd(NO(3))(2).4H(2)O] in synthetic soft water (hardness=20 mg l(-1) as CaCO(3), alkalinity=15 mg l(-1) as CaCO(3), pH 7.2) for 30 days. Mortality was minimal for all treatments (up to 14% for 0.11 microg l(-1) Cd). No significant effects of chronic Cd exposure were seen in growth rate, swimming performance (stamina), routine O(2) consumption, or whole body/plasma ion levels. In contrast to the hard water study, no acclimation occurred in either exposure group in soft water, with no significant increases in 96-h LC(50) values. Cadmium accumulated in a time-dependent fashion to twice the control levels in the gills and only marginally in the liver by 30 days. No significant Cd accumulation occurred in the gall bladder or whole body. Cadmium uptake/turnover tests were run using radioactive 109Cd for acute (3 h) exposures. Saturation of the gills occurred for control fish but not for Cd-exposed fish when exposed to up to 36 microg l(-1) Cd for 3 h. Cd-exposed trout accumulated less 'new' Cd in their gills compared to controls and they internalized less 109Cd than control fish. This effect of lowered Cd uptake by the gills of acclimated trout was earlier seen for the fish acclimated to 10 microg l(-1) Cd in hard water. The affinity of the gill for Cd was greater in hard water (logK(Cd-gill)=7.6) than in soft water (logK(Cd-gill)=7.3) but the number of binding sites (B(max)=0.20 microg g(-1) gill) was similar in both media. In addition, there was a shift in affinity of the gill for Cd (i.e. lowered logK(Cd-gill)) and increased B(max) with chronic Cd exposure in both soft water and hard water. We conclude that the present gill modelling approach (i.e. acute gill surface binding model or Biotic Ligand Model) does work for soft and hard water exposures but there are complications when applying the model to fish chronically exposed to cadmium.     

Modes of metal toxicity and impaired branchial ionoregulation in rainbow trout exposed to mixtures of Pb and Cd in soft water

Models such as the Biotic Ligand Model (BLM) predict how natural organic matter (NOM) and competing ions (e.g., Ca(2+), H(+) and Na(+)) affect metal bioavailability and toxicity in aquatic organisms. However, such models focus upon individual metals, not metal mixtures. This study determined whether Pb and Cd interact at the gill of rainbow trout (Oncorhynchus mykiss) when trout were exposed to environmentally relevant concentrations of these metals (Cd<100nmolL(-1); Pb<500nmolL(-1)) in soft (<100mumolCa(2+)L(-1)), moderately acidic (pH 6.0) water. The 96-h LC50 for Pb was 482nmolL(-1), indicating that Pb was one-order of magnitude more toxic in soft, acidic water than in harder, circumneutral pH waters. The LC50 for Cd alone was also low, 6.7nmolL(-1). Surprisingly, fish acclimated to soft water had multiple populations of Pb-gill and Cd-gill binding sites. A low capacity, high affinity population of Pb-gill binding sites had a B(max) of 18.2nmolg(-1) wet weight (ww) and apparent logK(Pb-gill)=7.05, but a second low affinity population could not be saturated up to free Pb concentrations approaching 4000nmolL(-1). Two populations of Cd-gill binding sites were characterized: a high affinity, low capacity population with an apparent logK(Cd-gill)=7.33 and B(max)=1.73nmolg(-1) ww, and a low affinity, high capacity population with an apparent logK(Cd-gill)=5.86, and B(max)=13.7nmolg(-1) ww. At low concentrations, Cd plus Pb accumulation was less than additive because Cd out-competed Pb for gill binding sites, which were likely apical Ca(2+)-channels. While disturbances to Ca(2+) influx were caused by Cd alone, Pb alone had no effect. However, Pb exacerbated Cd-induced disturbances to Ca(2+) influx demonstrating that, although Pb- plus Cd-gill binding was less than additive due to competition, the effects (ionic disturbances) were more than additive (synergistic). Pb was also likely binding to intracellular targets, such as branchial carbonic anhydrase, which led to inhibited Na(+) influx. This ionic disturbance was exacerbated by Cd. We conclude that exposure to environmentally relevant concentrations of Pb plus Cd results in less than additive metal-gill binding in soft, moderately acidic waters. However, ionic disturbances caused by Cd plus Pb are greater than additive, and this may ultimately increase the toxicity of Cd-Pb mixtures to fishes. Our findings suggest that it may be necessary to re-evaluate water quality criteria and assumptions of the BLM for fish exposed to mixtures of Pb and Cd in the acidic, soft waters found in the Canadian Shield, Scandinavia and other sensitive regions.     

