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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.)

Microplastics pollution threatens to marine organisms, particularly bivalves that actively ingest and accumulate microplastics of certain sizes, potentially disrupting intestinal homeostasis. This study investigated the microplastic abundance in wild and farmed mussels around Singapore, and examined the size-dependent effects of nano- to micro-scale polystyrene (0.5 µm/5 µm/50 µm) on the mussel intestinal microbiome in the laboratory. The field investigation revealed higher microplastic abundance in farmed mussels compared to wild ones. Experimentally, mussels exposed to 0.6 mg/L of microplastics for 7 days, followed by a 7-day depuration period, showed substantial impacts on Spirochaetes and Proteobacteria, facilitating the proliferation of pathogenic species and differentially affecting their pathogenic contributions. Metagenomics analysis revealed that microplastic exposure reduced Spirochaeta's contribution to virulence and pathogenicity loss, did not affect Vibrio and Oceanispirochaeta's pathogenicity, and increased Treponema and Oceanispirochaeta's contributions to pathogenicity loss. Moreover, microplastics increased transmembrane transporters and impacted oxidative phosphorylation enzymes, impairing energy metabolism. These effects persisted after depuration, indicating lack of resilience in the microbiome. Nano- and micro-scale plastics perturbed the mussel microbiome composition and functions in a size-dependent manner, with nano-plastics being the most disruptive. The increasing use and sale of aquaculture equipment of plastic may exacerbate the intestinal dysbiosis in bivalves, which threatens consumers' health., 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 © 2024 Elsevier B.V. All rights reserved.)

Harmful algal blooms (HABs) of Alexandrium pacificum have affected the Marlborough Sounds in New Zealand since 2010, posing a threat to green-lipped mussel (GLM, Perna canaliculus) farming. Previous studies have shown A. pacificum has negative effects GLM embryos and larvae. To further investigate these toxic mechanisms, in vitro bioassays were conducted on GLM spermatozoa, hemocytes, and the diatom, Chaetoceros muelleri. The three cell types were exposed to several treatments of A. pacificum for 2 h and responses were measured using flow cytometry and pulse amplitude-modulated fluorometry. Significant spermatozoa mortality was recorded in treatments containing A. pacificum cells or fragments, while hemocyte and C. muelleri mortality was recorded in cell-free treatments of A. pacificum which contained paralytic shellfish toxins (PSTs). Variation in sensitivity between cell types as well as the sublethal effects observed, emphasise the diverse toxic mechanisms of A. pacificum on co-occurring species in the environment., 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 © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The presence and persistence of microplastics (MPs) in diverse aquatic environments are of global concern. Microplastics can impact marine organisms via direct physical interaction and the release of potentially harmful chemical additives incorporated into the plastic. These chemicals are physically bound to the plastic matrix and can leach out. The hazards associated with chemical additives to exposed organisms is not well characterized. We investigated the hazards of plastic additives leaching from plastic. We used the common plasticizer dibutyl phthalate (DBP) as a chemical additive proxy and the New Zealand green-lipped mussel (Perna canaliculus) as a model. We used early-adult P. canaliculus exposed to combinations of virgin and DBP-spiked polyvinyl chloride (PVC), MPs, and DBP alone for 7 days. Whole transcriptome sequencing (RNA-seq) was conducted to assess whether leaching of DBP from MPs poses a hazard. The differences between groups were evaluated using pairwise permutational multivariate analysis of variance (PERMANOVA), and all treatments were significantly different from controls. In addition, a significant difference was seen between DBP and PVC MP treatment. Transcriptome analysis revealed that mussels exposed to DBP alone had the most differentially expressed genes (914), followed by PVC MP + DBP (448), and PVC MP (250). Gene ontology functional analysis revealed that the most enriched pathway types were in cellular metabolism, immune response, and endocrine disruption. Microplastic treatments enriched numerous pathways related to cellular metabolism and immune response. The combined exposure of PVC MP + DBP appears to cause combined effects, suggesting that DBP is bioavailable to the exposed mussels in the PVC MP + DBP treatment. Our results support the hypothesis that chemical additives are potentially an important driver of MP toxicity. Environ Toxicol Chem 2024;43:1604-1614. © 2024 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC., (© 2024 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.)

Cocaine (COC) is a powerful illicit drug frequently detected in the aquatic environment. COC acts by inhibiting the reuptake of dopamine (DOPA) and 5-hydroxytryptamine (5-HT - serotonin) and causes endocrine disturbances in mammals. This study investigated similar effects from cocaine exposure in the marine mussel Perna perna, as well as neurotoxicity and energy imbalances. Mussels were exposed to COC (0.2 μg.L -1 and 2 μg.L -1 ) for periods of 48, 96, and 168 h. Acetylcholinesterase (AChE) was measured in adductor muscle tissue to determine neurotoxicity, and neurotransmitter levels (DOPA and 5-HT), monoamine oxidase (MAO) and cyclooxygenase (COX) activity, and energy status (mitrochondrial electron transport, MET, and total lipids, TLP) were evaluated in the mussels' gonads. COC decreased AChE activity in the mussels exposed to 0.2 μg.L -1 and 2 μg.L -1 after 168 h, and all concentrations of COC increased neurotransmitter levels. Increases in MET (0.2 μg.L -1, for all exposure periods) and TLP (0.2 μg.L 1 after 48 h, and 2 μg.L -1 after 96 h and 168 h) were also observed. No significant change was detected in MAO activity. COC also decreased COX activity in the mussels exposed to 0.2 μg.L -1 (48 h and 96 h) and 2 μg.L -1 (96 h). These results suggest that COC may compromise neurotransmitter levels and COX activity. Furthermore, the changes in MET and LPT suggest that COC affects the energy balance of the mussels, and could negatively affect physiological processes such as metabolism, hormone production, and embryonic development., (Copyright © 2021 Elsevier Inc. All rights reserved.)

