Responses of antioxidant defenses in the clam Mactra veneriformis to 2,2′,4,4′‑tetrabromodiphenyl ether exposure

https://doi.org/10.1016/j.cbpc.2018.12.003Get rights and content

Highlights

  • BDE-47 was accumulated in a dose-dependent manner in M. veneriformis.

  • BDE-47 increased MDA content in a time- and dose-dependent manner.

  • SOD, CAT, and GPx responded to BDE-47 exposure with different order.

  • The correlation of mRNA expressions and enzymatic activities changed with exposure duration.

Abstract

Antioxidant enzymes play essential roles against oxidative stress caused by 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47), which is ubiquitous in marine environment and organisms. However, research on antioxidant responses to BDE-47 in marine bivalves is scarce. In this study, we identified the full-length cDNA of catalase (CAT), and glutathione peroxidase (GPx) in the clam Mactra veneriformis. Subsequently, the responses of CAT, GPx, and copper, zinc-superoxide dismutase (Cu, Zn-SOD) were investigated in the clams exposed to 0.1, 1, and 10 μg/L BDE-47 for 7 days, and then depurated in natural seawater for 3 days. MvCAT and MvGPx contained conserved sequences. The deduced amino acid sequences shared high similarity with CATs and GPxs in other mollusks. M. veneriformis accumulated BDE-47 in a dose-dependent manner and eliminated BDE-47 poorly. BDE-47 induced a time- and dose-dependent increase of malondialdehyde content. Both the dose and the duration had significant effect on mRNA expressions and activities of the three antioxidants. Cu, Zn-SOD responded to BDE-47 earlier than CAT and GPx. The antioxidant responses could recover after depuration. These results suggested that M. veneriformis could accumulate BDE-47 efficiently. Antioxidant enzymes were triggered to counter the oxidative stress generated by BDE-47. Cu, Zn-SOD acted as the first defense against oxidative stress, while CAT and GPx intervened later. This study is therefore helpful in understanding the antioxidant responses to PBDEs in marine bivalves.

Introduction

Polybrominated diphenyl ethers (PBDEs) are manufactured brominated flame retardants (BFRs), comprising 209 congeners. They have been used for many years in consumer products such as textiles, electronic devices, and furniture. PBDEs can readily enter the environment and become a ubiquitous pollution problem (Law et al., 2006). Due to the persistence, bioaccumulation, and toxicity, some PBDEs have been listed as persistent organic pollutants by the Stockholm Convention. The use of penta- and octa-BDEs was discontinued in Europe during 1998 and in the USA and Canada during 2004 (Stockholm Convention, 2018). However, PBDEs can still be detected in the environment in dust, air, water, sediment, and in organisms such as plants, fish, bivalves, and mammals (Jin et al., 2008; Krieger et al., 2016). 2,2′,4,4′-Tetrabromodiphenyl ether (BDE-47) is the most abundant congener of PBDEs detected in environments and organisms (Jin et al., 2008; Palacio-Cortés et al., 2017). As the ocean is the main destination of pollutants, marine pollution by PBDEs has attracted considerable attention worldwide, especially in China. In Laizhou Bay, where BFRs manufacturers are located, the PBDEs concentrations in sediments and in shellfish are as high as 1800 ng/g dry weight, and 720 ng/g lipid, respectively (Jin et al., 2008).

Bivalves are employed as bio-indicators of aquatic contamination due to their wide distribution and their ability to accumulate many types of contaminants. Shellfish monitoring programs have been launched throughout the world (Bricker et al., 2014). The biotransformation of pollutants and the physiological responses of shellfish have been studied to determine the environmental pollution status. An Asian Mussel Watch Program was implemented in 2003–2005, using mussels to monitor contamination by PBDEs and organochlorines in the coastal waters of Asian countries (Ramu et al., 2007). PBDEs concentrations in the mussels varied from 0.66 to 440 ng/g lipid wet weight. BDE-47, BDE-99, and BDE-100 were the dominant congeners among PBDEs. Higher concentrations of PBDEs were detected in samples from the coastal waters of Korea, China, and Philippines. Subsequently, the toxicological effects of PBDEs on mussels were investigated (Jiang et al., 2017). Mussels are attached to solid objects and are rarely distributed in tidal flat areas lacking substrates for attachment. Thus, other species are needed for biological monitoring in flat areas. Mactra veneriformis is an infaunal suspension-feeding bivalve with ubiquitous distribution in the tidal flats of China, Korea, and Japan. It is an economically important shellfish in China. M. veneriformis can efficiently accumulate pollutants and respond sensitively to pollutants (Wang et al., 2005; Fang et al., 2010; Shi et al., 2018). Studies on this species may be beneficial for national monitoring programs.

