Graphene oxide-silver nanoparticle hybrid material: an integrated nanosafety study in zebrafish embryos.

This work reports an integrated nanosafety study including the synthesis and characterization of the graphene oxide-silver nanoparticle hybrid material (GO-AgNPs) and its nano-ecotoxicity evaluation in the zebrafish embryo model. The influences of natural organic matter (NOM) and a chorion embryo membrane were considered in this study, looking towards more environmentally realistic scenarios and standardized nanotoxicity testing. The nanohybrid was successfully synthesized using the NaBH4 aqueous method, and AgNPs (~ 5.8 nm) were evenly distributed over the GO surface. GO-AgNPs showed a dose-response acute toxicity: the LC50 was 1.5 mg L-1 for chorionated embryos. The removal of chorion, however, increased this toxic effect by 50%. Furthermore, the presence of NOM mitigated mortality, and LC50 for GO-AgNPs changed respectively from 2.3 to 1.2 mg L-1 for chorionated and de-chorionated embryos. Raman spectroscopy confirmed the ingestion of GO by embryos; but without displaying acute toxicity up to 100 mg L-1, indicating that the silver drove toxicity down. Additionally, it was observed that silver nanoparticle dissolution has a minimal effect on these observed toxicity results. Finally, understanding the influence of chorion membranes and NOM is a critical step towards the standardization of testing for zebrafish embryo toxicity in safety assessments and regulatory issues.

TiO2-MWCNT nanohybrid: Cytotoxicity, protein corona formation and cellular internalisation in RTG-2 fish cell line.

Titanium dioxide nanoparticles-multiwalled carbon nanotubes (TiO2-MWCNT) nanohydrid has an enhanced photocatalytic activity across the visible light with promising applications in environmental remediation, solar energy devices and antimicrobial technologies. However, it is necessary to evaluate the toxicological effects of TiO2-MWCNT towards safe and sustainable development of nanohybrids. In this work, we studied the cytotoxicity, protein corona formation and cellular internalisation of TiO2-MWCNT on fibroblasts derived from gonadal rainbow trout tissue (RTG-2) for the first time. This nanohydrid did not show any toxicity effect on RTG-2 cells up to 100 mg L-1 after 24 h of exposure as monitored by alamar blue, neutral red and trypan blue assays (in presence or absence of foetal bovine serum, FBS). Futhermore, cryo-transmission electron microscopy analysis demonstrated that TiO2 particles is attached on nanotube surface after FBS-protein corona formation in cell culture medium. Raman spectroscopy imaging showed that TiO2-MWCNT can be internalised by RTG-2 cells. This work is a novel contribution towards better understanding the nanobiointeractions of nanohydrids linked to their in vitro effects on fish cells in aquatic nanoecotoxicology.

Chitosan nanoparticles containing the insecticide dimethoate: A new approach in the reduction of harmful ecotoxicological effects.

Organophosphate insecticides such as dimethoate (DMT) are widely used in agriculture. As a side effect, however, these insecticides contaminate bodies of water, resulting in damage to aquatic organisms. The development of nanopesticides may be an innovative alternative in the control of agricultural pests, increasing effectiveness and reducing their toxicological effects. Based upon this, the present study has investigated encapsulated DMT in alginate chitosan nanoparticles (nanoDMT) and evaluated its toxicological effects on non-target organisms. The nanoparticles were characterized by DLS, NTA and AFM, as well as being evaluated by the release profile. Nanoparticle toxicity was also evaluated in comparison with DMT, empty nanoparticles and DMT (NP + DMT), and commercial formulations (cDMT), in the embryos and larvae of Danio rerio (zebrafish) according to lethality, morphology, and behavior. The nanoparticle control (NP) showed hydrodynamic size values of 283 ± 4 nm, a PDI of 0.5 ± 0.05 and a zeta potential of -31 ± 0.4 mV. For nanoparticles containing dimethoate, the nanoparticles showed 301 ± 7 nm size values, a PDI of 0.45 ± 0.02, a zeta potential of -27.9 ± 0.2 mV, and an encapsulation of 75 ± 0.32%, with slow-release overtime (52% after 48 h). The AFM images showed that both types of nanoparticles showed spherical morphology. Major toxic effects on embryo larval development were observed in commercial dimethoate exposure followed by the technical pesticide, predominantly in the highest tested concentrations. With regard to the toxic effects of sodium alginate/chitosan, although there was an increase for LC50-96 h concerning the technical dimethoate, the behavior of the larvae was not affected. The data obtained demonstrate that nanoencapsulated dimethoate reduces the toxicity of insecticides on zebrafish larvae, suggesting that nanoencapsulation may be safer for non-target species, by eliminating collateral effects and thus promoting sustainable agriculture.

