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Global climate change is a major trigger of unexpected temperature fluctuations. The impacts of marine heatwaves (MHWs) and nano-titanium dioxide (nano-TiO2) on marine organisms have been extensively investigated. However, the potential mechanisms underlying their interactive effects on physiological processes and metabolism remain poorly understood, especially regarding periodic MHWs in real-world conditions. In this study, the effects of nano-TiO2 (at concentrations of 0, 25, and 250 µg/L) and periodic MHWs on the condition index (CI) and underlying metabolic mechanisms were investigated in mussels (Mytilus coruscus). The results showed that mussels try to upregulate their respiration rate (RR) to enhance aerobic metabolism (indicated by elevated succinate dehydrogenase) under short-term nano-TiO2 exposure. However, even at ambient concentration (25 µg/L), prolonged nano-TiO2 exposure inhibited ingestion ability (decreased clearance rate) and glycolysis (inhibited pyruvate kinase, hexokinase, and phosphofructokinase activities), which led to an insufficient energy supply (decreased triglyceride, albumin, and ATP contents). Repeated thermal scenarios caused more severe physiological damage, demonstrating that mussels are fragile to periodic MHWs. MHWs decreased the zeta potential of the nano-TiO2 particles but increased the hydrodynamic diameter. Additionally, exposure to nano-TiO2 and periodic MHWs further affected aerobic respiration (inhibited lactate dehydrogenase and succinate dehydrogenase activities), metabolism (decreased RR, activities of respiratory metabolism-related enzymes, and expressions of PEPCK, PPARγ, and ACO), and overall health condition (decreased ATP and CI). These findings indicate that the combined stress of these two stressors exerts more detrimental impact on the physiological performance and energy metabolism of mussels, and periodic MHWs exacerbate the toxicological effects of ambient concentration nano-TiO2. Given the potential worsening of nanoparticle pollution and the increase in extreme heat events in the future, the well-being of mussels in the marine environment may face further threats.

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Vibrio parahaemolyticus and microplastics are prevalent in the ocean. Bacteria attach onto plastic particles, forming harmful biofilms that collectively threaten bivalve health. This study investigates the interaction between polyamide microplastics (PA: particle size 38 ± 12 µm) and V. parahaemolyticus, as well as their combined impact on thick-shelled mussels (Mytilus coruscus). We introduced 1011 CFU/L of V. parahaemolyticus into varying PA concentrations (0, 5, 50, and 500 particles/L) to observe growth over 14 h and biofilm formation after 48 h. Our findings indicate that microplastics suppress biofilm formation and virulence gene expression. Four treatments were established to monitor mussel responses: a control group without PA or V. parahaemolyticus; a group with 50 particles/L PA; a group with 1011 CFU/L V. parahaemolyticus; and a co-exposure group with both 50 particles/L PA and 1011 CFU/L V. parahaemolyticus, over a 14-day experiment. However, combined stress from microplastics and Vibrio led to immune dysregulation in mussels, resulting in intestinal damage and microbiome disruption. Notably, V. parahaemolyticus had a more severe impact on mussels than microplastics alone, yet their coexistence reduced some harmful effects. This study is the first to explore the interaction between microplastics and V. parahaemolyticus, providing important insights for ecological risk assessments.

