Pearl Farming Micro-Nanoplastics Affect Oyster Physiology and Pearl Quality.

Pearl farming is crucial for the economy of French Polynesia. However, rearing structures contribute significantly to plastic waste, and the widespread contamination of pearl farming lagoons by microplastics has raised concerns about risks to the pearl industry. This study aimed to evaluate the effects of micro-nanoplastics (MNPs, 0.4-200 µm) on the pearl oyster (Pinctada margaritifera) over a 5-month pearl production cycle by closely mimicking ecological scenarios. MNPs were produced from weathered plastic pearl farming gear and tested at environmentally relevant concentrations (0.025 and 1 µg L-1) to decipher biological and functional responses through integrative approaches. The significant findings highlighted the impacts of MNPs on oyster physiology and pearl quality, even at remarkably low concentrations. Exposure to MNPs induced changes in energy metabolism, predominantly driven by reduced assimilation efficiency of microalgae, leading to an alteration in gene expression patterns. A distinct gene expression module exhibited a strong correlation with physiological parameters affected by MNP conditions, identifying key genes as potential environmental indicators of nutritional-MNP stress in cultured oysters. The alteration in pearl biomineralization, evidenced by thinner aragonite crystals and the presence of abnormal biomineral concretions, known as keshi pearls, raises concerns about the potential long-term impact on the Polynesian pearl industry.

Effect of an environmental microplastic mixture from the Seine River and one of the main associated plasticizers, dibutylphthalate, on the sentinel species Hediste diversicolor.

The aim of this study was to explore the adverse effects of a microplastic (MP) mixture obtained from litter accumulated in the Seine River (France) compared to those of their major co-plasticizer, dibutylphthalate (DBP), on the sentinel species Hediste diversicolor. A suite of biomarkers has been investigated to study the impacts of MPs (100 mg kg-1 sediment), DBP (38 µg kg-1 sediment) on worms compared to non-exposed individuals after 4 and 21 days. The antioxidant response, immunity, neurotoxicity and energy and respiratory metabolism were investigated using biomarkers. After 21 days, worms exposed to MPs showed an increasing aerobic metabolism, an enhancement of both antioxidant and neuroimmune responses. Energy-related biomarkers demonstrated that the energy reallocated to the defence system may come from proteins. A similar impact was depicted after DBP exposure, except for neurotoxicity. Our results provide a better understanding of the ecotoxicological effects of environmental MPs and their associated-contaminants on H. diversicolor.

Effect of aging of microplastics on gene expression levels of the marine mussel Mytilus edulis: Comparison in vitro/in vivo exposures.

In the present study, effects of aging MPs of polyethylene (PE) were investigated in the marine mussel Mytilus edulis, commonly used as bioindicator of aquatic ecosystem, using both in vitro and in vivo exposures, using concentrations found in marine waters (0.008, 10 and 100 µg.L-1). Changes in gene expression levels implicated in detoxification, immune system, cytoskeletton and cell cycle control were evaluated by quantitative RT-qPCR. Results demonstrated differential expression levels depending upon the state of plastic degradation (aged vs non-aged) and way of exposure (vitro vs vivo). This study highlighted the interest of using molecular biomarkers based on analysis of gene expression pattern in an ecotoxicological context that gives indication of relative slight changes between tested conditions as compared to other biochemical approaches (e.g. enzymatic activities). In addition, in vitro analysis could be used to generate large amount of data as regards to the toxicological effects of MPs.

Exposure to polyethylene microplastics alters immature gut microbiome in an infant in vitro gut model.

Infants are characterized by an immaturity of the gut ecosystem and a high exposure to microplastics (MPs) through diet, dust and suckling. However, the bidirectional interactions between MPs and the immature infant intestinal microbiota remain unknown. Our study aims to investigate the impact of chronic exposure to polyethylene (PE) MPs on the gut microbiota and intestinal barrier of infants, using the new Toddler mucosal Artificial Colon coupled with a co-culture of epithelial and mucus-secreting cells. Gut microbiota composition was determined by 16S metabarcoding and microbial activities were evaluated by gas, short chain fatty acid and volatolomics analyses. Gut barrier integrity was assessed via evaluation of intestinal permeability, inflammation and mucus synthesis. Exposure to PE MPs induced gut microbial shifts increasing α-diversity and abundance of potentially harmful pathobionts, such as Dethiosulfovibrionaceae and Enterobacteriaceae. Those changes were associated to butyrate production decrease and major changes in volatile organic compounds profiles. In contrast, no significant impact of PE MPs on the gut barrier, as mediated by microbial metabolites, was reported. For the first time, this study indicates that ingestion of PE MPs can induce perturbations in the gut microbiome of infants. Next step would be to further investigate the potential vector effect of MPs.