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.

Recent studies have shown that organisms including humans are exposed to microplastics directly or indirectly. The present study aims to examine the ingestion of these microplastics and the consequences of the same by studying the accumulation behavior of weathered Polyethylene (wPE) microplastics. The Perna viridis were exposed chronically to three different environmentally relevant concentrations of wPE for 30 days, followed by a one-week depuration phase. There was no mortality observed in the control and exposed groups, but the feeding rate was observed to have substantially decreased in the group exposed to higher concentration (3 μgL -1 ) of wPE. It was also observed that a higher number of wPE particles accumulated in the intestine of exposed organisms. Interestingly, the present study revealed the presence of the substantial number of wPE particles in exposed organisms, which may adversely affect the internal organs as well as growth and reproduction. This study perceived that accumulation is marginally influenced by size of wPE. Similarly, biomarker analysis showed that wPE exposure significantly altered both the metabolism and histology of the internal organs of the exposed organisms. Overall, the study confirmed that the intestine was the most sensitive organ followed by gills, adductor muscles, and foot tissue adding new insights into the adverse effects of wPE in the marine ecosystem., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Marine bivalves have been widely applied as environmental contamination bioindicators, although studies concerning tropical species are less available compared to temperate climate species. Assessments regarding Perna perna mytilid mussels, in particular, are scarce, even though this is an extremely important species in economic terms in tropical countries, such as Brazil. To this end, Perna perna mytilids were sampled from two tropical bays in Southeastern Brazil, one anthropogenically impacted and one previously considered a reference site for metal contamination. Gill metallothionein (MT), reduced glutathione (GSH), carboxylesterase (CarbE) and lipid peroxidation (LPO) were determined by UV-vis spectrophotometry, and metal and metalloid contents were determined by inductively coupled plasma mass spectrometry (ICP-MS). Metalloprotein metal detoxification routes in heat-stable cellular gill fractions were assessed by size exclusion high performance chromatography (SEC-HPLC) coupled to an ICP-MS. Several associations between metals and oxidative stress endpoints were observed at all four sampling sites through a Principal Component Analysis. As, Cd, Ni and Se contents, in particular, seem to directly affect CarbE activity. MT is implicated in playing a dual role in both metal detoxification and radical oxygen species scavenging. Differential SEC-HPLC-ICP-MS metal-binding profiles, and, thus, detoxification mechanisms, were observed, with probable As-, Cu- and Ni-GSH complexation and binding to low molecular weight proteins. Perna perna mussels were proven adequate tropical bioindicators, and further monitoring efforts are recommended, due to lack of data regarding biochemical metal effects in tropical species. Integrated assessments, as performed herein demonstrate, are invaluable in evaluating contaminated aquatic environments, resulting in more accurate ecological risk assessments., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

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.)

The toxicity of heavy metals commonly impacts the survival of crustacean and bivalve larvae in hatchery culture, and this has led to the widespread use of EDTA to decrease this toxicity. Since EDTA has a very poor biodegradability leading to potential persistent environmental effects, alternative methods to prevent heavy metal toxicity to shellfish larvae are needed. EDDS is a biodegradable potential alternative to EDTA for this application and was tested as a treatment of the seawater used for rearing aquaculture Greenshell™ mussel (Perna canaliculus) larval embryos in this study. Mussel embryos reared with EDTA or EDDS had significantly better survival than without. The concentrations and spatial distributions of heavy metals in D-veliger larvae as determined by Inductively Coupled Plasma Mass Spectrometry (ICP-MS) and X-ray Fluorescence Microscopy (XFM) suggested that chelating agents increased the levels of calcium in larvae while they reduced the concentration of zinc. In addition, where decreased accumulation of the other heavy metals was not observed, chelating agents affected their distribution within the larvae, especially for copper and arsenic. This is the first study to test the use of EDDS for aquaculture hatchery application and shows that EDDS is an effective biodegradable alternative to EDTA that can mitigate the effects of heavy metals for shellfish larval rearing., (Copyright © 2020 Elsevier B.V. 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.)

Bivalve molluscs rely only on an innate immune system to execute cellular and humoral processes. Haemocytes, the haemolymph circulating cells, play a major role in this type of immunity, principally regarding cellular defences. Considering that environmental pollutants can affect the immune system of invertebrates, this work evaluated the effects of the antifouling biocide 4,5-dicloro-2-n-octil-4-isotiazolin-3-ona (DCOIT) on the haemocytes of mussels Perna perna. Individuals were exposed to 0 (control), 0.1 μg L -1 and 10 μg L -1 of DCOIT for up to 96 h. The analysed parameters included: total (THC) and differential (DHC) haemocyte count, cellular viability, adhesion capacity, phagocytic activity, levels of reactive oxygen species and DNA damage. Moreover, the stress on stress (SOS) response of mussels was analysed as a general stress index. The results show that DCOIT increased the haemocyte adhesion capacity and caused a decrease in THC and in the haemocyte viability after 24 h of exposure. After 96 h of exposure, DCOIT only affected the haemocyte adhesion capacity, which was decreased by biocide exposure. Moreover, exposure to DCOIT for 96 h did not affect the capacity for air survival of mussels. These results indicate that DCOIT interferes in important parameters associated with the innate immunity of P. perna, mainly after 24 h of exposure. It is suggested that the animals were able to develop some compensatory response strategy, making them more resistant to the biocide., 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 Inc. 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.)

