Polychaete Hediste Diversicolor Research Articles

Biosynthesis of long-chain polyunsaturated fatty acids in the nereid polychaete Hediste diversicolor: Molecular cloning and functional characterisation of three fatty acyl elongases and two front-end desaturases

Aquaculture of carnivorous fish species heavily relies on marine ingredients to guarantee the provision of essential nutrients, including long-chain (C20–24) polyunsaturated fatty acids (LC-PUFA) such as arachidonic acid (ARA, 20:4n-6), eicosapentaenoic acid (EPA, 20:5n-3), and docosahexaenoic acid (DHA, 22:6n-3). These nutrients are crucial for fish growth and neural development, offering significant health benefits to consumers. The rapid growth of global aquaculture production in recent decades, has driven the exploration of alternative and sustainable feeding sources that ensure n-3 LC-PUFA content. Within the context of circular economy, polychaete worms arise as promising candidates for aquafeed formulations, due to the enzymatic capacity for de novo LC-PUFA biosynthesis. The present study focused on the molecular and functional characterisation of elongases (Elovl) and front-end desaturases (Fed) involved in the LC-PUFA biosynthesis of the nereid polychaete Hediste diversicolor. The sequence and phylogenetic analysis established the orthology of three Elovl, identified as Elovl2/5, Elovl4 and Elovl1/7, that have putative functions in the LC-PUFA biosynthesis. Functional characterisation in yeast demonstrated their roles as PUFA elongases, exhibiting affinity towards C18 and C20 PUFA substrates (Elovl2/5) and towards C18, C20 and C22 (Elovl4 and Elovl1/7). Additionally, H. diversicolor possesses two front-end desaturases (Fed1 and Fed2) with Δ5 and Δ6/Δ8 desaturation activities, respectively, thus encompassing all the necessary desaturation reactions involved in the LC-PUFA biosynthesis pathways. These findings, along with the previously characterised methyl-end desaturases, show that H. diversicolor possesses the full enzymatic repertoire necessary to produce LC-PUFA like ARA and EPA. This highlights the potential of polychaetes and other annelids for producing nutrient-rich biomass suitable for aquaculture.

Transfer and bioaccumulation of chemical and biological contaminants in the marine polychaete Hediste diversicolor (OF müller 1776) when reared on salmon aquaculture sludge

Side streams from aquaculture production such as fish sludge poses ample opportunities for biological upcycling, as the sludge contains high amounts of nutrients, energy and valuable biochemicals, making it an ideal food for extractive species. Sludge has been proposed as a feed stock for polychaete production, which in turn can be utilized live in shrimp aquaculture or as an aquafeed ingredient. However, the biosafety of such value chains has not yet been addressed. We conducted an experiment exposing the polychaete Hediste diversicolor to aquaculture sludge spiked with four different fish pathogens (Mycobacterium salmoniphilum, Yersinia ruckeri, Infectious Pancreatic Necrosis (IPN) and Infectious Salmon Anaemia (ISA)) known to cause diseases in Atlantic salmon (Salmo salar L.). Moreover, we assessed whether heavy metals and other potentially hazardous elements present in fish sludge bioaccumulates in the polychaetes. Neither of the bacteria nor viruses could be detected in the polychaetes after 14 days of continuous exposure. Seven of the 15 elements we analysed showed bioaccumulation factors significantly below one, meaning biodilution, while the other eight did not differ from one, meaning no bioaccumulation. None of the elements showed a significant bioaccumulation. Further on, none of the heavy metals found in the polychaetes at the end of our experiment exceeded the EU regulatory maximum levels for fish feed ingredients. The current results suggest that a H. diversicolor can reared on aquaculture sludge, and aquaculture sludge may serve as feed stock for polychaete production without the product exceeding EU regulations for contaminants in animal feed.

