Environmental safety of second and third generation bioplastics in the context of the circular economy.

Bioplastics derived from organic materials other than crude oil are often suggested as sustainable solutions for tackling end-of-life plastic waste, but little is known of their ecotoxicity to aquatic species. Here, we investigated the ecotoxicity of second and third generation bioplastics toward the freshwater zooplankton Daphnia magna. In acute toxicity tests (48 h), survival was impacted at high concentrations (g.L-1 range), within the range of salinity-induced toxicity. Macroalgae-derived bioplastic induced hormetic responses under chronic exposure (21 d). Most biological traits were enhanced from 0.06 to 0.25 g.L-1 (reproduction rate, body length, width, apical spine, protein concentration), while most of these traits returned to controls level at 0.5 g.L-1. Phenol-oxidase activity, indicative of immune function, was enhanced only at the lowest concentration (0.06 g.L-1). We hypothesise these suggested health benefits were due to assimilation of carbon derived from the macroalgae-based bioplastic as food. Polymer identity was confirmed by infra-red spectroscopy. Chemical analysis of each bioplastic revealed low metal abundance whilst non target exploration of organic compounds revealed trace amounts of phthalates and flame retardants. The macroalgae-bioplastic disintegrated completely in compost and biodegraded up to 86 % in aqueous medium. All bioplastics acidified the test medium. In conclusion, the tested bioplastics were classified as environmentally safe. Nonetheless, a reasonable end-of-life management of these safer-by-design materials is advised to ensure the absence of harmful effects at high concentrations, depending on the receiving environment.

Short Depuration of Oysters Intended for Human Consumption Is Effective at Reducing Exposure to Nanoplastics.

Nanoplastics (NPs; <1 µm) have greater availability to marine organisms than microplastics (1-5000 µm). Understanding NP uptake and depuration in marine organisms intended for human consumption is imperative for food safety, but until now it has been limited due to analytical constraints. Oysters (Crassostrea gigas) were exposed to polystyrene NPs doped with palladium (Pd), allowing the measurements of their uptake into tissues by inductively coupled plasma mass spectrometry (ICP-MS) combined with electron microscopy. Oysters were exposed for 6 days (d) to "Smooth" or "Raspberry" NPs, followed by 30 d of depuration with the aim of assessing the NP concentration in C. gigas following exposure, inferring the accumulation and elimination rates, and understanding the clearance of Pd NPs during the depuration period. After 6 d, the most significant accumulation was found in the digestive gland (106.6 and 135.3 µg g-1 dw, for Smooth and Raspberry NPs, respectively) and showed the most evident depuration (elimination rate constant KSmooth = 2 d-1 and KRaspberry = 0.2 d-1). Almost complete depuration of the Raspberry NPs occurred after 30 d. While a post-harvesting depuration period of 24-48 h for oysters could potentially reduce the NP content by 75%, more research to validate these findings, including depuration studies of oysters from the field, is required to inform practices to reduce human exposure through consumption.

Long-Term Exposure to Environmentally Relevant Doses of Large Polystyrene Microplastics Disturbs Lipid Homeostasis via Bowel Function Interference.

The question of whether long-term chronic exposure to microplastics (MPs) could induce dose- and size-dependent adverse effects in mammals remains controversial and poorly understood. Our study explored potential health risks from dietary exposure to environmentally relevant doses of polystyrene (PS) MPs, through a mouse model and integrated analyses of the interruptions of fecal microbial metagenomes and plasma lipidomes. After 21 weeks of exposure to the MPs (40-100 µm), mice mainly exhibited gut microbiota dysbiosis, tissue inflammation, and plasma lipid metabolism disorder, although no notable accumulation of MPs was observed in the gut or liver. The change of the relative abundance of microbiota was strongly associated with the exposure dose and size of MPs while less significant effects were observed in gut damage and abnormal lipid metabolism. Moreover, multiomics data suggested that the host abnormal lipid metabolism was closely related to bowel function disruptions, including gut microbiota dysbiosis, increased gut permeability, and inflammation induced by MPs. We revealed for the first time that even without notable accumulation in mouse tissues, long-term exposure to MPs at environmentally relevant doses could still induce widespread health risks. This raises concern on the health risks from the exposure of humans and other mammals to environmentally relevant dose MPs.