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The homologous E6-AP carboxy-terminal structural domain (HECT) contained in E3 ubiquitin ligases (E3s) is a key factor in protein degradation and maintenance of cellular homeostasis in animals. However, the functional roles and evolutionary aspects of the HECT gene family in bivalve mussels remain unclear and warrant further investigation. In this study, we identified 22 HECT genes within the genome of Mytilus coruscus Gould, all containing a conserved HECT structural domain derived from dispersed repeats, distributed unevenly across 11 chromosomes. Phylogenetic analysis classified M. coruscus HECT genes into six major classes, with amino acid sequences within the same evolutionary clade displaying similar conserved motifs. Homology analysis with HECT genes of four bivalve species revealed that M. coruscus and Mytilus galloprovincialis possessed the largest number of homologous gene pairs, showing a significant correlation between the two in the evolution of the HECT gene family. Homology analysis with HECT genes of four bivalve species revealed that M. coruscus and M. galloprovincialis possessed the largest number of homologous gene pairs, showing a significant correlation between the two in the evolution of the HECT gene family. M. coruscus exhibited pronounced and specific expression in gills and blood tissues. Notably, Mco_UPL3 gene expression was significantly upregulated after 12 h of acute heat stress (33 °C) and 24 h of Vibrio injection (0.4 OD). Gene ontology analysis of the HECT genes in M. coruscus revealed that it is primarily enriched in protein modification and degradation functions. This suggests that HECT genes may play a key role in protein degradation and immunomodulation in M. coruscus. These findings offer valuable insights for the breeding of stress-tolerant traits in M. coruscus. In summary, our data shed light on the potential functions of HECT E3 ligases in response to heat stress and Vibrio infection, providing practical guidance for enhancing resilience through breeding in M. coruscus.

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In the marine environment, nanoparticles play a role in adsorbing and catalytically degrading organic pollutants, thereby mitigating their toxic effects on aquatic organisms. This study aimed to investigate the impact of nano titanium dioxide (nTiO2) and tris (2-chloropropyl) phosphate (TCPP) on the hemolymph and digestive function of the thick-shell mussel Mytilus coruscus. Mussels were divided into a control group, a group exposed to TCPP alone, a group exposed to a combination of TCPP and 0.5 mg/L nTiO2, and a group exposed to a combination of TCPP and 1 mg/L nTiO2. After 14 days of exposure, oxidative stress responses, including superoxide dismutase (SOD) activity and malondialdehyde (MDA) content, immune defense responses, including acid phosphatase (ACP) and alkaline phosphatase (AKP) activities, and gene expression, including HSP70 expression, were measured in the hemolymph and digestive glands of the mussels. Compared to the control group, mussels solely exposed to 100 µg/L TCPP exhibited a significant reduction in SOD activity in the hemolymph. When TCPP was co-exposed with 0.5 mg/L nTiO2, there were significant increases in MDA content and AKP activity in both the digestive gland and hemolymph compared to the control group. Upon co-exposure of TCPP with 1 mg/L nTiO2, MDA content and AKP activity in the digestive gland significantly decreased, while SOD, ACP, and AKP activity in the hemolymph significantly increased and MDA content significantly decreased, returning to the control group levels. Furthermore, in the combined exposure, HSP70 gene expression significantly decreased as the nTiO2 concentration increased from 0.5 mg/L to 1 mg/L. In summary, TCPP impacted the hemolymph and digestive function of mussels, whereas a concentration of 1 mg/L nTiO2 effectively alleviated the toxic effects of TCPP. This study is crucial for assessing the ecological risks of nanoparticles and emerging organic pollutants in marine environments, and provides new insights into the interaction between nTiO2 and TCPP, as well as the influence of nTiO2 concentration on mitigating TCPP toxicity.

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Phthalate esters (PAEs), as a major plasticizer with multi-biotoxicity, are frequently detected in marine environments, and potentially affecting the survival of aquatic organisms. In the study, three typical PAEs (dimethyl phthalate [DMP], dibutyl phthalate [DBP] and di(2-ethylhexyl) phthalate [DEHP]) were selected to investigate the accumulation patterns and ecotoxicological effects on Mytilus coruscus (M. coruscus). In M. coruscus, the accumulation was DEHP>DBP>DMP, and the bioaccumulation in tissues was digestive glands>gills>gonads>muscles. Meanwhile, the activities of superoxide dismutase (SOD) and catalase (CAT) showed an activation-decrease-activation trend of stress, with more pronounced concentration effects. Glutathione reductase (GSH) activity was significantly increased, and its expression was more sensitive to be induced at an early stage. The metabolic profiles of the gonads, digestive glands and muscle tissues were significantly altered, and DEHP had a greater effect on the metabolic profiles of M. coruscus, with the strongest interference. PAEs stress for 7 d significantly altered the volatile components of M. coruscus, with potential implications for their nutritional value. This study provides a biochemical, metabolomic, and nutritional analysis of DMP, DBP, and DEHP toxic effects on M. coruscus from a multidimensional perspective, which provides support for ecotoxicological studies of PAEs on marine organisms. ENVIRONMENTAL IMPLICATION: Phthalate esters (PAEs), synthetic compounds from phthalic acid, are widespread in the environment, household products, aquatic plants, animals, and crops, posing a significant threat to human health. However, the majority of toxicological studies examining the effects of PAEs on aquatic organisms primarily focus on non-economic model organisms like algae and zebrafish. Relatively fewer studies have been conducted on marine organisms, particularly economically important shellfish. So, this study is innovative and necessary. This study provides a biochemical, metabolomic, and nutritional analysis of DMP, DBP, and DEHP toxic effects on mussels, and supports the ecotoxicology of PAEs on marine organisms.