Chlorothalonil is an effective fungicide used in agriculture and formulations of antifouling paints, which use and possible toxicity has been generating great concern. Thus, the present study investigated the effects of chlorothalonil on the antioxidant defense system (ADS) of the mussel Perna perna. The ADS was evaluated in gills and digestive gland after 24 h and 96 h of exposure to environmental relevant levels of chlorothalonil (0.1 and 10 μg/L). The activity of the enzymes superoxide dismutase (SOD), catalase (CAT), glutamate cysteine-ligase (GCL) and glutathione S-transferase (GST), levels of non-enzymatic defenses, represented by glutathione (GSH), and lipoperoxidation (LPO) and protein carbonyls (PCO) were evaluated. Results indicated that exposure to chlorothalonil is affecting the ADS in both tissues. While the activity of SOD increased and GST and GSH were not altered in gills, they decreased in digestive gland after 24 h of exposure to 10 μg/L of chlorothalonil. The contrasting results indicate that gills and digestive gland presented different patterns of responses after exposure to chlorothalonil. Moreover, a tissue-specific response to chlorothalonil was observed. Gills could be acting as the first line of defense, presenting higher enzymatic levels with minor effects on the parameters analyzed. On the other hand, digestive gland, with lower levels of antioxidant defenses, was the most affect organ by chlorothalonil. It also should be highlighted that the fungicide reduced the glutathione metabolism in the digestive gland, which can lead to an imbalance of the redox state within the cells of animals., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The present study evaluates the enzyme activities and histopathological changes in the post larvae (PL) of shrimp (Penaeus monodon), green mussel (Perna viridis) and fingerlings of crescent perch (Terapon jarbua) exposed to sublethal gradient concentrations of Nickel (Ni). The median lethal concentration (LC 50 ) values were 2.49, 66.03 and 43.92 mg Ni L -1 derived for the PL of shrimp, green mussel and fish fingerlings respectively. No Observed Effect Concentration (NOEC), Lowest Observed Effect Concentration (LOEC) and chronic values of the PL of shrimp were 46.5, 73.0 and 58.3 μg Ni L -1 derived for the 21-d survival endpoint. The NOEC, LOEC and chronic values for the 30-d survival endpoint of the green mussels and fish fingerlings were 4.6, 6.32, 5.4 and 1.95, 2.6, 2.25 mg Ni L -1 respectively. The isoforms of esterase, superoxide dismutase and malate dehydrogenase activities in the whole body tissues of test organisms were studied by native polyacrylamide gel electrophoresis after exposure to Ni. Histological examination of compound eye sections of shrimp revealed deformation, compression, fusion and detachement in the corneal cells from the corneal facet of the ommatidia indicating cellular anomalies due to Ni toxicity. Gill sections of the green mussel witnessed reduced haemolymph in sinuses of gill filaments, degenerative changes in interfilamentous junction and necrosis of frontal ciliated epithelial cells with vacuoles after exposure to Ni. Nickel affects the vision of shrimp and fish fingerlings, gills and byssus of green mussels., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The presence of cocaine and its metabolites and by-products has been identified in different aquatic matrices, making crack cocaine the target of recent studies. The aim of this study was to evaluate the sublethal effects of crack on the brown mussel Perna perna. Mussels were exposed to three concentrations of crack cocaine (0.5, 5.0, and 50.0 μg L -1 ) for 168 h. Gills, digestive glands, and hemolymph were extracted and analyzed after three different exposure times using a suite of biomarkers (EROD, DBF, GST, GPX, LPO, DNA damage, ChE, and lysosomal membrane stability [LMS]). After 48 and 96 h of exposure, EROD, DBF, GST, GPX activities and DNA strand breaks in the gills increased significantly after 48 and 96 h of exposure. Alterations in LMS were also observed in the mussels exposed to all crack concentrations after 96 and 168 h. Our results demonstrated that crack cocaine is metabolized by CYP-like and GST activities in the gills. GPX was not able to prevent primary genetic damage, and cytotoxic effects in the hemocytes were also observed in a dose- and time-dependent response. Our study shows that the introduction of illicit drugs into coastal ecosystems must be considered a threat to marine organisms.