Induction of micronuclei and binuclei in blood, gill and liver cells of fishes subchronically exposed to cadmium chloride and copper sulphate.

Common carp (Cyprinus carpio), Prussian carp (Carassius gibelio) and Peppered cory (Corydoras paleatus) were evaluated as target species to perform genotoxicity tests for heavy metals. Fishes were exposed to different doses of cadmium (0.005-0.1 mg/L) and copper (0.01-0.25 mg/L) for 21 days. Hexavalent chromium at a single dose of 5 mg/L was used as a positive control. Frequencies of micronuclei and binuclei were evaluated comparatively in peripheral blood erythrocytes, gill epithelial cells and liver cells. As a result it was observed that, fish species and their tissues showed differential sensitivity to the heavy metal treatment. In general, frequencies of micronucleated and binucleated cells significantly increased following the exposure for 21 days to copper, cadmium and chromium. On the other hand, gill and liver cells showed higher frequencies of micronuclei and binuclei than erythrocytes. Our results indicated the formation of micronuclei and binuclei in fish cells caused by their exposure to cadmium, copper and chromium, thus verifying results obtained earlier on mammals, which indicated that these heavy metals have cytotoxic and genotoxic effects. The suitability of the micronucleus assay in native fish species for the screening of aquatic genotoxicants is highlighted and the importance of target tissue selection in the piscine micronucleus test is emphasized.     

Gill and kidney histopathology in the freshwater fish Cyprinus carpio after acute exposure to deltamethrin

The histopathological effects of deltamethrin on the gill and kidney tissues of the common carp, Cyprinus carpio were determined by light microscopy. The fish were exposed to 0.029mgl(-1) (50% of 96h LC(50)) and 0.041mgl(-1) (70% of 96h LC(50)) solutions of deltamethrin for short-term (96h). The most common gill changes at all doses of deltamethrin were desquamation and necrosis. Besides, aneurism in secondary lamellae, lifting of the lamellar epithelium, oedema, epithelial hyperplasia and fusion of the secondary lamellae were reported. Lesions in the kidney tissues of fish exposed to deltamethrin were characterized by degeneration in the epithelial cells of renal tubule, pycnotic nuclei in the hematopoietic tissue, dilation of glomerular capillaries, degeneration of glomerulus, intracytoplasmatic vacuoles in epithelial cells of renal tubules with hypertrophied cells and narrowing of the tubular lumen.     

Physiological and behavioural responses of Gammarus pulex (Crustacea: Amphipoda) exposed to cadmium

The aim of this study was to investigate the effects of cadmium on physiological and behavioural responses in Gammarus pulex. In a first experiment, cadmium LC50s for different times were evaluated in 264 h experiment under continuous mode of exposure (LC50(96 h)=82.1 microgL(-1), LC50(120 h)=37.1 microgL(-1), LC50(168 h)=21.6 microgL(-1), LC50(264 h)=10.5 microgL(-1)). In a second experiment, the physiological and behavioural responses of the amphipod exposed to cadmium (0, 7.5 and 15 microgL(-1)) were investigated under laboratory conditions. The mortality and the whole body cadmium concentration of organisms exposed to cadmium were significantly higher than in controls. Concerning physiological responses, cadmium exposure exerted a significant decrease on osmolality and haemolymph Ca(2+) concentration, but not on haemolymph Na(+) and Cl(-) concentrations, whereas the Na(+)/K(+)-ATPase activity was significantly increased. Behavioural responses, such as feeding rate, locomotor and ventilatory activities, were significantly reduced in Cd exposed organisms. Mechanism of cadmium action and consequent energetic reallocation in favour of maintenance functions (i.e., osmoregulation) are discussed. The results of this study indicate that osmolality and locomotor activity in G. pulex could be effective ecophysiological/behavioural markers to monitor freshwater ecosystem and to assess the health of organisms.     

