Impact of the glyphosate-based commercial herbicide, its components and its metabolite AMPA on non-target aquatic organisms.

Glyphosate (GLY) is the active ingredient of several herbicide formulations widely used to control weeds in agricultural and non-agricultural areas. Due to the intensive use of GLY-based herbicides and their direct application on soils, some of their components, including the active ingredient, may reach the aquatic environment through direct run-off and leaching. The present study assessed the acute toxicity and genotoxicity of the GLY-based formulation Atanor 48 (ATN) and its major constituents GLY, surfactant polyethoxylated tallow amine (POEA), as well as the main metabolite of GLY aminomethylphosphonic acid (AMPA) on non-target aquatic organisms. The toxic effects of these chemicals were evaluated in the fish embryo acute toxicity test with zebrafish (Danio rerio), while genotoxic effects were investigated in the comet assays with cells from zebrafish larvae and rainbow trout gonad-2 (RTG-2). GLY and AMPA caused no acute toxic effect, while ATN and POEA induced significant lethal effects in zebrafish (LC50-96 h 76.50 mg/L and 5.49 mg/L, respectively). All compounds were genotoxic in comet experiments with zebrafish larvae (LOEC 1.7 mg/L for GLY, ATN, AMPA and 0.4 mg/L for POEA). Unlike in vivo, only POEA induced DNA damage in RTG-2 cells (LOEC 1.6 mg/L), suggesting that it is a direct acting genotoxic agent. In summary, these data indicate that the lethal effects on zebrafish early-life stages can be ranked in the following order from most to least toxic: surfactant POEA > formulation ATN > active ingredient GLY ≈ metabolite AMPA. Genotoxic effects were observed in both RTG-2 cells (only POEA) and zebrafish (all test compounds) with the lowest tested concentrations. Therefore, it is important to evaluate different toxicological endpoints as well as use different non-target organisms to predict the hazards of GLY-based formulations and their components and breakdown product to aquatic biota.

Genotoxic and mutagenic assessment of iron oxide (maghemite-γ-Fe2O3) nanoparticle in the guppy Poecilia reticulata.

The environmental risk of nanomaterials (NMs) designed and used in nanoremediation process is of emerging concern, but their ecotoxic effects to aquatic organism remains unclear. In this study, the citrate-coated (maghemite) nanoparticles (IONPs) were synthesized and its genotoxic and mutagenic effects were investigated in the female guppy Poecilia reticulata. Fish were exposed to IONPs at environmentally relevant iron concentration (0.3 mg L-1) during 21 days and the animals were collected at the beginning of the experiment and after 3, 7, 14 and 21 days of exposure. The genotoxicity and mutagenicity were evaluated in terms of DNA damage (comet assay), micronucleus (MN) test and erythrocyte nuclear abnormalities (ENA) frequency. Results showed differential genotoxic and mutagenic effects of IONPs in the P. reticulata according to exposure time. The IONP induced DNA damage in P. reticulata after acute (3 and 7 days) and long-term exposure (14 and 21 days), while the mutagenic effects were observed only after long-term exposure. The DNA damage and the total ENA frequency increase linearly over the exposure time, indicating a higher induction rate of clastogenic and aneugenic effects in P. reticulata erythrocytes after long-term exposure to IONPs. Results indicated that the P. reticulata erythrocytes are target of ecotoxicity of IONPs.

Effects of triclosan on zebrafish early-life stages and adults.

BACKGROUND, AIM AND SCOPE: The biocide triclosan (TCS) is commonly used in personal care, acrylic, plastic, and textiles products. TCS has been detected in surface water in several countries, and its ecological impact is largely unknown. In this work, the toxicity of TCS in zebrafish (Danio rerio), embryos and adults was studied. Several lethal and sub-lethal endpoints were analysed in organisms exposed to TCS such as mortality, embryo development and behaviour, hatching, micronuclei and biochemical markers (cholinesterase (ChE), glutathione S-transferase (GST) and lactate dehydrogenase (LDH)). MATERIALS AND METHODS: Embryo/larvae assay followed the OECD guideline on Fish Embryo Toxicity Test. Embryos were exposed at nominal concentrations of 0.1, 0.3, 0.5, 0.7 and 0.9 mg/l of TCS for 6 days and were inspected daily with the help of a stereomicroscopy for mortality, developmental parameters (otolith formation, eye and body pigmentation, somite formation, heart beat, tail circulation, detachment of the tail-bud from the yolk sac) and hatching. A similar test was run to obtain larvae for ChE, GST and LDH analysis. The adult test followed the OECD Guideline TG 203 in semi-static conditions. Adult zebrafish of similar length and age were exposed to nominal concentrations of 0.1, 0.2, 0.3, 0.4 and 0.5 mg/l of TCS for 96 h and were inspected daily for mortality and behaviour alterations. A second test was run to obtain organs for biomarkers analysis: Heads, muscles and gills were isolated and snap-frozen in eppendorfs and used for ChE, LDH and GST determinations, respectively. Adult zebrafish testing also comprised a third test for micronucleus analysis in which the nominal concentrations of 0, 0.175 and 0.350 mg/l were used. Peripheral blood was obtained by cardiac puncture and used for the analysis. RESULTS: TCS showed acute toxicity for embryo/larvae (96 h LC(50) = 0.42 mg/l) and delayed hatching. Moreover, embryo toxicity was evident: Delay on the otolith formation and eye and body pigmentation were found, and malformations were also evident, including spine malformations, pericardial oedema and undersize. Biomarkers levels were affected: ChE and LDH activity were increased in larvae exposed to 0.25 mg/l, and GST activity was increased in larvae exposed to 0.25 and 0.35 mg/l. TCS also demonstrated acute toxicity to adult zebrafish (96 h LC(50) = 0.34 mg/l). However, TCS did not change biomarkers levels and did not elicit a micronucleus in adults. DISCUSSIONS: Despite the fact that similar 96 h LC(50) values have been found for D. rerio embryos and adults (0.42 and 0.34 mg/l, respectively), the embryo assay was much more informative, showing important effects at several levels, including teratogenic response, hatching delay and alteration of biomarker levels. TCS does not seem to be genotoxic for adult fish or to interfere with biomarkers levels at the concentrations tested. CONCLUSIONS: TCS has deleterious effects on zebrafish adults and during early stages, (including embryotoxicity, hatching delay and alterations of biomarkers levels). The range of endpoints used on the embryo test allows an integrated analysis that contributes to a better understanding of the toxicity and mode of action of TCS. RECOMMENDATIONS AND PERSPECTIVES: Future works should focus on a deeper investigation of TCS modes of action on zebrafish early-life stages. As embryo testing was revealed to be so informative, a refinement of the test could be made, including other endpoints such as different biochemical markers as well as DNA microarrays to assess a gene expression level for the effect of exposure to TCS. In the perspective of risk assessment, these endpoints should be explored in order to assess their usefulness as early warning signs and links should be sought between these short-term tests and effects of long-term exposures as it is observed in more realistic scenarios.