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This study evaluated the toxicity of cadmium (Cd) on the green mussel Perna viridis, aiming to identify toxicological endpoints and investigate its responses across physiological, bioenergetic, and biochemical parameters. The 96-hour LC 50 value for Cd in P. viridis was 3.03 ± 0.12 mg L -1 , with a 95% confidence interval of 2.35-3.91 mg L -1 . Chronic toxicity tests revealed a No Observable Effect Concentration (NOEC), Lowest Observable Effect Concentration (LOEC), and chronic toxicity values of 0.20, 0.37, and 0.29 mg L -1 , respectively. Cadmium accumulation in treated mussels increased 46- to 215-fold compared to the control group. Superoxide dismutase, catalase, glutathione S-transferase, and glutathione peroxidase levels in exposed mussels exhibited a significant increase compared to the control group. The redox index ratio, acetylcholinesterase activity, and lysosomal membrane stability decreased with increasing exposure concentrations. Levels of reduced and oxidized glutathione, glutathione reductase, lipid peroxidation, and metallothionein-like proteins increased in exposed mussels. Clearance rate, respiration rate, and excretion rate decreased in a dose-dependent manner. Protein, carbohydrate, and lipid levels decreased with increasing exposure concentration (p < 0.001). Mitochondrial electron transport system activity increased, while cellular energy allocation (p < 0.001) and scope for growth decreased in a dose-dependent manner (p < 0.01). The significant increase in antioxidants suggests heightened oxidative stress in mussels under Cd exposure. The physiological activities of the mussels were severely affected, ultimately leading to a reduced scope for growth. The toxicological data generated in this study contribute to the development of seawater quality criteria for the metal Cd., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Ocean acidification causes severe shell dissolution and threats the survival of marine molluscs. The periostracum in molluscs consists of macromolecules such as proteins and polysaccharides, and protects the inner shell layers from dissolution and microbial erosion. Moreover, it serves as the primary template for shell deposition. However, the chemical composition and formation mechanism of the periostracum is largely unknown. In this study, we applied transcriptomic, proteomics, physical, and chemical analysis to unravel the mysteries of the periostracum formation in the green mussel Perna viridis Linnaeus. FTIR analysis showed that the periostracum layer was an organic membrane mainly composed of polysaccharides, lipids, and proteins, similar to that of the shell matrix. Interestingly, the proteomic study identified components enriched in tyrosine and some enzymes that evolved in tyrosine oxidation, indicating that tyrosine oxidation might play an essential role in the periostracum formation. Moreover, comparative transcriptomics suggested that tyrosine-rich proteins were intensively synthesized in the periostracum groove. After being secreted, the periostracum proteins were gradually tanned by oxidation in the seawater, and the level of crosslink increased significantly as revealed by the ATR-FTIR. Our present study sheds light on the chemical composition and putative tanning mechanism of the periostracum layer in bivalve molluscs. SIGNIFICANCE: The periostracum layer, plays an essential role in the initiation of shell biomineralization, the protection of minerals from dissolution for molluscs and especially ocean acidification conditions in the changing global climate. However, the molecular mechanism underlying the periostracum formation is not fully understood. In this study, we revealed that the oxidation and cross-link of tyrosine-rich proteins by tyrosinase are involved in periostracum formation in the green mussel Perna viridis. This study provides some insights into the first step of mussel shell formation and the robust adaptation of P. viridis to diverse habitats. These findings also help to reveal the potential acclimation of bivalves to the projected acidifying seawater., Competing Interests: Declaration of competing interest The authors declare that they have no financial and non-financial competing interests., (Copyright © 2024. Published by Elsevier B.V.)

As a ubiquitous environmental pollutant in China, triazophos (TP) is known to have neurotoxicity, oxidative stress, and reproductive toxicity to mussels. To investigate the molecular mechanisms of TP toxicity, metabolic changes in the digestive glands of Perna viridis in different sexes were examined after treated with 35 μg/L TP. Notably, 158 significant different metabolites (SDMs) were detected in TP-treated mussels and more than half of the SDMs were lipids and lipid-like molecules, which suggested that TP disturbed the lipid metabolism of P. viridis. In addition, metabolites associated with neurotoxicity and reproductive disturbance were also detected in female and male mussels. Moreover, a larger number of SDMs were found in male mussels (120 SDMs) than females (99 SDMs), and 60 common metabolites exhibited consistent variation tendency and similar magnitude in both sexes. The metabolic alternations in female and male mussels displayed similar protective mechanisms and also sex-specific responses, male mussels were more sensitive to TP exposure. This research provided new data about the molecular mechanisms of TP toxicity and the gender specific changes in mussels after treated by chemicals., Competing Interests: Declaration of Competing Interest None., (Copyright © 2023. Published by Elsevier Inc.)

The occurrence of pathogenic bacteria has emerged as a plausible key component of summer mortalities in mussels. In the current research, four bacterial isolates retrieved from moribund Greenshell࣪ mussels, Perna canaliculus, from a previous summer mortality event, were tentatively identified as Vibrio and Photobacterium species using morpho-biochemical characterization and MALDI-TOF MS and confirmed as V. celticus, P. swingsii, P. rosenbergii, and P. proteolyticum using whole genome sequencing. These isolates were utilized in a laboratory challenge where mussels were injected with cell concentrations ranging from 105 to 109 CFU/mussel. Of the investigated isolates, P. swingsii induced the highest mortality. Additionally, results from quantitative polymerase chain reaction analysis, focusing on known virulence genes were detected in all isolates grown under laboratory conditions. Photobacterium rosenbergii and P. swingsii showed the highest expression levels of these virulence determinants. These results indicate that Photobacterium spp. could be a significant pathogen of P. canaliculus, with possible importance during summer mortality events. By implementing screening methods to detect and monitor Photobacterium concentrations in farmed mussel populations, a better understanding of the host-pathogen relationship can be obtained, aiding the development of a resilient industry in a changing environment., (© The Author(s) 2022. Published by Oxford University Press on behalf of FEMS.)