Discussion about suitable applications for biodegradable plastics regarding their sources, uses and end of life.

This opinion paper offers a scientific view on the current debate of the place of biodegradable plastics as part of the solution to deal with the growing plastic pollution in the world's soil, aquatic, and marine compartments. Based on the current scientific literature, we focus on the current limits to prove plastic biodegradability and to assess the toxicity of commercially used biobased and biodegradable plastics in natural environments. We also discuss the relevance of biodegradable plastics for selected applications with respect to their use and end of life. In particular, we underlined that there is no universal biodegradability of plastics in any ecosystem, that considering the environment as a waste treatment system is not acceptable, and that the use of compostable plastics requires adaptation of existing organic waste collection and treatment channels.

Microplastics: What happens in the human digestive tract? First evidences in adults using in vitro gut models.

Microplastics (MPs) are ubiquitous in the environment and humans are inevitably exposed to them. However, the effects of MPs in the human digestive environment are largely unknown. The aim of our study was to investigate the impact of repeated exposure to polyethylene (PE) MPs on the human gut microbiota and intestinal barrier using, under adult conditions, the Mucosal Artificial Colon (M-ARCOL) model, coupled with a co-culture of intestinal epithelial and mucus-secreting cells. The composition of the luminal and mucosal gut microbiota was determined by 16S metabarcoding and microbial activities were characterized by gas, short chain fatty acid, volatolomic and AhR activity analyses. Gut barrier integrity was assessed via intestinal permeability, inflammation and mucin synthesis. First, exposure to PE MPs induced donor-dependent effects. Second, an increase in abundances of potentially harmful pathobionts, Desulfovibrionaceae and Enterobacteriaceae, and a decrease in beneficial bacteria such as Christensenellaceae and Akkermansiaceae were observed. These bacterial shifts were associated with changes in volatile organic compounds profiles, notably characterized by increased indole 3-methyl- production. Finally, no significant impact of PE MPs mediated by changes in gut microbial metabolites was reported on the intestinal barrier. Given these adverse effects of repeated ingestion of PE MPs on the human gut microbiota, studying at-risk populations like infants would be a valuable advance.

Cryogrinding and sieving techniques as challenges towards producing controlled size range microplastics for relevant ecotoxicological tests.

The impact of microplastics (MP) has attracted much attention from the scientific community and many laboratory assessments have been made of their effects on aquatic organisms. To produce MP from real environmental plastic waste, which would enable more realistic experiments, we used plastic pearl farming equipment from French Polynesian lagoons. Here, the pearl oyster Pinctada margaritifera could encounter MP coming from their breakdown in its surrounding environment. We tested an established method based on mechanical cryogenic grinding and liquid sieving. Our desired size range was 20-60 µm, corresponding to the optimal particle size ingested by P. margaritifera. The protocol was effective, generating MP particles of 20-60 µm (∼17,000-28,000 MP µg-1), but also produced too many smaller particles. The peak in the desired size range was thus flattened by the many small particles <3 µm (∼82,000-333,000 MP µg-1; 53-70% of total analysed particles), visible at the limit of Coulter counter analysis (cut-off point: 2 µm). Laser diffraction analysis (cut-off point: 0.4 µm) provided greater detail, showing that ∼80-90% of the total analysed particles were <1 µm. Diverging particle size distributions between those expected based on sieving range and those really observed, highlight the need to perform fine-scaled particle size distribution analyses to avoid underestimating the number of small micro- and nanoplastics (MNP) and to obtain an exact estimation of the fractions produced. Size and microstructure characterization by scanning electron microscopy suggested spontaneous particle self-assembly into crystal superstructures, which is the supposed cause of the divergence we observed. Overall, our results emphasize that particle self-assembly is a technical hurdle requiring further work and highlight the specific need to finely characterize the size distribution of MNP used in ecotoxicological experiments to avoid overestimating effects.