Metabolomic and biochemical disorders reveal the toxicity of environmental microplastics and benzo[a]pyrene in the marine polychaete Hediste diversicolor

Recently, the abundance of environmental microplastics (MPs) has become a global paramount concern. Besides the danger of MPs for biota due to their tiny size, these minute particles may act as vectors of other pollutants. This study focused on evaluating the toxicity of environmentally relevant concentrations of MPs (10 and 50 mg/kg sediment) and benzo[a]pyrene (B[a]P, 1 µg/kg sediment), alone and in mixture, for 3 and 7 days in marine polychaete Hediste diversicolor, selected as a benthic bioindicator model. The exposure period was sufficient to confirm the bioaccumulation of both contaminants in seaworms, as well as the potential capacity of plastic particles to adsorb and vehiculate the B[a]P. Interestingly, increase of acidic mucus production was observed in seaworm tissues, indicative of a defense response. The activation of oxidative system pathways was demonstrated as a strategy to prevent lipid peroxidation. Furthermore, the comprehensive Nuclear Magnetic Resonance (NMR)-based metabolomics revealed significant disorders in amino acids metabolism, osmoregulatory process, energetic components, and oxidative stress related elements. Overall, these findings proved the possible synergic harmful effect of MPs and B[a]P even in small concentrations, which increases the concern about their long-term presence in marine ecosystems, and consequently their transfer and repercussions on marine fauna.

Sea urchin waste as a promising nutritional source for the polychaete Hediste diversicolor (OF Müller, 1776)

In aquaculture, species diversification emerges as an imperative sustainable strategy, essential for mitigating both biological and financial risks. In this context, the cultivation of sea urchins has gained prominence as an emerging trend. However, its commercial viability still faces several challenges, namely limited growth performance. Integrated multi-trophic aquaculture (IMTA) represents a potentially more profitable model by integrating extractive species to mitigate ecological impacts and promote circular economy principles. The use of polychaetes in IMTA has proven effective in recycling waste from fish aquaculture production, however, their potential should be further explored in low-trophic aquaculture solutions. This study evaluated the potential of reared sea urchin biodeposits as a nutritional feed source for the polychaete Hediste diversicolor (OF Müller, 1776). H. diversicolor juveniles were fed three diets, over 50 days: sea urchin waste (SUW), fish waste (FW) and fish feed (FF). Survival, growth, and protein and fatty acid (FA) contents were assessed. Total survival was observed in all treatments except for FW (98%). Polychaetes fed SUW and FF presented similar specific growth rates. The protein level in the polychaetes fed SUW was significantly lower, reflecting the diet's composition. The FA profile of H. diversicolor was also influenced by the respective diets. The absolute content (μg mg−1 DW) and FA profile of polychaetes were significantly different among treatments, reflecting the composition of the diets. Saturated FA (SFA) represented the dominant class in the polychaetes fed with waste diets, while in FF polychaetes polyunsaturated fatty acids (PUFA) were the major group. Nevertheless, polychaetes fed the waste diets demonstrated the PUFA biosynthetic capacity of this species, showing a considerably higher proportion of essential long-chain FA, compared to diets. Overall, these preliminary results suggest, for the first time, the potential for recycling sea urchin biodeposits into a polychaete biomass with high commercial value, presenting a promising opportunity for low-trophic integrated aquaculture models.

Retention of prions in the polychaete Hediste diversicolor and black soldier fly, Hermetia illucens, larvae after short-term experimental immersion and feeding with brain homogenate from scrapie infected sheep

Finding alternative protein and lipid sources for aquafeeds is crucial for the sustainable growth of fed aquaculture. Upcycling industrial side streams and byproducts using extractive species can reduce waste and help reduce the sector's dependence on fish meal and fish oils. Polychaete worms (Hediste diversicolor) and black soldier fly (Hermetia illucens) larvae (BSFL) are promising candidates for converting waste materials into valuable protein and lipid sources. However, further research and evaluations are needed to ensure the safety and regulatory compliance of these alternative feed sources, especially regarding prions spreading potential in the unlikely case that prions would be introduced in the value chain via feedstocks. In the present investigation, BSFL and juvenile polychaetes that had received a massive dose of scrapie prions through immersion and oral inoculation were found to harbour detectable prions using an ultrasensitive amplification method known as PMCA. This observation suggests that both H. diversicolor and BSFL have the potential to serve as mechanical vectors for prions diseases. However, it is important to note that insects, lacking the prion protein gene, are incapable of propagating prions. Therefore, the quantity of prions present in the larvae will inevitably be lower than the amount of prions they encountered. This is the first study to report on the fate of prions through ingestion by these marine and terrestrial invertebrate species.

