Combined toxic effects of nanoplastics and norfloxacin on antioxidant and immune genes in mussels.

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.

The combined effects of phenanthrene and micro-/nanoplastics mixtures on the cellular stress responses of the thick-shell mussel Mytilus coruscus.

Pollution with complex mixtures of contaminants including micro- and nano-plastics (MNPs) and organic pollutants like polycyclic aromatic hydrocarbons (PAH) poses a major threat to coastal marine ecosystems. Toxic mechanisms of contaminant mixtures are not well understood in marine organisms. We studied the effects of single and combined exposures to polycyclic aromatic hydrocarbon phenanthrene (Phe) and MNPs mixture with sizes of 70 nm, 5 µm and 100 µm on the immune health and oxidative stress parameters in the thick-shell mussel Mytilus coruscus. Immune cells (hemocytes) were more sensitive to the pollutant-induced oxidative stress than the gills. In hemocytes of co-exposed mussels, elevated mortality, lower lysosomal content, high production of reactive oxygen species (ROS) and decrease mitochondrial were found. Disparate responses of antioxidant enzymes in the hemolymph (e.g. increased superoxide dismutase (SOD) activity without a corresponding increase in catalase (CAT) in Phe exposures and an increase in CAT without a change in SOD in MNPs exposures) suggests misbalance of the antioxidant defense in the pollutant-exposed mussels. Gill lacked pronounced oxidative stress response showing a decline in ROS and antioxidant levels. Tissue-specific single and combined effects of Phe and MNPs suggest variation in bioavailability and/or different sensitivity to these pollutants in the studied tissues. Notably, the combined effects of MNPs and Phe were additive or antagonistic, showing that MNPs do not enhance and occasionally mitigate the toxic effects of Phe on the hemocytes and the gills of the mussels. Overall, our study sheds light on the impact of long-term exposure to MNPs and Phe mixtures on mussels, showing high sensitivity of the immune system and modulation of the Phe toxicity by MNPs co-exposure. These findings that may have implications for understanding the impacts of combined PAH and MNPs pollution on the health of mussel populations from polluted coastal habitats.

Bio-based microplastic polylactic acid exerts the similar toxic effects to traditional petroleum-based microplastic polystyrene in mussels.

Microplastics (MPs) have accumulated in the oceans, causing adverse effects on marine organisms and the environment. Biodegradable polylactic acid (PLA) is considered as an excellent substitute for traditional petroleum-based plastics, but it is difficult to degrade completely and easily become MPs in the marine environment. To test the ecological risk of bio-based PLA, we exposed thick-shelled mussels (Mytilus coruscus) to bio-based PLA and petroleum-based polystyrene (PS) (at 102, 104, and 106 particles/L) for 14 days. The significant increase in enzyme activities related to oxidative stress and immune response showed that mussels were under physiological stress after MP ingestion. While enzyme activities of nerve conduction and energy metabolism were significantly disturbed after exposure. Meanwhile, normal physiological activities in respiration, ingestion and assimilation were also suppressed in association with enzyme changes. The negative effects of PS and PLA in mussels were not differentiated, and further integration analysis of integrated biomarker response (IBR) and principal component analysis (PCA) also showed that PLA would induce adverse effects in mussels and ecological risks as PS, especially at environmental concentrations. Therefore, it is necessary to pay more attention to the environmental and ecological risk of bio-based MP PLA accumulating in the marine environment.

Physiological responses of the microalga Isochrysis galbana exposed to polystyrene microplastics with different particle sizes.

Due to continuous increase in marine plastic waste, microplastics are ubiquitous in the marine environment. However, there are few studies on the harmful effects caused by microplastics with different particle sizes, and the interaction between particle size and concentration requires further investigation. This study explored the differences in physiological and biochemical responses, photosynthesis and oxidative stress damage of the microalga Isochrysis galbana exposed to three different particle size microplastics. It was found that different particle sizes and concentrations of microplastics resulted in significant differences (p < 0.05) in the growth rate, photosynthesis, and oxidative stress level of I. galbana. With the decrease of the particle size and lowering concentration of microplastics, the growth rate, photosynthesis and oxidative stress levels of I. galbana were reduced. Significant differences in photosynthesis and oxidative stress levels were observed when I. galbana was exposed to smallest particle size and lowest concentration of microplastics. This study provides new insights about whether polystyrene microplastics of different particle sizes and concentrations exhibit complex effects on microalgae, and explores the underlying reasons for such effects. In short, this study predicts the exacerbating adverse effects of microplastic pollution on the primary productivity, with significant implications for marine food webs and ecosystem health.

