Ecotoxicology

Correction to: Toxicity prediction and assessment of an environmentally realistic pesticide mixture to Daphnia magna and Raphidocelis subcapitata Acknowledgements This work was supported by LEAF (UID/AGR/04129/2013) and CESAM (UID/AMB/50017/2013) from FCT/MEC through national funds, and the cofunding by FEDER (POCI-01-0145-FEDER-007638), within the PT2020 Partnership Agreement and Compete2020.

Acute toxic and genotoxic effects of formalin in Danio rerio (zebrafish) Abstract 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 toxicity of leaches from hydrothermal sulfide deposits by means of a delayed fluorescence-based bioassay with the marine cyanobacterium Cyanobium sp. NIES-981 Abstract The commercial use of metals such as copper, lead, and zinc has markedly increased in recent years, resulting in increased interest in deep-sea mining of seafloor hydrothermal sulfide deposits. However, the full extent of the impact of deep-sea mining at hydrothermal field deposits on the environment remains unclear. In addition to impacting the deep sea, the leaching of heavy metals from extracted sulfide mineral may also affect the upper ocean zones as the sulfide rock is retrieved from the seafloor. Here, we used a delayed fluorescence-based bioassay using the marine cyanobacterium Cyanobium sp. NIES-981 to evaluate the toxicity of three sulfide core samples obtained from three drill holes at the Izena Hole, middle Okinawa Trough, East China Sea. Leaches from two of the cores contained high concentrations of zinc and lead, and they markedly inhibited delayed fluorescence in Cyanobium sp. NIES-981 compared with control. By examining the toxicity of artificial mixed-metal solutions with metal compositions similar to those of the leaches, we confirmed that this inhibition was a result of high zinc and lead concentrations into the leaches. In addition, we conclude that this delayed fluorescence-based bioassay is a viable method for use by deep-sea mining operations because it is quicker and requires less laboratory space and equipment than the standard assay.

Hydrolytic enzymes mediated lipid-DNA catabolism and altered gene expression of antioxidants under combined application of lead and simulated acid rain in Fenugreek ( Trigonella foenum graecum L.) seedlings Abstract Understanding ill effects of simultaneous existence of various abiotic stresses, commonly observed due to various anthropogenic activities and global climate change these days, over plants growth, metabolic activity and yield responses are important for continued agricultural productivity and food security. In the present study, seedlings of Fenugreek (Trigonella foenum graecum L.) were subjected to lead (Pb, 1200 ppm) and/or simulated acid rain (SAR, pH 3.5) for 30 days, and were then analysed. The results revealed reduced growth, and total lipid and DNA contents, while enhanced Pb accumulation, biological concentration factor, biological accumulation coefficient, translocation factor, lipase activity, and levels of free fatty acid, conjugated diene, lipid hydroperoxide, DNA oxidation and DNase activity under Pb and/or SAR exposure. Additionally, activities and gene expression levels of antioxidants (superoxide dismutase, catalase, guaiacol peroxidase and ascorbate peroxidase) were enhanced in response to applied treatments. The results also suggested that inhibitions/ accelerations determined under joint addition of Pb and SAR were comparatively more profound than those measured under their single application. Additionally, root was more sensitive to Pb treatment, compared to both leaf and shoot. Hence, under simultaneous presence of two or more number of abiotic stresses, the strategy opted by plants for survival is chiefly governed by the interaction between prevailing stressors, which is then conceived by plants as a new state of stress.