Impact of fertilizer (urea) on oxygen consumption and feeding energetics in the fresh water fish Oreochromis mossambicus

The effects of urea on survival, food utilization and oxygen consumption of the fresh water fish Oreochromis mossambicus were studied. The percentage survival of O. mossambicus when exposed to different concentrations of urea at 24, 48, 72 and 96h exposures was noted and it was found that 22,000 and 38,000mgL(-1) urea concentration were sublethal and lethal, respectively. The median lethal concentration, which killed 50% of the fishes during 96h exposure, was 28,000mgL(-1). Rearing the fish in increasing sublethal concentrations of urea, it was found that the feeding rate decreased from 34.341±7.067mgglivefish(-1)d(-1) (control) to 13.921±2.315mgglivefish(-1)d(-1) at the highest concentration of urea (22,000mgL(-1)). Growth rate was drastically reduced. The consumption of oxygen in O. mossambicus diminished from 0.962±0.208 to 0.645±0.118mgglivefish(-1)h(-1) when reared in the highest sublethal concentration of urea.     

The effect of starving and feeding on copper toxicity and uptake in Cu acclimated and non-acclimated carp

Common carp (Cyprinus carpio) were fed two different food rations: 0.5% body weight (low ration, LR) and 5% body weight (high ration, HR) and were either acclimated to sublethal copper (1 microM) for 28 days in softened Antwerp city tap water or not acclimated. Fish were exposed for 10 days to high Cu levels using four different concentrations (3.5, 6, 10, and 15 microM) before and after the Cu acclimation. Fish tolerance against Cu exposure was evaluated, and gill, liver, and carcass Cu and sodium levels were measured in dead and surviving fish. HR fish were twice as sensitive as LR fish in both tests. The 96h median lethal concentration (LC50) values for the non-acclimated LR and HR fish were 8.46+/-2.79 and 4.34+/-0.82 microM, respectively. The fish became more resistant to low Cu concentrations after Cu acclimation and the LC50 values were slightly increased, reaching 9.20+/-1.56 microM and 5.01+/-1.93 in LR and HR fish accordingly. Cu concentrations in the gills, liver, and carcass were significantly elevated in response to the short-term Cu exposure, and were significantly higher in LR fish than in HR fish. High Cu levels caused a net loss of sodium resulting in a severe ion regulatory disturbance. The rate of sodium loss increased linearly with increasing exposure concentrations. Cu acclimation resulted in reduced sodium loss and increased the resistance and tolerance to Cu toxicity.     

In vivo genotoxicity of mercury chloride and lead acetate: Micronucleus test on acridine orange stained fish cells.

The genotoxic effects of mercury chloride and lead acetate were evaluated in vivo using the micronucleus (MN) assay on acridine-orange (AO) stained peripheral blood erythrocytes, gill and fin epithelial cells of Carassius auratus auratus. Fish were exposed to three different concentrations of mercury chloride (MC) (1 microg/, 5 microg/L and 10 microg/L) and lead acetate (LA) (10 microg/L, 50 microg/L and 100 microg/L) for 2, 4 and 6 days. A single dose of 5 mg/L cyclophosphamide was used as a positive control. In addition to micronuclei, nuclear buds (NBs) were assessed in the erythrocytes. The ratio of polychromatic and normochromatic erythrocytes (PCE/NCE) in peripheral blood was also evaluated to assess cytotoxicity. MN frequencies in all three tissues were elevated in fish exposed to both LA and MC. However, NBs showed different sensitivity to metal treatments. MN frequencies in both control and treated fish were highest in gill cells and generally lower in erythrocytes and fin cells. PCE/NCE rations decreased in relation to MC and LA treatments. The results of this study indicate that LA and MC have genotoxic and cytotoxic damage in fish and confirmed that AO staining is a suitable technique for in vivo MN test in fish.     