Assessing the safety of Pseudomonas putida introduction in the environment: an overview of ecotoxicological tests.

Risk assessment guidelines for the environmental release of microbial agents are performed in a tiered sequence which includes evaluation of exposure effects on non-target organisms. However, it becomes important to verify whether environmental risk assessment from temperate studies is applicable to tropical countries, as Brazil. Pseudomonas putida is a bacteria showing potential to be used for environmental applications as bioremediation and plant disease control. This study investigates the effects of this bacteria exposure on rodents and aquatic organisms (Daphnia similis) that are recommended to be used as non-target organism in environmental risk assessments. Also, the microbial activity in three different soils under P. putida exposure was evaluated. Rats did not show clinical alterations, although the agent was recovered 16h after the exposure in lung homogenates. The bacteria did not reduce significantly the reproduction and survival of D. similis. The soil enzymatic activities presented fluctuating values after inoculation with bacteria. The measurement of perturbations in soil biochemical characteristics is presented as an alternative way of monitoring the overall effects of the microbial agent to be introduced even in first stage (Tier I) of the risk assessment in tropical ecosystems.

Influence of Pseudomonas putida AF7 inoculation on soil enzymes.

There has been some concern about the environmental impact of microbial agents. Pseudomonas may be used as bioremediator and as biopesticide. In this study, we report the use of soil enzyme assays as biological indicator of possible negative effects in soil functioning after the P. putida AF7 inoculation. For that, P. putida AF7 was originally isolated from the rizosphere of rice and was inoculated on three soil types: Rhodic Hapludox (RH), Typic Hapludox (TH); and Arenic Hapludult (AH). The acid phosphatase, beta-glucosidase and protease enzymes activities were measured for three period of evaluation (7, 14 and 21 days). In general, the enzymatic activities presented variation among the tested soils. The highest activities of beta-glucosidase and acid phosphatase were observed in the RH and AH soils, while the protease activity was higher in the TH soil. Also, the soil characteristics were measured for each plot. The activity of enzymes from the carbon cycle was positively correlated with the N and the P and the enzyme from the nitrogen cycle was negatively correlated with N and C.org. The presented data indicate that soil biochemical properties can be an useful tool for use as an indicator of soil perturbations by microbial inoculation in a risk assessment.

Assessments of Motor Abnormalities on the Grid-Walking and Foot-Fault Tests From Undernutrition in Wistar Rats.

This study was designed to verify whether different lactation conditions influenced nervous system development. The authors used motor tasks to verify changes in exploratory activity and muscle strength of weaned rats from different litter sizes and evaluated the applicability of the grid-walking test for assessing motor abnormalities caused by undernutrition. Alterations in litter size during the suckling period perturbed the nutritional status of pups, which exhibited body weight differences between the groups. Large-litter (L) pups showed significant delays in achieving developmental milestones and neurological reflexes compared to the small-litter (S) and medium-litter (M) pups. The S, M, and L group pups exhibited similar exploratory responses and muscle strength. In the grid-walking and foot-fault tests, the L group pups traveled shorter distances and, consequently, had less footsteps. However, the percentages of foot faults in the L group were higher than S and M groups. These results reflect delayed maturation of structures responsible for sensorimotor responses, such as the cerebellum, because much cerebellar maturation takes place postnatally. This is the first study to report that early undernutrition in pups resulted in suboptimal performances on the grid-walking and foot-fault tests and that the former test was sensitive to alterations caused by nutritional deficiency.

Developmental toxicity potential of paclobutrazol in the rat.

Paclobutrazol is used as a fungicide and as a plant growth regulator, but there are few studies about its potential toxic effects. Experiments were conducted to determine whether this compound has adverse effects on the reproduction and development of offspring. The influence of 1.0 mg/kg paclobutrazol (10 x the acceptable daily intake-ADI), by oral exposure during gestational organogenesis of rats, was evaluated on maturational and behavioural aspects of offspring development. This dose did not promote evidence of maternal toxicity and the weight of gravid uterus, fetus, and ovary on days 16 and 20 of pregnancy were not affected. Also, the pesticide did not affect body weight gain of the dams and offspring. However, the pups' survival at weaning was impaired by paclobutrazol. Beyond that, study of the functional state of rat pups' nervous system at different stages of postnatal development revealed some differences in treated ones. Damage was observed in the expression of acoustic startle reflex and altered locomotion and/or rearing in an open-field apparatus in treated pups depending on their age. There were no observed alterations in swimming behaviour. The data support further studies of the potentially toxic effects that exposing mothers to this pesticide may have on their litters.