Integrative time series of cellular, humoral and molecular response revealed immunotoxicity of bifenthrin to Chinese rare minnow (Gobiocypris rarus) following Pseudomonas fluorescens challenge.

Bifenthrin is a common pesticide that is widespread in aquatic environments. Although it has been shown to be toxic to aquatic organisms, its immunotoxicity and mechanism are unclear. Herein, we reported the immunotoxicity of bifenthrin on adult Chinese rare minnow (Gobiocypris rarus) after 28 days of exposure to different concentrations of bifenthrin (0.1, 0.3, and 1.0 µg/L) and 36-h Pseudomonas fluorescens challenge. Bifenthrin inhibited the fish humoral immune response to bacteria by altering the lymphocyte and neutrophil ratios and decreasing the production of lysozyme, complement component 3, immunoglobulin M, and C-reactive protein, particularly were 1.0 µg/L. Bifenthrin caused intestinal damage and significantly reduced the volume of intestinal mucus at 12 and 36 hours postinjection (hpi) (p < 0.05). Moreover, 1.0 µg/L bifenthrin significantly increased the fish mortality and bacterial loads at 12 and 36 hpi (p < 0.05). RNA-seq analysis revealed several enriched genes involved in pathogen attachment and recognition, inflammatory responses, and complement system at the early-to-mid stage of infection (4-12 hpi). Overall, our results corroborated that bifenthrin induced immunotoxicity in Gobiocypris rarus, resulting in immune dysfunction of fish and increasing their sensitivity to bacterial infection and accelerating mortality. Moreover, 4-12 hpi was better than 36 hpi for analyzing immune responses against pathogen infection in fish exposed to bifenthrin.

Comparative toxicogenomics of benzotriazole ultraviolet stabilizers at environmental concentrations in Asian clam (Corbicula fluminea): Insight into molecular networks and behavior.

Benzotriazole ultraviolet stabilizers (BUVSs) are widespread emerging pollutants, which can pose exposure risks to benthic organisms. However, the toxicity and mechanisms of BUVSs congeners in benthic clams are far from elucidated. In this study, Asian clams (Corbicula fluminea) were exposed to one of UV-234, UV-326, UV-329, or UV-P at environmentally relevant levels (0.1, 1, and 10 µg/L) for 21 days. Filtration rate (FR) was increased in clams exposed to all BUVSs and there were notable histopathologic changes, including irregular digestive lumen, lipid droplet vacuolation, and degraded epithelial cells. To determine the molecular underpinnings following BUVSs exposure, the transcriptome responses in digestive glands were compared. Differentially expressed genes shared among BUVSs treatments were associated with focal adhesion, TNF-α/NF-κB proinflammatory pathways, and apoptosis. Following this, biochemical analysis of biomarkers related to apoptosis were conducted to further validate response. Exposure to BUVSs inhibited anti-oxidant enzyme activity and induced oxidative stress. Heat shock proteins were also triggered with exposure, and there was an induction of caspase-3 and caspase-9 activity. Molecular responses were not identical in the digestive gland of C. fluminea when comparing responses to BUVSs; nevertheless conserved mechanism (impairment of the oxidative defense system, immune system disruption, and induction of apoptosis) among BUVSs congeners was noted. This study provides novel insight into the toxicity and hazards of BUVSs in benthic organisms.

Environmental concentrations of 2, 4-DTBP cause immunotoxicity in zebrafish (Danio rerio) and may elicit ecological risk to wildlife.

