Comet assay in Aegla platensis (Decapoda: Anomura) using a non-lethal hemolymph field sampling for in situ monitoring of freshwater genotoxicity.

This study aimed to apply the comet assay on Aegla platensis crabs as a suitable non-destructive approach for in situ monitoring of freshwater genotoxicity. Animals were captured during four sampling periods in a stream under minor anthropogenic impacts in Southern Brazil. Crabs were captured with a hand net, then the hemolymph samples were collected, and the animals were released into the stream after a 20-min recovery time. Hemolymph samples were transported to the laboratory and used to perform the alkaline comet assay. Results showed an intermediate level in the DNA damage index (range 107.3-165.0 arbitrary unit). No significant differences were observed among the different sampling periods. Hemolymph was successfully used as a non-lethal source of biological samples, and the comet assay using A. platensis proved to be a feasible approach for genotoxicity studies.

Treated tannery effluent and its impact on the receiving stream water: physicochemical characterization and cytogenotoxic evaluation using the Allium cepa test.

Tanneries are considered some of the most polluting industries due to the heavy use of toxic compounds, most of which are released into water bodies, thus exerting adverse effects on aquatic biota. However, the effects on organisms of treated effluents when released into the natural environment are rarely evaluated. This study aims to assess the physicochemical parameters of a tannery effluent after treatment (TE) at a Common Effluent Treatment Plant as well as the water of the receiving stream and to evaluate cytogenotoxic effects in Allium cepa. Three sampling sites (A: TE discharge point; B: 100 m downstream from site A along the receiving stream; C: 100 m upstream from site A along the stream) were selected. Onion bulbs were exposed to TE (100%, 80%, 60% v/v), water samples from sites B and C, and tap water for 72 h. Chromosomal aberration and mitotic index were analyzed on the root cells of A. cepa. The TE was above the standard limits for ammoniacal nitrogen, COD, and total nitrogen. No cytogenotoxicity was observed in A. cepa exposed to samples from sites A and C. However, the stream water sampled downstream from the TE discharge site significantly reduced the mitotic index, indicating a cytotoxic effect. Therefore, this demonstrates the effects of interactions between the receiving water and the complex chemical mixtures in the TE. The findings thus showed that the toxicity assessment of treated effluents along with the receiving water body would provide valuable and more realistic information about the joint toxicity of chemical pollutants in aquatic environments.

Repeated dose of meloxicam induces genotoxicity and histopathological changes in cardiac tissue of mice.

Meloxicam is the non-steroidal anti-inflammatory drug most used in small animals; however, studies on genotoxicity, oxidative stress, and histopathologic alterations in cardiac tissue are limited, especially at therapeutical doses used in these animals. This study evaluated the toxic effects caused by the treatment involving repeated low at higher doses of meloxicam in mice, by genotoxicity, oxidative stress, and histopathological parameters. Mice (CF1, male) received, by gavage, meloxicam at the therapeutic dose indicated for small animals (0.1 mg/kg) and at higher doses (0.5 and 1 mg/kg) for 28 days. Later, they were euthanized for blood and organ analysis. Oxidative stress was analyzed by the plasma ferric reduction capacity (FRAP) and catalase, and genotoxicity, by the comet assay and the micronucleus test. Heart, liver, lung, and kidney tissues were analyzed by the histology, and stomach and duodenum were analyzed with a magnifying glass. The relative weight of organs did not present significant alterations. However, congestion of duodenum vessels was observed at the three tested doses and caused hyperemia of stomach mucosa at 1 mg/kg. In the heart histology there was a reduction in the number of cardiomyocytes, accompanied by an increase in cell diameter (possible cell hypertrophy) dose-dependent. The highest tested dose of meloxicam also increased the DNA damage index, without alterations in the micronucleus test. Meloxicam did not affect the catalase activity but increased the FRAP (1 mg/kg). Meloxicam at the dose prescribed for small animals could potentially cause cardiac histopathologic alterations and genotoxic effects.

Evaluation of hematological, biochemical parameters and thiol enzyme activity in chrome plating workers.