The Role of Life Stages in the Sensitivity of Hediste diversicolor to Nanoplastics: A Case Study with Poly(Methyl)Methacrylate (PMMA).

The presence of plastic particles in oceans has been recognized as a major environmental concern. The decrease in particle size increases their ability to directly interact with biota, with particles in the nanometer size range (nanoplastics-NPs) displaying a higher ability to penetrate biological membranes, which increases with the decrease in particle size. This study aimed to evaluate the role of life stages in the effects of poly(methyl)methacrylate (PMMA) NPs on the polychaete Hediste diversicolor, a key species in the marine food web and nutrient cycle. Thus, behavioral (burrowing activity in clean and spiked sediment) and biochemical endpoints (neurotransmission, energy reserves, antioxidant defenses, and oxidative damage) were assessed in juvenile and adult organisms after 10 days of exposure to spiked sediment (between 0.5 and 128 mg PMMA NPs/Kg sediment). Overall, the results show that H. diversicolor is sensitive to the presence of PMMA NPs. In juveniles, exposed organisms took longer to burrow in sediment, with significant differences from the controls being observed at all tested concentrations when the test was performed with clean sediment, whereas in PMMA NP-spiked sediment, effects were only found at the concentrations 8, 32, and 128 mg PMMA NPs/Kg sediment. Adults displayed lower sensitivity, with differences to controls being found, for both sediment types, at 8, 32, and 128 mg PMMA NPs/Kg sediment. In terms of Acetylcholinesterase, used as a marker of effects on neurotransmission, juveniles and adults displayed opposite trends, with exposed juveniles displaying increased activity (suggesting apoptosis), whereas in adults, overall decreased activity was found. Energy-related parameters revealed a generally similar pattern (increase in exposed organisms) and higher sensitivity in juveniles (significant effects even at the lower concentrations). NPs also demonstrated the ability to increase antioxidant defenses (higher in juveniles), with oxidative damage only being found in terms of protein carbonylation (all tested NPs conditions) in juveniles. Overall, the data reveal the potential of PMMA NPs to affect behavior and induce toxic effects in H. diversicolor, with greater effects in juveniles.

Sludge from sea-based Atlantic salmon (Salmo salar L.) production: quantification, composition, and potential application in integrated multi-trophic aquaculture

Future growth of Atlantic salmon (Salmo salar L.) in Norway is tied to finding solutions for major ecological challenges connected to salmon lice, escapees, and nutrient emissions from sea cages. At the same time, nutrient-rich sludge from salmon production comprises a valuable resource for the cultivation of lower trophic species using an integrated multi-trophic aquaculture (IMTA) approach. This study aimed to quantify the sedimentation of aquaculture sludge under sea cages of an Atlantic salmon aquaculture site and to qualify the composition of this sludge. Additionally, the study evaluated the hypothetical use of sludge from sea-based aquaculture as a feed source for polychaetes Hediste diversicolor. Using sediment traps, sludge samples were collected under two different Atlantic salmon sea cages, at two different depths, and three different sampling dates. Subsequently, they were quantified, and their composition was assessed with regards to carbon (C), nitrogen (N), phosphorus (P), lipid, fatty acid (FA), protein, amino acid (AA), and ash content as well as elemental ratios and composition of FAs and AAs. The quantity of collected sludge was significantly different between sea cages, with a strong positive correlation between feed input and collected sludge (R2 = 0.98, p < 0.05). Sampling depth did not affect the quantity of collected sludge in the sediment traps (2215 ± 480 mg DW day−1), and no significant difference in sedimented sludge as a proportion of theoretically produced sludge (12.94 ± 2.16%) was found when comparing the different cages and sampling depths. Furthermore, the composition of collected sludge was similar at all sampling points. The overall nutritional value was lower compared to sludge from land-based aquaculture; regardless, sludge from sea-based salmon production can in theory be considered a potential feed resource to be used for the production of polychaetes H. diversicolor.