Synthetic phenolic antioxidant 2,4-di-tert-butylphenol (2,4-DTBP) has gained growing concerns due to relatively high concentrations in aquatic ecosystems. There are, however, significant knowledge gaps regarding its potential toxicity to aquatic organisms. In this study, zebrafish (Danio rerio) larvae were exposed to 0.01, 0.1, or 1 µM 2,4-DTBP for 6 d. Transcriptomic analysis of larvae revealed that biological processes related to anti-inflammatory function of macrophage M2 lineage were inhibited by 0.01 µM 2,4-DTBP. Decreases of transcripts related to the IL1B-MYD88-NF-κB pathway (i.e., il1b, il1rl1, myd88, irak4, irak1, traf6, ikbkg, nfkbia, nfkb) and protein levels of NF-κB in larvae intestine confirmed anti-inflammatory effects of 2,4-DTBP. Subsequently, larvae exposed to 2,4-DTBP were challenged with E. coli and showed higher survival rate, suggesting sustained activation of inflammation via LPS can be attenuated by 2,4-DTBP. Moreover, histological examination revealed that intestine barrier was compromised and there was an imbalance of intestine macrophage homeostasis. Food intake was also reduced following exposure to 0.1 and 1 µM 2,4-DTBP. In addition, a risk assessment revealed that 2,4-DTBP in surface water pose low to high ecological risks to aquatic organisms. Taken together, exposure to environmentally relevant concentrations of 2,4-DTBP could negatively affect immune response in zebrafish and may elicit ecological risk in fish population.

Long-Term Exposure to SSRI Citalopram Induces Neurotoxic Effects in Zebrafish.

Residual antidepressants are of increasing concern worldwide, yet critical information on their long-term neurotoxic impacts on nontarget aquatic animals is lacking. Here, we investigated the long-term effects (from 0 to 150 days postfertilization) of the selective serotonin reuptake inhibitor citalopram (0.1-100 µg/L) on motor function, learning, and memory in zebrafish over two generations and explored the reversibility of the effect in F1 larvae. Unlike F0+ larvae, we found that F1+ larvae displayed decreased sensorimotor performance when continuously exposed to citalopram at 100 µg/L. No adverse effects were found in F1- larvae after they were transferred to a clean medium. Whole-mount immunofluorescence assays suggested that the motor impairments were related to axonal projections of the spinal motor neurons (MNs). For F0+ adults, long-term citalopram exposure mainly caused male-specific declines in motor, learning, and memory performance. Analysis of serotonergic and cholinergic MNs revealed no significant changes in the male zebrafish spinal cord. In contrast, the number of glutamatergic spinal MNs decreased, likely associated with the impairment of motor function. Additionally, treatment with 100 µg/L citalopram significantly reduced the number of dopaminergic neurons, but no significant neuronal apoptosis was observed in the adult telencephalon. Overall, this study provides neurobehavioral evidence and novel insights into the neurotoxic mechanisms of long-term citalopram exposure and may facilitate the assessment of the environmental and health risks posed by citalopram-containing antidepressant drugs.

Transcriptome profiling reveals toxicity mechanisms following sertraline exposure in the brain of juvenile zebrafish (Danio rerio).

Sertraline (SER) is one of the most commonly detected antidepressants in the aquatic environment that can negatively affect aquatic organisms at low concentrations. Despite some knowledge on its acute toxicity to fish, the effects of chronic SER exposure remain poorly understood along with any underlying mechanisms of SER-induced toxicity. To address this knowledge gap, the effects of chronic exposure to three SER concentrations from low to high were investigated in zebrafish. Juvenile zebrafish were exposed to three concentrations of 1, 10, or 100 µg/L of SER for 28 d, after which indicators of oxidative stress and neurotoxicity in the brain were measured. Superoxide dismutase (SOD) activity was significantly enhanced by SER at 1 up to 100 µg/L, and catalase (CAT) activity was significantly induced by SER at 1 or 10 µg/L. The activity of acetylcholinesterase (AChE) was significantly induced by 10 and 100 µg/L of SER, and the serotonin (5-HT) level was significantly increased by all three concentrations of SER. To ascertain mechanisms of SER-induced toxicity, transcriptomics was conducted in the brain of zebrafish following 100 µg/L SER exposure. The molecular signaling pathways connected with circadian system and the immune system were significantly altered in the zebrafish brain. Based on transcriptomic data, the expression levels of six circadian clock genes were measured, and three genes were significantly altered in relative abundance in fish from all experimental treatments with SER, including cryptochrome circadian regulator 2 (cry2), period circadian clock 2 (per2), and period circadian clock 3 (per3). We hypothesize that the circadian system may be related to SER-induced neurotoxicity and oxidative stress in the central nervous system. This study reveals potential mechanisms and key events (i.e., oxidative stress and neurotoxicity) associated with SER-induced toxicity, and improves understanding of the molecular and biochemical pathways putatively perturbed by SER.