Quantitative Analysis of Selected Plastics in High-Commercial-Value Australian Seafood by Pyrolysis Gas Chromatography Mass Spectrometry.

Microplastic contamination of the marine environment is widespread, but the extent to which the marine food web is contaminated is not yet known. The aims of this study were to go beyond visual identification techniques and develop and apply a simple seafood sample cleanup, extraction, and quantitative analysis method using pyrolysis gas chromatography mass spectrometry to improve the detection of plastic contamination. This method allows the identification and quantification of polystyrene, polyethylene, polyvinyl chloride, polypropylene, and poly(methyl methacrylate) in the edible portion of five different seafood organisms: oysters, prawns, squid, crabs, and sardines. Polyvinyl chloride was detected in all samples and polyethylene at the highest total concentration of between 0.04 and 2.4 mg g-1 of tissue. Sardines contained the highest total plastic mass concentration (0.3 mg g-1 tissue) and squid the lowest (0.04 mg g-1 tissue). Our findings show that the total concentration of plastics is highly variable among species and that microplastic concentration differs between organisms of the same species. The sources of microplastic exposure, such as packaging and handling with consequent transference and adherence to the tissues, are discussed. This method is a major development in the standardization of plastic quantification techniques used in seafood.

Nanoplastics Promote Microcystin Synthesis and Release from Cyanobacterial Microcystis aeruginosa.

Although the fate of nanoplastics (<100 nm) in freshwater systems is increasingly well studied, much less is known about its potential threats to cyanobacterial blooms, the ultimate phenomenon of eutrophication occurrence worldwide. Previous studies have evaluated the consequences of nanoplastics increasing the membrane permeability of microbes, however, there is no direct evidence for interactions between nanoplastics and microcystin; intracellular hepatotoxins are produced by some genera of cyanobacteria. Here, we show that the amino-modified polystyrene nanoplastics (PS-NH2) promote microcystin synthesis and release from Microcystis aeruginosa, a dominant species causing cyanobacterial blooms, even without the change of coloration. We demonstrate that PS-NH2 inhibits photosystem II efficiency, reduces organic substance synthesis, and induces oxidative stress, enhancing the synthesis of microcystin. Furthermore, PS-NH2 promotes the extracellular release of microcystin from M. aeruginosa via transporter protein upregulation and impaired cell membrane integrity. Our findings propose that the presence of nanoplastics in freshwater ecosystems might enhance the threat of eutrophication to aquatic ecology and human health.

Microplastic ingestion ubiquitous in marine turtles.

Despite concerns regarding the environmental impacts of microplastics, knowledge of the incidence and levels of synthetic particles in large marine vertebrates is lacking. Here, we utilize an optimized enzymatic digestion methodology, previously developed for zooplankton, to explore whether synthetic particles could be isolated from marine turtle ingesta. We report the presence of synthetic particles in every turtle subjected to investigation (n = 102) which included individuals from all seven species of marine turtle, sampled from three ocean basins (Atlantic [ATL]: n = 30, four species; Mediterranean (MED): n = 56, two species; Pacific (PAC): n = 16, five species). Most particles (n = 811) were fibres (ATL: 77.1% MED: 85.3% PAC: 64.8%) with blue and black being the dominant colours. In lesser quantities were fragments (ATL: 22.9%: MED: 14.7% PAC: 20.2%) and microbeads (4.8%; PAC only; to our knowledge the first isolation of microbeads from marine megavertebrates). Fourier transform infrared spectroscopy (FT-IR) of a subsample of particles (n = 169) showed a range of synthetic materials such as elastomers (MED: 61.2%; PAC: 3.4%), thermoplastics (ATL: 36.8%: MED: 20.7% PAC: 27.7%) and synthetic regenerated cellulosic fibres (SRCF; ATL: 63.2%: MED: 5.8% PAC: 68.9%). Synthetic particles being isolated from species occupying different trophic levels suggest the possibility of multiple ingestion pathways. These include exposure from polluted seawater and sediments and/or additional trophic transfer from contaminated prey/forage items. We assess the likelihood that microplastic ingestion presents a significant conservation problem at current levels compared to other anthropogenic threats.

Effects of Nylon Microplastic on Feeding, Lipid Accumulation, and Moulting in a Coldwater Copepod.

