The sublethal effects of (2,4-Dichlorophenoxy) acetic acid (2,4-D) on narrow-clawed crayfish (Astacus leptodactylus Eschscholtz, 1823).

2,4-D is a widely used phenoxy herbicide, potentially toxic to humans and biota. The objective of the present study was to reveal short term sublethal effects of 2,4-D on narrow-clawed freshwater crayfish (Astacus leptodactylus Eschscholtz, 1823), based on histology, total haemocyte counts, selected haemolymph parameters, and oxidative stress parameters. In the laboratory conditions crayfish specimens were exposed to 9 mg L-1 of 2,4-D for one week. Experiments were conducted under semi-static conditions in 20 L-capacity aquaria where 10 freshwater crayfish were stocked per aquarium. Exposure (experimental) and control groups were used and the experiments were repeated two times. No mortality and behavioural changes were recorded during the experiments. Total haemocyte counts decreased significantly, while haemolymph glucose levels increased (P<0.05), when compared to the control group. Haemolymph levels of calcium, chloride, sodium, potassium, magnesium, total protein, and lactate did not change. Exposure resulted with increased levels of malondialdehyde (MDA) only in hepatopancreas. However, results of gill FOX assay showed a significant decrease in oxidative stress parameters (P<0.05). MDA levels of gill and abdominal muscle tissues and FOX levels of hepatopancreas and abdominal muscle tissues did not change when compared to the control group. Significant histopathological alterations were observed both in hepatopancreas (multifocal deformations in tubule lumen) and gill tissue (melanisation of gill lamella). Exposure of crayfish even to a sublethal concentration of 2,4-D alters histopathology and lipid peroxidation due to stress. Biomarkers studied here seem to be useful for the assessment of adverse/toxic effects of pesticides on non-target, indicator aquatic organisms.

Evaluation of genotoxic and antigenotoxic effects of boron by the somatic mutation and recombination test (SMART) on Drosophila.

OBJECTIVE: In this study, different concentrations of boron have been evaluated for genotoxic and antigenotoxic properties by using the somatic mutation and recombination test (SMART) on Drosophila melanogaster. STUDY DESIGN: The treatment concentrations were chosen to a pretest. Third-instar larvae trans-heterozygous for two genetic markers, multiple wing hair (mwh) and flare (flr3), were treated at different concentrations (0.1, 5, 10, 20, and 40 mg/mL) of boron. In addition to investigating antigenotoxic effects, the same boron concentrations were co-administered with 0.1 mM Ethyl Methane Sulfonate (EMS). Distilled water was used as a negative control; 0.1 mM of EMS was used as a positive control. For the chronic feeding study, small plastic vials were prepared with 1.5 g of dry Drosophila Instant Medium and 5 mL of the respective test solution. Hundreds of trans-heterozygous larvae were embedded into this medium. Feeding ended with pupation of the surviving larvae. After metamorphosis, all surviving flies were collected and stored in a 70% ethanol solution. Preparation and microscopic analyses of wing were made after the treatment. Then the observed mutations were classified according to size and type of mutation per wing. RESULTS: Results indicated that there is no significant genotoxic effect with all of the boron concentrations. In addition, the antigenotoxic activities of boron against EMS were tested. Results indicated that all boron concentrations (0.1, 5, 10, 20 and 40 mg/mL) were able to abolish the genotoxic effects induced by the EMS. CONCLUSION: It is suggested that the observed effects can be linked to the antioxidant properties of boron. Moreover, these in vivo results will contribute to the antigenotoxicity database of boron.

Detection of transfluthrin and metofluthrin genotoxicity in the ST cross of the Drosophila Wing Spot Test.