Environmentally relevant concentrations of fenvalerate induces immunotoxicity and reduces pathogen resistance in Chinese rare minnow (Gobiocypris rarus).

Fenvalerate is a broadly used type II pyrethroid with a potential toxic effect in fish. However, information on the immunotoxicity of fenvalerate in fish is scarce. Here, to discover the immunotoxicity of fenvalerate and its underlying mechanism in fish, adult Chinese rare minnow was exposed to fenvalerate at 0, 0.3, 1, and 3 µg/L for 28 days and then subjected to Pseudomonas fluorescens (P. fluorescens) challenge. Fenvalerate induced significant pathological changes, with disintegration of cell boundaries in the intestine, epithelial hyperplasia in gills, and vacuolation of hepatocytes at 3 µg/L treatment. Additionally, the pathological characteristics were more serious during P. fluorescens infection after fenvalerate exposure. A significant increase in neutrophil counts was observed after 3 µg/L fenvalerate exposure for 28 days (p < 0.05), whereas significantly increased monocyte and neutrophil counts and greatly decreased lymphocyte counts were detected at 24 h post-injection (hpi) with P. fluorescens (p < 0.05). Furthermore, obvious decreases in LYS, IgM, ALP, and C3 levels were detected in plasma after 3 µg/L fenvalerate exposure for 28 days, which was consistent with the results at 24 and 48 hpi. Notably, fish exposed to fenvalerate suppressed the transcription of TLR-NF-κB signaling pathway-relevant genes in response to P. fluorescens, accompanied by high mortalities and bacterial loads. Therefore, our results demonstrate that fenvalerate at environmentally relevant concentrations caused immunotoxicity in fish. This study highlights the importance of considering the combined effects of chemicals and pathogens to refine our ability to predict the effects of environmental contaminants on aquatic organisms.

Low doses and lifecycle exposure of waterborne antidepressants in zebrafish model: A survey on sperm traits, reproductive behaviours, and transcriptome responses.

Venlafaxine and citalopram have been commonly found in surface water and may disrupt fish reproduction, yet the long-term impact and the underlying mechanism are largely unknown. Here, zebrafish were exposed to 0.1-100 µg/L venlafaxine and citalopram for their entire life cycle from embryo to adult, respectively. After exposure for 180 days, the lowest observable effective concentration (LOEC) of venlafaxine and citalopram to significantly reduce the mean number of egg production in adults were 10 and 1 µg/L, respectively, whereas the fertilization rate displayed no significant changes. Further, we investigated the impacts of venlafaxine and citalopram in a reproductive context, including sperm quality and reproductive behaviour. In contrast, venlafaxine and citalopram exposure did not affect sperm quality but caused a reduction of reproductive behaviour (e.g., mating duration and mating interval) of adults exposed to 1-10 µg/L of the antidepressant. Transcriptomic profiling of the whole ovary revealed that lifecycle venlafaxine and citalopram exposure significantly affected the Na+/Cl- dependent neurotransmitter transporters signaling. Moreover, immune system-mediated ovarian regeneration and creatine metabolism regulated energy metabolism were proposed as the novel mechanism in the observed effects. Taken together, our results highlight the risk of lifecycle venlafaxine and citalopram exposure to fish reproduction and provide novel perspectives for unveiling the mechanism of female reproductive dysfunction.

Environmentally relevant concentrations of benzophenones triggered DNA damage and apoptosis in male Chinese rare minnows (Gobiocypris rarus).