Polyamide microplastics can mitigate the effects of pathogenic bacterium on the health of marine mussels.

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.

Tris(1-chloro-2-propyl) phosphate enhances the adverse effects of biodegradable polylactic acid microplastics on the mussel Mytilus coruscus.

Microplastics (MPs) and organophosphate flame retardants (OPFRs) have recently become ubiquitous and cumulative pollutants in the oceans. Since OPFRs are added to or adsorbed onto MPs as additives, it is necessary to study the composite contamination of OPFRs and MPs, with less focus on bio-based PLA. Therefore, this study focused on the ecotoxicity of the biodegradable MP polylactic acid (PLA) (5 µm, irregular fragments, 102 and 106 particles/L), and a representative OPFRs tris(1-chloro-2-propyl) phosphate (TCPP, 0.5 and 50 µg/L) at environmental and high concentrations. The mussel Mytilus coruscus was used as a standardised bioindicator for exposure experiments. The focus was on examining oxidative stress (catalase, CAT, superoxide dismutase, SOD, malondialdehyde, MDA), immune responses acid (phosphatase, ACP, alkaline phosphatase, AKP, lysozyme, LZM), neurotoxicity (acetylcholinesterase, AChE), energy metabolism (lactate dehydrogenase, LDH, succinate dehydrogenase, SDH, hexokinase, HK), and physiological indices (absorption efficiency, AE, excretion rate, ER, respiration rate, RR, condition index, CI) after 14 days exposure. The results of significantly increased oxidative stress and immune responses, and significantly disturbed energy metabolism and physiological activities, together with an integrated biomarker response (IBR) analysis, indicate that bio-based PLA MPs and TCPP could cause adverse effects on mussels. Meanwhile, TCPP interacted significantly with PLA, especially at environmental concentrations, resulting in more severe negative impacts on oxidative and immune stress, and neurotoxicity. The more severe adverse effects at environmental concentrations indicate higher ecological risks of PLA, TCPP and their combination in the real marine environment. Our study presents reliable data on the complex effects of bio-based MP PLA, TCPP and their combination on marine organisms and the environment.

Combined effects of norfloxacin and polystyrene nanoparticles on the oxidative stress and gut health of the juvenile horseshoe crab Tachypleus tridentatus.

Pollution with anthropogenic contaminants including antibiotics and nanoplastics leads to gradual deterioration of the marine environment, which threatens endangered species such as the horseshoe crab Tachypleus tridentatus. We assessed the potential toxic mechanisms of an antibiotic (norfloxacin, 0, 0.5, 5 µg/L) and polystyrene nanoparticles (104 particles/L) in T. tridentatus using biomarkers of tissue redox status, molting, and gut microbiota. Exposure to single and combined pollutants led to disturbance of redox balance during short-term (7 days) exposure indicated by elevated level of a lipid peroxidation product, malondialdehyde (MDA). After prolonged (14-21 days) exposure, compensatory upregulation of antioxidants (catalase and glutathione but not superoxide dismutase) was observed, and MDA levels returned to the baseline in most experimental exposures. Transcript levels of molting-related genes (ecdysone receptor, retinoic acid X alpha receptor and calmodulin A) and a molecular chaperone (cognate heat shock protein 70) showed weak evidence of response to polystyrene nanoparticles and norfloxacin. The gut microbiota T. tridentatus was altered by exposures to norfloxacin and polystyrene nanoparticles shown by elevated relative abundance of Bacteroidetes. At the functional level, evidence of suppression by norfloxacin and polystyrene nanoparticles was found in multiple intestinal microbiome pathways related to the genetic information processing, metabolism, organismal systems, and environmental information processing. Future studies are needed to assess the physiological and health consequences of microbiome dysbiosis caused by norfloxacin and polystyrene nanoparticles and assist the environmental risk assessment of these pollutants in the wild populations of the horseshoe crabs.