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Mussel byssal proteins are of biomimetic importance for the development of novel underwater bio-adhesive agents. It is important to maintain a reduced state during the process of byssus adhesion. There are 19 mussel foot proteins (MFPs) have been reported in previous studies, among which only MFP-6 had been confirmed as an antioxidant protein in mussel byssus due to the function of cysteines, and playing an essential role in the redox balance of mussel byssus during adhesion process. Although the other four MFPs (MFP-16 ~ MFP-19) also have abundant cysteines, their function is still unknown. In this study, a novel mussel foot protein, named MFP-20, was identified from Mytilus coruscus foot. The sequential features, expression profile, and function of recombinant MFP-20 were verified. The results showed that MFP-20 has more abundant cysteines than other MFPs, the relative expression of mfp-20 was upregulated in Fe3+ stress and low pH seawater. In addition, different adhesive substrates induced significant changes of expression level of mfp-20. Furthermore, rMFP-20 showed strong antioxidant capacity in the DPPH assay, and the abundant cysteines in its sequence may play vital roles in the antioxidation activity. Our findings revealed the possible function of MFP-20 with a totally different sequence from the reported MFP-6 and provided new clues for exploring the redox balance of mussel byssus during the adhesion process.

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Environmental stressors such as salinity fluctuations can significantly impact the ecological dynamics of mussel beds. The present study evaluated the influence of hyposalinity stress on the detachment and survival of attached mussels by simulating a mussel farming model in a laboratory setting. Byssus production and mechanical properties of thread in response to varying salinity levels were assessed, and histological sections of the mussel foot were analyzed to identify the changes in the byssus secretory gland area. The results showed that hyposalinity stress (20 and 15 psu) led to a significant decrease in mussel byssus secretion, delayed initiation of new byssus production, and reduced plaque adhesion strength and breaking force of byssal threads compared to the control (30 psu) (p < 0.05). The complete suppression of byssal thread secretion in mussels under salinity conditions of 10 and 5 psu, leading to lethality, indicates the presence of a blockade in byssus secretion when mussels are subjected to significant physiological stressors. Histological analysis further demonstrated a decrease in the percentage of foot secretory gland areas in mussels exposed to low salinities. However, contrary to expectations, the study found that mussels did not exhibit marked detachment from ropes in response to the reduced salinity levels during one week of exposure. Hyposalinity stress exposure reduced the byssal secretion capacity and the mechanical properties of threads, which could be a cause for the detachment of suspension-cultured mussels. These results highlight the vulnerability of mussels to hyposalinity stress, which significantly affects their byssus mechanical performance.

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Exposing marine organisms to contemporary contaminants, such as perfluorooctanoic acid (PFOA) and nano-titanium dioxide (nano-TiO2), can induce multifaceted physiological consequences. Our investigation centered on the responses of the mussel, Mytilus coruscus, to these agents. We discerned pronounced disruptions in gill filament connections, pivotal structures for aquatic respiration, suggesting compromised oxygen uptake capabilities. Concurrently, the respiratory rate exhibited a marked decline, indicating a respiratory distress. Furthermore, the mussels' clearance rate, a metric of their filtration efficacy, diminished, suggesting the potential for bioaccumulation of deleterious substances. Notably, the co-exposure of PFOA and nano-TiO2 exhibits interactive effects on the physiological performance of the mussels. The mussels' digestive performance waned in the face of heightened PFOA and nano-TiO2 concentrations, possibly hampering nutrient assimilation and energy accrual. This was mirrored in the noticeable contraction of their energy budget, suggesting long-term growth repercussions. Additionally, the dysregulation of the gut microbiota and the reduction in its diversity further confirm alterations in intestinal homeostasis, subsequently impacting its physiological functions and health. Collectively, these findings underscore the perils posed by escalated PFOA and nano-TiO2 levels to marine mussels, accentuating the need for a deeper understanding of nanoparticle-pollutant synergies in marine ecosystems.