The New Zealand Greenshell™ mussel (Perna canaliculus) is an endemic bivalve species with cultural importance, that is harvested recreationally and commercially. However, production is currently hampered by increasing incidences of summer mortality in farmed and wild populations. While the causative factors for these mortality events are still unknown, it is believed that increasing seawater temperatures and pathogen loads are potentially at play. To improve our understanding of these processes, challenge experiments were conducted to investigate the combined effects of increased seawater temperature and Vibrio infection on the immune and metabolic responses of adult mussels. Biomarkers that measure the physiological response of mussels to multiple-stressors can be utilised to study resilience in a changing environment, and support efforts to strengthen biosecurity management. Mussels acclimated to two temperatures (16 °C and 24 °C) were injected with either autoclaved, filtered seawater (control) or Vibriosp. DO1 (infected). Then, haemolymph was sampled 24 h post-injection and analysed to quantify haemocyte immune responses (via flow-cytometry), antioxidant capacity (measured electrochemically) and metabolic responses (via gas chromatography-mass spectrometry) to bacterial infection. Both seawater temperature and injection type significantly influenced the immune and metabolite status of mussels. A lack of interaction effects between temperature and injection type indicated that the effects of Vibrio sp. 24 h post-infection were similar between seawater temperatures. Infected mussels had a higher proportion of dead haemocytes and lower overall haemocyte counts than uninfected controls. The proportion of haemocytes showing evidence of apoptosis was higher in mussels held at 24 °C compared with those held at 16 °C. The proportion of haemocytes producing reactive oxygen species did not differ between temperatures or injection treatments. Mussels held at 24 °C exhibited elevated levels of metabolites linked to the glycolysis pathway to support energy production. The saccharopin-lysine pathway metabolites were also increased in these mussels, indicating the role of lysine metabolism. A decrease in metabolic activity (decreases in BCAAs, GABA, urea cycle metabolites, oxidative stress metabolites) was largely seen in mussels injected with Vibrio sp. Itaconate increased as seen in previous studies, suggesting that antimicrobial activity may have been activated in infected mussels. This study highlights the complex nature of immune and metabolic responses in mussels exposed to multiple stressors and gives an insight into Vibrio sp. infection mechanisms at different seawater temperatures., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Some marine organisms can resist to aqueous tidal environments and adhere tightly on wet surface. This behavior has raised increasing attention for potential applications in medicine, biomaterials, and tissue engineering. In mussels, adhesive forces to the rock are the resultant of proteinic fibrous formations called byssus. We present the solution structure of Pvfp-5β, one of the three byssal plaque proteins secreted by the Asian green mussel Perna viridis, and the component responsible for initiating interactions with the substrate. We demonstrate that Pvfp-5β has a stably folded structure in agreement with the presence in the sequence of two EGF motifs. The structure is highly rigid except for a few residues affected by slow local motions in the µs-ms time scale, and differs from the model calculated by artificial intelligence methods for the relative orientation of the EGF modules, which is something where computational methods still underperform. We also show that Pvfp-5β is able to coacervate even with no DOPA modification, giving thus insights both for understanding the adhesion mechanism of adhesive mussel proteins, and developing of biomaterials., (© 2022. The Author(s).)

Microplastics (MPs) can be defined as small pieces of plastics that are less than five millimetres in diameter. MPs can be consumed and may be accumulated by filter-feeding organisms such as mussels. Therefore, this study aimed to determine the acute effects of different types, sizes and concentrations of artificially synthesized MPs on the mortality rate and MP accumulation of the green mussel Perna viridis. The samples were exposed to 66, 333, 666, and 1333 items/L of small MPs (<30 μm), medium MPs (30-300 μm), and large MPs (300-1000 μm) polystyrene (PS), polypropylene (PP), and polybutylene succinate (PBS) for 96 h. MPs accumulation in the soft tissue of mussels and mortality effects from MPs ingestion were assessed. There was no mortality observed in the control group. Small PP particles can lead to more mortality than PS and PBS particles of the same size. However, medium- and large PS caused a higher mortality percentage than the same size particles of PP and PBS. Large PS, PP, and PBS showed higher mortality potential than other sizes. MPs largely accumulated in the soft tissues rather than in gill tissues following the 96-hour exposure period. Increased accumulation of the three types of MPs was accompanied by an increase in the percentage of mussel mortality. The study highlights how particle size and type are key factors in plastic particulate toxicity., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The present study has investigated the distribution of microplastics in sediment and its impact on histological, ultrastructural, and oxidative stress mechanisms in Perna viridis (P. viridis) from Kasimedu, Chennai, India. The results confirmed that fibers were the predominant type of microplastics observed, followed by spheres, flakes, sheets, and fragments. The observed microplastics were confirmed as polyester, polypropylene, polyethylene, cellophane, and rayon using μ-FT-IR. Microplastic particles entangled in gills caused abrasion of ciliated structure and hemocyte infiltration in the hemolymph vessels. The digestive gland showed a shrunken nucleus, dark inclusions, and damage in the nucleoid core structure. Enlarged vacuoles and the presence of clusters of vesicles presumably represented the transformed golgi cisternae. Further, the results confirmed that oxidative stress markers were significantly high in gills and digestive diverticula of P. viridis. Overall, the results indicated that microplastics induced different toxic physiological and structural alterations in gills and digestive diverticula of P. viridis. These findings highlighted the necessity to focus on exposure studies to understand the absolute magnitude of the problem due to microplastic pollution in the urban estuarine ecosystems of Chennai, Tamil Nadu, India., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Diarrheal shellfish toxins (DSTs) are among the most widely distributed phytotoxins, and are associated with diarrheal shellfish poisoning (DSP) events in human beings all over the world. Therefore, it is urgent and necessary to identify an effective method for toxin removal in bivalves. In this paper, we found that curcumin (CUR), a phytopolylphenol pigment, can inhibit the accumulation of DSTs (okadaic acid-eq) in the digestive gland of Perna viridis after Prorocentrum lima exposure. qPCR results demonstrated that CUR inhibited the induction of DSTs on the aryl hydrocarbon receptor (AhR), hormone receptor 96 (HR96) and CYP3A4 mRNA, indicating that the CUR-induced reduction in DSTs may be correlated with the inhibition of transcriptional induction of AhR, HR96 and CYP3A4. The histological examination showed that P. lima cells caused severe damage to the digestive gland of P. viridis , and the addition of curcumin effectively alleviated the damage induced by P. lima . In conclusion, our findings provide a potential method for the effective removal of toxins from DST-contaminated shellfish.

Background: The New Zealand Green-lipped mussel industry is well-established providing vastly to aquaculture exports. To assess mussel health and reproduction status, visual examination of organs and/or collection of haemolymph is commonly applied. Anesthetics, such as magnesium chloride (MgCl 2 ) can be utilized to prevent muscle contraction and keep shells open during sampling. The specific effects of muscle relaxing agents on baseline metabolism in invertebrates is unknown, but it is evident that molecular, cellular and physiological parameters are altered with these chemical applications. To this end, metabolomics approaches can help elucidate the effects of relaxing agents for better assessment of their use as a research tool., Methods: Adult Green-lipped mussels were anaesthetized for 3 h in a MgCl 2 bath, whereafter haemolymph samples were collected and analyzed via gas chromatography-mass spectrometry applying methyl chloroformate alkylation derivatization., Results: Anesthetized mussels were characterized as non-responsive to manual manipulation, with open valves, and limited siphoning function. Metabolite profiling revealed significant increases in the abundances of most metabolites with an array of metabolic activities affected, resulting in an energy imbalance driven by anaerobic metabolism with altered amino acids acting as neurotransmitters and osmolytes., Conclusion: This research is the first to use a metabolomics approach to identify the metabolic consequences of this commonly used bivalve relaxing technique. Ultimately the use of MgCl 2 anesthetization as a sampling strategy should be carefully evaluated and managed when performing metabolomics-related research., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Diarrhetic shellfish toxins (DSTs), some of the most important phycotoxins, are distributed almost all over the world, posing a great threat to human health through the food chain. Therefore, it is of great significance to find effective methods to reduce toxin accumulation in shellfish. In this paper, we observed the effects of four phytochemicals including cinnamaldehyde (CA), quercetin, oridonin and allicin on the accumulation of DSTs in the digestive gland of Perna viridis after exposure to the DSTs-producing Prorocentrum lima . We found that, among the four phytochemicals, CA could effectively decrease the accumulation of DSTs (okadaic acid-eq) in the digestive gland of P. viridis . Further evidence demonstrated that CA could reduce the histological alterations of the digestive gland of a mussel caused by DSTs. RT-qPCR showed that CA could suppress the CYP3A4 induction by DSTs, suggesting that the DSTs' decrease induced by CA might be related to the inhibition of CYP3A4 transcription induction. However, further studies on the underlying mechanism, optimal treatment time, ecological safety and cost should be addressed before cinnamaldehyde is used to decrease the accumulation of DSTs in field.