It is generally known that oxidative stress mediates pollutant-induced toxicity (Shenai-Tirodkar et al., 2017). Studies have demonstrated that PBDEs can induce the production of reactive oxygen species (ROS) in aquatic organisms (Jiang et al., 2017; Park et al., 2017). Antioxidant enzymes such as superoxide dismutase (SOD, EC1.15.1.1), catalase (CAT, EC1.11.1.6), and glutathione peroxidase (GPx, EC1.11.1.9) scavenge ROS and maintain metabolic homeostasis. SOD eliminates superoxide radicals by converting them into hydrogen peroxide (H2O2) and oxygen via dismutation. Subsequently, H2O2 can be removed by CAT and GPx. Due to their rapid and sensitive responses to pollutants, antioxidant enzymes have been proposed as early biomarkers of pollution. The responses of these antioxidant enzymes to PBDEs have been illustrated in aquatic animals. mRNA expressions of SOD and CAT were up-regulated as the response to BDE-47 in the freshwater bivalve Anodonta woodiana (Xia et al., 2016). In zebra mussel Dreissena polymorpha, SOD and GPx activities were induced by BDE-154, while CAT activity was depressed (Parolini et al., 2012). SOD, CAT, and CAT activities decreased with the increase in BDE-47 concentration in the rotifer Brachionus plicatilis (Jian et al., 2017). GPx activity showed dose-dependent response to BDE-47 in Mytilus galloprovincialis mussels (Vidal-Liñán et al., 2015). However, the responses of antioxidant enzymes to PBDEs, especially the transcriptional responses are yet to be fully explored in the marine bivalves. More information is required for the risk assessment of PBDEs in marine ecosystem.

In this study, M. veneriformis clams were exposed to BDE-47 for 7 days and then subjected to a depuration period for 3 days. The objectives of this study were (1) to illustrate the cascade responses of SOD, CAT, and GPx to BDE-47 over time and the recovery of these responses after depuration, (2) to elucidate the accumulation and elimination of BDE-47 in test clams, and (3) to explore the correlation between antioxidant responses and BDE-47 concentrations. It is helpful in understanding the antioxidant responses to PBDEs in bivalves and facilitating national monitoring programs.

Section snippets

Clams and BDE-47 exposure

Adult M. veneriformis were collected along Dongying coast and acclimated in our laboratory at 16 °C for 2 weeks before exposure to BDE-47. The number of these clams was about 2000. The biological characteristics of these clams were as follows: average shell length was 3.46 ± 0.24 cm, height was 3.06 ± 0.21 cm, width was 2.38 ± 0.32 cm, and wet mass was 12.45 ± 0.82 g. After acclimatization, healthy and regular-shaped clams were transferred to 30 aquariums with each aquarium filled with 30 L of

BDE-47 concentrations in seawater and clams

The concentration of BDE-47 in seawater in the control and solvent control groups was under the MDLs (Table 1). The actual BDE-47 concentrations in BDE-47 exposed groups were 66.40–75.64% of the nominal concentrations.

BDE-47 concentrations in the digestive glands of M. veneriformis in control and solvent control groups were under the MDLs (Table 1). BDE-47 concentration in clams increased as the concentrations in seawater increased. BDE-47 accumulation was maximal in clams in 10 μg/L BDE-47

Accumulation of BDE-47 in M. veneriformis

As the exposure concentration of BDE-47 increased, the accumulation of BDE-47 in M. veneriformis increased accordingly. The accumulation of BDE-47 in test clams depended on the exposure dose. Similar results were also reported in other aquatic species. After 7 days of exposure to 2, 4, and 15 μg/L BDE-47, the BDE-47 accumulation in M. galloprovincialis was 6858.18, 17,460.21, and 75,688.79 μg/kg ww, respectively (Vidal-Liñán et al., 2015). The BDE-47 accumulation in gammarid Gammarus pulex

Conclusions

The cloned sequences of MvCAT and MvGPx had characteristics of the corresponding gene family sequences. M. veneriformis could accumulate BDE-47 efficiently and depurate BDE-47 poorly. CAT, GPx, and Cu, Zn-SOD cooperated to combat the oxidative stress induced by BDE-47. The three antioxidants responded to BDE-47 at different time intervals and levels.

Conflict of interest

We declare that we have no conflicts of interest.

Acknowledgements

This study was supported by grants from the National Natural Science Foundation of China (No. 41206120, No. 31502191, and No. 31502173), the Development Plan of Science and Technology in Yantai (No. 2011062), the Ludong University Research Funding (No. 210-32040301), and the Innovative Project of Undergraduate (201710451328 and ld171133).

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