Comparative genomic damage among three shark species with different habits: Sublethal impacts of human origin in a protected island environment in the South Atlantic.

Elasmobranchs can bioaccumulate and biomagnify pollutants. However, few studies are directed to the effects of pollutants on the health of these animals, and in most cases, are limited to the analysis of biochemical markers. Thus, the incidence of genomic damage among shark species inhabiting a protected ocean island in the South Atlantic was investigated in association with the analysis of pollutants in seawater sample. High levels of genomic damage were identified, especially in Negaprion brevirostris and Galeocerdo cuvier, in addition to interspecific variations that may be related to characteristics such as animal size, metabolism and habits. High concentrations of Surfactants were observed in seawater sample, in addition to low concentrations of Cadmium, Lead, Copper, Chromium, Zinc, Manganese, and Mercury. The results evidenced the potential of shark species as a bioindicator of environmental quality and allowed assessing the anthropic impact on the archipelago, which currently drives its economy through tourism.

Contrasting tourism regimes due to the COVID-19 lockdown reveal varied genomic toxicity in a tropical beach in the Southern Atlantic.

Tourist occupancy in coastal environments threatens the stability of various coastal ecosystems and is thus a cause for concern for the environmental sector. As such, it is important to perform environmental monitoring in a way that analyses and quantifies the environmental impact of coastal ecosystems. Porto de Galinhas beach (Pernambuco - Brazil) has one of the highest visitation rates in Brazil and suffered from restrictions to human mobility due to the COVID-19 pandemic. These restrictions allowed for the evaluation of the impact of tourism on Porto de Galinhas beach and the effects that the lack of tourist occupancy had during the lockdown period of 2020. Blood samples from the species Abudefduf saxatilis were collected monthly over a period of 1 year and during the lockdown quarter, in order to perform micronucleus (MN) and nuclear morphological alteration (NMA) tests, and data were analyzed at a seasonal level (dry/rainy period) using a comet assay. For the control group, A. saxatilis samples were collected in an environmentally protected area on Tamandaré beach (68 km from Porto de Galinhas). The MN and NMA tests showed a greater frequency of genomic damage when there was greater tourist flow. In relation to rain seasonality, the comet assay showed a greater incidence of genomic damage during the dry period, where there was a higher rate of tourist migration, compared to the rainy period. The lockdown period presented a lower incidence of genotoxic damage compared to the period without restrictions on human mobility and the control. The results show that tourism has been causing a significant environmental impact on Porto de Galinhas beach. The data collected during the lockdown period demonstrated how the absence of human movement results in changes that are favorable to environmental recuperation, as illustrated by the lower frequency of genomic damage.

DNA damage as indicator of the environmental vulnerability of bats in Brazil's Caatinga drylands.

Brazil's Caatinga drylands is under extensive environmental deterioration, with 38% of its natural cover already lost. There is a need for a better understanding of the effects of such degradation on Caatinga's rich and singular biota. Bats form a large part of this biota, and are pointed as good bioindicators. Here, we used the micronucleus test -an easy-to-use, accessible and cost-effective in vivo approach- to detect DNA damage in cells from bats of different species and feeding habits in three protected areas in the Caatinga, comparing them with samples from an industrial sugarcane plantation. We hypothesized that environmental disturbance would reflect in DNA damage, with lower levels of damage in the less disturbed protected areas. The frequency of micronucleated cells differed significantly between sites and feeding habits (carnivores > insectivores > frugivores > nectarivores > hematophagous) but did not differ between sexes. Alarmingly, the highest levels of DNA damage were in two strictly protected areas (Seridó and Raso da Catarina Ecological Stations). Glossophaga and Anoura were the genera with more damaged cells, and had, respectively, 1.48 and 3.53 times more micronucleated cells (average of 19.33 and 22.67 cells, respectively) than individuals from the same genera from the area with least damaged cells (average of 7.80 and 5.00 cells, respectively). Our analysis is a warning call for an in-depth investigation on the effects of both genotoxic contamination and environmental stressors on bats and other species in Brazil's Caatinga, including the role that historical human-induced processes -like the intense use of agrochemicals- may have had in the region.