Diarrhetic shellfish poisoning (DSP) toxins are key shellfish toxins that cause diarrhea, vomiting and even tumor. Interestingly, bivalves such as Perna viridis have been reported to exhibit some resistances to alleviate toxic effects of DSP toxins in a species-specific manner. Nevertheless, the molecular mechanisms underlying the resistance phenomenon to DSP toxins, particularly the mechanistic role of CYP450 is scant despite its crucial role in detoxification. Here, we exposed P. viridis to Prorocentrum lima and examined the expression pattern of the CYP450 and our comprehensive analyses revealed that P. lima exposure resulted in unique expression pattern of key CYP450 genes in bivalves. Exposure to P. lima (2 × 10 5  cells/L) dramatically orchestrated the relative expression of CYP450 genes. CYP2D14-like mRNA was significantly down-regulated at 6 h in gill, but up-regulated at 2 h in digestive gland compared with control counterparts (p < 0.05), while CYP3A4 mRNA was increased at 12 h in gill. After exposure to P. lima at 2 × 10 6  cells/L, the expression of CYP3A4 mRNA was significantly increased in digestive gland at 2 h and 12 h, while CYP2D14-like was up-regulated at 6 h. Besides, CYP3L3 and CYP2C8 also exhibited differential expression. These data suggested that CYP3A4, CYP2D14-like, and even CYP3L3 and CYP2C8 might be involved in DSP toxins metabolism. Besides, provision of ketoconazole resulted in significant decrement of CYP3A4 in digestive gland at 2 h and 12 h, while the OA content significantly decreased at 2 h and 6 h compared to control group without ketoconazole. These findings indicated that ketoconazole could depress CYP3A4 activity in bivalves thereby altering the metabolic activities of DSP toxins in bivalves, and also provided novel insights into the mechanistic role of CYP3A4 on DSP toxins metabolism in bivalves., (Copyright © 2019. Published by Elsevier Inc.)

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.)

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.

Prorocentrum lima is a widely distributed marine benthic dinoflagellate that produces diarrhetic toxins, okadaic acid (OA) and its analogs, that may promote damage on bivalve tissues and cellular responses. Cultivation of the brown mussel Perna perna represents an important economic activity in the tropical and subtropical regions, where mussels may co-occur with P. lima. This study aimed to assess the behavioral, cellular immune responses, and pathological condition of P. perna following a short-term experimental exposure to P. lima. The toxic dinoflagellate treatment was compared to a non-toxic exposure to the chlorophyte Tetraselmis sp. at similar concentrations. The prevalence of pathological conditions and parasites were assessed, and a pathological index was applied by scoring the prevalences into four levels. Reaction time and the number of stimuli necessary for shell-valve closure response significantly increased after 72 h of P. lima exposure. Circulating hemocyte concentration was significantly lower in P. lima exposed mussels than in control mussels at 48- and 96 h of incubation, while hemocyte relative size in exposed mussels was significantly higher than that in control mussels. Comparatively, phagocytic activity and ROS production by hemocytes was significantly higher in mussels exposed to P. lima at 48- and 96 h of incubation, respectively. In addition, exposed mussels significantly presented exacerbated hemocytic infiltration in digestive organs, higher prevalence of moderate to severe atrophy in digestive tubules, and higher pathological index which suggests an impairment of mussel immunologic responses. A lower prevalence of Rickettsia-like organisms (RLOs), trematodes and copepods in P. lima exposed mussels suggests a direct toxic effect of OA on parasites. The exposure of mussels to P. lima is likely to occur frequently and may lead to constraints on mussel behavior, physiology, and pathological condition., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

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.)

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.

The environmental impact of microplastics is a challenging theme, especially under realistic experimental conditions. We investigated physiological responses to 0.1-1.0 μm PVC particles intake by the mussel Perna perna after a relative long-term exposure (90 days) at a less extreme concentration compared with previous studies (0.125 g/L). Microplastic intake was inferred by the presence of PVC in the feces of mussels, and physiological damages were assessed through ingestion rate, assimilation efficiency, growth rate, cellular and molecular biomarkers (lysosomal integrity, lipid peroxidation, and DNA damage), and condition index. All physiological responses showed nonsignificant effects of the microplastics on the exposed mussels. We suggest that, despite the experimental concentration of microplastics, mussels were able to acclimate to the exposure through their abilities for long-term recovery and tolerance to stresses. These data have positive implications for environmental health and in terms of human food resource because mussel farming is a worldwide practice that heavily relies on plastic materials, increasing the chances of microplastic exposure and mussels contamination.

The study was carried out to evaluate enzyme activities and histopathological changes due to the effect of acute and chronic definitive toxicity of selenium (Se) on the post larvae (PL) of giant tiger shrimp (Penaeus monodon), and green mussel (Perna viridis). The 96-h Median Lethal concentration (LC 50 ) for the PL of shrimp was 3.36 mg L -1 and the chronic value for the long-term survival endpoint in a 21-d exposure was 0.10 mg L -1 . The green mussel 96-h LC 50 was 28.41 mg L -1 and the chronic value for the long-term survival endpoint in a 30-d exposure was 3.06 mg L -1 . Native polyacrylamide gel electrophoresis revealed altered diverse isoforms of esterase, superoxide dismutase and malate dehydrogenase activities in the PL of shrimp and green mussel exposed to sublethal concentration of Se. Cellular anomalies such as deformation and fusion of corneal cells, detachment of corneal cells from cornea facet and increased space between ommatidia were observed in the compound eye of PL of shrimp exposed to Se for 21-d. Shrinkage and clumping of mucous gland, degenerative changes in phenol gland, and ciliated epithelium were observed in the foot of green mussel exposed to Se for 30-d. This study shows that cellular anomalies in the compound eye of PL of P. monodon and foot tissues of P. viridis described would affect the vision of shrimp and byssus thread formation in green mussel., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The immunotoxicity of 4 commonly detected perfluoroalkyl substances (PFASs), namely, perfluorooctanesulfonate (PFOS), perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), and perfluorodecanoic acid (PFDA) was investigated by measuring biomarkers of the immune profile of green mussels, Perna viridis. The biomarkers included neutral red retention, phagocytosis, and spontaneous cytotoxicity, all of which were tested on mussel hemocytes. Hemocytes are an important component of the invertebrate immune system. We found that exposure to PFASs could lead to reduced hemocyte cell viability and suppress immune function by up to 50% of normal performance within the experimental exposure range. The results indicate that PFASs have an immunotoxic potential and thus could pose severe health risks to aquatic organisms. The reported immunotoxicity is likely to result from the compounds' direct and indirect interactions with the hemocyte membrane, and therefore likely to affect the functionality of these cells. The immunotoxic response was found to be related to the organism's burden of PFASs, and was reversible when the compounds were removed from the test organisms. Based on this relationship, models using an organism's PFAS concentration and bioaccumulation factor (BAF) as the independent variables were established to quantify PFAS-induced immunotoxicity. The models help us to gain a better understanding of the toxic mechanism of PFASs, and provide a tool to evaluate adverse effects for the whole group of compounds with one mathematical equation. Environ Toxicol Chem 2018;37:1138-1145. © 2018 SETAC., (© 2018 SETAC.)