Effects of Reduced Seawater pH and Oil Contamination on Bacterial Communities and Biochemical Markers of Estuarine Animal Hosts

Ecosystem functioning depends on complex interactions between microorganisms, hosts, and the environment. Changes in environmental conditions (e.g., ocean acidification) in combination with anthropogenic pollution have been shown to affect the composition and function of free-living microbial communities, but little is known about the effects these stressors on host-associated communities. This study aims to characterize the response of host-associated bacterial communities of the bottom-dwelling polychaete Hediste diversicolor and the epibenthic gastropod Peringia ulvae to oil contamination and reduced seawater pH. The independent and interactive effects of both stressors were simulated under controlled conditions. The response of host-associated bacterial communities was assessed using the high-throughput sequencing of the 16S rRNA gene and several biochemical markers related to host metabolic pathways, e.g., neurotransmission, anaerobic metabolism, biotransformation, oxidative stress, and energy consumption. In H. diversicolor, reduced seawater pH was associated with a high relative abundance of Cyanobacteria, while in P. ulvae oil contamination was associated with a reduction in the relative abundance of Chitinophagales. In P. ulvae, enrichment with oil hydrocarbon-degrading bacteria suggests a possible role of these organisms in the dispersion of oil hydrocarbon degraders. Furthermore, oil supplementation shifted some specific biochemical markers of gastropods related to oxidative stress and energy consumption, which suggests host stress. In general, the bacterial communities and biochemical markers of the gastropod were more affected by stressors than those of the polychaete. Overall, this study contributes to a better understanding of the response of host-associated bacterial communities of benthic macrofauna to anthropogenic contamination and environmental change.

Upcycling of carbon, nitrogen, and phosphorus from aquaculture sludge using the polychaete Hediste diversicolor (OF Müller, 1776) (Annelida: Nereididae)

IntroductionFocusing on circular bioeconomy and finding solutions for managing finite and scarce resources such as phosphorus are issues that need to be addressed across sectors to attain food security. Land-based aquaculture of Atlantic salmon (Salmo salar L.) smolt and post-smolt results in a large output of aquaculture sludge. These nutrient-rich side streams from aquaculture production constitute a valuable resource for cultivation of polychaetes Hediste diversicolor. The aim of this study was to evaluate the capacity of H. diversicolor to upcycle carbon (C), nitrogen (N), and phosphorus (P) contained in aquaculture sludge from Norwegian salmon production.Material and methodsWe investigated the effect of two diets, smolt sludge and post-smolt sludge, which were supplied at four different feed levels, on the gain of C, N, and P in H. diversicolor and established uptake rates for the upcycling of these nutrients. Additionally, we examined how different feed composition (type of diet) and supply (feel level) influenced C, N, and P content as well as the elemental ratios of H. diversicolor.Results and discussionPolychaetes fed with both types of sludge were shown to incorporate C, N, and P from their diets into biomass. Incorporation of C, N, and P in polychaete biomass was highest at the highest feed levels, following specific growth rates. Uptake of diet C, N, and P into polychaete biomass was similar for both diets and across feed levels. On average, C and N uptake rates amounted to 2–10% and 2–15%, respectively, while values for P uptake were lower at 0.5–1.5%. C, N, and P content of polychaetes was not affected by different diets or feed levels while the C:N ratio increased with increasing feed supply. Overall, we found that H. diversicolor can successfully take up nutrients contained in aquaculture sludge and thus can be beneficial for upcycling of side streams from Atlantic salmon production.

Effects of polymethylmethacrylate nanoplastics on the polychaete Hediste diversicolor: Behavioural, regenerative, and biochemical responses