Genotoxic activity of L-asparaginase produced by Streptomyces ansochromogenes UFPEDA 3420.

L-asparaginase is an enzyme capable of hydrolyzing the substrate asparagine in aspartic acid and ammonia. Due to this mechanism of action observed, L-asparaginase is widely used in the treatment of Acute Lymphoblastic Leukemia, since these cells use asparagine for their survival. Because it is frequently used as an antineoplastic, it is necessary to evaluate its genotoxic effects. The aim of the present study was to evaluate cellular DNA damage after exposure to L-asparaginase produced by Streptomyces ansochromogenes UFPEDA 3420. NCIH-292, MCF-7 and MOLT-4 neoplastic cell lines and normal PBMC cells were used. L-Asparaginase used in this study was produced by actinobacteria S. ansochromogenes UFPEDA 3420, isolated and purified by chromatographic methods. L-Asparaginase induced micronucleus formation in PBMC cells and tumor lines when compared to the negative control. These data suggest that L-Asp appears to have a genotoxic effect very close to the positive control in normal cells (p < 0.05). The level of genomic damage measured by DNA breaks in alkaline SCGE assay was detected from the lowest concentration (12.5 µg/mL) to the highest concentration (50 µg/mL) for tumor cell lines and PBMC. In view of the above, new genotoxic studies will be carried out to better elucidate L-Asparaginase and its mutagenic potential, still unknown, enough for this drug to be safely used in conventional antineoplastic therapies.

Genomic damage in Mugil curema (Actinopterygii: Mugilidae) reveals the effects of intense urbanization on estuaries in northeastern Brazil.

The largest metropolitan centers in northeastern region of Brazil are all located near the coast, and industrial, tourist, and agro-industrial activities are the principal causes of water contamination due to discharges of untreated sewage. Adverse environmental conditions can often be detected by analyzing the genetic material of organisms exposed to pollutants, and furnish an overview of environmental quality. We evaluated possible damage to the DNA of one of the fish resources most widely consumed and commercialized by coastal communities in northeastern Brazil, Mugil curema ("tainha"). Erythrocytes from M. curema were analyzed by the presence of micronuclei and by comet assay (single cell gel electrophoresis, SCGE). Statistical comparisons to both tests revealed considerably greater genomic damage in polluted estuaries than in the control site (p < 0.05), suggesting strong genotoxic impacts on the specimens evaluated, principally among those taken near localities with dense demographic and industrial development.

Characterization of brain acetylcholinesterase of bentonic fish Hoplosternum littorale: Perspectives of application in pesticides and metal ions biomonitoring.

Acetylcholinesterase (AChE; EC 3.1.1.7) is a serine hydrolase, whose main function is to modulate neurotransmission at cholinergic synapses. It is, therefore, the primary target of some pesticides and heavy metals. Its inhibition in aquatic organisms has been used as an indicator of the presence of these pollutants in water bodies. The present study aimed to characterize physicochemical and kinetic parameters of brain AChE in the benthic fish Hoplosternum littorale and to analyze the in vitro effects of pesticides (dichlorvos, diazinon, chlorpyrifos, parathion-methyl, temephos, carbaryl, carbofuran, aldicarb, diflubenzuron, novaluron and pyriproxyfen) and metal ions (As3+, Cd2+, Cu2+, Fe2+, Mn2+, Mg2+, K+, Pb2+, Hg2+, Zn2+) investigating the potential of this enzyme as environmental biomarker based on current regulations. Specific substrates and inhibitors have indicated AChE to be the predominant cholinesterase (ChE) in the brain of H. littorale. Peak activity was observed at pH 8.0 and 30 °C. The enzymatic activity is otherwise moderately thermostable (≈ 50% activity at 45 °C). The enzyme can reduce the activation energy of acetylthiocholine hydrolysis reaction to 8.34 kcal mol-1 while reaching a rate enhancement of 106. Among the pesticides under study, dichlorvos presented an IC50 value below the maximum concentrations allowed by legislation. This study presents the first report on the inhibition of brain AChE activity from Siluriformes by the pesticides novaluron and pyriproxyfen. Mercury ion also exerted a strong inhibitory effect on its enzymatic activity. The H. littorale enzyme thus has the potential to function as an in vitro biomarker for the presence of the pesticide dichlorvos as well as mercury in areas of mining and industrial discharge.