Benzophenone-type ultraviolet (UV) filters (BPs) are commonly used as sunscreen agents, fragrance enhancers and plastic additives, and are great threats to aquatic organisms due to their high detected concentrations in the aquatic environment. However, few studies on their toxicity and mechanism in fish have been clearly reported. In this study, Chinese rare minnows (Gobiocypris rarus) were exposed to benzophenone (BP), 2,4-dihydroxybenzophenone (BP-1), and 5-benzoyl-4-hydroxy-2-methoxybenzenesulfonic acid (BP-4) at 5, 50, 500 µg/L for 28 d to assess their toxicity. Transcriptomics screening showed that cell cycle, DNA replication and repair were significantly altered pathways (p < 0.05). The altered transcripts were similar to those identified by RNA-seq. DNA damage and 8-OHdG levels were significantly increased at 50 and 500 µg/L groups (p < 0.05). The DNA methylcytosine level was not significantly changed exposure to BP, BP-1 and BP-4. TUNEL assays indicated that hepatic apoptosis was significantly improved at 500 µg/L BP and BP-4 and 50 and 500 µg/L BP-1 (p < 0.05), with the significantly increasing the activity of caspase-3, -8 and -9 (p < 0.05). Molecular docking analysis revealed that BP, BP-1 and BP-4 could bind differently to caspase-3 through different binding interactions. Therefore, BP-1 induced more serious oxidative DNA damage and apoptosis by activating caspase-3 than BP and BP-4, which will provide theoretical basis and data support for ecological evaluation of aquatic organisms induced by BPs.

Toxicity of waterborne vortioxetine, a new antidepressant, in non-target aquatic organisms: From wonder to concern drugs?

Vortioxetine is increasing in popularity as a treatment for major depressive disorder and has been detected in wastewater effluent. However, information on the toxicity and environmental risk of vortioxetine in non-target organisms is scarce. Here, embryonic and juvenile zebrafish (Danio rerio) were used to assess the toxicity of vortioxetine (0, 1, 10, 30, 100, 300, and 1000 µg/L) after 120 h and 7 d of exposure, respectively. Vortioxetine induced significant toxicity during embryonic development, including effects on survival, hatching, basal heart rate, spontaneous tail coiling and developmental abnormalities, and inhibited larval locomotor activity at concentrations higher than 30 µg/L. Additionally, vortioxetine evoked anxiolytic-like behavior and caused histopathological changes to multiple organs (gills, heart, liver and intestine) in juvenile zebrafish. Significant increase in 5-HT content was observed in whole zebrafish larvae and juvenile brain tissues from animals treated with 1 or 100 µg/L vortioxetine. Notably, the lowest effective concentrations of vortioxetine for zebrafish were mainly in the range of 10-30 µg/L, which were slightly lower than the vortioxetine therapeutic concentrations. Risk quotients assuming conservative exposure assessments were above one in European countries indicating moderate risk for the behavioral endpoints assessed. We believe that these results highlight the adverse effects of vortioxetine on non-target organisms and that further investigations will be required to provide a higher confidence.

Identification of toxicity factors and causal analysis of toxicity in surface sediments from Liaohe river basin, Northeast China using an effect guidance strategy.

Sediments play a pivotal role in maintaining the aquatic ecological status of rivers. However, the determination of the key toxicants that consider the combined effects of all sediment-related contaminants are still challenging and necessary for an appropriate sediment risk assessment. The effects of sediments on aquatic organisms have been reported in Liaohe River, but their key toxicity factors are not well known. To determine the key toxicity factors, twenty-six surface sediment samples from Liaohe River tributaries in Northeast China were collected. Acute toxicity test of midge larvae results showed that 6 of 26 tributaries had obvious toxic effects, with survival rates of 37%-57% (p < 0.05). The masking test showed that the main pollutants in the surface sediments of T7 and T16 were metals, that of T8 was an organic pollutant, those of T19 and T26 were organic pollutants and ammonia, and those of T17 were heavy metal and ammonia. Chemical analysis showed that the relatively high concentrations of ammonia were only presented in surface sediments of T17, T19, and T26, with PTU of 1.5, 1.2 and 1.1, respectively, whereas heavy metals were markedly high in surface sediments from T7 and T16, with PTU of 0.92 and 0.61, respectively. Interestingly, the observed toxicity in surface sediments agreed with the toxicity predicted by chemical analysis Moreover, the significant correlation between the survival and volume ratio of the sediment and overlying water confirmed ammonia nitrogen was key toxicity factor in T17, T19, and T26, whereas Cu was the key toxicity factor in T7 that cause the biological toxicity. In conclusion, the major toxic factors of ammonia and copper in the sediments were identified. Moreover, our study suggested that effect guidance strategy was an effective method for sediment quality assessment.