Pharmaceutical discharges into the aquatic ecosystem are of environmental concern and sewage treatment plants (STPs) have been pointed out as the major source of these compounds to coastal zones, where oceanic disposal of sewage occurs through submarine outfalls. Diclofenac (DCF) is one of the most frequently detected pharmaceuticals in water, but little is known about the effects on marine organisms. In this study, we employed a tiered approach involving the determination of environmental concentrations of DCF in marine water and the adverse biological effects for fertilization, embryo-larval development and biomarker responses of the mussel Perna perna. Results indicate that effects in fertilization rate and embryo-larval development were found in the order of mg·L -1 . However, low concentrations of DCF (ng·L -1 ) significantly decreased the lysosomal membrane stability and COX activity, as well as triggered DNA damage, oxidative stress and changes in antioxidant defenses. Our results point to an environmental hazard at coastal ecosystems and suggest the need for improvements in the treatment of domestic wastewater aiming to reduce DCF concentrations, as well as regulation on current environmental legislation and monitoring of aquatic ecosystems., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Offshore oil exploration creates threats to coastal ecosystems, including increasing urbanization and associated effluent releases. Genotoxicity biomarkers in mussels were determined across a gradient of coastal zone influences of offshore petroleum exploration in southeastern Brazil. Coastal ecosystems such as estuaries, beaches and islands were seasonally monitored for genotoxicity evaluation using the brown mussel Perna perna. The greatest DNA damage (5.2% ± 1.9% tail DNA and 1.5‰  ± 0.8‰ MN) were observed in urban estuaries, while Santana Archipelago showed levels of genotoxicity near zero and is considered a reference site. Mussels from urban and pristine beaches showed intermediate damage levels, but were also influenced by urbanization. Thus, mussel genotoxicity biomarkers greatly indicated the proposed oil exploration and urbanization scenarios that consequently are genetically affecting coastal organisms.

This study was aimed to investigate the antifouling (AF) potentials and toxic properties of methanol extract from leaves of mangrove Excoecaria agallocha. Antimicrofouling activity results inferred that this extract strongly inhibited fouling bacterial and microalgal growth. This extract had also inhibited the settlement of brown mussel Perna indica and larvae of barnacle Balanus amphitrite. Further, EC 50 < LC 50 and therapeutic ratio > 1 together propagated non-toxic nature of the extract. Mollusk foot adherence assay result showed complete inhibition of foot spreading and loss of attachment of common rocky fouler Patella vulgata to the substrata. Field assay results affirmed that this extract effectively deterred settlement of biofoulers. Purification and GC-MS analysis of bioassay-guided active spot evidenced presence of three major compounds (> 85%) responsible for the promising AF activity. The identified lead compounds subjected to an estimation (BIOWIN™) program developed by United States Environmental Protection Agency (USEPA) predicts that they are biodegradable in nature. Graphical abstract.

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.)

Our study aimed to evaluate crack cocaine effects in different life stages of the marine mussel Perna perna. For this purpose, fertilization rate, embryo-larval development, lysosomal membrane stability and DNA strand breaks were assessed. Effect concentrations in gametes and in larval development were found after 1h (IC 50 =23.53mg·L -1 ) and 48h (IC 50 =16.31mg·L -1 ), respectively. The highest tested concentration showing no acute toxicity (NOEC) was 10mg·L -1 , while the lowest observed effect concentration (LOEC) was 20mg·L -1 . NOEC concerning embryo-larval development was 0.625mg·L -1 , while the LOEC was 1.25mg·L -1 . Cyto-genotoxic effects were evidenced in mussels exposed to crack cocaine concentrations ranging from 5 to 500μg·L -1 . Our results report the first data on effects of an illicit drug to marine organisms and should encourage further ecotoxicological studies of these contaminants of emerging concern in coastal ecosystems., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Marine ecosystems are subjected to a variety of contaminants. Antifouling paints, for example, have been extensively used to protect ship surfaces from marine biofouling, but their toxicity has generated great concern. Thus, we evaluated the effect of the biocide chlorothalonil on the immune system of Perna perna mussels. The mussels were exposed to 0 (control), 0.1μg/L and 10μg/L of chlorothalonil for up to 96h. After 24h and 96h of exposure, the following immune-related parameters were analyzed in the hemolymph of mussels: total hemocyte count, cell adhesion, phagocytic activity, level of reactive oxygen species, cell viability and comet assay. After 24h and 96h of chlorothalonil exposure, cellular adhesion increased and the hemocyte viability reduced. Moreover, an increase in phagocytic activity was also observed after 96h of exposure to cholorothalonil. The exposure to 10μg/L of chlorothalonil for 96h reduced the air survival capacity of mussels. Total hemocyte count, ROS generation and DNA damage were not affected by the contaminant exposure. Our results indicate that chlorothalonil affected important immune responses of the bivalves, demonstrating that this biocide has effects on non-target species. This modulation of immune system reduced the health status of mussels, which could compromise their ability to survive in the environment., (Copyright © 2017. Published by Elsevier B.V.)