The most commonly used solution in chrome plating bath is chromic acid (hexavalent Cr), and a considerable amount of mists is released into the air and consequently produce hazards to workers. Thus, the aim of this study was to evaluate whether the biomarker of exposure to metals, specially Cr levels, presents associations with hematological and biochemical parameters and if they can alter the activity of enzymes that contain thiol groups such as pyruvate kinase, creatine kinase, adenylate kinase, and δ-aminolevulinate dehydratase. Fifty male chrome plating workers were used for exposed group and 50 male non-exposed workers for control group. For that, biological monitoring was performed through quantification of metals on total blood and urine by inductively coupled plasma mass spectrometry (ICP-MS) and enzyme activity was performed by spectrometry in erythrocytes. In addition, chromium levels in water was quantified and ecotoxicology assay was performed with Allium cepa test. The results demonstrated that blood and urinary chromium levels in exposed group were higher than the control group (p < 0.0001). Furthermore, decreased activity of enzymes was found in those that contain thiol groups from exposed group when compared with the control group (p < 0.001). The water analysis did not present a statistical difference between control and exposed groups (p > 0.05), demonstrating that water did not seem to be the source of contamination. In summary, our findings indicated some toxicology effects observed in the exposed group, such as thiol enzyme inhibition, mainly associated with occupational exposure in chrome plating and besides the presence of other metals, and Cr demonstrated to influence the activity of the enzymes analyzed in this research.

In situ monitoring of the Sinos River, southern Brazil: water quality parameters, biomarkers, and metal bioaccumulation in fish.

The Sinos River is an important water supply in Southern Brazil and receives industrial, agricultural, and domestic effluents which may affect aquatic biota. Water physicochemical and microbiological analyses, biomarker responses (scaled mass index (SMI), gill histopathology, and micronucleus and nuclear abnormality (MN and NA) frequencies), and metal bioaccumulation in muscle were assessed in the fish species Bryconamericus iheringii (Characidae) captured at three sampling sites (S1, S2, and S3) in four sampling periods. The mean values of five parameters (total phosphorus, thermotolerant coliforms, aluminum, iron, and lead) exceeded the limits established by the Brazilian legislation at the three sampling sites. Although physicochemical analysis indicated higher impacts at S3, in some samples, significantly higher MN frequencies and bioaccumulation of manganese in fish muscle were observed at S1, whereas low SMI and higher concentrations of aluminum and zinc in fish muscle were found at S2. Histopathological alterations in gills were observed in fish collected at the three sampling sites; however, no spatial differences were observed, indicating similar environmental conditions with respect to this biomarker. Moreover, temporal variation of biomarker responses and metal bioaccumulation were found at all sampling sites. Furthermore, the consumption of fish from the Sinos River should be avoided given the concentrations of chromium (all samples), cadmium, and lead in fish muscle above the threshold for safe human consumption.

Monitoring the Genotoxic and Cytotoxic Potential and the Presence of Pesticides and Hydrocarbons in Water of the Sinos River Basin, Southern Brazil.

The Sinos River is one of the most polluted rivers in Brazil. The purpose of this work was to monitor the presence of some pesticides and hydrocarbons as well as the genotoxic and cytotoxic potential on HEp-2 cells from water samples collected at seven sites in the Sinos River Basin (SRB), southern Brazil. Nine samples were taken from the three main rivers in the SRB and used as a solution to dilute the HEp-2 cell culture medium after microfiltration. Twenty-four pesticides and 19 hydrocarbons were measured. Cytotoxicity was assessed by methyl thiazolyl tetrazolium (MTT) and neutral red (NR) assays, in which cells were exposed to different concentrations of the water samples for 24 h. Genotoxicity of the microfiltrated raw water samples was assessed by comet assay after 6 and 24 h of exposure. Among the chemicals analyzed, only the 2,4-D, dichloromethane, tetrachloroethene, chloroform, bromodichloromethane, styrene, and toluene were detected, but they were all lower than the limit established by Brazilian regulations. Twenty samples from a total of 60 had a cytotoxic effect in the MTT assay and 30 in the NR assay. The comet assay indicated the presence of genotoxic substances in the water at the seven locations monitored. Temporal and spatial variation was observed in the cytotoxicity and genotoxicity assays. Results indicated that the water in all stretches of the SRB is contaminated and it can cause harmful effects to humans and to the aquatic biota. This HEp-2 cell-line approach can be an additional tool for environmental monitoring.

DNA Damage in Wistar Rats Exposed to Organophosphate Pesticide Fenthion.