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Histones and their N-terminal or C-terminal derived peptides have been studied in vertebrates and presented as potential antimicrobial agents playing important roles in the innate immune defenses. Although histones and their derived peptides had been reported as components of innate immunity in invertebrates, the knowledge about the histone derived antimicrobial peptides (HDAPs) in invertebrates are still limited. Using a peptidomic technique, a set of peptide fragments derived from the histones was identified in this study from the serum of microbes challenged Mytilus coruscus. Among the 85 identified histone-derived-peptides with high confidence, 5 HDAPs were chemically synthesized and the antimicrobial activities were verified, showing strong growth inhibition against Gram-positive bacteria, Gram-negative bacteria, and fungus. The gene expression level of the precursor histones matched by representative HDAPs were further tested using q-PCR, and the results showed a significant upregulation of the histone gene expression levels in hemocytes, gill, and mantle of the mussel after immune stress. In addition, three identified HDAPs were selected for preparation of specific antibodies, and the corresponding histones and their derived C-terminal fragments were detected by Western blotting in the blood cell and serum of immune challenged mussel, respectively, indicating the existence of HDAPs in M. coruscus. Our findings revealed the immune function of histones in Mytilus, and confirmed the existence of HDAPs in the mussel. The identified Mytilus HDAPs represent a new source of immune effector with antimicrobial function in the innate immune system, and thus provide promising candidates for the treatment of microbial infections in aquaculture and medicine.

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Perfluorooctanoate (PFOA) is widely used as a surfactant and has metabolic, immunologic, developmental, and genetic toxicity on marine organisms. However, the effects of PFOA on individual defense functions in mussels in the presence of titanium dioxide nanoparticles (nano-TiO2) are poorly understood. To investigate the defense strategies and regulatory mechanisms of mussels under combined stressors, the thick-shell mussels Mytilus coruscus were exposed to different PFOA concentrations (0, 2 and 200 µg/L) and nano-TiO2 (0 and 0.1 mg /L, size: 25 nm) for 14 days. The results showed that, compared to the control group, PFOA and nano-TiO2 significantly reduced the number of byssal threads (NBT), byssal threads length (BTL), diameter of proximal threads (DPB), diameter of middle threads (DMB), diameter of distal byssal threads (DDB), adhesive plaque area (BPA), and breaking force of byssal threads (N). Under the influence of PFOA and nano-TiO2, the morphological surface smoothness of the fractured byssal threads surface increased, concurrently inducing an increased surface roughness in the adhesive plaques. Additionally, under the presence of PFOA and nano-TiO2, the foot displayed dispersed tissue organization and damaged villi, accompanied by an increased incidence of cellular apoptosis and an upregulation of the apoptosis gene caspase-8. Expression of the adhesion gene mfp-3 and byssal threads strength genes (preCOL-D, preCOL-NG) was upregulated. An interactive effect on the performance of byssal threads is observed under the combined influence of PFOA and nano-TiO2. Under co-exposure to PFOA and nano-TiO2, the performance of the byssal threads deteriorates, the foot structure is impaired, and the genes mRNA expression of byssal thread secretory proteins have compensated for the adhesion and byssal threads strength by up-regulation. Within marine ecosystems, organic and particulate contaminants exert a pronounced effect on the essential life processes of individual organisms, thereby jeopardizing their ecological niche within community assemblages and perturbing the dynamic equilibrium of the overarching ecosystem. ENVIRONMENTAL IMPLICATION: Perfluorooctanoic acid (PFOA) is prone to accumulate in marine organisms. TiO2 nanoparticles (nano-TiO2) are emerging environmental pollutants frequently found in marine environment. The effects of PFOA and nano-TiO2 on marine mussels are not well understood, and their toxic mechanisms remain largely unknown. We investigated the impacts of PFOA and nano-TiO2 on mussel byssus defense mechanisms. By assessing byssus performance indicators, morphological structures of the byssus, subcellular localization, and changes in byssal secretion-related genes, we revealed the combined effects and mechanisms through which these two types of pollutants may affect the functional capabilities and survival of mussels in the complex marine ecosystem.