Physiological response of mussel to rayon microfibers and PCB's exposure: Overlooked semi-synthetic micropollutant?

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".

Combined effects of microfibers and polychlorinated biphenyls on the immune function of hemocytes in the mussel Mytilus coruscus.

Numerous studies have shown that microplastics can interact with other pollutants in the environment to produce synergistic effects, leading to more serious impacts. To date, there is little consensus on the combined effects of microfibers (MFs) and polychlorinated biphenyls (PCBs, Aroclor 1254), two legacy and alarming environmental pollutants. There is an urgent need to assess the impact of combined exposures on bivalve immune defences. In this study, we assessed the immune response of the mussels (Mytilus coruscus) hemocyte to MFs and PCBs alone and in combination by using flow cytometry. M. coruscus were exposed to MFs (1000 pieces/L) and PCBs (PCBs) (100 ng/L and 1000 ng/L) alone or in combination for 14 consecutive days and recovered for 7 days. The hemocyte of M. coruscus was collected on day 7, 14 and 21. MF exposure alone had no effect on the hemocyte. The total hemocyte count (THC), esterase (EA), lysosomal contents (LC), mitochondrial number (MN) and mitochondrial membrane potential (MMP) of mussels showed a decreasing trend with increasing PCB concentrations, both individually and in combination; The decreases in EA, MN and MMP were associated with the induction of reactive oxygen species (ROS). Hemocyte mortality (HM) was associated with a decrease in THC. Combined exposure to MFs and PCBs would exacerbate the effects on hemocyte immunity. These new findings improve our understanding of the toxic effects of MFs and organic chemical pollutants, and demonstrate the potential mechanism of PCBs to bivalves through changes in hemolymph immunity-related indicators.

Response of moulting genes and gut microbiome to nano-plastics and copper in juvenile horseshoe crab Tachypleus tridentatus.

Nanoplastics (NPs) and heavy metals are typical marine pollutants, affecting the gut microbiota composition and molting rate of marine organisms. Currently, there is a lack of research on the toxicological effects of combined exposure to horseshoe crabs. In this study, we investigated the effects of NPs and copper on the expression of molt-related genes and gut microbiome in juvenile tri-spine horseshoe crabs Tachypleus tridentatus by exposing them to NPs (100 nm, 104 particles L-1) and/or Cu2+ (10 µgL-1) in seawater for 21 days. Compared with the control group, the relative mRNA expression of ecdysone receptor (EcR), retinoid x receptor (RXR), calmodulin-A-like isoform X1 (CaM X1), and heat shock 70 kDa protein (Hsp70) were significantly increased under the combined stress of NPs and Cu2+. There were no significant differences in the diversity and abundance indices of the gut microbial population of horseshoe crabs between the NPs and/or Cu2+ groups and the control group. According to linear discriminant analysis, Oleobacillus was the most abundant microorganism in the NPs and Cu2+ stress groups. These results indicate that exposure to either NPs stress alone or combined NPs and Cu2+ stress can promote the expression levels of juvenile molting genes. NPs exposure has a greater impact on the gut microbial community structure of juvenile horseshoe crabs compared to Cu2+ exposure. This study is helpful for predicting the growth and development of horseshoe crabs under complex environmental pollution.

A short review on the recent method development for extraction and identification of microplastics in mussels and fish, two major groups of seafood.