Modulation of growth, ascorbate-glutathione cycle and thiol metabolism in rice ( Oryza sativa L. cv. MTU-1010) seedlings by arsenic and silicon Abstract Arsenic is a carcinogenic metalloid, exists in two important oxidation states—arsenate (As–V) and arsenite (As-III). The influence of arsenate with or without silicate on the growth and thiol metabolism in rice (Oryza sativa L. cv. MTU-1010) seedlings were investigated. Arsenate was more toxic for root growth than shoot growth where the root lengths were short, characteristically fragile and root tips turned brown. The multiple comparison analysis using Tukey’s HSD (honest significant difference) tests indicated that the rate of arsenate accumulation and its conversion to arsenite by arsenate reductase were significantly increased in all arsenate treated seedlings while in seedlings treated jointly with arsenate and silicate, arsenate accumulation and its conversion to arsenite decreased. Silicate content was detected in the seedlings treated with silicate alone and under co-application of arsenate with silicate. In the test seedlings arsenic toxicity increased ascorbate and glutathione contents along with the activities of their regulatory enzymes, viz., ascorbate peroxidase, glutathione reductase, glutathione peroxidase and glutathione–s-transferase to reduce the toxicity level induced by arsenic whereas ascorbate oxidase activity was decreased to maintain sufficient ascorbate pool under arsenate treatment. Phytochelatins production were increased in both root and shoot of the test seedlings under arsenate exposure to alter the detrimental effects of arsenic by chelation with arsenite and their subsequent sequestration into vacuole. Thus, joint application of silicate along with arsenate showed significant alterations on all the parameters tested compared to arsenate treatment alone due to less availability of arsenic in the tissue leading to better growth and metabolism in rice seedlings. Thus use of silicon in arsenic contaminated medium may help to grow rice with improved vigour.

Production of a biosurfactant from Bacillus methylotrophicus UCP1616 for use in the bioremediation of oil-contaminated environments Abstract The aim of the present study was to produce a microbial biosurfactant for use in the bioremediation of environments contaminated with petroleum products. Bacillus methylotrophicus was isolated from seawater taken from a port area and cultivated using industrial waste as substrate (corn steep liquor and sugarcane molasses [both at 3%]). Surface tension measurements and motor oil emulsification capacity were used for the evaluation of the production of the biosurfactant, which demonstrated stability in a broad range of pH and temperature as well as a high concentration of saline, with the reduction of the surface tension of water to 29 mN/m. The maximum concentration of biosurfactant (10.0 g/l) was reached after 144 h of cultivation. The biosurfactant was considered to be a lipopeptide based on the results of proton nuclear magnetic resonance and Fourier transformed infrared spectroscopy. The tests demonstrated that the biosurfactant is innocuous and has potential for the bioremediation of soil and water contaminated by petroleum products. Thus, the biosurfactant described herein has a low production cost and can be used in environmental processes.

Female masculinization and reproductive success in Cnesterodon decemmaculatus (Jenyns, 1842) (Cyprinodontiforme: Poeciliidae) under anthropogenic impact Abstract Aquatic organisms are exposed to a myriad of chemical compounds, with particular concerns focused on endocrine disruptors. Growing scientific evidence indicates that these compounds interfere with normal endocrine function and could affect the reproductive system of humans and wildlife. We analyzed the proportion of masculinized females, defined by elongation and fusion of the anal fin rays, and the extent of masculinization, masculinization index, defined by anal fin length divided by the standard length, of Cnesterodon decemmaculatus resident to areas of agricultural and urban-industrial activities in the Arroyo Colorado basin. Additionally, a bioassay was carried out to assess the potential effects of masculinization on reproductive success, measured as the number of viable progenies, using pregnant females from the site downstream of the urban–industrial zone. Masculinized females were observed in all sampling sites, particularly downstream of the urban–industrial area, where over 80% of females presented abnormal sexual characteristics and the highest masculinization index was registered. In the laboratory, masculinized adult females showed male mating behavior, and survival of their progeny was lower than those of normal females. To our knowledge, this is the first report of endocrine disruption in field-collected C. decemmaculatus, and the first evaluation of the reproductive success of masculinized females. Finally, our results support C. decemmaculatus as an excellent sentinel species due to its wide distribution, easy culture in laboratory conditions, and its potential capability to respond to sources of pollution, particularly endocrine disruptors.

