Due to worldwide production, sales and application, neonicotinoids dominate the global use of insecticides. While, neonicotinoids are considered as pinpoint neurotoxicants that impair cholinergic neurotransmission in pest insects, the sublethal effects on nontarget organisms and other neurotransmitters remain poorly understood. Thus, we investigated long-term neurological outcomes in the decomposer nematode Caenorhabditis elegans. In the adult roundworm the neonicotinoid thiacloprid impaired serotonergic and dopaminergic neuromuscular behaviors, while respective exposures to thiamethoxam showed no effects. Thiacloprid caused a concentration-dependent delay of the transition between swimming and crawling locomotion that is controlled by dopaminergic and serotonergic neurotransmission. Age-resolved analyses revealed that impairment of locomotion occurred in young as well as middle-aged worms. Treatment with exogenous serotonin rescued thiacloprid-induced swimming deficits in young worms, whereas additional exposure with silica nanoparticles enhanced the reduction of swimming behavior. Delay of forward locomotion was partly caused by a new paralysis pattern that identified thiacloprid as an agent promoting a specific rigidity of posterior body wall muscle cells and peripheral neuropathy in the nematode (lowest-observed-effect-level 10 ng/ml). On the molecular level exposure with thiacloprid accelerated protein aggregation in body wall muscle cells of polyglutamine disease reporter worms indicating proteotoxic stress. The results from the soil nematode Caenorhabditis elegans show that assessment of neurotoxicity by neonicotinoids requires acknowledgment and deeper research into dopaminergic and serotonergic neurochemistry of nontarget organisms. Likewise, it has to be considered more that different neonicotinoids may promote diverse neural end points.

The interactions of plastics and soil organisms are complex and inconsistent observations on the effects of plastics on soil organisms have been made in published studies. In this study, we assessed the effects of plastic exposure on plants, fauna and microbial communities, with a meta-analysis. Using a total of 2936 observations from 140 publications, we analysed how responses in plants, soil fauna and microorganisms depended on the plastic concentration, size, type, species and exposure media. We found that overall plastics caused substantial detrimental effects to plants and fauna, but less so to microbial diversity and richness. Plastic concentration was one of the most important factors explaining variations in plant and faunal responses. Larger plastics (>1 μm) caused unfavourable changes to plant growth, germination and oxidative stress, while nanoplastics (NPs; ≤ 1 μm) only increased oxidative stress. On the contrary, there was a clear trend showing that small plastics adversely affected fauna reproduction, survival and locomotion than large plastics. Plant responses were indifferent to plastic type, with most studies conducted using polyethylene (PE) and polystyrene (PS) plastics, but soil fauna were frequently more sensitive to PS than to PE exposure. Plant species played a vital role in some parameters, with the effects of plastics being considerably greater on vegetable plants than on cereal plants.

6-benzylaminopurine (6-BA) is one of the first synthetic hormones and has been widely used in fruit cultivation, gardening and agriculture. However, excessive use of 6-BA will cause potential harm to the environment and humans. Therefore, our research focused on assessing the impact of 6-BA on the development and neurobehavior of zebrafish. The results showed that 6-BA had little effect on the embryos from 2 hpf to 10 hpf. However, delayed development, decreased survival and hatchability were observed under 30 and 40 mg/L 6-BA from 24 hpf. 6-BA also reduced surface tension of embryonic chorions at 24 hpf. In addition, 6-BA caused abnormal morphology and promoted the accumulation of oxidative stress. Transcription of genes in connection with development and oxidative stress was also strikingly altered. Results of movement assay showed that zebrafish were less active and their behavior was significantly inhibited under the 20 and 30 mg/L 6-BA treatments. Locomotion-related genes th and mao were down-regulated by gradient, while the transcription of dbh was upregulated at a low concentration (2 mg/L) but decreased as the concentration increased. Moreover, 6-BA exposure caused increased arousal and decreased sleep. Sleep/wake related genes hcrt and hcrtr2 were upregulated, but decreased at 30 mg/L, while the mRNA level of aanat2 was reduced in a concentration-dependent manner. To sum up, our results showed that 6-BA induced developmental toxicity, promoted the accumulation of oxidative stress, and damaged locomotion and sleep/wake behavior.