Assessment of benthic invertebrate diversity and river ecological status along an urbanized gradient using environmental DNA metabarcoding and a traditional survey method.

Benthic invertebrate diversity is one of the most commonly used bioindicators for assessing aquatic ecosystem health in river systems. Although an increasing number of studies have focused on assessing benthic invertebrate diversity using environmental DNA metabarcoding and traditional survey methods, benthic invertebrate diversity and ecological status assessments performed across different landscapes within river systems have not been well documented. Here, the diversity and ecological status of benthic invertebrates and the influence of water quality on the invertebrate assemblage distribution along an urbanization gradient in rivers from the Jingjinji (JJJ) region, China, were investigated using eDNA metabarcoding and the traditional method. With the combination of the two methods, 395 benthic invertebrates from 6 phyla, 27 orders, 94 families, and 222 genera were identified. The species richness of the benthic invertebrate community in the mountain area was significantly higher than that in the urban and agricultural areas. Compared to the traditional results, eDNA metabarcoding obtained a significantly greater number of species from every sampling site (P = 0.000) and detected a notably higher abundance in Annelida (P = 0.000). Furthermore, the nonmetric multidimensional scaling (NMDS) and permutational multivariate analysis of variance (PERMANOVA) based on the Bray-Curtis dissimilarity index indicated that the benthic invertebrate communities from the different habitats were discriminated more accurately and easily using eDNA metabarcoding (P = 0.038) than with the traditional method (P = 0.829). Additionally, the assemblages identified by eDNA metabarcoding were more closely linked to water quality and could be realistically used to assess the ecological status of rivers. Our findings highlight that eDNA metabarcoding could represent a rapid and reliable method for estimating benthic invertebrate diversity and ecological status in river systems.

Estimating aquatic plant diversity and distribution in rivers from Jingjinji region, China, using environmental DNA metabarcoding and a traditional survey method.

Traditional survey methods (TSMs) are difficult to use to perform a census of aquatic plant diversity completely in river ecosystems, and improved aquatic plant community monitoring programs are becoming increasingly crucial with a continuous decline in diversity. Although environmental DNA (eDNA) metabarcoding has been applied successfully to assess aquatic biodiversity, limited work has been reported regarding aquatic plant diversity in rivers. In this study, the efficiency of eDNA to estimate the aquatic plant diversity and spatial distribution of rivers from the Jingjinji (JJJ) region was evaluated by comparing results obtained by the TSM. Based on a combination of the two methods, 157 aquatic plant species, including 24 hydrophytes, 61 amphibious plants, and 72 mesophytes, were identified. The spatial patterns in species richness and abundance by eDNA exhibited agreement with the TSM results with a gradual decline from the mountain area (MA) to the agricultural area (AA) and then to the urban area (UA). Compared to the TSM, eDNA identified a significantly greater number of species per site (p < 0.01) and obtained a significantly higher abundance in hydrophytes (p < 0.01), supplementing the unavailable abundance data from the TSM. Furthermore, the aquatic plant assemblages from the different areas were discriminated well using eDNA (p < 0.05), but they were better discriminated by the TSM (p < 0.01). Thus, our study provides more detailed data on aquatic plant diversity in rivers from the JJJ region, which is essential for biodiversity conservation. Our findings also highlight that eDNA can be reliable for evaluating aquatic plant diversity and has the potential to respond to landscape heterogeneity in river ecosystems.

Environmentally relevant concentrations of clozapine induced lipotoxicity and gut microbiota dysbiosis in Chinese rare minnow (Gobiocypris rarus).