The prevalence of microplastics in the marine environment poses potential health risks to humans through seafood consumption. Relevant data are available but the diverse analytical approaches adopted to characterise microplastics have hampered data comparison among studies. Here, the techniques for extraction and identification of microplastics are summarised among studies of marine mussels and fish, two major groups of seafood. Among the reviewed papers published in 2018-2021, the most common practice to extract microplastics was through tissue digestion in alkaline chemicals (46 % for mussels, 56 % for fish) and oxidative chemicals (28 % for mussels, 12 % for fish). High-density solutions such as sodium chloride could be used to isolate microplastics from other undigested residues by flotation. Polymer analysis of microplastics was mainly carried out with Fourier-transform infrared (FTIR) spectroscopy (58 % for both mussels and fish) and Raman spectroscopy (14 % for mussels, 8 % for fish). Among these methods, we recommend alkaline digestion for microplastic extraction, and the automated mapping approach of FTIR or Raman spectroscopy for microplastic identification. Overall, this study highlights the need for a standard protocol for characterising microplastics in seafood samples.

Combined toxic effects of nanoplastics and norfloxacin on mussel: Leveraging biochemical parameters and gut microbiota.

Antibiotics and nanoplastics (NPs) are among the two most concerned and studied marine emerging contaminants in recent years. Given the large number of different types of antibiotics and NPs, there is a need to apply efficient tools to evaluate their combined toxic effects. Using the thick-shelled mussel (Mytilus coruscus) as a marine ecotoxicological model, we applied a battery of fast enzymatic activity assays and 16S rRNA sequencing to investigate the biochemical and gut microbial response of mussels exposed to antibiotic norfloxacin (NOR) and NPs (80 nm polystyrene beads) alone and in combination at environmentally relevant concentrations. After 15 days of exposure, NPs alone significantly inhibited superoxide dismutase (SOD) and amylase (AMS) activities, while catalase (CAT) was affected by both NOR and NPs. The changes in lysozyme (LZM) and lipase (LPS) were increased over time during the treatments. Co-exposure to NPs and NOR significantly affected glutathione (GSH) and trypsin (Typ), which might be explained by the increased bioavailable NOR carried by NPs. The richness and diversity of the gut microbiota of mussels were both decreased by exposures to NOR and NPs, and the top functions of gut microbiota that were affected by the exposures were predicted. The data fast generated by enzymatic test and 16S sequencing allowed further variance and correlation analysis to understand the plausible driving factors and toxicity mechanisms. Despite the toxic effects of only one type of antibiotics and NPs being evaluated, the validated assays on mussels are readily applicable to other antibiotics, NPs, and their mixture.

Adverse impacts of high-density microplastics on juvenile growth and behaviour of the endangered tri-spine horseshoe crab Tachypleus tridentatus.

The impacts of high-density microplastics, namely polyamine 6,6 (nylon), polymethyl methacrylate (PMMA) and polyethylene terephthalate (PET), on growth and behaviour of the endangered tri-spine horseshoe crab Tachypleus tridentatus were investigated for 100 days. Negative changes in wet weight and prosomal width of the juveniles were observed in all treatments of microplastics, but significant difference was only detected in prosomal width between control and PMMA. T. tridentatus became significantly less active upon exposure to nylon and PET. The extent of burrowing by T. tridentatus did not significantly differ among the treatments but was overall significantly reduced towards day 100. T. tridentatus exposed to PET significantly showed the lowest survival probability (30 %), compared to the other treatments (70-90 %). In conclusion, high-density microplastics compromised growth and behaviour of juvenile horseshoe crabs. Among the polymers that were tested, PET was considered more harmful and associated with higher mortality.

Microplastics can aggravate the impact of ocean acidification on the health of mussels: Insights from physiological performance, immunity and byssus properties.

Ocean acidification may increase the risk of disease outbreaks that would challenge the future persistence of marine organisms if their immune system and capacity to produce vital structures for survival (e.g., byssus threads produced by bivalves) are compromised by acidified seawater. These potential adverse effects may be exacerbated by microplastic pollution, which is forecast to co-occur with ocean acidification in the future. Thus, we evaluated the impact of ocean acidification and microplastics on the health of a mussel species (Mytilus coruscus) by assessing its physiological performance, immunity and byssus properties. We found that ocean acidification and microplastics not only reduced hemocyte concentration and viability due to elevated oxidative stress, but also undermined phagocytic activity of hemocytes due to lowered energy budget of mussels, which was in turn caused by the reduced feeding performance and energy assimilation. Byssus quality (strength and extensibility) and production were also reduced by ocean acidification and microplastics. To increase the chance of survival with these stressors, the mussels prioritized the synthesis of some byssus proteins (Mfp-4 and Mfp-5) to help maintain adhesion to substrata. Nevertheless, our findings suggest that co-occurrence of ocean acidification and microplastic pollution would increase the susceptibility of bivalves to infectious diseases and dislodgement risk, thereby threatening their survival and undermining their ecological contributions to the community.