In the last several decades, exposure to pesticides has become a concern to environmental and human health. Many pesticides are environmentally persistent and are characterized by varying degrees of toxicity and adverse effects, including DNA damage. The present study was undertaken to evaluate the genotoxic potential of organophosphate pesticide fenthion in Wistar rats, as assessed by the comet assay. Adult male Wistar rats were treated with a solution of fenthion at a concentration of 40 mg/kg/day, administered intraperitoneally for 18 consecutive days. Rats were killed 24 hours after the last pesticide administration, and the comet assay was performed in peripheral blood cells. The comet assay results revealed that the damage index (19.29 ± 3.59 vs. 7.80 ± 2.25) and the damage frequency (17.00 ± 3.46 vs. 7.5 ± 2.46) found in fenthion-treated rats were significantly higher than those found in the control group (p = 0.001 and p = 0.0006, respectively). The results show that fenthion affects the DNA integrity of rat cells and may induce DNA damage in exposed organisms.

Environmental and biological monitoring of occupational formaldehyde exposure resulting from the use of products for hair straightening.

The evaluation of formaldehyde (FD) exposure in beauty salons, due to the use of hair straightening products, and its relation with genotoxicity biomarkers was performed in this study. Regardless of official recommendations, the inappropriate use of homemade hair creams has became a popular practice in Brazil, and high formaldehyde content in the "progressive straightening" creams can contain mutagens that could increase the incidence of neoplasia in those people who use them. Damage to DNA was assessed by conducting a micronuclei test (MNT) on buccal cells and the comet assay on heparinized venous blood samples. A total of 50 volunteers were recruited at six different beauty salons (labeled A to F). At two salons that used products that did not contain FD (salons D and E), environmental FD concentrations were 0.04 and 0.02 ppm. In contrast, the products used at salons A, B, C, and F contained 5.7, 2.61, 5.9, and 5.79% of FD, and these salons had environmental FD concentrations of 0.07, 0.14, 0.16, and 0.14 ppm, respectively. Comparison of the beauty salon workers from each of the six beauty salons revealed significant differences in urinary formic acid (FA) concentration before exposure (p = 0.016), urinary FA after exposure (p = 0.004), variation in FA concentration before and after exposure (p = 0.018), environmental FD concentration (p < 0.001), cytogenetic damage detected by the comet assay according to both damage index (p < 0.001) and frequency of damage (p < 0.001), and for karyorrhexis only according to the MNT (p = 0.001).

Evaluation of genotoxicity in workers exposed to low levels of formaldehyde in a furniture manufacturing facility.

Formaldehyde (FA) is a chemical widely used in the furniture industry and has been classified as a potential human carcinogen. The purpose of this study was to evaluate the occupational exposure of workers to FA at a furniture manufacturing facility and the relationship between environmental concentrations of FA, formic acid concentration in urine, and DNA damage. The sample consisted of 46 workers exposed to FA and a control group of 45 individuals with no history of occupational exposure. Environmental concentrations of FA were determined by high-performance liquid chromatography. Urinary formic acid concentrations were determined by gas chromatography with flame ionization detector. DNA damage was evaluated by the micronucleus (MN) test performed in exfoliated buccal cells and comet assay with venous blood. The 8-h time-weighted average of FA environmental concentration ranged from 0.03 ppm to 0.09 ppm at the plant, and the control group was exposed to a mean concentration of 0.012 ppm. Workers exposed to higher environmental FA concentrations had urinary formic acid concentrations significantly different from those of controls (31.85 mg L(-1) vs. 19.35 mg L(-), p ≤ 0.01 Mann-Whitney test). Significant differences were found between control and exposed groups for the following parameters: damage frequency and damage index in the comet assay, frequency of binucleated cells in the MN test, and formic acid concentration in urine. The frequency of micronuclei, nuclear buds, and karyorrhexis did not differ between groups. There was a positive correlation between environmental concentrations of FA and damage frequency (Spearman's rank correlation coefficient [r s] = 0.24), damage index (r s = 0.21), binucleated cells (r s = 0.34), and urinary formic acid concentration (r s = 0.63). The results indicate that, although workers in the furniture manufacturing facility were exposed to low environmental levels of FA, this agent contributes to the observed increase in cytogenetic damage. In addition, urinary formic acid concentrations correlated strongly with occupational exposure to FA.

Assessment of DNA damage in floriculturists in southern Brazil.

The aim of this study was to assess possible genotoxic effects on floriculturists in a region of the state of Rio Grande do Sul, in the south of Brazil, using the micronucleus test (MN) and comet assay. Thirty-seven floriculturists and 37 individuals not exposed to pesticides participated in the study. The micronucleus test was performed with epithelial cells of the oral mucosa. In the microscopic analysis, 2000 cells were evaluated per subject, verifying the frequency of MN and the frequency of other nuclear abnormalities (nuclear buds, binucleated cells, and karyorrhexis). For the comet assay in the peripheral blood lymphocytes, 100 cells were classified in five classes, according to the migration of DNA fragments, thereby generating the frequency of damaged cells and the damage index. There was no difference between the exposed and control groups in the frequencies of MN and other nuclear abnormalities in the epithelial cells of the oral mucosa. However, the comet assay showed that both the frequency of DNA damaged cells and the damage index were significantly greater in the exposed group. The results therefore indicate that floriculturists are exposed to mixtures of pesticides with genotoxic potential.