Seasonal and spatial distribution of microplastics in sediments by FTIR imaging throughout a continuum lake - lagoon- beach from the Tunisian coast.

Plastics pollution in marine environment has become an issue of increasing scientific concern. This work aims to study the temporal and spatial distribution of plastics in sediments from three different Tunisian ecosystems; Rimel Beach, Bizerta lagoon and Ichkeul lake. Sediment sampling was conducted in surface (2 cm) and depth (15 cm) during spring, summer and winter. Plastics debris were separated by size fractions: macro (>5 mm), meso (1-5 mm) and microplastics (<1 mm) to optimize the time necessary for their characterisation. Macroplastics and mesoplastics were identified using an IR Attenuated total reflectance (ATR) and microplastics with Imaging Fourier transform infrared spectroscopy (FTIR) spectroscopy after an optimized extraction protocol. Results indicate that, the lowest contamination degree with macroplastics was recorded in Ichkeul lake, 2 macro debris/m2 (marine protected area, national parc of Ichkeul). Mesoparticles were only detected in lagoon of Bizerte in large quantities (4900 item/kg of sediment in surface and 680 item/kg of sediment in depth) and were identified principally as paint products. For microplastics, the repartition was quite homogeneous between the three sites with an average abundance was 130.55 ± 65.61 items/kg for all seasons. The variations of microplastics abundances on the three sites could not be clearly related to the seasons. Whereas the polymer characterisation in the surface and depth sediments of the three studied areas were principally due to eight types of polymers (PVC, PET, PP, PE PS, Polyamide (PA) and polymeric methyl methacrylate (PMMA)) as reported in many other studies, surprisingly all MPs recovered in the study were smaller than 300 µm, >70 % being fragments. This study brings new results as regards to the state of plastic contamination in Tunisian coast and shows the importance of investigating different ecosystems in such studies.

Metabolite Quantification by Fourier Transform Infrared Spectroscopy in Diatoms: Proof of Concept on Phaeodactylum tricornutum.

Diatoms are feedstock for the production of sustainable biocommodities, including biofuel. The biochemical characterization of newly isolated or genetically modified strains is seminal to identify the strains that display interesting features for both research and industrial applications. Biochemical quantification of organic macromolecules cellular quotas are time-consuming methodologies which often require large amount of biological sample. Vibrational spectroscopy is an essential tool applied in several fields of research. A Fourier transform infrared (FTIR) microscopy-based imaging protocol was developed for the simultaneous cellular quota quantification of lipids, carbohydrates, and proteins of the diatom Phaeodactylum tricornutum. The low amount of sample required for the quantification allows the high throughput quantification on small volume cultures. A proof of concept was performed (1) on nitrogen-starved experimental cultures and (2) on three different P. tricornutum wild-type strains. The results are supported by the observation in situ of lipid droplets by confocal and brightfield microscopy. The results show that major differences exist in the regulation of lipid metabolism between ecotypes of P. tricornutum.

Chemicals sorbed to environmental microplastics are toxic to early life stages of aquatic organisms.

Microplastics are ubiquitous in aquatic ecosystems, but little information is currently available on the dangers and risks to living organisms. In order to assess the ecotoxicity of environmental microplastics (MPs), samples were collected from the beaches of two islands in the Guadeloupe archipelago, Petit-Bourg (PB) located on the main island of Guadeloupe and Marie-Galante (MG) on the second island of the archipelago. These samples have a similar polymer composition with mainly polyethylene (PE) and polypropylene (PP). However, these two samples are very dissimilar with regard to their contamination profile and their toxicity. MPs from MG contain more lead, cadmium and organochlorine compounds while those from PB have higher levels of copper, zinc and hydrocarbons. The leachates of these two samples of MPs induced sublethal effects on the growth of sea urchins and on the pulsation frequency of jellyfish ephyrae but not on the development of zebrafish embryos. The toxic effects are much more marked for samples from the PB site than those from the MG site. This work demonstrates that MPs can contain high levels of potentially bioavailable toxic substances that may represent a significant ecotoxicological risk, particularly for the early life stages of aquatic animals.

An Irgafos® 168 story: When the ubiquity of an additive prevents studying its leaching from plastics.