Biological fouling is an unwanted phenomenon that results in economic losses to the shipping industry. To prevent fouling, antifouling paints are used. DCOIT (4,5- dichloro-2-n-octyl-4-isothiazolin-3-one) is a biocide present in many antifouling paint formulations, and is toxic to a wide range of organisms. The aim of the present study was to evaluate the effects of DCOIT on oxidative stress indicators of the brown mussel, Perna perna. Molecular (SOD-like, GSTO-like and MGST-like mRNA levels) and biochemical (activities of superoxide dismutase (SOD), catalase (CAT) and glutathione S-transferase (GST), and levels of glutathione (GSH), reactive oxygen species (ROS) and protein carbonyls (PCO)) components were evaluated. Further, levels of biomarkers were assessed in the gills and digestive glands of mussels. Bivalves were exposed to DCOIT (control, 0.1 μg/L and 10 μg/L) for up to 96 h. DCOIT exposure decreased GSH content in gills. Moreover, exposure to DCOIT also decreased CAT activity in the gills and digestive glands of mussels. GST activity increased in digestive gland after exposure for 24 h to both concentrations of DCOIT tested. SOD activity, ROS levels and PCO content were not affected by exposure to the contaminant. Regarding the molecular biomarkers evaluated, DCOIT exposure altered mRNA levels of SOD-like in both tissues after 24 and 96 h of exposure, and decreased MGST-like mRNA levels in the digestive gland after 96 h of exposure to the chemical. These findings suggested that exposure to DCOIT may alter the biochemical and molecular functioning of P. perna, which may harm the species., (Copyright © 2020. Published by Elsevier Inc.)

Increasing water temperatures due to climate change have resulted in more frequent high mortality events of New Zealand Greenshell™ mussels (Perna canaliculus Gmelin 1791). These events have significant impacts within mussel farms which support a major shellfish industry for New Zealand. The present study investigates metabolic responses of farmed mussels during a summer mortality event in order to identify health impacts and elucidate mechanistic effects of external stressors on mussels. A gas chromatography-mass spectrometry (GC-MS)-based metabolomics approach was used to identify metabolic perturbations and flow cytometry assays were used to assess viability, oxidative stress and apoptosis of haemocytes from healthy and unhealthy mussels during a summer mortality event. The results showed significantly higher mortality and apoptosis of haemocytes in unhealthy mussels compared to healthy mussels. Reactive oxygen species (ROS) production, which is an indicator of oxidative stress was very high in both mussel groups, but no differences were observed between the two mussel groups. Metabolomics revealed alterations of many metabolites in both haemolymph and hepatopancreas (digestive gland) of unhealthy mussels compared to healthy mussels, reflecting perturbations in several molecular pathways, including energy metabolism, amino acid metabolism, protein degradation/tissue damage and oxidative stress. An increased level of itaconic acid which is an antimicrobial metabolite and biomarker of pathogen infection was observed in haemolymph, but not in hepatopancreas samples. This investigation provides the first detailed metabolic characterization of mussel immune responses to a summer mortality event and illustrates the benefits of using an integrated metabolomics and flow cytometry workflow for mussel health assessment and biomarker identification for summer mortality early detection., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Microplastics (MPs) are of increasing concern for filter feeding marine and freshwater species. Additionally MPs can sorb hydrophobic contaminants from the water, potentially providing an additional pathway of exposure of aquatic species to contaminants. An acute 48 h laboratory study was conducted to investigate the effects of microplastics and triclosan, both individually and combined, on New Zealand's green-lipped mussel, Perna canaliculus. Biomarkers included clearance rate, oxygen uptake, byssus production; and superoxide dismutase (SOD) activity, glutathione-S-transferase (GST) activity and lipid peroxidation in the gill tissue. Microplastics and triclosan, both individually and combined significantly decreased oxygen uptake and byssus production. These physiological responses were not observed when the microplastics were spiked with triclosan. Triclosan, both alone and spiked to microplastics, increased mussel oxidative stress markers including SOD activity and lipid peroxidation. An enhanced effect was observed on the SOD enzyme activity when mussels were exposed to microplastics spiked with triclosan. No effects on the biochemical biomarkers were observed for mussels exposed to microplastic only. Microplastics enhanced the uptake of triclosan in mussel tissue compared with triclosan only treatments indicating that microplastics potentially provide an additional pathway of exposure to hydrophobic contaminants., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020. Published by Elsevier Inc.)

Gills are considered a key player in organism defenses against environmental pollutants. Since it is the major site of uptake of waterborne chemicals, the modulation of important cellular defenses is expected in this tissue. Chlorothalonil, a fungicide presented in herbicides and antifouling paints, might be responsible for toxicity in marine biota. In this context, mussels were exposed to 0.1 μgL -1 and 10 μgL -1 of chlorothalonil for 24 h and 96 h. Genes from biotransformation and antioxidant defense pathways were investigated. Overall, we report, for the first time, an increase in the transcripts of the AhR-like, SULT1A1-like, CYP1A2-like, GSTO-like, MGST-like and SOD-like genes in the gills of the brown mussel Perna perna. This up-regulation was observed mostly after 96 h of exposure to chlorothalonil. Those results reinforce the important role of gills in xenobiotic metabolism and suggest the involvement of the mentioned genes in the detoxification of the compound. Throughout biotransformation and antioxidant defenses pathway, mussels exposed to chlorothalonil are activating mechanisms of defense against this contaminant., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Diarrheic shellfish poisoning (DSP) toxins are produced by harmful microalgae and accumulate in bivalve mollusks, causing various toxicity. These toxic effects appear to abate with increasing DSP concentration and longer exposure time, however, the underlying mechanisms remain unclear. To explore the underlying molecular mechanisms, de novo transcriptome analysis of the digestive gland of Perna viridis was performed after Prorocentrum lima exposure. RNA-seq analysis showed that 1886 and 237 genes were up- and down-regulated, respectively after 6 h exposure to P. lima, while 265 genes were up-regulated and 217 genes were down-regulated after 96 h compared to the control. These differentially expressed genes mainly involved in Nrf2 signing pathways, immune stress, apoptosis and cytoskeleton, etc. Combined with qPCR results, we speculated that the mussel P. viridis might mainly rely on glutathione S-transferase (GST) and ABC transporters to counteract DSP toxins during short-term exposure. However, longer exposure of P. lima could activate the Nrf2 signaling pathway and inhibitors of apoptosis protein (IAP), which in turn reduced the damage of DSP toxins to the mussel. DSP toxins could induce cytoskeleton destabilization and had some negative impact on the immune system of bivalves. Collectively, our findings uncovered the crucial molecular mechanisms and the regulatory metabolic nodes that underpin the defense mechanism of bivalves against DSP toxins and also advanced our current understanding of bivalve defense mechanisms., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Polybrominated diphenyl ether (PBDE) contamination has become a major concern over the effects on human health. In the present study, we collected widely consumed green mussels (Perna viridis) samples from the northern South China Sea (NSCS) to investigate the occurrence, spatial distribution, congener profiles as well as potential risk of 18 PBDEs. All the target PBDEs were detected in green mussel samples, indicating their ubiquitous distribution. The concentrations of the total 18 PBDE S (ΣPBDEs) in all samples varied from 6.96 to 55.6 ng/g lipid weight (lw), with BDE-47 and BDE-209 being the predominant PBDE congeners. Overall, the ΣPBDEs pollution in green mussels from NSCS was at a moderate to high level in comparison with the PBDEs pollution worldwide. The dietary exposure of the local population in South China to PBDEs via consuming green mussels was estimated to be 0.30-0.80 ng/kg body weight (bw)/day. Evaluation of the exposure risk for BDE-47, 99, 153 and 209 indicated that health risks due to green mussel consumption are substantially lower than the U.S. EPA minimum concern level., (Copyright © 2019 Elsevier B.V. All rights reserved.)