Is microplastic an oxidative stressor? Evidence from a meta-analysis on bivalves.

Microplastic pollution is a major threat to the marine environment attracting attention from scientific and public communities. Although we have sufficient evidence that microplastic is ubiquitous in all ecosystems, the question of the harmfulness of microplastic exposure is still under debate. Filter feeders like bivalves are commonly exposed to microplastics in water and sediments and thus can serve as excellent biological indicators for microplastic pollution. A relatively rich toxicological literature has been focusing on microplastic effects on bivalves but we have yet to reach an agreement on the toxic effects and mechanisms of microplastics. Here, we conducted a meta-analysis and bibliometrics analysis of the microplastic studies in bivalves. The bibliometric analysis (used to evaluate the general research trends) showed that the investigation of microplastic distribution in the marine environment and the molecular mechanisms of microplastic toxicity are the two major hot spots of research. Based on analyses of ecologically and environmentally relevant microplastics concentrations, particle sizes and polymer types, we discuss the physiological effects of microplastics on bivalves, and the severity and direction of the effects at the cellular, tissue, organ and organismal levels. The meta-analysis results show that microplastics can induce time-dependent oxidative stress in bivalves. Generally, the activities of antioxidant enzymes, such as glutathione peroxidase (GPx), glutathione-S-transferase (GST) and superoxide dismutase (SOD) increased during short-term exposure but declined after long-term exposure to microplastics. Non-linear response of GPx, GST and SOD enzymes to MP exposure over time indicate that these enzymes are not good biomarkers of MPs effects in marine bivalves. The tissue glutathione levels and catalase (CAT activity) showed an increase during both short- and long term MP exposures and thus can be used as oxidative stress biomarkers of sublethal MPs effects in marine bivalves.

Microplastic accumulation via trophic transfer: Can a predatory crab counter the adverse effects of microplastics by body defence?

Microplastics are considered detrimental to aquatic organisms due to their potential accumulation along food chains. Thus, it is puzzling why some of them appear unaffected by microplastics. Here, we assessed the contribution of water filtration and food consumption to microplastic accumulation in a predatory marine crab (Charybdis japonica) and examined the associated impacts of microplastics (particle size: 5 µm) following ingestion for one week. Results showed that water filtration and food consumption contributed similarly to the accumulation of microplastics, which were distributed among organs in this order: hepatopancreas > guts > gills > muscles. Yet, biomagnification (i.e. accumulation through consumption of microplastic-contaminated mussels) did not occur possibly due to egestion of microplastics. The crabs upregulated detoxification capacity (EROD) and antioxidant defence (GST) in response to the microplastics accumulated in their tissues. However, these defence mechanisms collapsed when the microplastic concentration in hepatopancreas exceeded ~3 mg g-1, leading to severe hepatic injury (elevated AST and ALT) and impaired neural activity (reduced AChE). Our results suggest that marine organisms have an innate capacity to counter the acute effects of microplastics, but there is a limit beyond which the defence mechanisms fail and hence physiological functions are impaired. As microplastic pollution will deteriorate in the future, the fitness and survival of marine organisms may be undermined by microplastics, affecting the stability and functioning of marine ecosystems.

Ingestion of nano/micro plastic particles by the mussel Mytilus coruscus is size dependent.