Plastics, particularly microplastics (MPs) and nanoplastics (NPs), have been regarded as pollutants of emerging concern due to their effects on organisms and ecosystems, especially considering marine environments. However, in terms of NPs, there is still a knowledge gap regarding the effects of size and polymer on marine invertebrates, such as benthic organisms. Therefore, this study aimed to understand, regarding behavioural, physiological, and biochemical endpoints (neurotransmission, energy metabolism, antioxidant status, and oxidative damage), the effects of 50 nm waterborne polymethylmethacrylate (PMMA) NPs (0.5 to 500 µg/L) on the marine benthic polychaete Hediste diversicolor, a key species in estuarine and coastal ecosystems. Results demonstrated that worms exposed to PMMA NPs had a shorter burrowing time than control organisms. Nevertheless, worms exposed to PMMA NPs (0.5 and 500 µg/L) decreased cholinesterase activity. Energy metabolism was decreased at 50 and 500 µg/L, and glycogen content decreased at all concentrations of PMMA NPs. Enzymes related to the antioxidant defence system (superoxide dismutase and glutathione peroxidase) displayed increased activities in H. diversicolor specimens exposed to concentrations between 0.5 and 500 µg/L, which led to no damage at the cell membrane and protein levels. In this study, polychaetes also displayed a lower regenerative capacity when exposed to PMMA NPs. Overall, the data obtained in this study emphasize the potential consequences of PMMA NPs to benthic worms, particularly between 0.5 and 50 µg/L, with polychaetes exposed to 50 µg/L being the most impacted by the analysed NPs. However, since sediments are considered to be sinks and sources of plastics, further studies are needed to better understand the impacts of different sizes and polymers on marine organisms, particularly benthic species.

Hot and heavy: Responses of ragworms (Hediste diversicolor) to copper-spiked sediments and elevated temperature

Sediment contamination and seawater warming are two major stressors to macrobenthos in estuaries. However, little is known about their combined effects on infaunal organisms. Here we investigated the responses of an estuarine polychaete Hediste diversicolor to metal-contaminated sediment and increased temperature. Ragworms were exposed to sediments spiked with 10 and 20 mg kg−1 of copper at 12 and 20 °C for three weeks. No considerable changes were observed in the expression of genes related to copper homeostasis and in the accumulation of oxidative stress damage. Dicarbonyl stress was attenuated by warming exposure. Whole-body energy reserves in the form of carbohydrates, lipids and proteins were little affected, but the energy consumption rate increased with copper exposure and elevated temperature, indicating higher basal maintenance costs of ragworms. The combined effects of copper and warming exposures were mostly additive, with copper being a weak stressor and warming a more potent stressor. These results were replicable, as confirmed by two independent experiments of similar settings conducted at two different months of the year. This study suggests the higher sensitivity of energy-related biomarkers and the need to search for more conserved molecular markers of metal exposure in H. diversicolor.

Effects of Rearing Density, Substrate Height, and Feeding Frequency on Growth and Biomass Production of Hediste diversicolor

The polychaete Hediste diversicolor is a suitable species for industrial aquaculture; however, cost-effective culture techniques need to be developed for its intensive production. The aim of this study is to evaluate the effects of worm density and substrate height and their interaction, as well as feeding frequency, on the rearing performance of H. diversicolor. Two trials were conducted. In trial 1, the effects of two substrate heights—6 and 12 cm—and two rearing densities—1000 and 4000 individuals m−2—were assessed in terms of worm growth and biomass production. Worm initial wet weight was 48 mg, and specimens were fed with commercial fish feed during a 70-day assay. The results show no interaction between rearing density and substrate height, and confirm density as a key factor in growth; however, a density of 4000 individuals m−2 results in a significant increase in production (final biomass three times higher for the highest rearing density) without affecting survival. In trial 2, the effect of three levels of feeding frequency—seven days a week; three times a week; and once a week—on growth in individuals of three weight classes—small (25–50 mg); medium (100–150 mg); large (250–350 mg)—was evaluated in a 15-day growing assay. Feeding frequency showed a major influence on the smallest size class, with the best growth indicators obtained at the highest feeding frequency. This study shows that Hediste diversicolor can be reared at a high stocking density to obtain a higher biomass production, and that feeding frequency must be considered as an important factor and adapted to the culture phase.