In this study, different concentrations of transfluthrin and metofluthrin have been assayed for genotoxicity by using the Wing Spot Test on Drosophila melanogaster. Standard cross was used in the experiment. Third-instar larvae that were trans-heterozygous for the two genetic markers mwh and flr(3) were treated at different concentrations (0.0103mgmL(-1), 0.103mgmL(-1) for transfluthrin and 6µgmL(-1), 60µgmL(-1) for metofluthrin) of the test compounds. Feeding ended with pupation of the surviving larvae and the genetic changes induced in somatic cells of the wing's imaginal discs lead to the formation of mutant clones on the wing blade. Results indicated that two experimental concentrations of transfluthrin and 60µgmL(-1) metofluthrin showed mutagenic and recombinogenic effects in both the marker-heterozygous (MH) flies and the balancer-heterozygous (BH) flies.

Sublethal toxicity of esbiothrin relationship with total antioxidant status and in vivo genotoxicity assessment in fish (Cyprinus carpio L., 1758) using the micronucleus test and comet assay.

Esbiothrin, synthetic pyrethroid with quick activity against insects, is widely used against household pests and in public health. Despite widespread use, data on ecotoxicity and genotoxic effects are extremely scarce. The aim of the present study is to evaluate the genotoxic potential of esbiothrin on a model fish species Cyprinus carpio L., 1758 (Pisces: Cyprinidae, koi) using the micronucleus test and comet assay in peripheral blood erythrocytes. Effects of two sublethal exposure concentrations on plasma total antioxidant status (TAS mmol/L), and Hct values were examined. On the basis of the 96 h LC50 data from U.S. EPA ecotox database (32 µg/L) two sublethal exposure concentrations (5 and 10 µg/L) were used together with ethyl methanesulfonate (EMS) (5 mg/L) as positive control. Five fish were used for each dose/duration group (24, 48, and 72 h) under controlled laboratory conditions. The fish showed behavioral changes at the higher dose. Plasma TAS (mmol/L) levels decreased in 24 h; an increase was observed slightly for 48 and obviously for 72 h in both exposure doses. Similarly, hematocrit (Hct) values differed between exposure duration but no significant differences in mean values were found between groups of the same exposure time. The general trend was a rise after 48 h, which decreased afterwards. Our results revealed significant increases in the frequencies of micronuclei and levels of DNA strand breaks and thus demonstrated the genotoxic potential of this pesticide on fish, a nontarget organism of the aquatic ecosystem. To our knowledge this is the first study to report observable genotoxic effects of esbiothrin on fish.

Evaluation of potential genotoxicity of five food dyes using the somatic mutation and recombination test.

In this study, different concentrations of five food dyes (amaranth, patent blue, carminic acid, indigotine and erythrosine) have been evaluated for genotoxicity in the Somatic Mutation and Recombination Test (SMART) of Drosophila melanogaster. Standard cross was used in the experiment. Larvae including two linked recessive wing hair mutations were chronically fed at different concentrations of the test compounds in standard Drosophila Instant Medium. Feeding ended with pupation of the surviving larvae. Wings of the emerging adult flies were scored for the presence of spots of mutant cells which can result from either somatic mutation or somatic recombination. For the evaluation of genotoxic effects, the frequencies of spots per wing in the treated series were compared to the control group, which was distilled water. The present study shows that carminic acid and indigotine demonstrated negative results while erythrosine demonstrated inconclusive results. In addition 25 mg mL(-1) concentration of patent blue and 12.5, 25 and 50 mg mL(-1) concentrations of amaranth demonstrated positive results in the SMART.

Genotoxicity assessment of carp (Cyprinus carpio L.) fingerlings by tissue DNA damage and micronucleus test, after environmental exposure to fenitrothion.