Plastic particles are thought to accumulate in aquatic organisms and cause potential physiological effects. The uneven sizes of plastic particles may affect the ingestion by marine filter feeding bivalves and may lead to differential further physiological effects. To tackle this scientific question, we investigated the size dependent ingestion and dynamic accumulation of nano/micro plastic particles with different diameters (0.07, 0.5, 5, 10 and 100 µm) in the thick shell mussel Mytilus coruscus. The accumulation of particles in gill, digestive tract and mantle of mussels was measured after 3, 15, 87 h exposure and following 87 h depuration. The results showed that particle ingestion was negatively size dependant and positively related to time in digestive tract. In mantle, particles accumulated over the depuration time with a delay, indicating the translocation of particles. Moreover, our results showed that gill was not a target tissue for steady particle accumulation but the digestive tract was. This study showed size dependent and dynamic ingestion of nano/micro particles in mussels which are one of the main marine organisms for accumulating microplastics.

Physiological effects of plastic particles on mussels are mediated by food presence.

Plastic particles cause toxic effects on marine organisms but whether food particles can affect the toxic effects of plastic particles on filter feeding animals remains unknown. To evaluate the intake and physiological effects of different size particles and their exposure ways, the thick shell mussels Mytilus coruscus were exposed to polystyrene (PS) nanoplastics (NPs, 70 nm) and microplastics (MPs, 10 µm) respectively for two weeks by mixing NPs/MPs with microalgae or exposed to MNPs individually after feeding. Intake of particles and their physiological effects including energy budget, digestive enzymes and oxidative responses were assessed after exposure. Results indicated food presence mediate the effects while MPs decrease the energy budget and increase the catalase activity and malondialdehyde levels. Moreover, exposure way significantly affected energy budget and size of particle had a significant impact on enzyme activities. Our results showed MPs induce more significant effects than NPs on mussels, emphasized the importance of particle exposure way and suggested that mixture exposure with microalgae alleviate the influences on mussels caused by plastic particles alone. This study emphasized that we need to take the food particles into account for evaluating the toxic effects of plastic particles on filter feeding animals in the natural environment.

Improved Raman spectroscopy-based approach to assess microplastics in seafood.

Microplastics represent an emerging environmental issue and have been found almost everywhere including seafood, raising a great concern about the ecological and human health risks they pose. This study addressed the common technical challenges in the assessment of microplastics in seafood by developing an improved protocol based on Raman spectroscopy and using the green-lipped mussel Perna viridis and the Japanese jack mackerel Trachurus japonicus as the test models. Our findings identified a type of stainless-steel filter membranes with minimal Raman interference, and a combination of chemicals that achieved 99-100% digestion efficiency for both organic and inorganic biomass. This combined chemical treatment reached 90-100% recovery rates for seven types of microplastics, on which the surface modification was considered negligible and did not affect the accuracy of polymer identification based on Raman spectra, which showed 94-99% similarity to corresponding untreated microplastics. The developed extraction method for microplastics was further combined with an automated Raman mapping approach, from which our results confirmed the presence of microplastics in P. viridis and T. japonicus collected from Hong Kong waters. Identified microplastics included polypropylene, polyethylene, polystyrene and poly(ethylene terephthalate), mainly in the form of fragments and fibres. Our protocol is applicable to other biological samples, and provides an improved alternative to streamline the workflow of microplastic analysis for routine monitoring purposes.

Determination of microplastics in the edible green-lipped mussel Perna viridis using an automated mapping technique of Raman microspectroscopy.

Microplastics are prevalent in marine environments and seafood and thus can easily end up in human diets. This has raised serious concerns worldwide, particularly in Hong Kong where the seafood consumption per capita can be three times higher than the global average. This study focused on the green-lipped mussel Perna viridis, a popular seafood species which is subject to a high risk of contamination by microplastics due to its filter-feeding nature. P. viridis was collected from five mariculture sites in Hong Kong and assessed for its body load of microplastics using an automated Raman mapping approach. Microplastics were found in all sites, with an average of 1.60-14.7 particles per mussel per site, or 0.21-1.83 particles per g wet weight. Polypropylene, polyethylene, polystyrene and polyethylene terephthalate were detected among the microplastics, mainly as fragments or fibres in the size range of 40-1000 µm. It was estimated that through consumption of P. viridis, the population in Hong Kong could ingest up to 10,380 pieces of microplastics per person per year. These estimated rates were high compared to the values reported worldwide, suggesting the potential human health risk of microplastics in Hong Kong and adjacent areas.