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.

The present study investigated the toxicity of several emerging contaminants: the pharmaceutical drug carbamazepine (CBZ), the plasticizer bisphenol A (BPA), and the herbicide atrazine (ATZ) in a marine bivalve. Green mussels (Perna viridis) were exposed to different concentrations of CBZ, BPA, and ATZ, either individually or as mixtures over a 7-d period, and a suite of molecular and cellular biomarkers were analyzed: biomarkers of immunotoxicity (total hemocyte count, phagocytosis, extracellular lysozyme), genotoxicity (Comet assay), neurotoxicity (inhibition of acetylcholinesterase [AChE]), endocrine disruption (vitellin-like proteins), and detoxification enzymes (cytochrome P4501A [CYP1A], 7-ethoxyresorufin O-deethylase [EROD], and glutathione-S-transferase [GST]). Results of the single-chemical exposure tests highlighted the relatively low toxicity of CBZ because most biomarker responses observed were recorded at concentrations well above environmental levels. Bisphenol A exposure at environmentally realistic concentrations resulted in clear immunomodulatory, genotoxic, and endocrine-disruptive effects. Similarly, 3 of the 10 biomarkers tested on green mussels (genotoxicity, inhibition of AchE, and EROD) responded after exposure to ATZ at environmentally relevant doses or below, and confirmed the potency of this herbicide to marine bivalves. Exposure tests using mixtures of CBZ, BPA, and ATZ also revealed that these 3 substances were generally acting in an additive manner on the selected biomarkers, at environmental doses, with some exceptions (antagonism and/or synergy) at low and high concentrations. The present study also confirms that most of the biomarkers used are suitable for biomonitoring studies with green mussels. Environ Toxicol Chem 2017;36:429-441. © 2016 SETAC., (© 2016 SETAC.)

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.

The green mussel Perna viridis is an ideal biomonitor to evaluate marine environmental pollution. Benzo(a)pyrene (BaP) is a typical polycyclic aromatic hydrocarbon (PAH), which is well known for the mutagenic and carcinogenic characteristics. However, the toxicological effects of BaP on Perna viridis embryo are still unclear. In this study, we investigated the embryo transcriptomic profile of Perna viridis treated with BaP via digital gene expression analysis. A total of 92,362,742 reads were produced from two groups (control and BaP exposure) by whole transcriptome sequencing (RNA-Seq). Gene Ontology (GO) and Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway analysis were used on all genes to determine the biological functions and processes. Genes involved in various molecular pathways of toxicological effects were enriched further. The differential expression genes (DEGs) were related to stress response, infectious disease and innate immunity. Quantitative real-time PCR (qRT-PCR) measured expressional levels of six genes confirmed through the DGE analysis. This study reveals that RNA-seq for transcriptome profiling of P. viridis embryo can better understand the embryo toxic effects of BaP. Furthermore, it also suggests that RNA-seq is a superior tool for generating novel and valuable information for revealing the toxic effects caused by BaP at transcriptional level., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

This study aims to test if RNA/DNA ratio and various energy reserve parameters (i.e., glycogen, lipid, protein content and total energy reserves) are sensitive biomarkers for indicating stresses induced by metal contaminants in the green-lipped mussel Perna viridis, a common organism for biomonitoring in Southeast Asia. This study was, therefore, designed to examine the effects of cadmium (Cd) and copper (Cu) on these potential biomarkers in two major energy storage tissues, adductor muscle (AM) and hepatopancreas (HP), of P. viridis after sub-lethal exposure to either metal for 10 days. The results showed that neither Cd nor Cu treatments affected the RNA/DNA ratio, glycogen and protein contents in AM and HP. As the most sensitive biomarker in P. viridis, the total lipid content in both AM and HP was significantly decreased in the treatment of 5μg Cu/L and 0.01-0.1μgCd/L, respectively. However, soft-tissue body burdens of Cu or Cd did not significantly correlate with each of the four biomarkers regardless of the tissue type. In addition, AM generally stored more glycogen than HP, whereas HP stored more lipids than AM. We proposed that multiple biomarkers may be employed as an integrated diagnostic tool for monitoring the health condition of the mussels., (Copyright © 2016 Elsevier Inc. All rights reserved.)