Graphene (GR) and graphene oxide (GO) are widely being used as promising candidates for biomedical applications, as well as for bio-sensing, drug delivery, and anticancer therapy. However, their undesirable side effects make it necessary to assess further the toxicity and safety of using these materials. The main objective of the current study was to investigate the toxicities of GR and GO in predicted environmental relevant concentrations in adult zebrafish (Danio rerio), particularly on their behaviors, and conducted biochemical assays to elucidate the possible mechanism that underlies their toxicities. Zebrafish was chronically (∼14 days) exposed to two different doses of GR (0.1 and 0.5 ppm) or GO (0.1 and 1 ppm). At 14 ± 1 days, a battery of behavioral tests was conducted, followed by enzyme-linked immunosorbent assays (ELISA) test on the following day to inspect the alterations in antioxidant activity, oxidative stress, and neurotransmitters in the treated zebrafish brain. An alteration in predator avoidance behavior was observed in all treated groups, while GR-treated fish exhibited abnormal exploratory behavior. Furthermore, altered locomotor activity was displayed by most of the treated groups, except for the high concentration of the GR group. From the ELISA results, we discovered a high concentration of GR exposure significantly decreased several neurotransmitters and cortisol levels. Meanwhile, elevated reactive oxygen species (ROS) were displayed by the group treated with low and high doses of GR and GO, respectively. These significant changes would possibly affect zebrafish behaviors and might suggest the potential toxicity from GR and GO exposures. To sum up, the present study presented new evidence for the effects of GR and GO in zebrafish behavioral dysregulation. We hope these assessments can contribute to our understanding of graphene and graphene oxide biosafety.

Freshwater planarians have been gaining relevance as experimental animals for numerous research areas given their interesting features, such as high regeneration potential, shared features with the vertebrates’ nervous system or the range of endpoints that can be easily evaluated in response to contaminants. Ecotoxicological research using these animals has been steadily increasing in the past decades, as planarians’ potentialities for this research area are being recognized. In this work, we used polycyclic aromatic hydrocarbons (PAHs) as model contaminants and evaluated effects of exposure to phenanthrene, pyrene and benzo[a]pyrene (B[a]P) in planarians. The freshwater planarian Girardia tigrina was chosen and mortality, cephalic regeneration (during and post-exposure), behavioral endpoints and presence of PAHs in tissues, were evaluated. Mortality was only observed in planarians exposed to phenanthrene, with an estimated LC₅₀ of 830 μg L⁻¹. Results indicate that planarian behavioral endpoints were very sensitive in response to sub-lethal concentrations of PAHs, showing a greater sensitivity towards B[a]P and pyrene. Briefly, post-exposure locomotion and post-exposure feeding were significantly impaired by sub-lethal concentrations of all compounds, whereas regeneration of photoreceptors was only significantly delayed in planarians exposed to pyrene. Moreover, levels of PAH-type compounds in planarian tissues followed a concentration-dependent increase, showing uptake of compounds from experimental solutions. The present results highlight the importance of studying alternative and complementary endpoints, such as behavior, not only because these may be able to detect effects at lower levels of contamination, but also due to their ecological relevance. The simplicity of evaluating a wide range of responses to contaminants further demonstrates the utility of freshwater planarians for ecotoxicological research.

With increasing release of nanoparticles (NPs) into the environment, soil organisms likely suffer from high dose and long duration of NPs contamination, while the effect of NPs across multiple generations in soil is rarely studied. Herein, we investigated how multigenerational exposure to different crystal forms (anatase, rutile, and their mixture) of TiO₂ NPs (nTiO₂) affected the survival, behavior, physiological and biochemical traits, and lifespan of nematodes (C. elegans) in a paddy soil. The soil property changed very slightly after being spiked with nTiO₂, and the toxicities of three nTiO₂ forms were largely comparable. The nTiO₂ exposure adversely influenced the survival and locomotion of nematodes, and increased intracellular reactive oxygen species (ROS) generation. Interestingly, the toxic effect gradually attenuated and the lifespan of survived nematodes increased from the P0 to F3 generation, which was ascribed to the survivor selection and stimulatory effect. The lethal effect and the increased oxidative stress may continuously screen out offspring possessing stronger anti-stress capabilities. Moreover, key genes (daf-2, age-1, and skn-1) in the insulin/IGF-like signaling (IIS) pathway actively responded to the nTiO₂ exposure, which further optimized the selective expression of downstream genes, increased the antioxidant enzyme activities and antioxidant contents, and thereby increased the stress resistance and longevity of survived nematodes across successive generations. Our findings highlight the crucial role of bio-responses in the progressively decreased toxicity of nTiO₂, and add new knowledge on the long-term impact of soil nTiO₂ contamination.

Hydrocarbons contamination and hypoxia are two stressors that can coexist in coastal ecosystems. At present, few studies evaluated the combined impact of these stressors on fish physiology and behavior. Here, we tested the effect of the combination of hypoxia and petrogenic hydrocarbons on the anti-predator locomotor performance of fish. Specifically, two groups of European sea bass (Dicentrarchus labrax) were exposed to clean water (Ctrl) or oil-contaminated water (Oil). Subsequently, fish of both groups were placed in normoxic (norx) or hypoxic (hyp) experimental tanks (i.e. four groups of fish were formed: Ctrl norx, Ctrl hyp, Oil norx, Oil hyp). In these tanks, escape response was elicited by a mechano-acoustic stimulus and recorded with a high speed camera. Several variables were analyzed: escape response duration, responsiveness (percentage of fish responding to the stimulation), latency (time taken by the fish to initiate a response), directionality (defined as away or toward the stimulus), distance-time variables (such as speed and acceleration), maneuverability variables (such as turning rate), escape trajectory (angle of flight) and distancing of the fish from the stimulus. Results revealed (i) effects of stressors (Ctrl hyp, Oil norx and Oil hyp) on the directionality; (ii) effects of Oil norx and Oil hyp on maneuverability and (iii) effects of Oil hyp on distancing. These results suggest that individual stressors could alter the escape response of fish and that their combination could strengthen these effects. Such an impact could decrease the probability of prey escape success. By investigating the effects of hydrocarbons (and the interaction with hypoxia) on the anti-predator behavior of fish, this work increases our understanding of the biological impact of oil spill. Additionally, the results of this study are of interest for oil spill impact evaluation and also for developing new ecotoxicological tools of ecological significance.