Plastic pollution is a source of chemical to the environment and wildlife. Despite the ubiquity of plastic pollution and thus plastic additive in the environment, plastic additives have been studied to a limited extend. As a prerequisite to a study aiming to evaluate the leaching of a common additive used as an antioxidant (Irgafos® 168) from polyethylene microparticles, an inventory of the potential background contamination of the laboratory workplace was done. In this study, Irgafos® 168 (tris(2,4-ditert-butylphenyl) phosphite) and its oxidized form (tris (2,4-ditert-butylphenyl) phosphate) were quantified in different laboratory reagents, including the plastic packaging and the powders, using Pyrolysis-GC/MS. At least one form of Irgafos® 168 was detected in all tested laboratory reagents with higher concentrations in caps and bottles as compared to the powders. Additionally, oxidized Irgafos® 168 was also found in the reverse osmosed and deionized water container used in the laboratory. The same profile of contamination, i.e. higher concentration of the oxidized form and higher concentrations in acidic reagents, was observed when comparing the reagent and their respective containers suggesting that the additive is leaching from the container into the powder. Overall, this study demonstrates that the antioxidant additive Irgafos® 168 is ubiquitous in the laboratory workplace. Plastic additives such as Irgafos® 168 can therefore largely interfere and biased ecotoxicological and toxicological studies especially using environmentally relevant concentrations of microplastics. The source, fate and effects of plastic additive from plastic debris should be carefully considered in future studies that require setting up methods to overcome these contaminations.

Interactions between polystyrene nanoparticles and Chlamydomonas reinhardtii monitored by infrared spectroscopy combined with molecular biology.

For several decades, use of nanoparticles (NP) on a global scale has been generating new potential sources of organism disruption. Recent studies have shown that NP can cause modifications on the biochemical macromolecular composition of microalgae and raised questions on the toxicity of plastic particles, which are widespread in the aquatic environment. Polystyrene (PS) particles are among the most widely used plastics in the world. In our experimentation, a combined approach of infrared spectroscopy and molecular biology (real-time PCR) has been applied in order to better apprehend the consequences of interactions between Chlamydomonas reinhardtii, freshwater microalgae and PS NP. Two references have been used, nitrogen deprivation -a well-documented stressor-, and gold nanoparticles (Au-NP). As regards biochemical composition, our experiments show a differing microalga response, according to the NP to which they have been exposed. Results with infrared spectroscopy and gene expression methods are consistent and illustrate variation among several carbohydrates (galactose…). Furthermore, PS-NP seem to react in the same direction as nitrogen limitation, thereby supporting the hypothesis that PS-NP can induce response mechanisms to environmental changes in microalgae. This study highlighted the interest of combining infrared spectroscopy and gene expression as means of monitoring microalgae response to nanoplastics.

Infrared spectroscopy as a tool to monitor interactions between nanoplastics and microalgae.

The unicellular photosynthetic organisms known as microalgae are becoming one of the most important models for aquatic system studies. Among them, Chlamydomonas reinhardtii is widely used as a bioindicator of pollution or of different changes in the environment. Numerous pollutants are present in aquatic environments, particularly plastics and nanoplastics. Physiological variations after an environmental change highlight variation in the macromolecular composition of microalgae (proteins, nucleic acids, lipids and carbohydrates). Recently, Fourier transform infrared vibrational spectroscopy has been described as a reliable tool, sensitive and allowing rapid measurement of macromolecular composition of microalgae. Coupled with preprocessing and principal component analysis, it is well adapted to monitoring the effect of environmental stress on biochemical composition. In this study, infrared spectroscopy, combined with multivariate analysis, has been tested first on known environmental stresses such as light intensity variation and nitrogen limitation. Then, this technique has been applied to monitor the interaction and potential impacts of polystyrene nanoparticles on microalgae. The results showed slight variations on protein and carbohydrates bands in the presence of nanoplastics, suggesting that their presence led to modifications in the biochemical composition of the microalgae. To confirm the interaction between microalgae and nanoplastics, visualization by confocal microscopy and cytotoxicity measurement has been carried out. Results showed that polystyrene nanoparticles seemed to adsorb on microalgae surface, leading to a loss of plasma membrane integrity. The resulting chemical modifications, even if moderate, could be detected by infrared spectroscopy' showing that this tool could be very helpful in the understanding of nanoparticle-microalgae interaction mechanisms.