It is an open question whether adverse habitat conditions, characteristic for many anthropogenically impacted coastal habitats, can determine resistance to abiotic stress in populations of residing invertebrates. We tested experimentally for differences in stress tolerance between individuals of the Asian green mussel Perna viridis stemming from the heavily impacted Jakarta Bay and from two natural sites, Lada Bay and Pelabuhan Ratu, West Java. Mussel performance under hyposalinity and hypoxia was assessed in laboratory assays by measuring fitness-related response variables, e.g. body condition index, relative shell weight, byssus production, respiration rates and survival. We found stress-specific and population-specific differences in mussel resistance to adverse conditions: Individuals from the impacted Jakarta Bay performed better under hypoxia than their conspecifics from the natural sites, whereas the latter were more resistant to hyposalinity. We explain these differences by differential acclimation to environmental conditions in the respective habitats and by diverging degrees of food supply., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Mussels are important fouling organisms in the cooling water systems of coastal power plants. Continuous low-dose chlorination (CLDC) is being practiced as an effective method to control mussel biofouling in power plant cooling water systems. CLDC effectively controls mussel fouling by discouraging larval settlement rather than by killing the larvae or adults. Mussels are an integral part of the natural benthic community in the receiving water body where the coolant water is discharged. Hence, from a toxicological point of view, they can serve as both target and non-target organisms. Previous researchers have indicated that chlorine residual, rather than elevated temperature, can be the major stress factor in the effluents released from coastal power plants. However, very little data are available on the sub-lethal effects of low level chlorination on representative benthic fauna. In this study, we used native and transplanted mussels (Perna viridis) to study lethal and sub-lethal effects of chlorination in the cooling water circuit of an operating power plant. Experiments involving comet assay suggested that CLDC can cause DNA damage in treated mussels. However, activation of DNA repair appeared to get initiated after the accrued damage reached a threshold. The results indicate that, at chlorine residual levels observed at the discharge point, exposure to chlorinated effluents is unlikely to cause significant genetic damage to mussels in the recipient water body., (Copyright © 2016 Elsevier Inc. 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.)

Benzo(a)pyrene and dichlorodiphenyltrichloroethane are typical persistent organic pollutants, and also the widespread environmental estrogens with known toxicity towards green mussels Perna viridis. In this study, the toxicological effects of BaP and DDT and their mixture were assessed in green mussel gonads using proteomic and metabolomic approaches. Metabolomics by NMR spectroscopy revealed that BaP did not show obvious metabolite changes in the gonad of male green mussel. DDT mainly caused some disturbance of osmotic regulation and energy metabolism by changing BCAAs, alanine, threonine, arginine, etc., unknown metabolite (3.53 ppm), glycine, homarine and ATP at different levels. However, the mixture of BaP and DDT mainly caused some disturbance in osmotic regulation and energy metabolism by differentially altering branched chain amino acids, glutamate, alanine, arginine, unknown metabolite (3.53 ppm), glycine, 4-aminobutyrate, dimethylglycine, homarine and ATP. The results suggest that DDT alone may cause most of metabolites changes in the mixture exposed male mussel gonad, and the results also show that the male P. viridis gonad was more sensitive to DDT than BaP exposures. Proteomic study showed that BaP, DDT and their mixture may have different modes of action. Proteomic responses revealed that BaP induced signal transduction, oxidative stress, spermatogenesis, etc. in the male green mussel gonad; whereas DDT exposure altered proteins that were associated with signal transduction, oxidative stress, cytoskeleton and cell structure, cellular organization, energy metabolism, etc. However, the mixture of BaP and DDT affected proteins related to cytoskeleton and cell structure, oxidative stress, cellular organization, etc. This research demonstrated that metabolomic and proteomic approaches could better elucidate the underlying mechanism of environmental pollutants gonad toxicity., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

In Brazil B5 blend (5% biodiesel and 95% diesel oil) has been adopted as mandatory fuel since 2010 for automotive vehicles. Since little is known about the effects of B5 exposure can promote on antioxidant system of marine biota this study aimed to assess if B5 can generate modifications in antioxidant parameters of mussels Perna perna. To address this question mussels were exposed to two concentrations of B5 (0.01 mL L(-1) and 0.1 mL L(-1)) for 6h, 12h, 48 h and 168 h. Then the activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferase (GST) and glutathione reductase (GR) were evaluated in gills and digestive gland as well as the contents of glutathione (GSH) and lipid peroxidation by measuring the malondialdehyde concentration (MDA). In the gills, GST activity decreased after 48 h and GR after 12h of exposure to B5. In digestive glands, the activities of SOD, GPx and GR were changed due to treatments. GSH concentration increased in digestive gland after 6h and 12h and in gills after 48 h for B5 0.1 mL L(-1) and after 168 h in the digestive gland for B5 0.01 mL L(-1) treatment. No lipid peroxidation was detected. The integrated biomarker response index (IBR) evidenced a B5 effect in the digestive gland after 168 h of exposure. Regarding the experimental conditions and species used in this study, long-term exposure to B5 is apparently more likely to affect the parameters tested in P. perna mussels., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

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.)