The purpose of this study was to evaluate the adverse effects of sublethal doses of fenitrothion, an organophosphothionate insecticide on brain, gill, liver, and muscle tissues as a ratio of 8-OHdG to dG to indicate the DNA damage and erythrocyte micronucleus frequency for genotoxicity of carp (Cyprinus carpio L.) fingerlings. In our study, the mean weights and lengths of the fish (n = 4-12) were 31.13 ± 14.24 g and 12.53 ± 1.41, respectively. Before the experiment, fish were maintained in aerated dechlorinated tap water at 21.8 ± 1 °C and fed daily with commercial feed at a rate of 2% of their body weights. Experiments were conducted under static conditions in the aquaria. Technical grade (95%) fenitrothion was diluted in acetone to give a dosing solution of 10 mg/L. The increased lesions/106 DNA bases (p < 0.05) of liver tissue of exposure group (0.49 ± 0.18) was observed when compared to control group (0.28 ± 0.30). There was not any significant differences between brain tissues, no damage were detectable in gill and muscle tissues of control groups, and in exposure groups altered levels of damage were detected for gill (0.06 ± 0.05) and muscle (0.16 ± 0.21) tissues. The increased micronucleus frequencies (%) in erythrocytes of carp following the exposure to 48 h fenitrothion (6.43 ± 3.89; p<0.05) was observed when compared to control group (1.29 ± 1.03). The available data indicate that there is still lack of well-established dose-response relationships between occupational or environmental exposures and the induction of 8-OHdG. Such biomarkers may be used in assessing adverse/toxic effects of pesticides as environmental stressors.

The acute toxicity of fenitrothion on narrow-clawed crayfish (Astacus leptodactylus Eschscholtz, 1823) in association with biomarkers of lipid peroxidation.

The aim of this research was to evaluate the acute toxicity of fenitrothion to the crayfish (Astacus leptodactylus Eschscholtz, 1823), which is chosen as an alternative aquatic organism to fish by using the static test system and evaluate the basic lipid peroxidation parameters for the first 24 h. Crayfish of 27.3 ± 0.56 g mean weight and 10.0 ± 0.72 cm mean length were selected for the bioassay experiments. The experiments were repeated three times in 20 liters of tap water. The temperature of water was 21 ± 1°C. The data obtained were statistically evaluated by using a computer program developed by the United States Environmental Protection Agency, based on Finney's probit analysis method and the 96-h LC(50) value for crayfish was calculated to be 15.75 µg/L. The 95% lower and upper confidence limits for the LC(50) were 9.45 to 25.01 µg/L. In addition to the acute toxicity bioassay experiments, 24-h oxidative stress parameters such as malondialdehyde (MDA) levels and ferrous oxidation assay (FOX HP [hydrogen peroxide] equivalents) were also determined. Only MDA levels of hepatopancreas decreased at 5, 10, and 20 µg/L of fenitrothion doses. We can conclude that fenitrothion is highly toxic to crayfish, a nontarget organism in the ecosystem, and the lipid peroxidation indicators can be easily used for monitoring environmental effects.

Genotoxic evaluation of antiepileptic drugs by Drosophila somatic mutation and recombination test.

The study examines the potential genotoxicity of three antiepileptic drugs (phenytoin sodium, pregabalin, gabapentin) using the wing somatic mutation and recombination test (SMART) in Drosophila melanogaster. Trans-heterozygous (two genetic markers mwh and flr) third-instar larvae of D. melanogaster were treated with different concentrations of the test compounds. A positive correlation was observed between total mutations and the number of wings with morphologically detectable mutations. The observed mutations were classified according to size and type of mutation per wing. Phenytoin clearly increased the frequency of total spots at all concentrations above 1.25 microg/ml. Gabapentin also increased the frequency of total spots at concentrations of 40 and 80 microg/ml. This study shows that phenytoin and gabapentin have genotoxic effects according to the SMART test; however, pregabalin displays lower genotoxicity in the SMAR assay when compared with the other two antiepileptics. The results also show that all AED concentrations lower the survival rate of the flies.

Genotoxicity of two mouthwash products in the Drosophila Wing-Spot Test.

