Histological, physiological, and transcriptomic responses of hepatopancreas to air exposure in asian freshwater clam Corbicula fluminea.

Corbicula fluminea (C. fluminea) is an important freshwater economy shellfish in China, but it often suffers from air exposure during transportation. In this study, we investigated the histological, physiological (mainly including respiratory metabolism, antioxidant capacity, and immune function), and transcriptomic responses of hepatopancreas in C. fluminea to different times of air exposure. At histological level, air exposure caused vacuolation of digestive cells (24-96 h) and enlargement of digestive tubule lumen (6-96 h) in hepatopancreas. At physiological level, the activities of enzymes related to glycolysis (hexokinase and pyruvate kinase) and anaerobic respiration (lactate dehydrogenase) were increased first (6-24 h) of air exposure, then came back to normal level or even decreased. The activity of aerobic respiration-related enzyme (succinic dehydrogenase) began to reduce from 24 h of air exposure. The activities of antioxidant enzymes (superoxide dismutase and catalase) were enhanced during 6-48 h of air exposure and then returned to control level or even inhibited. The content of malondialdehyde (MDA) increased from 96 h of air exposure. The activities of immune-related enzymes (acid phosphatase and alkaline phosphatase) increased during 6-48 h, then returned to normal or began to decline. At transcriptome level, 44 differentially expressed genes (DEGs) in the hepatopancreas were identified after 96-h air exposure. Among these DEGs, 8 were associated with glycolysis, TCA cycle, immune, and antioxidant, and were downregulated after 96-h air exposure. Taken together, these findings illuminated the response of C. fluminea to air exposure at histological, physiological, and transcriptomic levels, which will be beneficial to the aquaculture and transportation of C. fluminea.

Fate and toxicity of legacy and novel brominated flame retardants in a sediment-water-clam system: Bioaccumulation, elimination, biotransformation and structural damage.

Due to the characteristics of persistent organic pollutants (POPs), some legacy brominated flame retardants (LBFRs) were prohibited from use, and then gradually replaced by novel brominated flame retardants (NBFRs). However, till now little research focused on the effects of NBFRs on the benthos. In the present study, 0.5, 5, and 50 mg/kg dw of pentabromotoluene (PBT), hexabromobenzene (HBB), 1,2-bis(2,4,6-tribromophenoxy) ethane (BTBPE), decabromodiphenyl ethane (DBDPE) and decabromodiphenyl ether (BDE209) were added into sediments to test freshwater clams (Corbicula fluminea). In the 35-day exposure experiment, C. fluminea had different enrichment behaviors in three treatment groups. It was conjectured that in the lower dose group, the clams ingested contaminants and tended to be stable over time. While in higher dose groups, the clams were induced by the chemicals, leading to the changes in physiological activities so that the concentrations showed a downward trend first and then went up. The half-lives of contaminants in freshwater clams were between 0.911 and 11.6 days. DBDPE showed stronger bioaccumulation ability than BDE209 in this study. Parabolic relationships were observed between log BSAF and log Kow values in clam tissues. Debromination, hydroxylation, and methoxylated products were detected. Additionally, the gill samples of C. fluminea exposed to 50 mg/kg dw of single substance were observed by scanning electron microscope (SEM), indicating that the adhesions, tissue hyperplasia, and messy cilia occurred on the surface. Our research potentially contributes to further evaluations of the environmental risks posed in sediments contaminated by PBT, HBB, BTBPE, DBDPE, and BDE209, particularly the benthic organisms.

Behaviors and biochemical responses of macroinvertebrate Corbicula fluminea to polystyrene microplastics.

Microplastic, a well-documented emerging contaminant, is widespread in aquatic environments resulting from the production and fragmentation of large plastics items. The knowledge about the chronic toxic effects and behavioral toxicity of microplastics, particularly on freshwater benthic macroinvertebrates, is limited. In this study, adult Asian clams (Corbicula fluminea) were exposed to gradient microplastic solutions for 42 days to evaluate behavioral toxicity and chronic biotoxicity. The results showed that microplastics caused behavior toxicity, oxidative stress, and tissue damage in high-concentration treatments. Siphoning, breathing, and excretion was significantly inhibited (p < 0.05) at high-concentration treatments, suggesting that high-concentration microplastics induced behavioral toxicity in C. fluminea. Malondialdehyde content, superoxide dismutase, catalase, and glutathione reductase activities were significantly enhanced (p < 0.05) and the acetylcholinesterase was significantly inhibited (p < 0.05) throughout the exposure period in high-concentration treatments. Enzymes associated with energy supply were significantly higher at high-concentration microplastics treatments on D7 and D21. However, they recovered to a normal level on D42. The instability of the enzymes indicated that high-concentration microplastics induced oxidative stress and disorder in neurotransmission and energy supply. The gills of C. fluminea in treatments underwent cilia degeneration, which indicated that microplastics caused tissue damage in the gills. The analysis of integrated biomarker response values revealed that high-concentration microplastics led to long-term effects on the health of C. fluminea. In conclusion, continuous exposure to microplastics (10 mg L-1) would damage physical behavior and the antioxidant system of C. fluminea.

Impacts of nanoplastics on bivalve: Fluorescence tracing of organ accumulation, oxidative stress and damage.