It is well documented that diarrhetic shellfish poisoning (DSP) toxins have strong genetic toxicity, cytotoxicity and oxidative damage to bivalve species. However, these toxic effects seem to decrease with the extension of exposure time and the increment of the toxin concentration, the mechanism involved remained unclear, though. In this paper, we found that expression of the genes related to cytoskeleton and Nrf2 signaling pathway displayed different changes over time in the gill of Perna viridis after exposure to DSP toxins-producing microalga Prorocentrum lima. During the short-term exposure (3 h and 6 h), KEAP1 gene expression was significantly up-regulated, coupled with up-regulation of MRP, ABCB1 and CAT transcriptions and down-regulation of GPx1 and NQO1 mRNA. After longer exposure to high density of P. lima, Nrf2 was significantly up-regulated, accompanied with up-regulation of Nrf2 pathway related genes such as NQO1, SOD, GST-ω and ABCB1, whereas KEAP1 was down-regulated. TUBA1C and TUBB1 transcripts were significantly down-regulated after short-term exposure of P. lima, but both of them were up-regulated at 96 h after exposure to high density of P. lima. Paraffin section demonstrated that P. lima had a strong damage on the gill of mussels during the short-term exposure. However, the negative effect to the gill decreased, and the gill restored after longer exposure (96 h). Taking together, we proposed that P. lima had a negative impact on cytoskeleton of mussel gill tissue, could cause oxidative damage to the gills. However, longer exposure of P. lima in high density could activate Nrf2 signaling pathway, thereby reducing the influence of toxin on mussel. Our study might provide a novel clue for the resistance mechanism of shellfish to DSP toxins., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Introduction: Itaconic acid (ITA) has recently been identified as an antimicrobial metabolite in mammalian immune cells. The presence of ITA was also reported in different tissues of marine molluscs, indicating its role as an endogenous metabolite of molluscs. In addition, the accumulation of ITA has been observed in different tissues of mussels following pathogen challenges. However, the concentration of ITA in mussel tissues and the possible role of this metabolite in the molluscan innate immune system remain unknown., Objectives: This study aims to quantitatively measure ITA levels in different tissues of marine mussels following an experimental challenge with Vibrio sp. DO1 isolate, and to identify the antimicrobial role of ITA in the innate immune system through the measurement of metabolic and immune alterations in tissues following the challenge., Methods: In this study, adult Perna canaliculus mussels were experimentally challenged with a pathogenic Vibrio sp. DO1 isolate. The metabolite profiles of five different tissues, including mantle, gill, muscle, hepatopancreas and haemolymph were obtained, and levels of ITA in each tissue were characterized using a gas chromatography-mass spectrometry (GC-MS) metabolomics approach. Flow cytometry was also employed to measure cell health parameters, including oxidative stress via reactive oxygen species (ROS) production, apoptosis via changes in mitochondrial membrane potential (MMP) and haemocyte viability., Results: The ITA levels in mantle, gill, muscle and hepatopancreas tissues at 18-h post infection (hpi) with Vibrio sp. were 40.31, 41.71, 11.61 and 41.66 ng mg -1 , respectively. In haemolymph, the level of ITA was significantly increased from 95.25 ng ml -1 at 6 hpi to 174.36 ng ml -1 at 18 hpi and 572.12 ng ml -1 at 60 hpi. In line with the accumulation of ITA, we observed increased levels of metabolites within the tricarboxylic acid (TCA) cycle, anti-inflammatory metabolites and alterations of other metabolites associated with immune responses of the host. The flow cytometry analyses revealed increases in ROS production, apoptotic cells and decreases in cell viability., Conclusions: We reported on the production of ITA in different tissues of P. canaliculus mussels challenged with a marine pathogen which confirmed ITA as an antimicrobial metabolite. The findings revealed insights into the biosynthesis of ITA and suggests its role in antimicrobial and anti-inflammatory activities in the innate immune system. This study also provided insights into the innate immune system of bivalves and highlighted the potential use of ITA as a biomarker for shellfish health assessment in aquaculture.

The trace elemental composition of biogenic calcium carbonate (CaCO 3 ) structures is thought to reflect environmental conditions at their time of formation. As CaCO 3 structures such as shell are deposited incrementally, sequential analysis of these structures allows reconstructions of animal movements. However, variation driven by genetics or ontogeny may interact with the environment to influence CaCO 3 composition. This study examined how genetics, ontogeny, and the environment influence shell composition of the bivalve Perna canaliculus. We cultured genetically distinct families at two sites in situ and in the laboratory. Analyses were performed on shell formed immediately prior to harvest on all animals as well as on shell formed early in life only on animals grown in the laboratory. Discriminant analysis using 8 elements (Co, Ti, Li, Sr, Mn, Ba, Mg, Pb, Ci, Ni) classified 80% of individuals grown in situ to their family and 92% to growth site. Generalised linear models showed genetics influenced all elements, and ontogeny affected seven of eight elements. This demonstrates that although genetics and ontogeny influence shell composition, environmental factors dominate. The location at which shell material formed can be identified if environmental differences exist. Where no environmental differences exist, genetically isolated populations can still be identified.

The Byssus, which is derived from the foot gland of mussels, has been proved to bind heavy metals effectively, but few studies have focused on the molecular mechanisms behind the accumulation of heavy metals by the byssus. In this study, we integrated high-throughput transcriptome and proteome sequencing to construct a comprehensive protein database for the byssus of Chinese green mussel (Perna viridis), aiming at providing novel insights into the molecular mechanisms by which the byssus binds to heavy metals. Illumina transcriptome sequencing generated a total of 55,670,668 reads. After filtration, we obtained 53,047,718 clean reads and subjected them to de novo assembly using Trinity software. Finally, we annotated 73,264 unigenes and predicted a total of 34,298 protein coding sequences. Moreover, byssal samples were analyzed by proteome sequencing, with the translated protein database from the foot transcriptome as the reference for further prediction of byssal proteins. We eventually determined 187 protein sequences in the byssus, of which 181 proteins are reported for the first time. Interestingly, we observed that many of these byssal proteins are rich in histidine or cysteine residues, which may contribute to the byssal accumulation of heavy metals. Finally, we picked one representative protein, Pvfp-5-1, for recombinant protein synthesis and experimental verification of its efficient binding to cadmium (Cd2+) ions., Competing Interests: The authors have declared that no competing interests exist.