In this study, genotoxicity of two mouthwash products (chlorexidin, benzidamine-HCl) were investigated in the Drosophila Wing-Spot Test which makes use of the wing cell markers multiple wing hairs (mwh) and flare (flr) and detects both mitotic recombination and various types of mutational events. Induced mutations are detected as single mosaic spots on the wing blade of surviving adults that show either the multiple wing hairs or flare phenotype. Induced recombination leads to mwh and flr twin spots and also, to some extent, to mwh single spots. Recording of the frequency and the size of different spots is allowed for a quantitative determination of the mutagenic and recombinogenic effects. Trans-heterozygous third-instar larvae were treated at different concentrations of the mouthwash products. Chlorexidin exposure concentrations were 0.5, 1 and 2mg/ml. Benzidamine-HCl exposure concentrations were 0.38, 0.75 and 1.5mg/ml. In addition, the observed mutations were classified according to size and type of mutation per wing. Both chlorexidin and benzidamine-HCl were genotoxic in terms of total mutations per wing at the highest doses. Survival rates of flies used in the experiments were significantly lower than those of the control group, with both mouthwash products showing toxic effects on Drosophila melanogaster larvae.

Genotoxic evaluation of sodium fluoride in the Somatic Mutation and Recombination Test (SMART).

In this study, genotoxic effect of sodium fluoride (NaF) was investigated in Drosophila melanogaster Somatic Mutation and Recombination Test. Third-instar larvae trans-heterozygous for two genetic markers mwh and flr, were treated at different concentrations (2.5 microg/ml, 5 microg/ml and 10 microg/ml) of the test compounds. After the treatment the observed mutations were classified according to size and type of mutation per wing. For the evaluation of genotoxic effects, the frequencies of spots per wing in the treated series were compared to the control group, which is distilled water. NaF has genotoxic and toxic effects for concentrations of 5 and 10 microg/ml. The present study shows that NaF may have genotoxic and toxic effects.

Sublethal cyfluthrin toxicity to carp (Cyprinus carpio L.) fingerlings: biochemical, hematological, histopathological alterations.

This study measures a whole series of biological endpoints, and histopathological and behavioral changes of carp (Cyprinus carpio L.) fingerlings in order to investigate the adverse effects of a sublethal concentration of 10 microg/L cyfluthrin for 48h and one week using the semi-static test system. The most significantly altered findings of this study was that total antioxidant status levels decreased in 48h, chloride levels decreased in one week, and sodium and phosphorous levels increased in the 48h-exposed group. Plasma cortisol increased in both the 48h-exposed and the one-week-exposed groups. The unique tissue was the brain that had an increased level of malondialdehyde after one week of exposure. Such biomarkers may be used in assessing adverse/toxic effects of pesticides as environmental stressors. Potential oxidative-stress-inducing effect of synthetic pyrethroid cyfluthrin, as lipid peroxidation, in addition to its sodium channel blocking, is reported for the first time as an additional mechanism of action.

Genotoxic assessment of oxcarbazepine and carbamazepine in drosophila wing spot test.

In this study, different concentrations of two antiepileptic drugs, carbamazepine (CBZ) and oxcarbazepine (OXC), have been evaluated for genotoxicity in the wing spot test of Drosophila melanogaster. The wing spot test detects different kinds of somatic mutations and allows detection of mitotic recombinations. Third-instar larvae trans-heterozygous for two genetic markers mwh and flr, were treated at different concentrations of the drugs. Oxcarbazepine exposure concentrations were 1.88, 3.75, 7.50 and 15microg/ml. Carbamazepine exposure concentrations were 5, 10, 20 and 40microg/ml. In addition, the observed mutations were classified according to size and type of mutation per wing. CBZ was genotoxic in terms of total mutations per wing in the highest two doses; the same was true for OXC in the highest three doses. Survival rates of flies used in the experiments were significantly lower than that of the control group showing both drugs to have toxic effects to Drosophila melanogaster larvae. Clone formation frequency for 10(5) cells was lower in OXC than CBZ. However this was lower than the critical genotoxicity frequency of 2.0.

Investigation of acute toxicity of fenitrothion on guppies Poecilia reticulata.