Clozapine (CLZ) is a neuroactive pharmaceutical that is frequently detected in aquatic environments. Although the cardiotoxicity, developmental toxicity, and neurotoxicity of CLZ in aquatic non-target organisms have been reported, its lipotoxicity and underlying mechanism are unknown. Therefore, in this study, 2-month-old Chinese rare minnows were exposed to 0, 0.1, 1, and 10 µg/L CLZ for 90 days. Overt dyslipidemia was observed after CLZ exposure, whereas the body weights of females significantly increased after CLZ exposure (p < 0.05). In addition, obvious hepatocyte vacuolization and hepatic lipid droplet accumulation were observed at all treatment groups (p < 0.05). The activities of sterol regulatory element binding proteins 1 (SREBP1) and fatty acid synthase (FAS) were significantly upregulated at the 1 and 10 µg/L CLZ treatment groups (p < 0.05). Moreover, evident cell boundary disintegration of the intestinal villi and increasing mucus secretion were observed at all treatment groups (p < 0.05). Furthermore, the diversity of the gut microbiota increased, whereas the relative abundances of Proteobacteria, Firmicutes and Bacteroidetes significantly increased after CLZ exposure (p < 0.05). Furthermore, significantly increased bacterial secondary bile acid biosynthesis activity in Chinese rare minnows was observed after 1 µg/L CLZ exposure (p < 0.05). Therefore, our findings confirmed that CLZ induced lipotoxicity by stimulating SREBP1 and affecting the bacterial secondary bile acid biosynthesis activity in Chinese rare minnows.

A review on China's constructed wetlands in recent three decades: Application and practice.

Constructed wetlands (CWs) have been introduced to and developed in China for environmental engineering over the most prosperous three decades (1990-2020). To study the origin, development process, and future trend of CWs, this review summarized a wide range of literatures between 1990 and 2020 by Chinese authors. Firstly, the publication number over years, research highlights, and the author contributions with the most published papers in this field were conducted through bibliometric analysis. Secondly, the most principal components of CWs, substrates and macrophytes were summarized and analyzed. Thirdly, the typical application cases from traditional CWs, pond systems to combined pond-wetland systems were presented. In China, CWs were predominately distributed in the east of the so-called 'Hu Huanyong Line'. Therefore CWs were limited by the socio-economic level and climatic conditions. It is unquestionable that the overall level of China's CWs has improved significantly, and one of the most prominent features has started towards the plural pattern development. There has been a trend of large-scale or low-cost CW application in the recent years. However, lifecycle research and management are required for better strategies in the future.

Risks to aquatic environments posed by 14 pharmaceuticals as illustrated by their effects on zebrafish behaviour.

The presence of pharmaceutical residues in aquatic ecosystems is a worldwide problem that may pose serious threats and challenges to the environment, especially to the safety of aquatic biota. In the present study, we investigated the effects of 14 environmentally relevant pharmaceutical compounds on individual and collective-related behaviours in juvenile zebrafish (Danio rerio) for 21 days. The tested concentrations of the compounds spanned three orders of magnitude. This study also compared the potential risks of these compounds in Chinese surface waters based on the data on their toxic effects or only on behavioural effects. In the case of individual behaviours, most antidepressants, but not anti-inflammatory agents or blood lipid-lowering agents, decreased fish locomotor activity (LMA) and individual social activity (IDS); however, all three classes of compounds induced significant disruptions in the light/dark transition locomotor response (LMR-L/D) performance, even at lower treatment levels (0.1-1 µg/L). Furthermore, collective behaviour (CLB) analysis suggested that most of the compounds significantly altered the group sociability of fish and frequently occurred at environmentally relevant concentrations. Finally, a risk assessment suggested that the presence of ibuprofen, fluoxetine, and venlafaxine in the surface waters of China poses a relatively high risk to fish, regardless of the risk ranking based on the data of the toxic or behavioural effects.

Evaluating environmental impact of STP effluents on receiving water in Beijing by the joint use of chemical analysis and biomonitoring.