Microplastic debris is a pervasive environmental contaminant that has the potential to impact the health of biota, although its modes of action remain somewhat unclear. The current study tested the hypothesis that exposure to fibrous and particulate microplastics would alter feeding, impacting on lipid accumulation, and normal development (e.g., growth, moulting) in an ecologically important coldwater copepod Calanus finmarchicus. Preadult copepods were incubated in seawater containing a mixed assemblage of cultured microalgae (control), with the addition of ∼50 microplastics mL-1 of nylon microplastic granules (10-30 µm) or fibers (10 × 30 µm), which are similar in shape and size to the microalgal prey. The additive chemical profiles showed the presence of stabilizers, lubricants, monomer residues, and byproducts. Prey selectivity was significantly altered in copepods exposed to nylon fibers (ANOVA, P < 0.01) resulting in a nonsignificant 40% decrease in algal ingestion rates (ANOVA, P = 0.07), and copepods exposed to nylon granules showed nonsignificant lipid accumulation (ANOVA, P = 0.62). Both microplastics triggered premature moulting in juvenile copepods (Bernoulli GLM, P < 0.01). Our results emphasize that the shape and chemical profile of a microplastic can influence its bioavailability and toxicity, drawing attention to the importance of using environmentally relevant microplastics and chemically profiling plastics used in toxicity testing.

Role of Marine Snows in Microplastic Fate and Bioavailability.

Microplastics contaminate global oceans and are accumulating in sediments at levels thought sufficient to leave a permanent layer in the fossil record. Despite this, the processes that vertically transport buoyant polymers from surface waters to the benthos are poorly understood. Here we demonstrate that laboratory generated marine snows can transport microplastics of different shapes, sizes, and polymers away from the water surface and enhance their bioavailability to benthic organisms. Sinking rates of all tested microplastics increased when incorporated into snows, with large changes observed for the buoyant polymer polyethylene with an increase in sinking rate of 818 m day-1 and for denser polyamide fragments of 916 m day-1. Incorporation into snows increased microplastic bioavailability for mussels, where uptake increased from zero to 340 microplastics individual-1 for free microplastics to up to 1.6 × 105 microplastics individual-1 when incorporated into snows. We therefore propose that marine snow formation and fate has the potential to play a key role in the biogeochemical processing of microplastic pollution.

Dynamics of nuclear receptor gene expression during Pacific oyster development.

BACKGROUND: Nuclear receptors are a highly conserved set of ligand binding transcription factors, with essential roles regulating aspects of vertebrate and invertebrate biology alike. Current understanding of nuclear receptor regulated gene expression in invertebrates remains sparse, limiting our ability to elucidate gene function and the conservation of developmental processes across phyla. Here, we studied nuclear receptor expression in the early life stages of the Pacific oyster, Crassostrea gigas, to identify at which specific key stages nuclear receptors are expressed RESULTS: We used quantitative RT-PCR to determine the expression profiles of 34 nuclear receptors, revealing three developmental key stages, during which nuclear receptor expression is dynamically regulated: embryogenesis, mid development from gastrulation to trochophore larva, and late larval development prior to metamorphosis. Clustering of nuclear receptor expression patterns demonstrated that transcriptional regulation was not directly related to gene phylogeny, suggesting closely related genes may have distinct functions. Expression of gene homologs of vertebrate retinoid receptors suggests participation in organogenesis and shell-formation, as they are highly expressed at the gastrulation and trochophore larval initial shell formation stages. The ecdysone receptor homolog showed high expression just before larval settlement, suggesting a potential role in metamorphosis. CONCLUSION: Throughout early oyster development nuclear receptors exhibited highly dynamic expression profiles, which were not confined by gene phylogeny. These results provide fundamental information on the presence of nuclear receptors during key developmental stages, which aids elucidation of their function in the developmental process. This understanding is essential as ligand sensing nuclear receptors can be disrupted by xenobiotics, a mode of action through which anthropogenic environmental pollutants have been found to mediate effects.

Fluoxetine Exhibits Pharmacological Effects and Trait-Based Sensitivity in a Marine Worm.