Do microplastics influence the long-term effects of ciprofloxacin on the polychaete Hediste diversicolor? An integrated behavioral and biochemical approach

Ciprofloxacin (CPX), the most commonly used fluoroquinolone antibiotic, and microplastics (MPs) are two classes of emerging contaminants with severe adverse impacts on aquatic organisms. Previous studies suggest that both CPX and MPs induce deleterious changes in exposed aquatic biota, but the characterization of a chronic and combined ecotoxicological response is not well known, especially in organisms from estuarine ecosystems. Thus, in this study, we investigated the behavioral and biochemical effects of environmentally relevant levels of CPX alone and in combination with polyethylene terephthalate (PET) microplastics over 28 days of exposure, using the polychaete Hediste diversicolor as a model. In addition to behavioral parameters, different biochemical endpoints were also evaluated, namely the levels of metabolic enzymes of phase I (7-ethoxy-resorufin-O-deethylase, EROD), and phase II (glutathione-S-transferase, GSTs), antioxidant defense (catalase, CAT; glutathione peroxidase, GPx; superoxide dismutase, SOD), oxidative damage (lipid peroxidation, by means of levels of thiobarbituric acid reactive substances [TBARS]) and acetylcholinesterase (AChE). Chronic exposure to ciprofloxacin caused a decrease in burrowing time and a significant increase in SOD activity. In animals exposed to the combination of CPX and PET MPs, effects on behavioral traits were also observed, with higher concentrations of MPs leading to a marked delay in the animals' burrowing time. In addition, these animals showed changes in their antioxidant defenses, namely, a significant increase in SOD activity, while GPx activity was severely compromised. For none of the experimental groups, significant alterations were observed in the metabolic enzymes, TBARS or AChE. These findings provide the first insights into the responses of H. diversicolor when exposed to the combination of CPX and PET MPs, highlighting that, although the here studied conditions, there was no evidence of oxidative damage or neurotoxicity, these organisms are not risk-free in co-exposure scenarios, even at low environmental relevant concentrations.

Diet rather than temperature determines the biochemical composition of the ragworm Hediste diversicolor (OF Müller, 1776) (Annelida: Nereidae)

The polychaete Hediste diversicolor is known to recycle side streams from aquaculture and biogas production. We conducted a feeding experiment to evaluate whether rearing temperature or mixtures of these two side streams enhances biomass production and fatty acid composition. We reared H. diversicolor along a 5-temperature gradient ranging from 5.8 °C to 17.1 °C and a 4-step gradient form 100% aquaculture sludge to 100% solid biogas digestate. Formulated fish feed served as a control diet. Polychaetes increased growth rate with increasing temperature, ranging from 0.01 at 5.8 °C to 0.14 at 17.1 °C, while survival was inversely affected by temperature with 100% survival at 5.8 °C and 70% survival at 17.1 °C. Diet had a less pronounced effect on polychaete survival, and no significant effect on growth rates. Contrasting to growth, the fatty acid composition of the polychaetes was not affected by temperature but was highly influenced by diet, as polychaetes did not cluster by rearing temperature but by the diets they received. In conclusion, H. diversicolor can be utilized as a recycler of aquaculture and biogas side streams, and production temperature can be optimized for growth without compromising fatty acid composition and quality of the polychaetes.

Biochemical and Behavioural Alterations Induced by Arsenic and Temperature in Hediste diversicolor of Different Growth Stages

Contamination with Arsenic, a toxic metalloid, is increasing in the marine environment. Additionally, global warming can alter metalloids toxicity. Polychaetes are key species in marine environments. By mobilizing sediments, they play vital roles in nutrient and element (including contaminants) cycles. Most studies with marine invertebrates focus on the effects of metalloids on either adults or larvae. Here, we bring information on the effects of temperature increase and arsenic contamination on the polychaete Hediste diversicolor in different growth stages and water temperatures. Feeding activity and biochemical responses-cholinesterase activity, indicators of cell damage, antioxidant and biotransformation enzymes and metabolic capacity-were evaluated. Temperature rise combined with As imposed alterations on feeding activity and biochemical endpoints at different growth stages. Small organisms have their antioxidant enzymes increased, avoiding lipid damage. However, larger organisms are the most affected class due to the inhibition of superoxide dismutase, which results in protein damage. Oxidative damage was observed on smaller and larger organisms exposed to As and temperature of 21 °C, demonstrating higher sensibility to the combination of temperature rise and As. The observed alterations may have ecological consequences, affecting the cycle of nutrients, sediment oxygenation and the food chain that depends on the bioturbation of this polychaete.