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BACKGROUND: Apoptosis is involved (directly and indirectly) in several physiological processes including tissue remodeling during the development, the turnover of immune cells, and a defense against harmful stimuli. The disordered apoptotic process participates in the pathogenesis of various diseases, such as neoplasms, and chronic inflammatory or systemic autoimmune diseases, which are associated with its inadequate regulation. Caspases are vital components of the apoptotic pathway that are involved in developmental and immune processes. However, genome-wide identification and functional analysis of caspase have not been conducted in Mytilus coruscus, which is an economically important bivalve. RESULTS: Here, 47 caspase genes were identified from the genomes of M. coruscus, and the expansion of caspase-2/9 and caspase-3/6/7 genes were observed. Tandem duplication acts as an essential driver of gene expansion. The expanded caspase genes were highly diverse in terms of sequence, domain structure, and spatiotemporal expression profiles, suggesting their functional differentiation. The high expression of the expanded caspase genes at the pediveliger larvae stage and the result of apoptosis location in the velum suggest that the apoptosis mediated by them plays a critical role in the metamorphosis of M. coruscus larvae. In gill, caspase genes respond differently to the challenge of different strains, and most caspase-2/9 and caspase-3/6/7 genes were induced by copper stress, whereas caspase-8/10 genes were suppressed. Additionally, most caspase genes were upregulated in the mantle under ocean acidification which could weaken the biomineralization capacity of the mantle tissue. CONCLUSIONS: These results provide a comprehensive overview of the evolution and function of the caspase family and enhanced the understanding of the biological function of caspases in M. coruscus larval development and response to biotic and abiotic challenges.

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Rayon microfibers, micro-sized semi-synthetic polymers derived from cellulose, have been frequently detected and reported as "micropollutants" in marine environments. However, there has been limited research on their ecotoxicity and combined effects with persistent organic pollutants (POPs). To address these knowledge gaps, thick-shell mussels (Mytilus coruscus) were exposed to rayon microfibers at 1000 pieces/L, along with polychlorinated biphenyls (PCBs) at 100 and 1000 ng/L for 14 days, followed by a 7-day recovery period. We found that rayon microfibers at the environmentally relevant concentration exacerbated the irreversible effects of PCBs on the immune and digestive systems of mussels, indicating chronic and sublethal impacts. Furthermore, the results of 16 s rRNA sequencing demonstrated significant effects on the community structure, species richness, and diversity of the mussels' intestinal microbiota. The branching map analysis identified the responsive bacteria to rayon microfibers and PCBs belonging to the Proteobacteria, Actinobacteriota, and Bacteroidota phyla. Despite not being considered a conventional plastic, the extensive and increasing use of rayon fibers, their direct toxicological effects, and their interaction with POPs highlight the need for urgent attention, investigation, and regulation to address their contribution to "micropollution".