Selective cytotoxic and genotoxic activities of 5-(2-bromo-5-methoxybenzylidene)-thiazolidine-2,4-dione against NCI-H292 human lung carcinoma cells.

BACKGROUND: Thiazolidine-2,4-dione ring system is used as a pharmacophore to build various heterocyclic compounds aimed to interact with biological targets. In the present study, benzylidene-2,4-thiazolidinedione derivatives (compounds 2-5) were synthesized and screened against cancer cell lines and the genotoxicity and cytotoxicity of the most active compound (5) was investigated on normal and lung cancer cell line. METHODS: For in vitro cytotoxic screening, the MTT assay was used for HL60 and K562 (leukemia), MCF-7 (breast adenocarcinoma), HT29 (colon adenocarcinoma), HEp-2 (cervix carcinoma) and NCI-H292 (lung carcinoma) tumor cell lines and Alamar-blue assay was used for non-tumor cells (PBMC, human peripheral blood mononuclear cells) were used. Cell morphology was visualized after Giemsa-May-Grunwald staining. DNA content, phosphatidylserine externalization and mitochondrial depolarization were measured by flow cytometry. Genotoxicity was assessed by Comet assay. RESULTS: 5-(2-Bromo-5-methoxybenzylidene)-thiazolidine-2,4-dione (5) presented the most potent cytotoxicity, especially against NCI-H292 lung cancer cell line, with IC50 value of 1.26µg/mL after 72h incubation. None of the compounds were cytotoxic to PBMC. After 48h incubation, externalization of phosphatidylserine, mitochondrial depolarization, internucleosomal DNA fragmentation and morphological alterations consistent with apoptosis were observed in NCI-H292 cells treated with compound (5). In addition, compound (5) also induced genotoxicity in NCI-H292 cells (2.8-fold increase in damage index compared to the negative control), but not in PBMC. CONCLUSION: Compound 5 presented selective cytotoxic and genotoxic activity against pulmonary carcinoma (NCI-H292 cells).

Acetylcholinesterase of mangrove oyster Crassostrea rhizophorae: A highly thermostable enzyme with promising features for estuarine biomonitoring.

Enzyme biomarkers from several aquatic organisms have been used for assessing the exposure to contaminants at sublethal levels. Amongst them, the cholinesterases are commonly extracted from several organisms to evaluate/measure organophosphate and carbamate neurotoxic effects. Acetylcholinesterase (AChE; EC 3.1.1.7) is an enzyme of the group of serine esterases that acts on the hydrolysis of the neurotransmitter acetylcholine allowing the intermittence of the nerve impulses responsible for the neuronal communication. This enzyme is the main target for the action of some pesticides and the inhibition of its activity in bivalve mollusks may be used as biomarker due to their filter-feeding habit. In this context, the present study aimed to characterize physicochemical and kinetic parameters of the AChE extracted from gills and viscera of the oyster Crassostrea rhizophorae and investigate the in vitro effect of pesticides (dichlorvos, diazinon, chlorpyrifos, methyl-parathion, temephos, carbaryl, carbofuran, aldicarb, diflubenzuron and novaluron) in search for assessing its potential as biomarker. Specific substrates and inhibitors evidenced the predominance of AChE in both tissues. The optimum pH found for gills and viscera AChE were 8.0 and 8.5, respectively. The maximum peak of activity occurred at 70 °C for gill AChE and 75 °C for viscera AChE. The enzymes of both tissues presented remarkable thermostability. The Michaelis-Menten constant for both enzymes were 1.32 ±â€¯0.20 mM for gills and 0.43 ±â€¯0.12 mM for viscera. The Vmax values for gills and viscera were 53.57 ±â€¯1.72 and 27.71 ±â€¯1.15 mU/mg, respectively. The enzymes were able to reduce the activation energy to 9.75 kcal mol-1 (gills) and 11.87 kcal mol-1 (viscera) obtaining rate enhancements of 3.57 × 105 and 1.01 × 104, respectively, in relation to non-catalyzed reactions. Among the pesticides under study, the carbamates carbaryl and carbofuran exerted the strongest inhibitory effects on the enzyme activity achieving important degrees of inhibition at concentrations below national and international current regulations. The first observation of the effects of benzoylurea pesticides (diflubenzuron and novaluron) on AChE from mollusks is reported here. The gills AChE of C. rhizophorae showed potential to be specific biomarker for the carbamate carbaryl while the viscera AChE showed it for carbofuran. According to their features, these enzymes may be proposed as promising tools for estuarine monitoring as well as biocomponent of biosensor devices.