Ecotoxicological tools in support of the aims of the European Water Framework Directive: A step towards a more holistic ecosystem-based approach

The Water Framework Directive (WFD) aims to attain ‘good quality’ status for all European water bodies through the achievement of good ecological status. To this purpose, the WFD advocates the creation of cost-effective monitoring tools to deliver appropriate data that help to create links between chemical and ecological indicators, as those from ecotoxicological research. Here, it was evaluated whether the integration of ecotoxicological tools, as bioassays and biomarkers, did (or did not) strengthen the robustness of the assessment of the ecological status obtained through well stablished biotic indices in two Atlantic estuaries. For that, a battery of in-situ bioassays, including five macroinvertebrate species (the crab Carcinus maenas, the amphipod Echinogammarus marinus, the isopod Cyathura carinata, the snail Peringia ulvae and the polychaete Hediste diversicolor, each one providing complementary information regarding key ecological functions) and a set of biomarkers, was used. In addition, the concentrations of heavy metals in sediments, and in macroalgae (Fucus), were determined, along with sediment granulometric, water physicochemical and nutrients characterization. We show that by interpreting the values of all indicators together, along with environmental components, it is possible to perform a more holistic description of the quality status of a waterbody – and, to begin to allude to factors limiting that quality. Ecotoxicological tools (in situ bioassays and biomarkers) appear to provide an added value of useful information for monitoring programmes regarding the true state - within which, both known and unknown contaminants are potentially present at concentrations sufficient to cause biological effects. Our results support the use of in situ bioassays and biomarkers within protocols aimed at fulfilling the goals of the WFD. In doing so, more complete and informative assessments of the ecological quality status of water bodies can take place.

Time-course distribution of fluorescent microplastics in target tissues of mussels and polychaetes

The majority of the plastic produced in the last century is accumulated in the environment, leading to an exacerbated contamination of marine environments due to transport from land to the ocean. In the ocean, mechanical abrasion, oxidation, and photodegradation degrade large plastics into microplastics (MPs) - 0.1 μm to 5 mm (EFSA, 2016) which are transported through water currents reaching the water surface, water column, and sediments. Further, they can be accumulated by aquatic and benthic species, entering the trophic chain and becoming a potential threat to humans. In the present research, we aimed to decipher the accumulation and distribution time-courses between different organs or target tissues of organisms inhabiting coastal areas such as mussels Mytilus galloprovincialis and polychaetes Hediste diversicolor. Both were exposed in microcosm experiments to fluorescent polystyrene MPs (1 μm) which were spiked at two doses (103 and 105 particles/mL) for 1, 4, 24, and 72 h. Mussels and polychaetes were digested with 10% KOH and filtered to quantify the number of MPs incorporated. Different anatomical parts of the body were selected and processed for cryosectioning and posterior microscopic localisation of MPs. Both species accumulate MPs spiked in water column, mainly after exposure to the highest dose. In mussels, particles were found in distinct parts of the digestive tract (stomach, digestive diverticula, ducts) and gills. Even if the majority of MPs were localised in the lumen of the digestive tract, in some cases, were inside the digestive epithelium. The identification of MPs and their internalization in the digestive system was studied using Raman spectroscopy. A decreasing trend with time regarding MPs number in the digestive tract (stomach) of mussels was observed while the opposite was recorded for polychaetes and sediments. The combination of microscopical observations of frozen sections and Raman, appeared to be accurate methodologies to address MPs abundances and to reveal their localisation in different organs. This work has enabled to understand the distribution and fate of MPs in different environmental compartments and it could contribute to gain knowledge about their impact after ingestion by coastal organisms.

Innovative in vivo and in vitro bioassays for the establishment of toxicity thresholds of pollutants in sediment quality assessment using polychaetes and their immune cells

