Does age pay off? Effects of three-generational experiments of nanodiamond exposure and withdrawal in wild and longevity-selected model animals.

Nanodiamonds (NDs) are considered a material with low toxicity. However, no studies describe the effects of ND withdrawal after multigenerational exposure. The aim was to evaluate ND exposure (in the 1st and 2nd generations) effects at low concentrations (0.2 or 2 mg kg-1) and withdrawal (in the 3rd generation) in the wild (H) and longevity-selected (D) model insect Acheta domesticus. We measured selected oxidative stress parameters, immunity, types of cell death, and DNA damage. Most of the results obtained in the 1st generation, e.g., catalase (CAT), total antioxidant capacity (TAC), heat shock proteins (HSP70), defensins, or apoptosis level, confirmed no significant toxicity of low doses of NDs. Interestingly, strain-specific differences were observed. D-strain crickets reduced autophagy, the number of ROS+ cells, and DNA damage. The effect can be a symptom of mobilization of the organism and stimulation of physiological defense mechanisms in long-living organisms. The 2nd-generation D-strain insects fed ND-spiked food at higher concentrations manifested a reduction in CAT, TAC, early apoptosis, and DNA damage, together with an increase in HSP70 and defensins. ROS+ cells and cells with reduced membrane potential and autophagy did not differ significantly from the control. H-strain insects revealed a higher number of ROS+ cells and cells with reduced membrane potential, decreased CAT activity, and early apoptosis. Elimination of NDs from the diet in the 3rd generation did not cause full recovery of the measured parameters. We noticed an increase in the concentration of HSP70 and defensins (H-strain) and a decrease in apoptosis (D-strain). However, the most visible increase was a significant increase in DNA damage, especially in H-strain individuals. The results suggest prolonged adverse effects of NDs on cellular functions, reaching beyond "contact time" with these particles. Unintentional and/or uncontrolled ND pollution of the environment poses a new challenge for all organisms inhabiting it, particularly during multigenerational exposure.

Chronic toxicity of nanodiamonds can disturb development and reproduction of Acheta domesticus L.

The use of nanodiamonds in numerous materials designed for industry and medicine is growing rapidly. Consequently health and environmental risks associated with the exposure of humans and other biota to nanodiamonds-based materials are of the utmost importance. Scarcity of toxicological data for these particles led us to examine the potentially deleterious effects of nanodiamonds in model insect species, Acheta domesticus (Orthoptera) chronically exposed to ND in its diet. Organism-level end-point indices (lifespan, body weight, consumption, caloric value of faeces, reproduction) revealed adverse changes in the treated crickets in comparison with the control. Preliminary studies of oxidative stress level in the offspring of ND-treated crickets suggest toxicity of these particles limited to the exposed individuals. EPR analysis showing increase of radical signal in the faeces of ND-fed crickets led us to propose novel mechanism of nanodiamonds toxicity that is discussed in the light of literature data. CAPSULE: Development and reproduction of Acheta domesticus can be disturbed by the chronic exposure to nanodiamonds.

Reduced fecundity and cellular changes in Acheta domesticus after multigenerational exposure to graphene oxide nanoparticles in food.

Despite the fact that the demand for graphene and its derivatives in commercial applications is still growing, many aspects of its toxicity and biocompatibility are still poorly understood. Graphene oxide, which is released into the environment (air, soil and water) as so-called nanowaste or nanopollution, is able to penetrate living organisms. It is highly probable that, due to its specific nature, it can migrate along food chains thereby causing negative consequences. Our previous studies reported that short-term exposure to graphene oxide may increase the antioxidative defense parameters, level of DNA damage, which results in numerous degenerative changes in the gut and gonads. The presented research focuses on reproductive dysfunction and cellular changes in Acheta domesticus after exposure to GO nanoparticles in food (concentrations of 20 and 200 µg·g-1 of food) throughout their entire life cycle. The results showed that long-term exposure to GO caused a significant decrease in the reproductive capabilities of the animals. Moreover, the next generation of A. domesticus had a lower cell vitality compared to their parental generation. It is possible that graphene oxide can cause multigenerational harmful effects.