Concerns are growing about the presence of fluoxetine (FLX) in environmental matrices, as well as its harmful effects on non-target organisms. FLX in aquatic ecosystems has been detected in a range varying from pg/L to ng/L, while adverse effects have been reported in several organisms inhabiting freshwater and marine environments. The present study quantifies FLX concentrations in seawater samples from Santos Bay, Brazil and assesses metabolic responses and sublethal effects on the tropical brown mussel Perna perna. Levels of ethoxyresorufin‑O‑deethylase, dibenzylfluorescein dealkylase, glutathione S-transferase, glutathione peroxidase, cholinesterase, lipoperoxidation, and DNA damage were assessed in the gills and digestive gland of these animals, and lysosomal membrane stability was also assessed in hemocytes. FLX altered phase I and II enzyme activities, caused cytogenotoxic effects, and negatively impacted the overall health of mussels exposed to environmentally relevant concentrations. These findings contribute to characterize the risks of introducing this drug into the marine environment., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The Asian green mussel, Perna viridis is a nutritious health food in the estuarine and coastal sea beds of the Arabian Sea along the west coast of India. In the present study, bioactivity-guided purification of the chloroform fraction of the methanolic extract of P. viridis was carried out. The isolated secondary metabolites were characterized by spectroscopic experiments, and their antioxidative/antiinflammatory properties were evaluated. The titled compounds were characterized as 3-hydroxy-13-vinyl-dodecahydro-11-phenanthrenone (1), 4,4,9-trimethyl-13-vinyl-dodecahydro-2-phenanthrenone (2), 11,20-dihydroxy-6,6-dimethyl-decahydro-5H-benzo[h]naphtho[1,2-c]chromene-16-carbaldehyde (3), 16-acetyl-20-hydroxy-6,6-dimethyl-dodecahydro-5H-benzo[h]naphtho[1,2-c]chromen-12-one (4), cholest-5-en-3β-3-yl-(30-hydroxy-3-methyl-36-methyleneundeca-30E,34E-dienoate) (5), and cholest-5-en-3β-3-yl-((E)-33-oxooct-31-enoate) (6). No significant differences in the antioxidant activities of the compounds with chromene-16-carbaldehyde (3) and chromen-12-one (4) functionalities (IC 50 0.52-0.68 mg/ml) vis-à-vis the positive control, α-tocopherol (IC 50 0.65-0.76 mg/ml) were registered. The studied compounds, 1-4, displayed potential antiinflammatory activities against pro-inflammatory 5-lipoxygenase (5-LOX) (IC 50  < 1 mg/ml). The balanced hydrophilic-lipophilic properties and lower steric values of the studied compounds, 1-4, were correlated with their bioactive potentials. PRACTICAL APPLICATIONS: The edible bivalve green mussels, P. viridis, are broadly available in the estuarine and coastal regions of the Indian Peninsula. The sequential chromatographic purification of the chloroform fraction of the methanolic extract of P. viridis led to the identification of six pure secondary metabolites. The metabolites with substituted chromene-16-carbaldehyde and chromen-12-one functionalities displayed potential antioxidative and antiinflammatory activities compared to other studied compounds. These bioactive metabolites could be used in functional food formulations and as antioxidant leads in medicinal food applications., (© 2018 Wiley Periodicals, Inc.)

Blooms of Aureococcus anophagefferens, referred to as brown tides are responsible for massive mortalities and recruitment failure of some bivalves. However, the molecular mechanisms underlying the toxicity remain elusive despite its biological significance, and the information currently available on the molecular effects is still insufficient. In this study, to evaluate the toxicity and associated mechanism of A. anophagefferens on bivalves, we analyzed the protein expression profiles in digestive glands of the A. anophagefferens-exposed Perna viridis by using iTRAQ. A total of 3138 proteins were identified in the digestive glands of A. anophagefferens-exposed P. viridis based on iTRAQ. Amongst, a repertoire of 236 proteins involved in cell, cell part, catalytic activity, metabolic process, biological regulation, immune system process, and response to stimulus were found to be differentially expressed. Functional analysis of the differentially expressed proteins demonstrated that innate immune system of P. viridis was activated, and some proteins associated with stress response and lipid metabolism were induced after exposure to A. anophagefferens. Additionally, MDA content, SOD activity and GSH-Px activity was increased significantly in the digestive gland of A. anophagefferens-exposed P. viridis. Taken together, our results indicated that the A. anophagefferens could induce oxidative stress, activate complement system and alter fat acid metabolism of P. viridis., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Contamination of toxic metals in P. viridis mussels has been prevalently reported; hence, health risk assessment for consuming this aquaculture product as well as the surrounding surface seawater at its harvesting sites appears relevant. Since Kampung Pasir Puteh, Pasir Gudang is the major harvesting site in Malaysia, and because the last heavy metal assessment was done in 2009, its current status remains unclear. Herein, flame atomic absorption spectrometry and flow injection mercury/hydride system were used to determine the concentrations of Pb, Cd, Cu and total Hg in P. viridis mussels and surface seawater (January-March 2015), respectively. Significantly higher concentrations of these metals were found in P. viridis mussels (p < 0.05) than that of surface seawater samples. The concentrations for Pb (4.27-6.55 μg/g) and Cd (1.55-2.21 μg/g) in P. viridis mussels exceeded the maximum permitted proportion prescribed by the Malaysian law. The concentrations of all metals in surface seawater also violated the Malaysia Marine Water Quality Criteria and Standards. Significant (p < 0.05) and high strength of association (r = 0.787) observed between Pb concentration in P. viridis mussel with the surface seawater indicates its possible application for inferring Pb concentrations in the mussel. Since both the calculated target hazard quotient and hazard index for Pb and Cd exceeded 1, the possible detrimental health impacts on human for consuming P. viridis mussels from this rearing site cannot be ignored. Hence, promoting continuous monitoring programmes and developing efficient toxic metal removal techniques prior to entering the market are required.

Massive mortalities due to pathogens are routinely reported in bivalve cultivation that have significant economic consequences for the global aquaculture industry. However, host-pathogen interactions and infection mechanisms that mediate these interactions are poorly understood. In addition, gender-specific immunological responses have been reported for some species, but the reasons for such differences have not been elucidated. In this study, we used a GC/MS-based metabolomics platform and flow cytometry approach to characterize metabolic and immunological responses in haemolymph of male and female mussels (Perna canaliculus) experimentally infected with Vibrio sp. Sex-based differences in immunological responses were identified, with male mussels displaying higher mortality, oxidative stress and apoptosis after pathogen exposure. However, central metabolic processes appeared to be similar between sexes at 24 h post injection with Vibrio sp. DO1. Significant alterations in relative levels of 37 metabolites were detected between infected and uninfected mussels. These metabolites are involved in major perturbations on the host's innate immune system. In addition, there were alterations of seven metabolites in profiles of mussels sampled on the second day and mussels that survived six days after exposure. These metabolites include itaconic acid, isoleucine, phenylalanine, creatinine, malonic acid, glutaric acid and hydroxyproline. Among these, itaconic acid has the potential to be an important biomarker for Vibrio sp. DO1 infection. These findings provide new insights on the mechanistic relationship between a bivalve host and a pathogenic bacterium and highlight the need to consider host sex as a biological variable in future immunological studies., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Examining the physiological responses of mussels to thermal stress is crucial to evaluate their biogeographic distribution and ability to adapt to a changing climate. In the present study, we investigated the effects of acute cold (8 °C and 15 °C) and heat (35 °C and 42 °C) stress on the mortality rate, reactive oxygen species (ROS) production, malondialdehyde (MDA) content, mitochondrial membrane potential (MMP) and antioxdative responses in the gill tissue of the green mussel species Perna viridis. Our results showed that cold and heat stress induced a temperature-dependent increase in mortality rate. ROS production increased significantly (p < 0.01) after both cold and heat stress. However, the activities of antioxidant enzymes, including SOD, CAT and GSH-Px, were greatly enhanced only after heat stress. In addition, MDA content and MMP increased significantly under both cold and heat stress. The up-regulation of Hsp70 transcripts was only detected after acute stress at 35 °C. However, p38-MAPK phosphorylation levels increased after both cold and heat stress. In addition, a moderate activation of caspase-3 was found after mussels were exposed to 8 °C and 42 °C stress. Our results suggest that both extreme cold and heat stress could induce ROS production in the gill tissue of P. viridis, which might result in lipid peroxidation and mitochondria dysfunction. Antioxidative enzymes and Hsp70 might be important in the heat stress response of animals, whereas p38-MAPK might be crucial in the acute response to both cold and heat stress. However, caspase-3 activation might be very weak under both cold and heat stress., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Copper is a common contaminant in aquatic environments, which may cause physiological dysfunction in marine organisms. However, the toxicity mechanisms of copper in marine bivalves is not fully understood. In this study, we applied an integrated approach that combines flow cytometry and Gas Chromatography-Mass Spectrometry (GC-MS)-based metabolomics to characterize cellular and molecular mechanisms of copper immunotoxicity in New Zealand Greenshell™ mussel (Perna canaliculus) haemolymph. Flow cytometric results showed significant increases in haemocyte mortality, production of reactive oxygen species and apoptosis (via alteration of caspase 3/7 and mitochondrial membrane potential) of haemocytes exposed to increasing total concentrations of Cu2+ (62.5, 125.0 and 187.5 μM) compared to a low Cu2+ concentration (25.0 μM) and control (0.0 μM). In addition to flow cytometric data, our metabolomics results showed alterations of 25 metabolites within the metabolite profile of Cu2+-exposed haemolymph (125 μM) compared to those of control samples. Changes in levels of these metabolites may be considered important signatures of oxidative stress (e.g., glutathione) and apoptosis processes (e.g., alanine, glutamic acid). This study provides insights into the cellular and molecular mechanisms of oxidative stress and apoptosis in marine bivalves and highlights the applicability and reliability of metabolomic techniques for immunotoxicological studies in marine organisms.