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Growth is a fundamental aspect of aquaculture breeding programs, pivotal for successful cultivation. Understanding the mechanisms that govern growth and development differences across various stages can significantly boost seedling production of economically valuable species, thereby enhancing aquaculture efficiency and advancing the aquaculture industry. Mytilus coruscus, a commercially vital marine bivalve, underscores this importance. To decipher the intricate molecular mechanisms dictating growth and developmental disparities in marine shellfish, we conducted transcriptome sequencing and meticulously analyzed gene expression variations and molecular pathways linked to growth traits in M. coruscus. This study delved into the molecular and gene expression variations across five larval development stages, with a specific focus on scrutinizing the differential expression patterns of growth-associated genes using RNA sequencing and quantitative real-time PCR analysis. A substantial number of genes-36,044 differentially expressed genes (DEGs)-exhibited significant differential expression between consecutive developmental stages. These DEGs were then categorized into multiple pathways (Q value < 0.05), including crucial pathways such as the spliceosome, vascular smooth muscle contraction, DNA replication, and apoptosis, among others. In addition, we identified two pivotal signaling pathways-the Hedgehog (Hh) signaling pathway and the TGF-beta (TGF-ß) signaling pathway-associated with the growth and development of M. coruscus larvae. Ten key growth-related genes were pinpointed, each playing crucial roles in molecular function and the regulation of growth traits in M. coruscus. These genes and pathways associated with growth provide deep insights into the molecular basis of physiological adaptation, metabolic processes, and growth variability in marine bivalves.

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Succinate dehydrogenase inhibitor fungicides (SDHIs) are frequently detected in the marine environment. However, studies on the toxicity of SDHIs to marine organisms, Mytilus coruscus (M. coruscus), are poorly reported. Therefore, the antioxidant activities and metabolomic response of four SDHIs, namely, boscalid (BC), thifluzamide (TF), fluopyram (FO), and bixafen (BIX), to (M. coruscus), were comprehensively investigated. The antioxidant activity of BC and TF was significantly increased (p<0.05), whereas those of FO and BIX were significantly decreased. Furthermore, metabolite discriminations among M. coruscus to four SDHIs were illustrated by an untargeted metabolomics approach. A total of 52, 50, 93, and 129 differential metabolites were obtained for BC, TF, FO, and BIX. KEGG of the different metabolites show that the four SDHIs had differential effects on the metabolic pathways of M. coruscus. The current study demonstrated four SDHIs triggered glucose metabolism, lipid metabolism, tricarboxylic acid cycle, and oxidative phosphorylation processes and caused the disruption of nutrient and energy conversion processes in mussels. Finally, five biomarkers were screened by analyzing common differential metabolites that emerged from the four SDHI exposures, which could be used for risk assessment of marine ecosystem exposure to SDHIs. Our results demonstrated the use of metabolomics to understand the potential mechanisms of toxicity of four SDHIs to mussels and to identify potential targets for future targeted risk assessment.

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Coastal habitats are exposed to increasing pressure of nanopollutants commonly combined with warming due to the seasonal temperature cycles and global climate change. To investigate the toxicological effects of TiO2 nanoparticles (TiO2 NPs) and elevated temperature on the intestinal health of the mussels (Mytilus coruscus), the mussels were exposed to 0.1 mg/L TiO2 NPs with different crystal structures for 14 days at 20 °C and 28 °C, respectively. Compared to 20 °C, the agglomeration of TiO2 NPs was more serious at 28 °C. Exposure to TiO2 NPs led to elevated mortality of M. coruscus and modified the intestinal microbial community as shown by 16S rRNA sequence analysis. Exposure to TiO2 NPs changed the relative abundance of Bacteroidetes, Proteobacteria and Firmicutes. The relative abundances of putative mutualistic symbionts Tenericutes and Fusobacteria increased in the gut of M. coruscus exposed to anatase, which have contributed to the lower mortality in this group. LEfSe showed the combined stress of warming and TiO2 NPs increased the risk of M. coruscus being infected with potential pathogenic bacteria. This study emphasizes the toxicity differences between crystal structures of TiO2 NPs, and will provides an important reference for analyzing the physiological and ecological effects of nanomaterial pollution on bivalves under the background of global climate change.