Biodegradation of chlorpyrifos and 3,5,6-trichloro-2-pyridinol by the epiphytic yeasts Rhodotorula glutinis and Rhodotorula rubra Abstract The possible involvement of the epiphytic yeasts Rhodotorula glutinis and Rhodotorula rubra in the biodegradation of the insecticide chlorpyrifos and its metabolite 3,5,6-trichloro-2-pyridinol (TCP), in pure cultures and in plant surfaces (tomato fruits) was investigated. Higher biodegradation rates were observed as the concentration of chlorpyrifos and the inoculum of the microorganisms were increased, while the yeasts proved to be more active at 25 and 15 °C. The presence of glucose in the mineral nutrient medium, as an extra source of carbon, delayed the biodegradation by Rhodotorula glutinis, while Rhodotorula rubra proved to be more active. The detection and quantification of the parent compound and TCP was successfully achieved using a LC/MS/MS chromatographic system. The in vitro enzymatic assays applied suggested that esterases may be involved in the biodegradation of chlorpyrifos, a fact that was further enhanced after the addition of the synergists triphenyl phosphate, diethyl maleate and piperonyl butoxide in the biodegradation trials. The decrease of chlorpyrifos residues on tomato fruits confirmed the corresponding on pure cultures, resulting in the suggestion that the yeasts R. glutinis and R. rubra can possibly be used successfully for the removal or detoxification of chlorpyrifos residues on tomatoes.

Antioxidant responses of Triticum aestivum plants to petroleum-derived substances Abstract Winter common wheat (Triticum aestivum L.) plants were cultivated on petroleum products contaminated soils with and without using biopreparation ZB-01. We determined the impact of soil contamination with petrol, diesel fuel and engine oil on selected antioxidant enzymes and the levels of antioxidants in the leaves of winter wheat. The impact of petroleum products on selected morphological characteristics of the plants, levels of nutrients and heavy metals was also assessed. Winter wheat was relatively resistant to soil contamination with petroleum products, and did not show a significant impact on the morphological characteristics of the plants. The levels of nutrients and heavy metals in the plants depended on the type of pollutant and the analyzed component.‬ Biopreparation ZB-01 generally resulted in an increase in calcium levels in the plants.‬ The winter wheat plants growing in soil contaminated with engine oil were characterized by higher levels of zinc, lead, manganese and cadmium than the control plants.‬ Biopreparation applied to the soil contaminated with petrol resulted in a slight increase in the levels of lead and zinc in the plants.‬ The petroleum products affected the activity of antioxidant enzymes and the levels of antioxidants in the plants.‬ The general markers of soil contaminated with diesel fuel and petrol were POD activity and proline levels. Use of the ZB-01 biopreparation caused an increase in the levels of proline and -SH groups and an increase in the levels of carbon and calcium in the plants and had no effect on the morphological characteristics of plants.‬

Cadmium toxicity degree on tomato development is associated with disbalances in B and Mn status at early stages of plant exposure Abstract Cadmium (Cd) toxicity is frequently coupled to its accumulation in plants, but not always the highest Cd concentration triggers the worst damages, indicating that additional events influence the magnitude of Cd side-effects. We investigated the early mechanisms behind the differential Cd-induced impacts on plant development of four tomato accessions with contrasting tolerance to Cd toxicity. At organ level, the highest Cd concentration was not associated with the largest biomass losses. In leaves, changes in superoxide dismutase and catalase activities were not related to differences in Cd concentration, which was unable to provoke H2O2 overproduction on the sixth day of plant exposure to this metal. Further investigation in the mineral profile revealed that magnitude of Cd toxicity depends probably on synergic effects from increased B status, in addition to the own Cd accumulation. Furthermore, disbalances in Mn status (i.e., excess in leaves and deficiency in roots) may enhance Cd toxicity degree. According to data, however, the low magnesium (Mg) status can be linked to tomato tolerance against Cd toxicity. In conclusion, the tomato tolerance degree under short-Cd exposure depends on actively, finely regulation of mineral homeostasis that results in different development of plant organs. The better understanding on the mode of action of Cd toxicity in plants can help in the establishment of strategies to mitigate its impacts on crop yield.