Global production of pharmacologically active compounds exceeds 100 000 tons annually, a proportion of which enters aquatic environments through patient use, improper medicine disposal, and production. These compounds are designed to have mode-of-action (MoA) effects on specific biological pathways, with potential to impact nontarget species. Here, we used MoA and trait-based approaches to quantify uptake and biological effects of fluoxetine, a selective serotonin reuptake inhibitor, in filter and deposit feeding marine worms (Hediste diversicolor). Worms exposed to 10 µg L(-1), accumulated fluoxetine with a body burden over 270 times greater than exposure concentrations, resulting in ∼10% increased coelomic fluid serotonin, a pharmacological effect. Observed effects included weight loss (up to 2% at 500 µg L(-1)), decreased feeding rate (68% at 500 µg L(-1)), and altered metabolism (oxygen consumption, ammonia excretion, and O/N from 10 µg L(-1)). Bioconcentration of fluoxetine was dependent on route of uptake, with filter feeding worms experiencing up to 130 times greater body burden ratios and increased magnitudes of effects than deposit feeders, a trait-based sensitivity likely as a consequence of fluoxetine partitioning to sediment. This study highlights how novel approaches such as MoA and trait-based methods can supplement environmental risk assessments of pharmaceuticals.

Effect of Microplastic on the Gills of the Shore Crab Carcinus maenas.

Microscopic plastic debris (microplastics, <5 mm in diameter) is ubiquitous in the marine environment. Previous work has shown that microplastics may be ingested and inhaled by the shore crab Carcinus maenas, although the biological consequences are unknown. Here, we show that acute aqueous exposure to polystyrene microspheres (8 µm) with different surface coatings had significant but transient effects on branchial function. Microspheres inhaled into the gill chamber had a small but significant dose-dependent effect on oxygen consumption after 1 h of exposure, returning to normal levels after 16 h. Ion exchange was also affected, with a small but significant decrease in hemolymph sodium ions and an increase in calcium ions after 24 h post-exposure. To further asses the effects on osmoregulation, we challenged crabs with reduced salinity after microplastic exposure. Neither microspheres nor natural sediments altered the crab's response to osmotic stress regardless of plastic concentration added. Carboxylated (COOH) and aminated (NH2) polystyrene microspheres were distributed differently across the gill surface, although neither had a significant adverse impact on gill function. These results illustrate the extent of the physiological effects of microplastics compared to the physiological resilience of shore crabs in maintaining osmoregulatory and respiratory function after acute exposure to both anthropogenic plastics and natural particles.

Microplastics Alter the Properties and Sinking Rates of Zooplankton Faecal Pellets.

Plastic debris is a widespread contaminant, prevalent in aquatic ecosystems across the globe. Zooplankton readily ingest microscopic plastic (microplastic, < 1 mm), which are later egested within their faecal pellets. These pellets are a source of food for marine organisms, and contribute to the oceanic vertical flux of particulate organic matter as part of the biological pump. The effects of microplastics on faecal pellet properties are currently unknown. Here we test the hypotheses that (1) faecal pellets are a vector for transport of microplastics, (2) polystyrene microplastics can alter the properties and sinking rates of zooplankton egests and, (3) faecal pellets can facilitate the transfer of plastics to coprophagous biota. Following exposure to 20.6 µm polystyrene microplastics (1000 microplastics mL(-1)) and natural prey (∼1650 algae mL(-1)) the copepod Calanus helgolandicus egested faecal pellets with significantly (P < 0.001) reduced densities, a 2.25-fold reduction in sinking rates, and a higher propensity for fragmentation. We further show that microplastics, encapsulated within egests of the copepod Centropages typicus, could be transferred to C. helgolandicus via coprophagy. Our results support the proposal that sinking faecal matter represents a mechanism by which floating plastics can be vertically transported away from surface waters.

Virgin microplastics cause toxicity and modulate the impacts of phenanthrene on biomarker responses in African catfish (Clarias gariepinus).