Realistic environmental exposure to microplastics does not induce biological effects in the Pacific oyster Crassostrea gigas.

The aim of the present study was to evaluate the presence and potential toxic effects of plastic fragments (<400 µm) of polyethylene and polypropylene on the Pacific oyster Crassostrea gigas. Oysters were exposed to environmentally relevant concentrations (0, 0.008, 10, 100 µg of particles/L) during 10 days, followed by a depuration period of 10 days in clean seawater. Effects of microplastics were evaluated on the clearance rate of organisms, tissue alteration, antioxidant defense, immune alteration and DNA damage. Detection and quantification of microplastics in oyster's tissues (digestive gland, gills and other tissues) and biodeposits using infrared microscopy were also conducted. Microplastics were detected in oyster's biodeposits following exposure to all tested concentrations: 0.003, 0.006 and 0.05 particles/mg of biodeposits in oysters exposed to 0.008, 10 and 100 µg of particles/L, respectively. No significant modulation of biological markers was measured in organisms exposed to microplastics in environmentally relevant conditions.

Current methods to monitor microalgae-nanoparticle interaction and associated effects.

Widespread use of nanoparticles for different applications has diffused their presence in the environment, particularly in water. Many studies have been conducted to evaluate their effects on aquatic organisms. Microalgae are at the base of aquatic trophic chains. These organisms which can be benthic or pelagic, meaning that they can enter into interaction with all kinds of particulate materials whatever their density, and constitute an interesting model study. The purpose of this review was to gather more than sixty studies on microalgae exposure to the different nanoparticles that may be present in the aquatic environment. After a brief description of each type of nanoparticle (metals, silica and plastic) commonly used in ecotoxicological studies, techniques to monitor their properties are presented. Then, different effects on microalgae resulting from interaction with nanoparticles are described as well as the parameters and techniques for monitoring them. The impacts described in the literature are primarily shading, ions release, oxidative stress, adsorption, absorption and disruption of microalgae barriers. Several parameters are proposed to monitor effects such as growth, photosynthesis, membrane integrity, biochemical composition variations and gene expression changes. Finally, in the literature, while different impacts of nanoparticles on microalgae have been described, there is no consensus on evidence of nanomaterial toxicity with regard to microalgae. A parallel comparison of different nanoparticle types appears essential in order to prioritize which factors exert the most influence on toxicity in microalgae cultures: size, nature, surface chemistry, concentration or interaction time.

From macroplastics to microplastics: Role of water in the fragmentation of polyethylene.

In this work, the artificial photodegradation of polyethylene films was studied in laboratory to compare the fragmentation pathways of this polymer at air and in water. Oxidation, surface mechanical properties, crystallinity and crack propagation were monitored to investigate their influence on fragmentation. Without any external stress, fragmentation only occurred in water despite a higher level of oxidation for films weathered at air. The cracking of the films did not appear correlated with the oxidation level and the presence of water appeared as a promoter of cracking propagation. The results also showed that the mechanical properties at the surface play a major role in the fragmentation pathway whereas the fabrication process may influence the propagation direction of the cracks. Consequently, the distribution in size of plastic fragments in the aquatic environment may be linked to the nature of the polymer but also to its manufacturing process. In this study, after 25 weeks of weathering in water, 90% of the fragments were >1 mm with very similar shapes showing that micrometric fragments were not yet abundant. These results suggest that long times of weathering in water and many steps of fragmentation appear necessary from macroplastics to reach sizes <1 mm in the aquatic environment. These results constitute a first attempt to understand the pathways leading from macroplastics to microplastics in water. They have to be confirmed for other polymers and the long-term behavior of the fragments needs to be studied to predict their decrease in size among time.

Raman Tweezers for Small Microplastics and Nanoplastics Identification in Seawater.