Short-term in vivo exposure to graphene oxide can cause damage to the gut and testis.

Graphene oxide (GO) has unique physicochemical properties and also has a potentially widespread use in every field of daily life (industry, science, medicine). Demand for nanotechnology is growing every year, and therefore many aspects of its toxicity and biocompatibility still require further clarification. This research assesses the in vivo toxicity of pure and manganese ion-contaminated GO that were administrated to Acheta domesticus with food (at 200mgkg-1 of food) throughout their ten-day adult life. Our results showed that short-term exposure to graphene oxide in food causes an increase in the parameters of oxidative stress of the tested insects (catalase - CAT, total antioxidant capacity - TAC), induces damage to the DNA at a level of approximately 35% and contributes to a disturbance in the stages of the cell cycle and causes an increase of apoptosis. Moreover, upon analyzing histological specimens, we found numerous degenerative changes in the cells of the gut and testis of Acheta domesticus as early as ten days after applying GO. A more complete picture of the GO risk can help to define its future applications and methods for working with the material, which may help us to avoid any adverse effects and damage to the animal.

Phenotypic Plasticity, Epigenetic or Genetic Modifications in Relation to the Duration of Cd-Exposure within a Microevolution Time Range in the Beet Armyworm.

In the case of the pests inhabiting metal polluted or fields where the use of pesticides is common, a natural selection of resistant individuals can occur. This may pose serious problems for humans, agriculture, as well as the economies of many countries. In this study, the hypothesis that multigenerational (120 generations) exposure to cadmium of a beet armyworm population could be a selecting factor toward a more efficient DNA protection was verified. The hemocytes of individuals from two culture strains (control and Cd-exposed) were treated with H2O2 (a DNA-damaging agent) or PBS (reference). The level of DNA damage was assessed using the Comet assay immediately and 5, 15 and 30 min. after the treatment. The immediate result of the contact with H2O2 was that the level of DNA damage in the hemocytes of the insects from both strains increased significantly. However, in the cells of the Cd-exposed individuals, the level of DNA damage decreased over time, while in the cells from the control insects it remained at the same level with no evidence of repair. These results suggest that efficient defense mechanisms may exist in the cells of insects that have prolonged contact with cadmium. Some evolutionary and trade-off aspects of the phenomenon are discussed. In a wider context, comparing the results obtained in the laboratory with field studies may be beneficial for understanding basic mechanisms of the resistance of an organism. To summarize, the high potential for the repair of DNA damage that was observed in the insects from the cadmium strain may confirm the hypothesis that multigenerational exposure to that metal may possibly contribute to the selection of insects that have a wider tolerance to oxidative stress. However, our investigations of polymorphism using AFLP did not reveal differences between the two main insect strains.

The level of DNA damage in adult grasshoppers Chorthippus biguttulus (Orthoptera, Acrididae) following dimethoate exposure is dependent on the insects' habitat.

The comet assay was used to study the DNA damage that was induced by dimethoate in the hemocyte cells of adult Chorthippus biguttulus grasshoppers (Insecta: Orthoptera) that originated from two sites with varying levels of pollution. The primary focus of the study was to examine whether continuous exposure to environmental stress can modify the effect of pesticides on genome stability. After three days of acclimation to laboratory conditions, the level of DNA damage in the hemocytes of Bow-winged grasshoppers was within a similar range in the insects from both areas. However, the level of DNA damage following dimethoate treatment was significantly higher in the insects from the reference area (Pogoria) than in the individuals from the heavily polluted location (Szopienice). Four hours after pesticide treatment, the Tail DNA (TDNA) in the hemocytes of the male and female specimens from Pogoria was as high as 75% and 50% respectively, whereas the values in males and females from Szopienice only reached 30% and 20%, respectively. A rapid decrease in DNA damage was observed in both populations 24 h after the pesticide application. The habitat of an insect (site), the administration of the dimethoate (treatment), and the period following the application of the pesticide (time), all significantly influenced the levels of DNA damage. No interactions related to TDNA were observed between the variables 'sex' and 'treatment'. Similarly, the variable 'sex', when analyzed alongside 'treatment' and 'site' (the area from which the insects were collected), or 'treatment' and 'time' had no influence on TL. Exposure to dimethoate undoubtedly contributed to the formation of DNA damage in the hemocytes of adult C. biguttulus. However, the level of damage was clearly dependent on the place where the insects were captured.

