Behavioral and biochemical consequences after chronic exposition to the herbicide atrazine in tadpoles.

Atrazine (ATZ) is the third most sold herbicide in Brazil, occupying the seventh position between most widely used pesticides. Due to its easy outflow, low reactivity and solubility, moderate adsorption to organic matter and clay, and long soil persistence, residual herbicide can be identified after long periods following application, and its usage has been prohibited in diverse countries. Amphibians are important bioindicators to assess impact of pesticide like atrazine, due to having a partial aquatic life cycle. This study had as objective to assess the response of bullfrog (Lithobates catesbeianus) tadpoles when exposed to this herbicide. Animals were exposed for a total of 168h to following concentrations: negative control, 40 µg/L, 200 µg/L, 2000 µg/L, 20000 µg/L of ATZ. Analysis of swimming activity was performed, and biochemical profile was assessed by analysis of blood and plasma glucose levels, urea, creatinine, cholesterol, HDL, triglycerides, glutamic pyruvic transaminase (GPT), alkaline phosphatase (AP), calcium, total proteins, phenol, peroxidase and polyphenol oxidase activity. Results exhibited malnutrition, anemia, likely muscle mass loss, and hepatic damage, indicating that ATZ can lead to an increase in energy to maintain homeostasis for animal survival.

Single and mixed exposure to distinct groups of pesticides suggests endocrine disrupting properties of imidacloprid in zebrafish embryos.

Due to their selective toxicity to insects, nicotinoid compounds have been widely used to control pests in crops and livestock around the world. However, despite the advantages presented, much has been discussed about their harmful effects on exposed organisms, either directly or indirectly, with regards to endocrine disruption. This study aimed to evaluate the lethal and sublethal effects of imidacloprid (IMD) and abamectin (ABA) formulations, separately and combined, on zebrafish (Danio rerio) embryos at different developmental stages. For this, Fish Embryo Toxicity (FET) tests were carried out, exposing two hours post-fertilization (hpf) zebrafish to 96 hours of treatments with five different concentrations of abamectin (0.5-11.7 mg L-1), imidacloprid (0.0001-1.0 mg L-1), and imidacloprid/abamectin mixtures (LC50/2 - LC50/1000). The results showed that IMD and ABA caused toxic effects in zebrafish embryos. Significant effects were observed regarding egg coagulation, pericardial edema, and lack of larvae hatching. However, unlike ABA, the IMD dose-response curve for mortality had a bell curve display, where medium doses caused more mortality than higher and lower doses. These data demonstrate the toxic influence of sublethal IMD and ABA concentrations on zebrafish, suggesting that these compounds should be listed for river and reservoir water-quality monitoring.

Toxic effects in Aphanomyces brasiliensis and zebrafish embryos caused by oomyceticides.

In fish farming, high losses occur during egg incubation and larviculture due to diseases caused by oomycetes. This study aimed to identify the oomycete species that occurs in zebrafish Danio rerio eggs and to evaluate the oomyceticidal effect of copper sulfate, bronopol and methylene blue on the mycelial growth of this organism, as well as to determine the lethal and sublethal toxicity of these compounds in embryos of D. rerio. The isolates were cultivated in yeast-starch medium to determine the concentration necessary to inhibit mycelial growth by 50% (IC50) and 100% (minimum oomyceticidal concentration) after a 96 h exposure to these compounds. In addition, tests with D. rerio eggs were conducted to determine the lethal concentrations for 50% of the organisms (96h-LC50), and the concentrations that inhibited 17% of the eggs hatching (96h-IC17) after 96 h. The organism responsible for the mortality of D. rerio eggs was classified by classical and molecular methods as Aphanomyces brasiliensis, representing the first report of this pathogen in zebrafish eggs. IC50 values could be determined for both bronopol and copper sulfate, whereas methylene blue had low effectiveness against the oomycete. Copper showed high toxicity to D. rerio eggs at low concentrations, while methylene blue and bronopol toxicity was low and similar to each other. The use of bronopol at a concentration of 4.8 mg l-1 for the treatment of zebrafish eggs allows controlling the pathology without causing deleterious effects to the treated organisms.

Acute toxic and genotoxic effects of formalin in Danio rerio (zebrafish).

Formalin is a readily soluble chemical used as a sanitizing agent in the home and hospital. Formaldehyde solutions are routinely used in aquaculture for the prophylaxis and treatment of parasites and fungi, but the adverse effects of their application need to be further investigated. Danio rerio or zebrafish has characteristics favorable to its handling and breeding, and it is highly sensitive to various chemicals, being an ideal experimental model for this type of investigation. Thus, the objective of this study was to verify the toxic and genotoxic effects of formalin and to determine the lethal concentrations of this chemical to support its safe use in disinfection processes. Acute and chronic tests were performed using methods in accordance with international protocols. The genotoxic effect of formalin was evaluated with the micronucleus test using blood samples, which were collected at 96 and 192 h of exposure. The LC50-96h of formalin in D. rerio was 45.73 mg L-1, demonstrating its high resistance compared to other species. Regarding the genotoxic effect, the sublethal concentrations of formalin showed a positive correlation with micronuclei according to the increase in its concentration independent of the time of exposure. The incidence of micronuclei increased with concentration, and the addition of 1 mg L-1 formalin corresponded to an increase of 2.9% in the average number of micronuclei. In other words, formalin at even sublethal concentrations caused genotoxic effects in peripheral blood erythrocytes of D. rerio. Therefore, we recommend further studies and other tests involving this chemical for its use at environmentally safe concentrations.

Evaluation of the potential teratogenic and toxic effect of the herbicide 2,4-D (DMA® 806) in bullfrog embryos and tadpoles (Lithobates catesbeianus).

The objective of this work was to evaluate the potential teratogenic and toxic effect of the herbicide 2,4-D (DMA® 806) on bullfrog embryos and tadpoles (Lithobates catesbeianus). We used the FETAX (Frog Embryo Teratogenesis Assay Xenopus) assay for embryos, and for tadpoles, we used acute (96 h) and chronic (49 days) toxicity tests and evaluated aspects of healthiness, hematology, and histopathology. The LC50-144h (Median Lethal Concentration), EC50-144h (Median Effective Concentration), MCIG (Minimum Concentration to Inhibit Growth) and TI (Teratogenic Index) for embryos were 792 mg/L, 593 mg/L, 150 mg/L of 2,4-D (DMA) and 1.34, respectively. For tadpoles, the LC50-96h was 700 mg/L of 2,4-D (DMA) and chronic test indicated an inflammatory process and erythrocytosis (with possible polycythemia), with consequent reduction of the spleen. This demonstrates physiological stress probably due to dehydration, which can be proven by the gill tufts widening intercellular space and gill tuft fusions. We also found injuries to the kidneys and skin of the animals even in the lowest concentration tested. Our results indicated that this pesticide is minimally teratogenic and has a low toxicity on L. catesbeianus embryos and tadpoles, but it can inhibit embryo growth in concentrations lower than those tested in this study. We hypothesized that the herbicide 2,4-D (DMA® 806) may be a respiratory allergen for L. catesbeianus tadpoles and recommend precautionary measures for prolonged exposure of aquatic organisms to this pesticide.