The outcomes of this research offer novel insights into the toxic effects of nanoparticles (i.e., nanoplastics or other nanomaterials) on the benthos. Herein, this study aimed to evaluate the accumulation pathway, distribution characteristics and potential biotoxicity of polystyrene nanoplastics in C. fluminea. The results revealed that nanoplastics could accumulate in the mantle through adherence, in the visceral mass through ingestion and in the gill through respiration. The gill, intestine and stomach were the main accumulation organs for nanoplastics. The aggregation of nanoplastics was observed in C. fluminea, which may exacerbate their biotoxicity. Moreover, oxidative stress was observed in the visceral mass, gill and mantle. Liver damage, neurotoxicity and intestinal inflammation were caused by imbalance in the antioxidation system. Analysis of IBR values showed that the visceral mass had a more effective response to oxidative stress than the gill and mantle after exposure to nanoplastics.

Halogen-free organophosphorus flame retardants caused oxidative stress and multixenobiotic resistance in Asian freshwater clams (Corbicula fluminea).

Halogen-free organophosphorus flame retardants are widespread in aquatic environments. Although it has been documented that they affect the behavior and reproduction of aquatic species, researches investigating cellular detoxification and the defense system in bivalves are scarce. In this study, adult Asian clams (C. fluminea) were exposed to tris (2-butoxyethyl) phosphate (TBEP) and tributyl phosphate (TBP) at 20, 200, and 2000 µg/L for 28 d. The results showed no noticeable difference in siphoning behavior. However, the siphoning behavior displayed a trend toward a slight decrease in the treatment groups. GR activity was markedly reduced compared with the control groups, whereas the levels of cyp4 significantly increased following the 2000 µg/L TBP treatments (p < 0.05). Moreover, the levels of gsts1 and gstm1 significantly decreased following all TBEP treatments and were significantly inhibited by 20 µg/L TBP (p < 0.05). The adverse effects on antioxidant enzymes suggested that C. fluminea mainly relies on the antioxidant system to reduce damage without an increase in MDA levels following exposure to a low concentration. Moreover, mRNA expression levels of heat shock proteins (hsp 22, 40, 60, 70, and 90) were significantly down-regulated with TBEP and TBP treatments lower than 200 µg/L (p < 0.05), whereas significant up-regulations were observed for hsp 22 and hsp 70 in response to 2000 µg/L TBP treatment (p < 0.05). Up-regulation of ATP-binding cassette (ABC) transporter genes (abcb1 and abcc1) showed that TBEP and TBP could activate the multixenobiotic resistance (MXR) system to discharge xenobiotics in C. fluminea, which kept its shell closed at high concentrations to prevent xenobiotic entry. Our results provide a new insight into the different mechanisms of cellular detoxification and the MXR system of C. fluminea in response to low and high concentrations of TBEP and TBP.

Effects of fluoxetine on behavior, antioxidant enzyme systems, and multixenobiotic resistance in the Asian clam Corbicula fluminea.

Fluoxetine (FLX), a selective serotonin reuptake inhibitor, is widespread in aquatic environments. Despite its reported effects on behavior and reproduction in aquatic species, little is known about the effects of FLX on cellular detoxification and defense system in bivalves. Here, the adult Asian clam (Corbicula fluminea) was exposed to 0.5, 5, and 50 µg L(-1) FLX for 30 d. Siphoning behavior was inhibited by treatment with 50 µg L(-1) FLX. Additionally, superoxide dismutase (SOD) activity in the gills and digestive glands significantly decreased (p<0.05) with 5 and 50 µg L(-1) FLX treatments, whereas catalase (CAT) activity and malondialdehyde (MDA) content markedly increased (p<0.05). Moreover, transcription of thioredoxin reductase (TR), glutathione peroxidase (GPx) and Glutathione S-transferase pi class (GSTpi) was significantly upregulated (p<0.05), whereas glutathione reductase (GR) was markedly downregulated (p<0.05). These findings suggest that FLX affects behavior and induces oxidative stress in C. fluminea. The downregulation of ATP-binding cassette (ABC) transportor genes (ABCB1, ABCC1 and ABCG2) transporter genes indicated that FLX might suppress the multixenobiotic resistance (MXR) system in C. fluminea. Our results provide new insights into the adverse effects of FLX on the cellular detoxification and MXR system of C. fluminea.

Dynamics of protein phosphatase gene expression in Corbicula fluminea exposed to microcystin-LR and to toxic Microcystis aeruginosa cells.

This study investigated the in vivo effects of microcystins on gene expression of several phosphoprotein phosphatases (PPP) in the freshwater clam Corbicula fluminea with two different exposure scenarios. Clams were exposed for 96 h to 5 µg L(-1) of dissolved microcystin-LR and the relative changes of gene expression of three different types of PPP (PPP1, 2 and 4) were analyzed by quantitative real-time PCR. The results showed a significant induction of PPP2 gene expression in the visceral mass. In contrast, the cyanotoxin did not cause any significant changes on PPP1 and PPP4 gene expression. Based on these results, we studied alterations in transcriptional patterns in parallel with enzymatic activity of C. fluminea for PPP2, induced by a Microcystis aeruginosa toxic strain (1 × 10(5) cells cm(-3)) during 96 h. The relative changes of gene expression and enzyme activity in visceral mass were analyzed by quantitative real-time PCR and colorimetric assays respectively. The clams exhibited a significant reduction of PPP2 activity with a concomitant enhancement of gene expression. Considering all the results we can conclude that the exposure to an ecologically relevant concentration of pure or intracellular microcystins (-LR) promoted an in vivo effect on PPP2 gene expression in C. fluminea.