Toxic effects of continuous low dose application of the antifouling biocide chlorine on marine benthic organisms were monitored using transplanted green mussels (Perna viridis) and a suite of biomarkers. Caged mussels were deployed in chlorinated and non-chlorinated sections of the cooling system of an operating electric power plant. Biomarkers indicative of general stress, oxidative stress (superoxide dismutase and catalase), and DNA integrity, along with expression of stress proteins, were studied to assess the effects. Deterioration in condition index with corresponding increase in DNA strand breaks was indicative of chlorine stress. Superoxide dismutase enzyme did not show any particular trend, but catalase activity was high during the initial days of exposure at the chlorinated site; later, it became almost equal to that at the control site. Similarly, expressions of stress proteins (HSP60, HSP70, HSP22, GSTS1, and CYP4) showed bell-shaped pattern during the period of study. Positive correlation among the endpoints indicated the utility of the multimarker approach to monitor the effects of continuous low dose chlorination on mussels.

Vibrio coralliilyticus is a bacterial pathogen which can affect a range of marine organisms, such as corals, fish and shellfish, with sometimes devastating consequences. However, little is known about the mechanisms involved in the host-pathogen interaction, especially within molluscan models. We applied gas chromatography-mass spectrometry (GC-MS)-based metabolomics to characterize the physiological responses in haemolymph of New Zealand Greenshell™ mussels (Perna canaliculus) injected with Vibrio sp. DO1 (V. coralliilyticus/neptunius-like isolate). Univariate data analyses of metabolite profiles in Vibrio-exposed mussels revealed significant changes in 22 metabolites at 6 h post-infection, compared to non-exposed mussels. Among them, 10 metabolites were up-regulated, while 12 metabolites were down-regulated in infected mussels. Multivariate analyses showed a clear distinction between infected and non-infected mussels. In addition, secondary pathway analyses indicated perturbations of the host innate immune system following infection, including oxidative stress, inflammation and disruption of the TCA cycle, change in amino acid metabolism and protein synthesis. These findings provide new insights into the pathogenic mechanisms of Vibrio infection of mussels and demonstrate our ability to detect detailed and rapid host responses from haemolymph samples using a metabolomics approach.

We evaluated the thermotolerance (LT 50 ) of adult green-lipped mussels (Perna canaliculus) following an acute thermal challenge in the summer of 2012 and the winter of 2013. Mussels were grouped into two treatments, naïve (N, no prior heat treatment) and heat-hardened (HH = 1 h at 29 °C, 12 h recovery at ambient) before being immersed for 3 h in water of varying temperature, i.e. Ambient (Control), 25, 29, 31, 33, and 35 °C with subsequent mortality monitored for 30 days. As expected, naïve mussels were less thermotolerant than heat-hardened i.e. Summer LT 50 , N = 31.9, HH = 33.5 °C; Winter LT 50 , N = 31.4, HH = 33.8 °C. Moreover, at 33 °C no heat-hardened mussels died compared to 100% mortality in naïve specimens. At 35 °C all mussels died regardless of treatment. For the 'Summer' mussels, metabolite abundances in gill tissues of both naïve and heat-hardened mussels were quantified. For mussels at 33 °C, succinic acid was significantly higher in naïve mussels than heat-hardened mussels, indicating perturbations to mitochondrial pathways in these thermally stressed mussels. Additionally, analysis of biochemical pathway activity suggested a loss of neural control i.e. significantly reduced GABAergic synapse activity, in naïve vs. heat-hardened mussels at 33 °C. Taken together these findings suggest that heat-hardening improves mussel survival at higher temperatures by delaying the onset of cellular anaerobic metabolism, and by maintaining inhibition of neural pathways. Such results offer new perspectives on the complex suite of sub-cellular stress responses operating within thermally stressed organisms., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

We examined spatiotemporal variations of metal levels and three growth related biomarkers, i.e., RNA/DNA ratio (RD), total energy reserve (Et) and condition index (CI), in green-lipped mussels Perna viridis transplanted into five locations along a pollution gradient in the marine environment of Hong Kong over 120days of deployment. There were significant differences in metal levels and biomarker responses among the five sites and six time points. Mussels in two clean sites displayed better CI and significantly lower levels of Ag, Cu, Pb and Zn in their tissues than the other sites. Temporal patterns of RD in P. viridis were found to be site-specific. Across all sites, Et decreased in P. viridis over the deployment period, though the rate of decrease varied significantly among the sites. Therefore, temporal variation of biomarkers should be taken to consideration in mussel-watch programs because such information can help discriminate pollution-induced change from natural variation., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Triazophos, as a lipophilic organophosphate pesticide, displays higher bioaccumulation in the gonads of shellfish. To study the reproductive toxicity of triazophos, we applied metabolomics to characterize the gender-specific metabolic responses in mussel Perna viridis exposed to triazophos. Metabolites were differently altered by triazophos in ovaries of mussel at different concentrations and time intervals, while basically similar metabolic response patterns were observed in male mussels at the two tested concentrations after exposure for 24 and 48h. The significant changes of metabolites in ovaries of mussel exhibited the disturbances in energy metabolism and osmotic regulation, while in male samples triazophos only affected the energy metabolism. Moreover, glycine, sn-glycero-3-phosphocholine, ethanol, aspartate, etc. exhibited consistent variation tendency in both male and female individuals. While the changes of homarine, betaine, taurine, hypotaurine, malonate, β-alanine, succinate, and choline showed obviously gender-specific responses. Overall, this study confirmed the gender-specific responses in gonad of P. viridis to triazophos exposure., (Copyright © 2017. Published by Elsevier Ltd.)