Our understanding of the fate and distribution of micro- and nano- plastics in the marine environment is limited by the intrinsic difficulties of the techniques currently used for the detection, quantification, and chemical identification of small particles in liquid (light scattering, vibrational spectroscopies, and optical and electron microscopies). Here we introduce Raman Tweezers (RTs), namely optical tweezers combined with Raman spectroscopy, as an analytical tool for the study of micro- and nanoplastics in seawater. We show optical trapping and chemical identification of sub-20 µm plastics, down to the 50 nm range. Analysis at the single particle level allows us to unambiguously discriminate plastics from organic matter and mineral sediments, overcoming the capacities of standard Raman spectroscopy in liquid, intrinsically limited to ensemble measurements. Being a microscopy technique, RTs also permits one to assess the size and shapes of particles (beads, fragments, and fibers), with spatial resolution only limited by diffraction. Applications are shown on both model particles and naturally aged environmental samples, made of common plastic pollutants, including polyethylene, polypropylene, nylon, and polystyrene, also in the presence of a thin eco-corona. Coupled to suitable extraction and concentration protocols, RTs have the potential to strongly impact future research on micro and nanoplastics environmental pollution, and enable the understanding of the fragmentation processes on a multiscale level of aged polymers.

Microplastic abundance and characteristics in French Atlantic coastal sediments using a new extraction method.

The ubiquitous presence of microplastics (MPs) has been demonstrated in all environmental compartments in the recent years. They are detected in air, freshwater, soil, organisms and particularly in marine ecosystems. Since sediments are known to be the major sink of many organic and inorganic pollutants, the aim of this study was to develop and validate a fast and cheap methodology to assess the MP contamination in intertidal sediments from the Gulf of Biscay (Pays de la Loire region, France). Sediments were sampled at three locations (Pays de la Loire region, France) and during two seasons: October 2015 and March 2016. The analytical protocol involved MP extraction from dried sediments using milliQ water and a centrifugation technique. After a filtration step of supernatants, MPs were detected and directly identified on the membrane filters using µFTIR spectroscopy in reflection mode. For the first time, the number of replicates allowing to obtain a satisfying representativeness of the whole sampled sediment was also evaluated at 10 replicates of 25 g each. The average number of MPs in sediments was 67 (±76) MPs/kg dw (N = 60) with no significant difference between sites and seasons. Ten different compositions of MPs were defined by µFT-IR with a high proportion of polypropylene (PP) and polyethylene (PE), 38 and 24%, respectively. Among MPs, mainly fragments (84%) were observed with main size classes corresponding to [>100 µm] and [50-100 µm] but no particles > 1 mm could be found suggesting that mainly small microplastics (<1 mm) were subject to vertical transport.

Factors influencing the microplastic contamination of bivalves from the French Atlantic coast: Location, season and/or mode of life?

Monitoring the presence of microplastics (MP) in marine organisms is currently of high importance. This paper presents the qualitative and quantitative MP contamination of two bivalves from the French Atlantic coasts: the blue mussel (Mytilus edulis) and the Pacific oyster (Crassostrea gigas). Three factors potentially influencing the contamination were investigated by collecting at different sampling sites and different seasons, organisms both wild and cultivated. Inter- and intra-species comparisons were also achieved. MP quantity in organisms was evaluated at 0.61±0.56 and 2.1±1.7MP per individual respectively for mussels and oysters. Eight different polymers were identified. Most of the MPs were fragments; about a half of MPs were grey colored and a half with a size ranging from 50 to 100µm for both studied species. Some inter-specific differences were found but no evidence for sampling site, season or mode of life effect was highlighted.

Quantification and characterization of microplastics in blue mussels (Mytilus edulis): protocol setup and preliminary data on the contamination of the French Atlantic coast.

Microplastics (MPs) constitute a main environmental issue due to their threat to marine organisms and so far to humans. The lack of a fast standard protocol in MP isolation and identification from living organisms bring to challenge for the science. In this paper, an optimized protocol using potassium hydroxide 10% (KOH 10%; m/v) for digestion of mussel soft tissues (Mytilus edulis) and multi-steps of sedimentation has been developed. Efficiency higher than 99.9% of organic and mineral matter elimination was shown by application on mussels sampled on the French Atlantic coast. The identification of MPs was performed by FTIR microscopy straight on the filter and the whole analysis can be compatible with a routine goal. Fourteen MPs of four different chemical natures were found and identified in 5 pools of 3 sampled mussels. Their size ranged from 30 to 200 µm. Further investigations are now needed to evaluate the potential risk of such particles within this marine bivalve species and other filter feeders.