Despite the ubiquity of microplastics (MPs) in aquatic environments and their proven ability to carry a wide variety of chemicals, very little is known about the impacts of virgin or contaminant-loaded MPs on organisms. The primary aim of this study was to investigate the impacts of virgin or phenanthrene (Phe)-loaded low-density polyethylene (LDPE) fragments on a suite of biomarker responses in juvenile African catfish (Clarias gariepinus). Virgin LDPE (50 or 500µg/L) were preloaded with one of two nominal Phe concentrations (10 or 100µg/L) and were exposed to the fish for 96h. Our findings showed one or both Phe treatments significantly increased the degree of tissue change (DTC) in the liver while decreased the transcription levels of forkhead box L2 (foxl2) and tryptophan hydroxylase2 (tph2) in the brain of C. gariepinus. Exposure to either levels of virgin MPs increased the DTC in the liver and plasma albumin: globulin ratio while decreased the transcription levels of tph2. Moreover, MPs modulated (interacted with) the impact of Phe on the DTC in the gill, plasma concentrations of cholesterol, high-density lipoprotein (HDL), total protein (TP), albumin, and globulin, and the transcription levels of fushi tarazu-factor 1 (ftz-f1), gonadotropin-releasing hormone (GnRH), 11 ß-hydroxysteroid dehydrogenase type 2 (11ß-hsd2), and liver glycogen stores. Results of this study highlight the ability of virgin LDPE fragments to cause toxicity and to modulate the adverse impacts of Phe in C. gariepinus. Due to the wide distribution of MPs and other classes of contaminants in aquatic environments, further studies are urgently needed to elucidate the toxicity of virgin or contaminant-loaded MPs on organisms.

Ingestion of Nanoplastics and Microplastics by Pacific Oyster Larvae.

Plastic debris is a prolific contaminant effecting freshwater and marine ecosystems across the globe. Of growing environmental concern are "microplastics"and "nanoplastics" encompassing tiny particles of plastic derived from manufacturing and macroplastic fragmentation. Pelagic zooplankton are susceptible to consuming microplastics, however the threat posed to larvae of commercially important bivalves is currently unknown. We exposed Pacific oyster (Crassostrea gigas) larvae (3-24 d.p.f.) to polystyrene particles spanning 70 nm-20 µm in size, including plastics with differing surface properties, and tested the impact of microplastics on larval feeding and growth. The frequency and magnitude of plastic ingestion over 24 h varied by larval age and size of polystyrene particle (ANOVA, P < 0.01), and surface properties of the plastic, with aminated particles ingested and retained more frequently (ANOVA, P < 0.01). A strong, significant correlation between propensity for plastic consumption and plastic load per organism was identified (Spearmans, r = 0.95, P < 0.01). Exposure to 1 and 10 µm PS for up to 8 days had no significant effect on C. gigas feeding or growth at <100 microplastics mL(-1). In conclusion, whil micro- and nanoplastics were readily ingested by oyster larvae, exposure to plastic concentrations exceeding those observed in the marine environment resulted in no measurable effects on the development or feeding capacity of the larvae over the duration of the study.

The impact of polystyrene microplastics on feeding, function and fecundity in the marine copepod Calanus helgolandicus.

Microscopic plastic debris, termed "microplastics", are of increasing environmental concern. Recent studies have demonstrated that a range of zooplankton, including copepods, can ingest microplastics. Copepods are a globally abundant class of zooplankton that form a key trophic link between primary producers and higher trophic marine organisms. Here we demonstrate that ingestion of microplastics can significantly alter the feeding capacity of the pelagic copepod Calanus helgolandicus. Exposed to 20 µm polystyrene beads (75 microplastics mL(­1)) and cultured algae ([250 µg C L(­1)) for 24 h, C. helgolandicus ingested 11% fewer algal cells (P = 0.33) and 40% less carbon biomass (P < 0.01). There was a net downward shift in the mean size of algal prey consumed (P < 0.001), with a 3.6 fold increase in ingestion rate for the smallest size class of algal prey (11.6­12.6 µm), suggestive of postcapture or postingestion rejection. Prolonged exposure to polystyrene microplastics significantly decreased reproductive output, but there were no significant differences in egg production rates, respiration or survival. We constructed a conceptual energetic (carbon) budget showing that microplastic-exposed copepods suffer energetic depletion over time. We conclude that microplastics impede feeding in copepods, which over time could lead to sustained reductions in ingested carbon biomass.

Ingestion of Plastic Microfibers by the Crab Carcinus maenas and Its Effect on Food Consumption and Energy Balance.

Microscopic plastic fragments (<5 mm) are a worldwide conservation issue, polluting both coastal and marine environments. Fibers are the most prominent plastic type reported in the guts of marine organisms, but their effects once ingested are unknown. This study investigated the fate of polypropylene rope microfibers (1-5 mm in length) ingested by the crab Carcinus maenas and the consequences for the crab's energy budget. In chronic 4 week feeding studies, crabs that ingested food containing microfibers (0.3-1.0% plastic by weight) showed reduced food consumption (from 0.33 to 0.03 g d(-1)) and a significant reduction in energy available for growth (scope for growth) from 0.59 to -0.31 kJ crab d(-1) in crabs fed with 1% plastic. The polypropylene microfibers were physically altered by their passage through the foregut and were excreted with a smaller overall size and length and amalgamated into distinctive balls. These results support of the emerging paradigm that a key biological impact of microplastic ingestion is a reduction in energy budgets for the affected marine biota. We also provide novel evidence of the biotransformations that can affect the plastics themselves following ingestion and excretion.