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Harmful algal blooms (HABs) and their associated phycotoxins are increasing globally, posing great threats to local coastal ecosystems and human health. Nutrients have been carried by the freshwater Yangtze River and have entered the estuary, which was reported to be a biodiversity-rich but HAB-frequent region. Here, in situ solid phase adsorption toxin tracking (SPATT) was used to monitor lipophilic shellfish toxins (LSTs) in seawaters, and extended local similarity analysis (eLSA) was conducted to trace the temporal and special regions of those LSTs in a one-year trail in a mussel culture ranch in the Yangtze River Estuary. Nine analogs of LSTs, including okadaic acid (OA), dinophysistoxin-1 (DTX1), yessotoxin (YTX), homoyessotoxin (homoYTX), 45-OH-homoYTX, pectenotoxin-2 (PTX2), 7-epi-PTX2 seco acid (7-epi-PTX2sa), gymnodimine (GYM) and azaspiracids-3 (AZA3), were detected in seawater (SPATT) or rope farmed mussels. The concentrations of OA + DTX1 and homoYTX in mussels were positively correlated with those in SPATT samplers (Pearson test, p < 0.05), indicating that SPATT (with resin HP20) would be a good monitoring tool and potential indicator for OA + DTX1 and homoYTX in mussel Mytilus coruscus. The eLSA results indicated that late summer and early autumn were the most phycotoxin-contaminated seasons in the Yangtze River Estuary. OA + DTX1, homoYTX, PTX2 and GYM were most likely driven by the local growing HAB species in spring and summer, while Yangtze River diluted water may impact the accumulation of HAB species, causing potential phycotoxin contamination in the Yangtze River Estuary in autumn and winter. Together, the results showed that the mussel harvesting season, late summer and early autumn, would be the season with the greatest phycotoxin risk and would be the most contaminated by local growing toxic algae. Routine monitoring sites should be set up close to the local seawaters.

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Micro-/nano-plastic particles (MNPs) are present in the ocean with potential detrimental impacts on marine ecosystems. Bivalves are often used as marine bioindicators and are ideal to evaluate the threat posed by various-sized MNPs. We exposed the mussel Mytilus coruscus to MNPs with different particle sizes (70 and 500 nm, 5, 10 and 100 µm) for 3, 72 h and 30 days. The antioxidant responses in digestive gland and the hemolymph were then evaluated. The time of exposure played a strong modulating role in the biological response. A 3-hour exposure had no significant impact on the digestive gland. After 72 h, an increase in oxidative stress was observed in the digestive gland, including increased hydrogen peroxide (H2O2) level, catalase (CAT), glutathione peroxidase (GPx) activities and malondialdehyde (MDA) production. After a 30-day exposure, the oxidative stress decreased while lipid peroxidation increased. A 30-day exposure increased hemocyte mortality (HM) and reactive oxygen species (ROS) levels in the hemolymph, while phagocytosis (PA), lysosome content (LC), mitochondrial number (MN) and mitochondrial membrane potential (MMP) significantly decreased. Longer-term exposure to MNPs caused oxidative stress in the digestive gland as well as impaired viability and immunity of hemocytes. Particle size also influenced the response with smaller particles having more severe effects. A depuration for 7 days was enough to reverse the negative effects observed on the digestive gland and hemolymph. This study provides new insights on the effects of small-sized MNPs, especially nanoplastic particles (NPs), on aquatic organisms, and provides a solid theoretical knowledge background for future studies on toxic effects of MNPs.

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Nanoplastics (NPs) and antibiotics (ABs) are two of the emerging marine contaminants that have drawn the most attention in recent years. Given the necessity of figuring out the effects of plastic and antibiotic contamination on marine organism life and population in the natural environment, it is essential to apply rapid and effective biological indicators to evaluate their comprehensive toxic effects. In this study, using mussel (Mytilus coruscus) as a model, we investigated the combined toxic effects of NP (80 nm polystyrene beads) and AB (Norfloxacin, NOR) at environmental-relevant concentrations on antioxidant and immune genes. In terms of the antioxidant genes, NPs significantly increased the relative expression of Cytochrome P450 3A-1 (CYP3A-1) under various concentrations of NOR conditions, but they only significantly increased the relative expression of CYP3A-2 in the high concentration (500 µg L-1 NOR) co-exposure group. In the NP-exposure group which exposed to no or low concentrations of NOR, nuclear factor erythroid 2-related factor 2 (Nrf2) was upregulated. In terms of the immune genes, interleukin-1 receptor-associated kinase (IRAK) -1 showed a significant increase in the low-concentration NOR group while a significant inhibition in the high-concentration NOR group. Due to the presence of NPs, exposure to NOR resulted in a significant increase in both IRAK-4 and heat shock protein (HSP) 70. Our findings indicate that polystyrene NPs can exacerbate the effects of NOR on the anti-oxidant and immune defense performance of mussels. This study delves into the toxic effects of NPs and ABs from a molecular perspective. Given the expected increase in environmental pollution due to NPs and ABs, future research is needed to investigate the potential synergistic effect of NPs and ABs on other organisms.