Oxidative stress and genotoxic effects of diamond nanoparticles.

Due to the unique and useful properties of nanodiamonds (ND), their production and use is rapidly increasing. Thus, more of these particles will be released into the environment and organisms will inevitably be exposed to them. The current knowledge about the toxicity of ND, especially in vivo toxicity, is fragmentary. In this study, the toxicity of nanodiamonds was assessed in Acheta domesticus following chronic exposure to different nominal concentrations of ND (20 and 200µgg(-1) food) administrated in food for the entire lifespan. The activity of oxidative stress enzymes (catalase, glutathione peroxidase), total antioxidant capacity, as well as the level of heat shock protein were determined. A significant increase in all of the measured parameters was observed after seven weeks of exposure in individuals exposed to higher concentrations of ND (200µgg(-1) food). In animals exposed to lower concentrations of ND (20µgg(-1) food), there were few significant changes to these parameters. Analysis of DNA damage performed after fourteen weeks using the comet assay revealed DNA instabilities in the insects, especially the ones that had been exposed to the higher doses of ND. These findings may suggest that the toxicity of ND is concentration dependent. While high doses interact in a toxic manner, trace amounts, which are more likely in the environment, might be safe for organisms. Extreme caution should be taken when handling nanodiamonds.

Ultrastructure of the gut epithelium in Acheta domesticus after long-term exposure to nanodiamonds supplied with food.

The biosafety of nanoparticles and the potential toxicity of nanopollutants and/or nanowastes are all currently burning issues. The increased use of nanoparticles, including nanodiamonds (ND), entails the real risk of their penetration into food chains, which may result in the contamination of animal and, as a result, human food. Knowledge about changes in the ultrastructure of tissues in organisms that have been exposed to ND is still very limited. The aim of the study was to describe the ultrastructure of the gut epithelium in Acheta domesticus after exposure to different concentrations of ND (0, 20 or 200 µg g(-1) - control, ND20 and ND200 groups, respectively) administered with food over a five-week period. The ultrastructure of the foregut, midgut and hindgut was assessed using Transmission Electron Microscopy (TEM). A number of changes in the structure of the gut in crickets that had consumed nanodiamond-contaminated food were observed. The epithelium of the midgut and hindgut were clearly damaged by ND, although the foregut was not affected. A positive relationship between the ND concentration in food and the degree of damage to the structure of epithelial cells was observed. Autophagy, especially mitophagy and reticulophagy, was activated in relation to the appearance of ND particles. A putative ND toxicity mechanizm is proposed. Extreme caution should be maintained when using nanodiamonds on a large scale.

Evaluation of in vivo graphene oxide toxicity for Acheta domesticus in relation to nanomaterial purity and time passed from the exposure.

Graphene and its oxidized form-graphene oxide (GO) have become exceptionally popular in industry and medicine due to their unique properties. However, there are suspicions that GO can cause adverse effects. Therefore, comprehensive knowledge on its potential toxicity is essential. This research assesses the in vivo toxicity of pure and manganese ion contaminated GO, which were injected into the hemolymph of Acheta domesticus. The activity of catalase (CAT) and gluthiathione peroxidases (GSTPx) as well as heat shock protein (HSP 70) and total antioxidant capacity (TAC) levels were measured at consecutive time points-1h, 24h, 48h and 72h after injection. Neither pure GO nor GO contaminated with manganese were neutral to the organism. The results proved the intensification of oxidative stress after GO injection, which was confirmed by increased enzyme activity. The organism seems to cope with this stress, especially in the first 24h after injection. In the following days, increasing HSP 70 levels were observed, which might suggest the synthesis of new proteins and the removal of old and damaged ones. With that in mind, the potential toxicity of the studied material, which could lead to serious and permanent damage to the organism, should still be taken into consideration.