Fenitrothion, an organophosphothionate insecticide (CAS number: 122-14-5) and potential toxic pollutant contaminating aquatic ecosystems, was investigated in the present study for acute toxicity. Guppy fish (Poecilia reticulata) were selected for the bioassay experiments. The experiments were repeated three times and the 96 h LC(50) was determined for the guppies. The static test method of acute toxicity test was used. The water temperature was regulated at 23 +/- 1 degrees C. In addition, behavioral changes at each fenitrothion concentration were observed for the individual fish. Data obtained from the fenitrothion acute toxicity tests were evaluated using the probit analysis statistical method. The 96 h LC(50) value for guppy was estimated as 3.28 mg l(-1). Values in the range of microg l(-1) and mg l(-1) have been reported for various other fish species.

How Sublethal Fenitrothion is Toxic in Carp (Cyprinus carpio L.) Fingerlings.

ABSTRACT The purpose of this study was to evaluate the effects of fenitrothion, an organophosphothionate insecticide, on selected serum hematological and biochemical parameters of carp (Cyprinus carpio L.) fingerlings and investigate histopathological changes after acute dose of exposure. Among those, cortisol and glucose are evaluated as responses of early life stages of carp to acute stress and hematological blood parameters are evaluated to investigate the mechanism of toxicity via histopathological changes to the standard test organism. No significant histological findings were observed in carp tissues (gills, liver, kidneys, spleen, gonads, brain, muscle, and skin) exposed to a sublethal concentration of 10 mg/L fenitrothion and controls. Hematocrit and erythrocyte counts were decreased significantly by exposure to fenitrothion when compared to the control group (25.26% +/- 0.82%, 1455 +/- 90.3 [10(3)/muL] vs. 28.83% +/- 1.60%, 1865 +/- 74.5 [10(3)/muL], P < 0.05), respectively. Leucocyte and thrombocyte counts did not change. Increases in plasma cortisol levels were found in the experimental group compared to controls (34.2 +/- 1.9 vs. 18.0 +/- 2.2, P < 0.05) respectively, while glucose, chloride, sodium, potassium, phosphorous, and brain MDA and FOX levels did not show significant differences from those of control (P > 0.05). As a result, we can conclude that early life stages of fish are very susceptible to the adverse effects of toxicants.

Investigation of acute toxicity of chlorpyrifos-methyl on guppy Poecilia reticulata.

Static bioassays were made to determine acute toxicity of chlorpyrifos-methyl, a wide spectrum organophosphorus insecticide and potential toxic pollutant of aquatic ecosystem, Guppy fish (Poecilia reticulata). Bioassays were made at a regulated temperature of 22+/-1 degrees C and were repeated three times. Lethal doses of the insecticides were determined using LC50 software programme of U.S. EPA based on Finney's Probit Analysis statistical method. The 96 h LC50 value and 95% confidence limit of chlorpyrifos-methyl for Guppy was estimated as 1.79 (1.47-2.10) mg/l. The fish exposed to chlorpyrifos-methyl exhibited behavioral changes in the form of neurotoxin toxicity: less general activity than control group, loss of equilibrium, erratic swimming and staying motionless at a certain location generally at mid-water level for prolonged periods. The 1 mg/l (lowest) concentration had similar behavior (NOEC) with the control group.

Genotoxicity testing of some organophosphate insecticides in the Drosophila wing spot test.

In this study, different concentrations of some organophosphate insecticides (methyl parathion, azamethiphos, dichlorvos and diazinon) have been evaluated for genotoxicity in the wing somatic mutation and recombination test (SMART) of Drosophila melanogaster. Third-instar larvae trans-heterozygous for two genetic markers mwh and flr, were treated at different concentrations (1 ppm, 3 ppm, 5 ppm, 7 ppm, 10 ppm) of the test compounds. A positive correlation was observed between total mutations and the number of wings having mutations. In addition, the observed mutations were classified according to size and type of mutation per wing. Chemicals used were ranked in decreasing order according to their genotoxic effects as diazinon, dichlorvos, methyl parathion, azamethiphos.