The conversion of waste into high-value materials is considered an important sustainability strategy in modern chemical industries. A large volume of shell waste is generated globally from mussel cultivation. In this work, mussel shell waste (Perna viridis) is transformed into individual calcium carbonate plates (ICCPs) and is applied as a support for a heterogeneous catalyst. Palladium nanoparticles (3-6 nm) are deposited with an even dispersion on the ICCP surface, as demonstrated by X-ray diffraction and scanning electron microscopy. Using this system, Sonogashira cross-coupling reactions between aryl iodides and terminal acetylenes were accomplished in high yields with the use of 1 % Pd/ICCP in the presence of potassium carbonate without the use of any copper metal or external ligand. The Pd/ICCP catalyst could also be reused up to three times and activity over 90 % was maintained with negligible Pd-metal leaching. This work demonstrates that mussel shell waste can be used as an inexpensive and effective support for metal catalysts in coupling reactions, as demonstrated by the successful performance of the Pd-catalyzed, copper-free Sonogashira cross-coupling process., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The Chinese green mussel, Perna viridis, is a marine bivalve with important economic values as well as biomonitoring roles for aquatic pollution. Byssus, secreted by the foot gland, has been proved to bind heavy metals effectively. In this study, using the RNA sequencing technology, we performed comparative transcriptomic analysis on the mussel feet with or without inducing by cadmium (Cd). Our current work is aiming at providing insights into the molecular mechanisms of byssus binding to heavy metal ions. The transcriptome sequencing generated a total of 26.13-Gb raw data. After a careful assembly of clean data, we obtained a primary set of 105,127 unigenes, in which 32,268 unigenes were annotated. Based on the expression profiles, we identified 9,048 differentially expressed genes (DEGs) between Cd treatment (50 or 100 μg/L) at 48 h and the control, suggesting an extensive transcriptome response of the mussels during the Cd stimulation. Moreover, we observed that the expression levels of 54 byssus protein coding genes increased significantly after the 48-h Cd stimulation. In addition, 16 critical byssus protein coding genes were picked for profiling by quantitative real-time PCR (qRT-PCR). Finally, we reached a primary conclusion that high content of tyrosine (Tyr), cysteine (Cys), histidine (His) residues or the special motif plays an important role in the accumulation of heavy metals in byssus. We also proposed an interesting model for the confirmed byssal Cd accumulation, in which biosynthesis of byssus proteins may play simultaneously critical roles since their transcription levels were significantly elevated.

This study investigated the effects of hypoxia on oxidative stress response and immune function in mussels Perna perna exposed to air for 6, 12, 24 and 48 h. In air-exposed mussels, the antioxidant enzymes superoxide dismutase (SOD), catalase, and glutathione reductase (GR) activities were lower in gill tissues (24-48 h) and digestive gland (12 h), while the glutathione peroxidase and GR activities were increased in the digestive gland (48 h). In both tissues, aerial exposure promoted a rapid (6 h) and persistent (up to 48 h) increase of glutathione levels. Decreased hemocyte count and viability, as well as increased phagocytic activity and cellular adhesion capacity were detected after prolonged aerial exposure (>12 h). In summary, induction of thiol pools, altered antioxidant enzyme activities, and activation of immune responses were detected in hypoxia exposed brown mussels, indicating hypoxia induced tissue-specific responses in both antioxidant and immune systems., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Research on the kinetics of Benzo[a]pyrene (B[a]P) bioaccumulation in the clam Pinctada martensii and mussel Perna viridis showed that the initial rate of uptake was directly related to the PAH concentrations in the ambient environment. The uptake and depuration rate constants were different at the four B[a]P exposure levels, which indicated that the toxicokinetic rate constants mainly depended on the exposure levels of pollutants to the environment. In addition, the uptake rate constants of B[a]P were higher than the depuration rate constants in the entire experiment. The comparison demonstrated that mussels release B[a]P more rapidly than clams. The bioconcentration factors (BCFs) of B[a]P varied from 3335 to 12892 in the clam and 2373-6235 in the mussel. These findings on the bioaccumulation kinetics for petroleum hydrocarbons, in association with the critical body residue, will be valuable when choosing sensitive organisms to assess the potential ecotoxicological risk to the marine environment.

The study of the uptake and distribution of elements in marine environments is of great interest for understanding their pathways and accumulation. Here, we investigated in laboratory experiments the accumulation behavior of Cd in the sea anemone Bunodosoma caissarum and the mussel Perna perna. Specimens were incubated with isotopically enriched 116 Cd in aquariums. Cd concentrations in the seawater and in the tissues of B. caissarum and P. perna were followed by inductively coupled plasma-mass spectrometry (ICP-MS) by means of isotope dilution analysis. Bioconcentration factors for B. caissarum and P. perna exposed to 0.9μg·L -1 of 116 Cd were determined to be 80.5 and 850, respectively. P. perna specimens exposed to 4.5μg·L -1 of 116 Cd reached 530. Cytosolic proteins associated with Cd from the tissues were extracted and further analyzed by size-exclusion chromatography coupled to ICP-MS. Cd accumulation could be detected in both organisms ranging from high-molecular to low-molecular species., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The study assesses the effects of carbon dioxide capture and storage (CCS) leaks and ocean acidification (OA) on the metal bioavailability and reproduction of the mytilid Perna perna. In laboratory-scale experiments, CCS leakage scenarios (pH 7.0, 6.5, 6.0) and one OA (pH 7.6) scenario were tested using metal-contaminated sediment elutriates and seawater from Santos Bay. The OA treatment did not have an effect on fertilisation, while significant effects were observed in larval-development bioassays where only 16 to 27 % of larva developed normally. In treatments that simulated CO 2 leaks, when compared with control, fertilisation success gradually decreased and no larva developed to the D-shaped stage. A fall in pH increased the bioavailability of metals to marine mussels. Larva shell size was significantly affected by both elutriates when compared with seawater; moreover, a significant difference occurred at pH 6.5 between elutriates in the fertilisation bioassay.

Green mussel protein hydrolysates (GMPH) utilization for the enrichment of gluten-free bread followed by characterization of flavour peptides using chromatography and electronic nose techniques have been done. The degree of hydrolysis was carried out in each protease digest, and the higher degree of hydrolysis was observed in pepsin digestion. Gluten-free (GF) bread was formulated by using buckwheat flour (BWF), rice flour (RF) and chickpea flour (CPF) (70:20:10) and GMPH were added in the range of 0-20% in the GF bread for enrichment with GMPH. Radar plot of the electronic nose analysis showed that the sensors P30/2, T30/1 and T70/2 had a higher response to the GF bread and GMPH. Consequently, the peptide sequence was obtained manually by ESI-MS spectra of GMPH (KGYSSYICDK) and F-II (SSYCIVKICDK). Flavour quality was 97% discriminately comparable to the GMPH and F-II fractions. Mussel flavoured GF bread can be included in the celiac diet., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Non-target screening analyses were conducted in order to identify a wide range of organic contaminants in sediment and animal tissue samples from Jakarta Bay. High concentrations of di-iso-propylnaphthalenes (DIPNs), linear alkylbenzenes (LABs) and polycyclic aromatic hydrocarbons (PAHs) were detected in all samples, whereas phenylmethoxynaphthalene (PMN), DDT and DDT metabolites (DDX) were detected at lower concentrations. In order to evaluate the uptake and accumulation by economic important mussel (Perna viridis) and fish species, contaminant patterns of DIPNs, LABs and PAHs in different compartments were compared. Different patterns of these contaminant groups were found in sediment and animal tissue samples, suggesting compound-specific accumulation and metabolism processes. Significantly higher concentrations of these three contaminant groups in mussel tissue as compared to fish tissue from Jakarta Bay were found. Because P. viridis is an important aquaculture species in Asia, this result is relevant for food safety., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Metallomic studies regarding environmental contamination by metals are of value in elucidating metal uptake, trafficking, accumulation and metabolism in biological systems. Many proven bioindicator species, such as bivalves, have not yet, however, been well-characterized regarding their metalloprotein expression in response to environmental contaminants. In this context, the aim of the present study was to investigate metalloprotein expressions in the thermostable protein fraction of muscle tissue and digestive glands from mussels (Perna perna) from three differentially metal-contaminated sites in Southeastern Brazil in comparison with a reference site. The thermostable protein fractions were analyzed by SDS-PAGE and SEC-HPLC-ICP-MS. Metal content was also determined in both the crude and the purified extracts. Several inter-organ differences were observed, which is to be expected, while inter-site differences regarding thermostable protein content were also verified, indicating accumulation of these elements in muscle tissue and digestive glands and disruption of homeostasis of essential elements, with detoxification attempts by metal-bound proteins, since all metalloproteins present in both matrices eluted bound to at least one non-essential metal. These results are also noteworthy with regard to the adopted reference site, that also seems to be contaminated by toxic metals., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