The potential adverse effects of nanoplastics, such as nanopolystyrene, have received the great attention recently. However, the molecular response of organisms to nanoplastics is still largely unknown. In this study, we employed Caenorhabditis elegans as an animal model to investigate the long non-coding RNAs (lncRNAs) in response to long-term exposure to low-dose nanopolystyrene (100 nm). Based on Hiseq 2000 sequencing and qRT-PCR confirmation, we identified 36 lncRNAs (21 down-regulated lncRNAs and 15 up-regulated lncRNAs) in response to nanopolystyrene (1 μg/L). Using intestinal reactive oxygen species (ROS) production and locomotion behavior as endpoints, we found that RNAi knockdown of linc-2, linc-9, or linc-61 induced a susceptibility to nanopolystyrene toxicity, and RNAi knockdown of linc-18 or linc-50 induced a resistance to nanopolystyrene toxicity. Meanwhile, nanopolystyrene (1 μg/L) increased expressions of linc-2, linc-9, linc-18, and linc-61 and decreased linc-50 expression, suggesting that these 5 lncRNAs mediated two different responses to nanopolystyrene exposure. Bioinformatical analysis implied that these 5 lncRNAs were associated with multiple biological processes and signaling pathways. Our results demonstrated the crucial roles of lncRNAs in response to long-term exposure to low-dose nanopolystyrene in organisms.

Nanoplastics are widely used in modern life, for example, in cosmetics and daily use products, and are attracting concern due to their potential toxic effects on environments. In this study, the uptake of nanopolystyrene particles by Caenorhabditis elegans (C. elegans) and their toxic effects were evaluated. Nanopolystyrene particles with sizes of 50 and 200 nm were prepared, and the L4 stage of C. elegans was exposed to these particles for 24 h. Their uptake was monitored by confocal microscopy, and various phenotypic alterations of the exposed nematode such as locomotion, reproduction and oxidative stress were measured. In addition, a metabolomics study was performed to determine the significantly affected metabolites in the exposed C. elegans group. Exposure to nanopolystyrene particles caused the perturbation of metabolites related to energy metabolism, such as TCA cycle intermediates, glucose and lactic acid. Nanopolystyrene also resulted in toxic effect including induction of oxidative stress and reduction of locomotion and reproduction. Collectively, these findings provide new insights into the toxic effects of nanopolystyrene particles.

At present, the majority of investigations concerning nanotoxicology in the nematode C. elegans address short-term effects. While this approach allows for the identification of uptake pathways, exposition and acute toxicity, nanoparticle-organism interactions that manifest later in the adult life of C. elegans are missed. Here we show that a microhabitat composed of liquid S-medium and live bacteria in microtiter wells prolongs C. elegans longevity and is optimally suited to monitor chronic eNP-effects over the entire life span (about 34 days) of the nematode. Silver (Ag) nanoparticles reduced C. elegans life span in concentrations ≥10 μg/mL, whereas nano ZnO and CeO₂ (1–160 μg/mL) had no effect on longevity. Monitoring of locomotion behaviors throughout the entire life span of C. elegans showed that Ag NPs accelerate the age-associated decline of swimming and increase of uncoordinated movements at concentrations of ≥10 μg/mL, whereas neuromuscular defects did not occur in response to ZnO and CeO₂ NPs. By means of a fluorescing reporter worm expressing tryptophan hydroxylase-1::DsRed Ag NP-induced behavioral defects were correlated to axonal protein aggregation and neurodegeneration in single serotonergic HSN as well as sensory ADF neurons. Notably, serotonergic ADF neurons represented a sensitive target for Ag NPs in comparison to GABAergic neurons that showed no signs of degeneration under the same conditions. We conclude that due to its analogy to the jellylike boom culture of C. elegans on microbe-rich rotting plant material liquid S-medium culture in spatially confined microtiter wells represents a relevant as well as practical tool for comparative identification of age-resolved nanoparticle effects and vulnerabilities in a significant target organism. Consistent with this, specifically middle-aged nematodes showed premature neuromuscular defects after Ag NP-exposure.