Gill EROD in monitoring of CYP1A inducers in fish: a study in rainbow trout (Oncorhynchus mykiss) caged in Stockholm and Uppsala waters

The gill filament 7-ethoxyresorufin O-deethylase (EROD) assay was evaluated as a monitoring tool for waterborne cytochrome P4501A (CYP1A) inducers using rainbow trout (Oncorhynchus mykiss) caged in urban area waters in Sweden. To compare the CYP1A induction response in different tissues, EROD activity was also analyzed in liver and kidney microsomes. Immunohistochemistry was used to localize CYP1A protein in gill and kidney. In two separate experiments fish were caged at sites with fairly high expected polyaromatic hydrocarbon (PAH) contamination. In the first experiment, gill EROD activities were analyzed in fish exposed for 1-21 days in a river running through Uppsala. The reference site was upstream of Uppsala. In the second, gill, liver and kidney EROD activities were analyzed in fish exposed for 1-5 days in fresh or brackish waters of Stockholm and in a reference lake 60km north of Stockholm. Fish exposed for 5 days followed by 2 days of recovery in tap water in the laboratory were also examined. The gill consistently showed a higher EROD induction compared with the liver and the kidney. After 1 day of caging, gill EROD activity was markedly induced (6-17-fold) at all sites examined. Induction in gill was pronounced (5-7-fold) also in fish caged at the reference sites. In the 21-day exposure study gill EROD activity remained highly induced throughout the experiment (26-fold at most) and the induced CYP1A protein was exclusively confined to the gill secondary lamellae. In the 5-day exposure experiment, EROD activity peaked after 1 day and then declined in both gill and liver, while CYP1A immunostaining in the gill remained intense over the 5-day period. In the kidney, CYP1A staining was weak or absent. We conclude that gill EROD activity is a more sensitive biomarker of exposure to waterborne CYP1A inducers than EROD activity in liver and kidney.     

Genotoxic effects of ochratoxin A in human-derived hepatoma (HepG2) cells.

Ochratoxin A (OTA) is a widespread mycotoxin that occurs in many commodities from grains to coffee beans all over the world. Evidence is accumulating that OTA may cause cancer in humans. The compound was tested in micronucleus (MN) and single-cell gel electrophoresis (SCGE) assays in human-derived hepatoma (HepG2) cells and caused pronounced dose-dependent effects at exposure concentrations of 5 microg/ml and greater. On the contrary, no induction of His(+) revertants was found in Salmonella microsome assays with strains TA98 and TA100 with HepG2-derived enzyme (S9) mix in liquid incubation assays under identical exposure concentrations. Taken together, our results indicate that OTA is clastogenic in the human-derived cells. These findings support the assumption that this mycotoxin may cause genotoxic effects in hepatic tissue of humans.     

Detection of micronucleus and abnormal nucleus in erythrocytes from the gill and kidney of Labeo bata cultivated in sewage-fed fish farms.

Determination of genotoxic effect in fish, micronucleus test as well as the study of the abnormal shape of nuclei is a suitable measure, in which the presence or absence of genotoxins can be detected in water. In the present study, micronuclei and abnormal nuclei frequencies were scored in the gill and kidney erythrocytes of the fish Labeo bata grown in the sewage-fed fish farms of East Calcutta wetlands. Three experimental sites were chosen, namely, Bantala, Chowbaga and Chingrihata (basically these sites have sewage-fed fishponds), which were compared with fishponds of no sewage influence as the control area. Highly significant differences (P<0.001) were noticed for micronucleus frequencies in the gill and kidney erythrocytes of experimental fishes, where kidney erythrocytes showed an increased value than gill erythrocytes without any statistical differences. The frequencies of nuclear abnormalities such as necrotic cells, apoptotic cells, notch nucleated cells and binucleated cells were also counted separately for gill and kidney erythrocytes, in which significantly (P<0.001, P<0.01, P<0.05) increased values were obtained in comparison to control populations. These genotoxicity results confirmed that the sewage-fed ponds contain genotoxic metals such as Cr, Zn, Cu, Pb, Mn, Fe through wastewater and sludge because of the direct use of sewage water without pretreatment which may lead to health risks among humans through chronic consumption of fish from these experimental fish ponds. Other vertebrates grown in sewage-fed ponds may also suffer a certain amount of genotoxic substances.