Bisphenol A and the risk of cardiometabolic disorders: a systematic review with meta-analysis of the epidemiological evidence.

Bisphenol A (BPA) is suspected to be associated with several chronic metabolic diseases. The aim of the present study was to review the epidemiological literature on the relation between BPA exposure and the risk of cardiometabolic disorders. PubMed and Embase databases were searched up to August 2014 by two independent investigators using standardized subject terms. We included observational studies (cohort, case-control and cross-sectional studies) carried out in children or adults, measuring urinary BPA (uBPA), including at least 100 participants and published in English. The health outcomes of interest were diabetes, hyperglycemia, measures of anthropometry, cardiovascular disease (CVD) and hypertension. Data were extracted and meta-analyzed when feasible, using a random-effects model. Thirty-three studies with sample size ranging from 239 to 4811 met the inclusion criteria, including five with a prospective design. Twelve studies reported on diabetes or hyperglycemia, 16 on anthropometry, 6 on CVD and 3 on hypertension. Evidence for a positive association between uBPA concentrations and diabetes, overweight, obesity, elevated waist circumference (WC), CVD and hypertension was found in 7/8, 2/7, 6/7, 5/5, 4/5 and 2/3 of the cross-sectional studies, respectively. We were able to conduct outcome-specific meta-analyses including 12 studies. When comparing the highest vs. the lowest uBPA concentrations, the pooled ORs were 1.47 (95% CI: 1.21-1.80) for diabetes, 1.21 (95% CI: 0.98-1.50) for overweight, 1.67 (95% CI: 1.41-1.98) for obesity, 1.48 (95% CI: 1.25-1.76) for elevated WC, and 1.41 (95% CI: 1.12-1.79) for hypertension. Moreover, among the five prospective studies, 3 reported significant findings, relating BPA exposure to incident diabetes, incident coronary artery disease, and weight gain. To conclude, there is evidence from the large body of cross-sectional studies that individuals with higher uBPA concentrations are more likely to suffer from diabetes, general/abdominal obesity and hypertension than those with lower uBPA concentrations. Given the potential importance for public health, prospective cohort studies with proper adjustment for dietary characteristics and identification of critical windows of exposure are urgently needed to further improve knowledge about potential causal links between BPA exposure and the development of chronic disease.

The nuclear receptor gene family in the Pacific oyster, Crassostrea gigas, contains a novel subfamily group.

BACKGROUND: Nuclear receptors are a superfamily of transcription factors important in key biological, developmental and reproductive processes. Several of these receptors are ligand- activated and through their ability to bind endogenous and exogenous ligands, are potentially vulnerable to xenobiotics. Molluscs are key ecological species in defining aquatic and terrestrial habitats and are sensitive to xenobiotic compounds in the environment. However, the understanding of nuclear receptor presence, function and xenobiotic disruption in the phylum Mollusca is limited. RESULTS: Here, forty-three nuclear receptor sequences were mined from the genome of the Pacific oyster, Crassostrea gigas. They include members of NR0-NR5 subfamilies, notably lacking any NR6 members. Phylogenetic analyses of the oyster nuclear receptors have been conducted showing the presence of a large novel subfamily group not previously reported, which is named NR1P. Homologues to all previous identified nuclear receptors in other mollusc species have also been determined including the putative heterodimer partner retinoid X receptor, estrogen receptor and estrogen related receptor. CONCLUSION: C. gigas contains a highly diverse set of nuclear receptors including a novel NR1 group, which provides important information on presence and evolution of this transcription factor superfamily in invertebrates. The Pacific oyster possesses two members of NR3, the sex steroid hormone receptor analogues, of which there are 9 in humans. This provides increasing evidence that steroid ligand specific expansion of this family is deuterostome specific. This new knowledge on divergence and emergence of nuclear receptors in C. gigas provides essential information for studying regulation of molluscan gene expression and the potential effects of xenobiotics.