Sediment toxicity testing has become a crucial component for assessing the risks posed by contaminated sediments and for the development of sediment quality assessment strategies. Commonly used organisms for bioassays with estuarine sediments include amphipods, Arenicola marina polychaetes and echinoids. Among the latter, the Sea Urchin Embryo test (SET) is the most widely used. However, one relevant limitation of this bioassay is the unavailability of gametes all year-round, particularly outside the natural spawning seasons. Consequently, the establishment of an appropriate and complementary model organism for a continuous assessment of sediment quality is recommended. A reliable assessment of the hazards resulting from pollutants in sediments or pore water, can be achieved with ecologically relevant species of sediment such as the polychaete Hediste diversicolor, which is widespread in estuaries and has the capacity to accumulate pollutants. The aim of this work was to develop reliable in vivo and in vitro bioassays with H. diversicolor and its coelomocytes (immune cells) to determine the toxicity thresholds of different contaminants bounded to sediments or resuspended into water. Polychaetes were exposed to sublethal concentrations of CuCl2 (in vivo) and a non-invasive method for collection of polychaetes coelomocytes was applied for the in vitro bioassay, exposing cells to a series of CuCl2 and AgNPs concentrations. Same reference toxicants were used to expose Paracentrotus lividus following the SET (ICES Nº 51; Beiras et al., 2012) and obtained toxicity thresholds were compared between the two species. In vivo exposure of polychaetes to high concentrations of Cu produced weight loss and histopathological alterations. After in vitro approaches, a significant decrease in coelomocytes viability was recorded for both toxicants, in a monotonic dose-response curve, at very short-exposure times (2 h). The toxicity thresholds obtained with polychaetes were in line with the ones obtained with the SET, concluding that their sensitivity is similar. In conclusion, in vivo and in vitro bioassays developed with H. diversicolor are accurate toxicity screenings of pollutants that could be bounded to sediments or dissolved in the pore water, and may complement the SET outside the spawning period of the echinoderms. The bioassays herein developed could be applied not only to establish the toxicity thresholds of individual compounds or mixtures, but also to assess the toxicity of field collected sediments.

Micro(nano)plastics and plastic additives effects in marine annelids: A literature review

Plastic debris are dispersed in the marine environment and are consequently available to many organisms of different trophic levels, including sediment-dwelling organisms such as polychaetae. Plastic degradation generates micro (MPs) and nanoplastics (NPs) and as well as releases bounded plastic additives, increasing the ecotoxicological risk for marine organisms. Therefore, this review summarizes current knowledge on the accumulation and effects of MPs and NPs and plastic additives in polychaetes, derived from laboratory and field evidences. Thirty-six papers (from January 2011 to September 2021) were selected and analysed: about 80% of the selected works were published since 2016, confirming the emerging role of this topic in environmental sciences. The majority of the analysed manuscripts (68%) were carried out in the laboratory under controlled conditions. These studies showed that polychaetes accumulate and are responsive to this contaminant class, displaying behavioural, physiological, biochemical and immunological alterations. The polychaetes Hediste diversicolor and Arenicola marina were the most frequent used species to study MPs, NPs and plastic additive effects. The consideration of field studies revealed that MP accumulation was dependent on the plastic type present in the sediments and on the feeding strategy of the species. Polychaetes are known to play an important role in coastal and estuarine food webs and exposure to MPs, NPs and plastic additives may impair their behavioural, physiological, biochemical and immunological responses. Thus, the estimated global increase of these contaminants in the marine environment could affect the health of these benthic organisms, with consequences at population and ecosystem levels.

MODELING THE DYNAMICS OF THE ECOLOGICAL NICHE OF STARRED STURGEON (&lt;i&gt;ACIPENSER STELLATUS&lt;/i&gt; PALLAS, 1771) IN THE NORTHERN PART OF THE CASPIAN SEA BASED ON SUMMER LONG-TERM MONITORING DATA (1992-2006)

Models of ecological niches of stellate sturgeon ( Acipenser stellatus Pallas, 1771) in the northern part of the Caspian Sea based on long-term monitoring data (1992-2006) are presented. The analysis of the results of the study of the long-term patterns of the spatial distribution of alien species of polychaete Hediste diversicolor , mollusc Abra ovata and crustaceans are reported. Estimates of the effect of abiotic and biotic factors on the formation of ecological niches of the stellate sturgeon in the eastern and western regions of the northern part of the Caspian Sea are presented. Graphical schemes of the spatial distribution of stellate sturgeon in different years of monitoring in the plane of two main components (PCA) are given. It is shown that the salinity of water and the density of crustaceans are the leading factors for constructing models of ecological niches.