Quantitative genotoxicity assays for analysis of medicinal plants: A systematic review.

ETHNOPHARMACOLOGICAL RELEVANCE: Medicinal plants are known to contain numerous biologically active compounds, and although they have proven pharmacological properties, they can cause harm, including DNA damage. AIM OF THE STUDY: Review the literature to evaluate the genotoxicity risk of medicinal plants, explore the genotoxicity assays most used and compare these to the current legal requirements. MATERIAL AND METHODS: A quantitative systematic review of the literature, using the keywords "medicinal plants", "genotoxicity" and "mutagenicity", was undertakenQ to identify the types of assays most used to assess genotoxicity, and to evaluate the genotoxicity potential of medicinal plant extracts. RESULTS: The database searches retrieved 2289 records, 458 of which met the inclusion criteria. Evaluation of the selected articles showed a total of 24 different assays used for an assessment of medicinal plant extract genotoxicity. More than a quarter of those studies (28.4%) reported positive results for genotoxicity. CONCLUSIONS: This review demonstrates that a range of genotoxicity assay methods are used to evaluate the genotoxicity potential of medicinal plant extracts. The most used methods are those recommended by regulatory agencies. However, based on the current findings, in order to conduct a thorough study concerning the possible genotoxic effects of a medicinal plant, we indicate that it is important always to include bacterial and mammalian tests, with at least one in vivo assay. Also, these tests should be capable of detecting outcomes that include mutation induction, clastogenic and aneugenic effects, and structural chromosome abnormalities. In addition, the considerable rate of positive results detected in this analysis further supports the relevance of assessing the genotoxicity potential of medicinal plants.

Differential genomic damage in different tumor lines induced by prodigiosin.

Prodigiosin is a secondary metabolite produced by Serratia marcercens. As this pigment is suggested to be a cancer drug, genotoxicity studies are necessary. The aim of the present investigation was to evaluate the genotoxic effects of prodigiosin on tumoral and normal cell lines, NCIH-292, MCF-7 and HL-60. A normal line BGMK was used as control. Genomic damage induced by prodigiosin was observed in all tumor lines as well as the control line. The pigment induced the formation of micronuclei in tumor cells. The present data confirm the antitumor potential of prodigiosin. However, these findings also raise concerns regarding its target-specific action, as genotoxic effects on normal cells also occurred.

Assessment of the association between micronuclei and the degree of uterine lesions and viral load in women with human papillomavirus.

Infection by human papillomavirus (HPV) is among the main etiologies of cervical cancer. The expression of oncogenic viral proteins enables the onset of the virus, which can trigger the carcinogenic process. One of the main characteristics of this process is the loss of genome stability, including chromosome stability. The micronucleus test is a cytogenetic method for the detection of genetic alterations that change chromosome behavior during cell division resulting in the formation of micronuclei. This method has been applied for the early detection of DNA damage in individuals with a greater likelihood of developing cancer. The aim of the present study was to assess the association between micronucleus expression and the degree of cytological lesions and viral load in patients with HPV. The micronucleus analysis revealed differences in the number micronuclei found in the groups, which ranged from 0.00067 to 0.00133 in the control group and 0.00267 to 0.02433 among patients with HPV. Statistically significant differences (p<0.05) were found in the number of micronucleated cervical cells between the patients and healthy women. Moreover, significant associations were found between micronucleus expression and both the degree of uterine lesions (r2=0.7237; r=0.8507; p=0.000002) and viral load (r2=0.7012; r=0.8374; p=0.000004). The findings demonstrate the efficacy of micronucleus analysis in monitoring risks to human health.