To evaluate the environmental impact of receiving water from the Qinghe River sewage treatment plant (STP) effluents in Beijing, we collected sediments and Bellamya aeruginosa (Up-site, Discharge-site, and Down-site) both in 2017 and 2018 and analyzed the samples via chemical analysis, biological responses and transcriptomics. In two years of data, our biological results showed that AChE activities presented different degrees of influence on B. aeruginosa captured at sampling points of the STP compared to control sites (P < 0.05). Additionally, indicators of the antioxidant system (e.g., SOD, CAT, GST, EROD activity) and MDA content were significantly increased in the whole tissue at the Up-site of the STP. Integration of the assessed biomarkers using the integrated biomarker response (IBR) index ranked the environmental impact at sites as Up-site > Discharge-site > Down-site. In terms of the transcriptome data, B. aeruginosa collected from the Discharge-site of the STP showed greater transcriptomic response than it did from all other sites. KEGG pathway analysis revealed that sewage significantly altered the expression of genes involved in xenobiotics by cytochrome P450, drug metabolism-cytochrome P450, glutathione metabolism, oxidative phosphorylation, citrate (TCA) cycle, glycolysis/gluconeogenesis, apoptotic and Parkinson's disease. The concentrations of 34 organic pollutants (17 PAHs, 10 PAEs, 7 EDCs) were measured. The chemical concentrations of pollutants decreased from Up-site to Down-site and were well correlated with enzyme activity, IBR, and transcriptomic results. Our results demonstrated that the combined use of chemical analysis, biological responses and transcriptome data is necessary to validate the efficacy of a battery of biomarkers chosen to detect environmental stress due to pollution.

Carbamazepine at environmentally relevant concentrations caused DNA damage and apoptosis in the liver of Chinese rare minnows (Gobiocypris rarus) by the Ras/Raf/ERK/p53 signaling pathway.

To assess genetoxicity and the underlying mechanisms of carbamazepine (CBZ) toxicity in fish, adult Chinese rare minnows (Gobiocypris rarus) were exposed to 1, 10, and 100 µg/L CBZ for 28 d. Comet assays indicated that hepatic DNA damage was significantly increased in groups of minnows exposed to CBZ at all concentrations in a dose-dependent manner compared to those of the control groups (p < 0.05). Liver levels of 8-hydroxydeoxyguanosine (8-OHdG) were significantly increased at 10 and 100 µg/L CBZ (p < 0.05). TUNEL assays indicated that the average apoptotic rates of the livers of female and male minnows were significantly increased following exposure to CBZ at all concentrations for 28 d (p < 0.05). Significant increases in caspase 3 and 9 activities after CBZ exposure at all concentrations and caspase 8 at 10 and 100 µg/L CBZ exposure reflected the presence of mitochondrial apoptosis (p < 0.05). The mRNA levels of gadd45a, mdm2, casp3 and casp9 in female and male minnows exposed to CBZ at all concentrations were significantly increased compared with those in the control groups (p < 0.05). Significant increases in the levels of p21 in female minnows exposed to 1 and 100 µg/L CBZ, p53 in female minnows at all CBZ treatments and bcl2 in male minnows exposed to 1 and 100 µg/L CBZ were observed, indicating p53 pathway activation. The inhibition of ras levels in females and males exposed to CBZ at all concentrations and increased levels of raf1 in males exposed to CBZ at all concentrations indicated Ras/Raf1/MAPK (ERK) activation. Therefore, the present study demonstrates that CBZ at environmentally relevant levels induces DNA damage and apoptosis in Chinese rare minnows by the Ras/Raf/ERK/p53 signaling pathway.

Environmentally relevant concentrations of carbamazepine induced lipid metabolism disorder of Chinese rare minnow (Gobiocypris rarus) in a gender-specific pattern.

Carbamazepine (CBZ), an anticonvulsant and mood stabilizer, is ubiquitous distributed in aquatic environment. Though the toxicity and endocrine disrupting effect of CBZ on non-target organisms have been studied, its lipotoxity are scarcely known. To assess the lipotoxicity of CBZ, 2-month-old Chinese rare minnow were exposed to 0, 1, 10, and 100 µg/L CBZ for 90 d. Obvious dyslipidemia was observed after 30 d and 90 d exposure, whereas overt hyperlipidemia was observed in males at 100 µg/L treatments. Severe lipid droplet accumulation in livers was observed at 10 and 100 µg/L treatments for 30 d and in females, whereas those was observed at all treatments in males. In addition, serious mitochondria damage was observed in males at 100 µg/L treatments. After 90 d exposure, the enzyme activities of FAS and ACCα were significantly increased at 10 and 100 µg/L treatments, whereas HMGCR were markedly increased at 100 µg/L treatments (p < 0.05). However, ACCß were markedly decreased in females at 10 and 100 µg/L treatments and in males at all treatments (p < 0.05). The transcription levels of fasn, accα, hmgcrα, fdft1, idi1, plin1, plin2, caveolin1, and caveolin2 were significantly increased at 100 µg/L treatments (p < 0.05). Moreover, the body weight was obviously increased at 10 and 100 µg/L treatments in males (p < 0.05). Our results confirmed that environmental relevant concentrations CBZ induced lipid metabolism disorder and mitochondria damage of Chinese rare minnow in a gender-specific pattern, which provided a new insight into the lipotoxicity mechanism of CBZ.