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Mytilus coruscus is an economically important marine bivalve that lives in estuarine sea areas with seasonal coastal acidification and frequently suffers shell injury in the natural environment. However, the molecular responses and biochemical properties of Mytilus under these conditions are not fully understood. In the present study, we employed tandem mass spectrometry combined with isobaric tagging to identify differentially expressed proteins in the mantle tissue of M. coruscus under different short-term treatments, including shell-complete mussels raised in normal seawater (pH 8.1), shell-damaged mussels raised in normal seawater (pH 8.1), and acidified seawater (pH 7.4). A total of 2694 proteins were identified in the mantle, and analysis of their relative abundance from the three different treatments revealed alterations in the proteins involved in immune regulation, oxidation-reduction processes, protein folding and processing, energy provision, and cytoskeleton. The results obtained by quantitative proteomic analysis of the mantle allowed us to delineate the molecular strategies adopted by M. coruscus in the shell repair process in acidified environments, including an increase in proteins involved in oxidation-reduction processes, protein processing, and cell growth at the expense of proteins involved in immune capacity and energy metabolism. SIGNIFICANCE: The impact of global ocean acidification on calcifying organisms has become a major ecological and environmental problem in the world. Mytilus coruscus is an economically important marine bivalve living in estuary sea area with seasonal coastal acidification, and frequently suffering shell injury in natural environment. Molecular responses of M coruscus under the shell damage and acute acidification is still largely unknown. For this reason, iTRAQ based quantitative proteomic and histological analysis of the mantle from M. coruscus under shell damage and acute acidification were performed, for revealing the proteomic response and possible adaptation mechanism of Mytilus under combined shell damage and acidified sea water, and understanding how the mussel mantle implement a shell-repair process under acidified sea water. Our study provides important data for understanding the shell repair process and proteomic response of Mytilus under ocean acidification, and providing insights into potential adaptation of mussels to future global change.

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The nuclear factor erythroid 2-related factor 2 (Nrf2) is a pivotal regulator of antioxidant gene expression in mammals, forming heterodimer complexes with small Maf proteins through its BZip domain. However, the underlying mechanism of Nrf2 action in molluscs remains poorly understood. The thick shell mussel, Mytilus coruscus, represents a model organism for the marine environment and molluscs interaction research. In this study, we used in silico cloning to obtain a small Maf homologue called McMafF_G_K from M. coruscus. McMafF_G_K possesses a typical BZip domain, suggesting its affiliation with the traditional small Maf family and its potential involvement in the Nrf2 signaling pathway. Transcriptional analysis revealed that McMafF_G_K exhibited a robust response to benzo[a]pyrene (Bap) in the digestive glands. However, this response was down-regulated upon interference with McMafF_G_K-siRNA. Interestingly, the expression levels of Nrf2, NAD(P)H: quinone oxidoreductase (NQO-1), and Glutathione Peroxidase (GPx), which are key players in oxidative stress response, showed a positive correlation with McMafF_G_K in digested adenocytes of M. coruscus. Furthermore, in vitro analysis of antioxidant capacity in digestive gland cells demonstrated that Bap exposure led to an increase in reactive oxygen species (ROS) levels, accompanied by an elevation in total antioxidant capacity (T-AOC), potentially counterbalancing the excessive ROS. Strikingly, transfection of McMafF_G_K siRNA resulted in a significant rise in ROS level and a down-regulation of T-AOC level. To validate the functional relevance of McMafF_G_K, a glutathione S-transferase (GST) pull-down assay confirmed its interaction with McNrf2, providing compelling evidence of their protein interaction. This study significantly contributes to our understanding of the functional role of McMafF_G_K in the Nrf2 signaling pathway and sheds light on its potential as a target for further research in oxidative stress response.

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