Concentrations of 11 elements were quantified in five marine species from different trophic levels of a food web (algae, mussel, shrimp and fish), representative for shallow Senegalese coastal waters, and including species of commercial importance. Significant differences in element concentrations and bioaccumulation were demonstrated, revealing the utility of employing a suite of organisms as bioindicators to monitor metal contamination in coastal areas. There was no clear seasonal pattern in concentration of elements, however inter-site differences were observed. Calculations of transfer factors for all the studied elements showed that transfer factors from water were greater than those from sediments. For shrimp and mussel, the concentrations of Pb and Cd were below the EU's maximum level for human consumption, however high concentrations of arsenic in shrimp were recorded at all sites., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Benzo(a)pyrene (BaP) and dichlorodiphenyltrichloroethane (DDT) are persistent organic pollutants and environmental estrogens (EEs) with known toxicity towards the green mussel, Perna viridis. In this study, the toxic effects of BaP (10 µg/L) and DDT (10 µg/L) and their mixture were assessed in green mussel gills with proteomic and metabolomic approaches. Metabolic responses indicated that BaP mainly caused disturbance in osmotic regulation by significantly decrease in branched chain amino acids, dimethylamine and dimethylglycine in gills of male green mussels after exposure for 7 days. DDT mainly caused disturbance in osmotic regulation and energy metabolism by differential alteration of betaine, dimethylamine, dimethylglycine, amino acids, and succinate in gills of male green mussels. However, the mixture of BaP and DDT didn't show obvious metabolite changes. Proteomic analysis showed different protein expression profiles between different treatment groups, which demonstrated that BaP, DDT and their mixture may have different modes of action. Proteomic responses revealed that BaP induced cell apoptosis, disturbance in protein digestion and energy metabolism in gills of green mussels, whereas DDT exposure altered proteins that were associated with oxidative stress, cytoskeleton and cell structure, protein digestion and energy metabolism. However, the mixture of BaP and DDT affected proteins related to the oxidative stress, cytoskeleton and cell structure, protein biosynthesis and modification, energy metabolism, growth and apoptosis., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

To contribute to the use of the tropical brown mussel Perna perna as a sentinel species for organochlorine pesticides (OCP) and polychlorinated biphenyls (PCB), the present study reports data on the toxicokinetics of these compounds in P. perna. Specifically, the authors present data on OCP and PCB bioaccumulation for eight sampling months from three bays (SE Brazil) and two transplant experiments (each 1 month long). Although seasonality is observed in the total lipid content of the whole soft tissue, with summer samples showing higher values, no such seasonality is observed in the OCP and PCB concentrations bioaccumulated by the mussel P. perna. Because no seasonal effect is observed in the annual OCP and PCB concentrations bioaccumulated by P. perna, the use of this species as a sentinel organism to monitor organochlorinated compounds is encouraged. One month of transplantation is not enough to allow the transplanted specimens to reach the concentrations observed in animals reared at the destination site. Nevertheless, P. perna showed a clear tendency to depurate the DDT metabolites p,p'-DDD and p,p'-DDE after 1 month of transplantation.

Marine mussels have long been used as biomonitors of contamination of trace elements, but little is known about whether variation in tissue trace elements is significantly associated with those of macronutrients and major cations. The authors examined the variability of macronutrients and major cations and their potential relationships with bioaccumulation of trace elements. The authors analyzed the concentrations of macronutrients (C, N, P, S), major cations (Na, Mg, K, Ca), and trace elements (Al, V, Mn, Fe, Co, Ni, Cu, Zn, As, Se, Mo, Cd, Ba, Pb) in the whole soft tissues of marine mussels Mytilus edulis and Perna viridis collected globally from 21 sites. The results showed that 12% to 84% of the variances in the trace elements was associated with major cations, and the tissue concentration of major cations such as Na and Mg in mussels was a good proxy for ambient seawater concentrations of the major cations. Specifically, bioaccumulation of most of the trace elements was significantly associated with major cations, and the relationships of major cations with trace cations and trace oxyanions were totally opposite. Furthermore, 14% to 69% of the variances in the trace elements were significantly associated with macronutrients. Notably, more than half of the variance in the tissue concentrations of As, Cd, V, Ba, and Pb was explained by the variance in macronutrients in one or both species. Because the tissue macronutrient concentrations were strongly associated with animal growth and reproduction, the observed coupling relationships indicated that these biological processes strongly influenced the bioaccumulation of some trace elements. The present study indicated that simultaneous quantification of macronutrients and major cations with trace elements can improve the interpretation of biomonitoring data., (© 2015 SETAC.)

Perna viridis is a recently introduced species to US coastal waters and have vigorously spread throughout the southeastern seaboard since their invasion. Little information regarding their response to local environmental factors has been reported including responses to the local HAB species, Karenia brevis. This study monitored the tissue toxin concentration of brevetoxins in P. viridis from existing populations throughout two consecutive natural K. brevis blooms. The results showed P. viridis to rapidly accumulate PbTx upon exposure to the bloom, far exceeding the peak tissue concentrations of oysters, Crassostrea virginica, sampled during the same period, 57,653 ± 15,937 and 33,462 ± 10,391 ng g(-1) PbTx-3 equivalent, respectively. Further, P. viridis retained high PbTx concentrations in their tissues post bloom remaining above the regulatory limit for human consumption for 4-5 months, significantly longer than the depuration time of 2-8 weeks for native oyster and clam species. In the second year, the bloom persisted at high cell concentrations resulting in prolonged exposure and higher PbTx tissue concentrations indicating increased bioaccumulation in green mussels. While this species is not currently harvested for human consumption, the threat for post bloom trophic transfer could pose negative impacts on other important fisheries and higher food web implications., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Perna viridis was used as biomonitor to assess heavy metal levels in the Chacopata-Bocaripo lagoon axis, Venezuela, during rain and drought seasons. The mussels were weighed and measured. The metal concentrations were determined by atomic absorption spectrophotometry. For rain period, the order of bioavailability was: Cu>Ni>Mn>Co>Cd>Pb, and for drought: Cu>Mn>Ni>Co>Pb>Cd. The concentrations of Ni, Co, Cd and Pb showed significant differences (P<0.05) in both periods. There was higher metal accumulation during drought season, possibly related to upwelling, since it produces an increase in primary productivity, which translates more food into organisms, making metals bioavailable for mussels. Only Cu and Mn showed significant relationships between the size and metal concentration, during drought period, it may be because of the organisms need for these essential metals in different physiological processes., (Copyright © 2014 Elsevier Ltd. All rights reserved.)