Environmentally relevant concentrations of bifenthrin induce changes in behaviour, biomarkers, histological characteristics, and the transcriptome in Corbicula fluminea.

Bifenthrin (BF) is an insecticide that is commonly used to control agricultural and domestic pests and is widespread in aquatic environments. Although previous studies have found that BF is toxic to aquatic organisms, such a comprehensive study of the mechanism of toxic effects in bivalves is not common. In this study, to assess the toxic effects of BF on bivalves, adult Corbicula fluminea (C. fluminea) were exposed to 0, 1, 5, and 25 µg/L BF for 15 days. Transcriptome analysis revealed that BF exposure significantly altered the expression of genes involved in detoxification, antioxidation, and metabolism. Moreover, the ROS content and GST activity at 25 µg/L treatments were significantly increased (p < 0.05), and significant increases of MDA concentration and CAT activity were observed at 5 and 25 µg/L treatments (p < 0.05). However, AChE activity was markedly inhibited at 25 µg/L treatments (p < 0.05). In addition, vacuolation in the digestive tubules and the hemolytic infiltration of connective tissue were observed at all treatments, and the degeneration of the digestive tubule was observed at 5 and 25 µg/L treatments. In the behavioural assay, the siphoning behaviour of C. fluminea was significantly inhibited at 25 µg/L treatments (p < 0.05), whereas no significant change in burrowing behaviour was observed. Our findings suggested that BF exposure caused changes in detoxification, antioxidation, and metabolism pathways, biomarker activity or concentrations and histopathological characteristics, resulting in changes in behaviour. Therefore, our findings provide a basis for further evaluation of the toxicity of pyrethroid insecticides in bivalves.

Comparison of the Toxicity Effects of Tris(1,3-dichloro-2-propyl)phosphate (TDCIPP) with Tributyl Phosphate (TNBP) Reveals the Mechanism of the Apoptosis Pathway in Asian Freshwater Clams (Corbicula fluminea).

To compare the toxicities of a chlorinated and a nonchlorinated organophosphorus flame retardant (OPFR) in this study, adult calms (Corbicula fluminea) were exposed to tris(1,3-dichloro-2-propyl)phosphate (TDCIPP) and tributyl phosphate (TNBP) at 20, 200, and 2000 µg/L for 30 days. Toxicity screening using transcriptomics indicated that the apoptosis pathway was significantly affected in the groups exposed to 2000 µg/L TDCIPP and TNBP (p ≤ 0.05), and this finding was further confirmed by the protein interaction network. The terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay suggested that TDCIPP and TNBP can cause apoptosis. The significant (p ≤ 0.05) increases in the activities of caspases 3 and 8 obtained with all treatments and in that of caspase 9 obtained with 2000 µg/L exposure treatments indicated the presence of mitochondria-dependent and mitochondria-independent apoptosis. Interestingly, a noticeable dose-dependent increase in DNA damage was observed in all treatments, resulting in apoptosis. Therefore, our results demonstrate that TDCIPP and TNBP induce DNA damage and apoptosis in C. fluminea, which indicates that these chemicals pose an ecological risk to benthic organisms. Moreover, through a similar mechanism of action in apoptosis, TDCIPP induced more serious toxicity than TNBP, which indicated that chlorination or differences in structure-specific metabolism could be key factors influencing toxicity.