Investigation of acute toxicity of (2,4-dichlorophenoxy)acetic acid (2,4-D) herbicide on larvae and adult Nile tilapia (Oreochromis niloticus L.).

A 48h LC(50) values of (2,4-dichlorophenoxy)acetic acid (2,4-D), a widely used agricultural herbicide, were determined on the larvae and adult Nile tilapia (Oreochromis niloticus L.). Each test was repeated three times. The data obtained were statistically evaluated by the use of the E.P.A computer program based on Finney's probit analysis method and a 48h LC(50) value for Nile tilapia (O. niloticus L.) larvae and adults were found to be 28.23mg/L and 86.90mg/L, respectively in a static bioassay test system. 95% lower and upper confidence limits for the LC(50) were 22.55-32.98 and 80.67-92.80mg/L, respectively. Water temperature was 24±1°C. Behavioral changes of both tilapia life forms were examined for various herbicide concentrations.

Genotoxicity testing of four food preservatives and their combinations in the Drosophila wing spot test.

In this study, four food preservatives (sodium nitrate, sodium nitrite, potassium nitrate and potassium nitrite) and there five combinations at a concentration of 25mM have been evaluated for genotoxicity in the somatic mutation and recombination test (SMART) of Drosophila melanogaster. Three-day-old larvae trans-heterozygous including two linked recessive wing hair mutations (multiple wing hairs and flare) were fed at different concentrations of the test compounds (25, 50, 75 and 100mM) in standard Drosophila Instant Medium. Wings of the emerging adult flies were scored for the presence of spots of mutant cells, which can result from either somatic mutation or mitotic recombination. Also lethal doses of food preservatives used were determined in the experiments. A positive correlation was observed between total mutations and the number of wings having mutation. In addition, the observed mutations in each wing were classified according to the size and type of the mutation. For the evaluation of genotoxic effects, the frequencies of spots per wing in the treated series were compared to the control group, which is distilled water. Chemicals used were ranked as sodium nitrite, potassium nitrite, sodium nitrate and potassium nitrate according to their genotoxic and toxic effects. Moreover, the genotoxic and toxic effects produced by the combined treatments were considerably increased, especially when the four chemicals were mixed. The present study shows that correct administration of food preservatives/additives may have a significant effect on human health.

Investigation of acute toxicity of fenitrothion on peppered corydoras (Corydoras paleatus) (Jenyns, 1842).

Fenitrothion, as an organophosphothionate insecticide, is a contact insecticide and selective acaricide, also used as a vector control agent for malaria in public health programs. A 96 h LC50 value of fenitrothion, a potential toxic pollutant contaminating aquatic ecosystems, was determined on the adult peppered corydoras (Corydoras paleatus). The experiments were repeated three times. The static test method of acute toxicity test was used. Water temperature was regulated at 23 +/- 1 degrees C. In addition, behavioral changes at each fenitrothion concentration were observed for the individual fish. Data obtained from acute toxicity tests were evaluated using the Probit Analysis Statistical Method. The 96 h LC50 value for peppered corydoras was estimated as 3.51 mg/l.

Investigation of acute toxicity and the effect of 2,4-D (2,4-dichlorophenoxyacetic acid) herbicide on the behavior of the common carp (Cyprinus carpio L., 1758; Pisces, Cyprinidae).

A 96-h LC(50) values of 2,4-D [(2,4-dichlorophenoxy)acetic acid], a common contaminating agricultural herbicide, were determined on the adult common carp (Cyprinus carpio L., 1758; Pisces, Cyprinidae). The study was conducted in two stages using 130 carp. The data obtained were statistically evaluated by the use of the EPA computer program based on Finney's Probit Analysis Method and a 96-h LC(50) value for C. carpio L., 1758 was found to be 63.24 mg/l in a static bioassay test system. 95% lower and upper confidence limits for the LC(50) were 55.03 and 71.92 mg/l, respectively. Water temperature was 17+/-1 degrees C. Behavioral changes of the above mentioned species were examined for various herbicide concentrations.