Toxicological evaluation of landfill leachate using plant (Allium cepa) and fish (Leporinus obtusidens) bioassays.

The disposal of municipal waste in landfills may pose an environmental problem because the product of the decomposition of these residues generates large volumes of leachate, which may present high toxicity. The aim of this study was to assess the toxic and genotoxic effects of a sample of untreated leachate in fish (Leporinus obtusidens) and onions (Allium cepa). The leachate was collected in a landfill located in the region of Vale do Rio dos Sinos, southern Brazil. The fish were exposed to raw leachate, at concentrations of 0.5%, 1.0%, 5%, 10% and 20% for 6 days, while the bulbs of A. cepa were exposed to concentrations of 5%, 10%, 25%, 50% and 100% for 48 h. For fish, the concentrations of 5%, 10% and 20% were lethal, thus indicating high toxicity; however, sublethal concentrations (0.5% and 1.0%) showed no genotoxicity by micronucleus test when compared with the control group. In the bioassays involving onions, high toxicity was observed, with significant reduction of root growth and mitotic index in bulbs exposed to the 100% concentration of the leachate. An increase in the frequency of chromosome abnormalities in the A. cepa root cells in anaphase-telophase was observed in accordance with the increase in the concentration of leachate (5%, 10%, 25% and 50%), with values significantly greater than the control, at the highest concentration. The results showed that the leachate contains toxic and genotoxic substances, thus representing a major source of environmental pollution if not handled properly.

In vivo genotoxicity of the pyrethroid pesticide beta-cyfluthrin using the comet assay in the fish Bryconamericus iheringii.

Environmental pollution by pesticide residues is a major environmental concern due to the extensive use of these substances in agriculture. The insecticide beta-cyfluthrin is a synthetic pyrethroid widely used in agricultural and other domestic activities. The aim of the present study was to assess the genotoxic effects of a sublethal exposure of the fish Bryconamericus iheringii (Characidae) to a commercial formulation of beta-cyfluthrin using the comet assay. Fish were exposed to sublethal concentrations (4.2 and 5.6 microg/L) of beta-cyfluthrin under static conditions during 24- and 48-h exposure periods. Fish in tap water were used as negative controls. Results obtained by the comet assay revealed genotoxic effects of the pyrethroid in the higher concentration and at the longer exposure period. The mean DNA damage index of fish exposed to 5.6 microg/L beta-cyfluthrin for 48 h was significantly higher (145.9 +/- 51.8) than in the control group (69.3 +/- 39.5). These findings indicate that native fish species might be at risk for genotoxic damage in waters contaminated with beta-cyfluthrin.

DNA damage in peripheral blood of patients with chronic obstructive pulmonary disease (COPD).

Chronic obstructive pulmonary disease (COPD) is a condition characterized by chronic airway inflammation and remodeling, lung parenchymal inflammation, and destruction resulting in expiratory airflow obstruction, hyperinflation of the lung with loss of elastic recoil, and impairment of gas exchange. Skeletal muscles in individuals with COPD generate free radicals at rest, and production increases during contractile activity. Overproduction of free radicals may result in oxidant-antioxidant imbalance in favor of oxidants. This study evaluated the levels of genetic damage in peripheral blood of patients with COPD using the cytokinesis-blocked micronucleus (CBMN) and the comet assays. The study was conducted with 25 patients with COPD and 25 controls matched for age and sex. Results of both comet and CBMN assays showed an increase in the level of DNA damage. In the group of patients with COPD, the mean frequency of binucleate cells with micronuclei was 6.72+/-3.02, and in the control group, 4.20+/-2.08 (p=0.00233). Mean comet value was 26.84+/-19.61 in patients with COPD and 7.25+/-7.57 in the control group (p=0.00004). The increased frequency of micronuclei in patients with COPD was primarily assigned to clastogenic events and DNA amplification because the frequency of nucleoplasmic bridges and buds was also increased. Oxidative stress in lung cells is a constant source of free radicals that damage genetic material of both lung and circulating cells.