Heavy metals contamination on the coast of Jakarta Bay has led to the level of pollution and can cause toxicity to organisms living in the sea, i.e., green mussels. Green mussels have the ability to detoxify metals entering their bodies. Their ability to accumulate metals is higher than other aquatic animals. This is due to their sedentary life which prevents them from avoiding the effects of pollution and their high tolerance to certain metals. The high concentration of metal content would be toxic to the cell because metal ions can act as oxidants and bind to organic and protein molecules. The results of the study showed that traces of heavy metals were detected in the hepatopancreas, gills, muscles and gonads organs of the mussels living in the waters of Muara Angke. Lead (Pb) and cadmium (Cd) were found in all four organs, while mercury (Hg) was not detected in the muscles. Traces of Hg and Cd were not detected in hepatopancreas, gills, muscles and gonads of green mussels in Panimbang, while Pb was detected by 0.00 1 in the male gonads and 0.01 in hepatopancreas. The concentration of Pb in the male gonads are still below the acceptable limit and concentration of Pb in the hepatopancreas is relatively equivalent to the acceptable limit. Metal detection in the organs above shows that the Muara Angke waters tend to be polluted and have an impact on the mussels weight loss as a result of heavy metal toxicity.

Background: Cryopreservation of P. canaliculus oocytes has not yet been achieved., Objective: The present study is to investigate whether the incorporation of: DMSO (0.09%), α-tocopherol (0.1 mM) plus taurine (1 mM) and ethylenediaminetetraacetic acid (EDTA; 0.1 mM), is beneficial during cryopreservation., Methods: These three additives were incorporated to both the cryoprotectant (CPA) and recovery media, and evaluated in terms of development and oxidative stress at three key stages of cryopreservation: 1) cryoprotectant addition [10% v/v ethylene glycol plus 0.2M trehalose; final concentration], 2) cooling to -6 degrees C, and 3) cooling to -35 degrees C and liquid nitrogen immersion., Results: Over all treatments (including controls) progressive cryopreservation steps resulted in a decrease in fertilization and development to D-larvae, an increase in macromolecular oxidative damage markers (protein carbonyls, lipid hydroperoxides and oxidized DNA), and a decrease in enzymatic (superoxide dismutase, catalase, glutathione S-transferase and glutathione reductase) and non-enzymatic antioxidants., Conclusion: Whilst results varied, the major effects of the additives were the improved percentage fertilization and a decrease in macromolecular damage.

Organic peroxide elimination in eukaryotes essentially depends on glutathione peroxidase (GPx) and peroxiredoxin (Prx) enzymes, which are supported by their respective electron donors, glutathione (GSH) and thioredoxin (Trx). This system depends on the ancillary enzymes glutathione reductase (GR) and thioredoxin reductase (TrxR) to maintain GSH and Trx in their reduced state. This study discusses the biological importance of GR and TrxR in supporting GPx and Prx during cumene hydroperoxide (CHP) exposure in brown mussel Perna perna. ZnCl2 or 1-chloro-2,4-dinitrobenze (CDNB) was used to decrease GR and TrxR activities in gills, as already reported with mammals and bivalves. ZnCl2 exposure lowered GR activity (28%), impaired the in vivo CHP decomposition and decreased the survival rates under CHP exposure. CDNB decreased GR (54%) and TrxR (73%) activities and induced glutathione depletion (99%), promoting diminished peroxide elimination and survival rates at a greater extent than ZnCl2. CDNB also increased the susceptibility of hemocytes to CHP toxicity. Despite being toxic and causing mortality at longer exposures, short (2 h) exposure to CHP promoted an up regulation of GSH (50 and 100 μM CHP) and protein-thiol (100 μM CHP) levels, which was blocked by ZnCl2 or CDNB pre-exposure. Results highlight the biological importance of GSH, GR and TrxR in supporting GPx and Prx activities, contributing to organic peroxides elimination and mussel survival under oxidative challenges. To our knowledge, this is the first work that demonstrates, albeit indirectly, the biological importance of GPx/GR/GSH and Prx/TrxR/Trx systems on in vivo organic peroxide elimination in bivalves., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Concerns regarding perfluorinated chemicals (PFCs) have risen in recent years because of their ubiquitous presence and high persistency. However, data on the environmental impacts of PFCs on marine organisms are very limited. Oxidative toxicity has been suggested to be one of the major toxic pathways for PFCs to induce adverse effects on organisms. To investigate PFC-induced oxidative stress and oxidative toxicity, a series of antioxidant enzyme activities and oxidative damage biomarkers were examined to assess the adverse effects of the following 4 commonly detected compounds: perfluoro-octanesulfonate, perfluoro-ocanoic acid, perfluorononanoic acid, and perfluorodecanoic acid, on green mussel (Perna viridis). Quantitative structure-activity relationship (QSAR) models were also established. The results showed that all the tested PFCs are able to induce antioxidant response and oxidative damage on green mussels in a dose-dependent manner. At low exposure levels (0 µg/L-100 µg/L), activation of antioxidant enzymes (catalase [CAT] and superoxide dismutase [SOD]) was observed, which is an adaptive response to the excessive reactive oxygen species induced by PFCs, while at high exposure levels (100 µg/L-10 000 µg/L), PFCs were found to inhibit some enzyme activity (glutathione S-transferase and SOD) where the organism's ability to respond in an adaptive manner was compromised. The oxidative stress under high PFC exposure concentration also led to lipid and DNA damage. PFC-induced oxidative toxicity was found to be correlated with the bioaccumulation potential of PFCs. Based on this relationship, QSAR models were established using the bioaccumulation factor (BAF) as the molecular descriptor for the first time. Compared with previous octanol-water partition coefficient-dependent QSAR models, the BAF-dependent QSAR model is more suitable for the impact assessment of PFCs and thus provides a more accurate description of the toxic behavior of these compounds., (© 2014 SETAC.)