Ecotoxicological risk of microplastics for marine organisms

La presenza di Microplastiche (MP) negli oceani rappresenta una problematica emergente per l’ecosistema marino; è ormai stato dimostrato che le MP possono essere ingerire da diverse specie di organismi, ma non è stata ancora dimostrata la capacità di trasferimento trofico e di accumulo di queste microparticelle, e sono ancora molto carenti le informazioni sui effetti avversi indotti. In questo lavoro di ricerca, il ruolo di alcuni polimeri plastici, come vettori di inquinanti chimici, è stato inizialmente valutato su campioni di macro-plastiche vergini, macro- e micro-plastiche spiaggiate. Tramite esperimenti di laboratorio, sono state invece studiate sia le cinetiche di adsorbimento del pirene e del cadmio in MP di polietilene (PE) e polistirene (PS), che la loro capacità di trasferire i composti adsorbiti ai mitili, Mytilus galloprovincialis. In questi organismi esposti sono stati analizzati diversi effetti a livello molecolare, biochimico e cellulare, incluse le risposte immunologiche, le alterazioni lisosomiali, la proliferazione perossisomiale, le difese antiossidanti, gli effetti neurotossici, genotossici oltre che il profilo di espressione genica. E’ stato inoltre ottimizzato e applicato un nuovo protocollo per l'estrazione e caratterizzazione (FT-IR) delle MP su diverse specie di pesci e invertebrati del Mar Mediterraneo. I livelli di IPA maggiori sono stati misurati nelle micro e macro-plastiche spiaggiate, mentre un efficiente assorbimento delle sostanze chimiche è stato confermato, sia per il PE che PS, con una modalità di adsorbimento tempo e dose-dipendente. Indagini istologiche hanno rivelato la presenza di MP nell’emolinfa, nelle branchie e nei tessuti digestivi dei mitili. Gli IPA adsorbiti sono stati rilasciati dalle MP e accumulati nei tessuti dei mitili che hanno mostrato alterazioni di diversi biomarker sia molecolari che cellulari. Il protocollo di estrazione sviluppato ha permesso di dimostrare la presenza di MP nello stomaco e, per la prima volta, nel fegato di cefali esposti. Studi sul campo hanno evidenziato la presenza di particelle nel 38% degli organismi selvatici analizzati, con una maggior frequenza di MP sotto forma di frammenti e linee, costituiti preferenzialmente da PE, PS e nylon. In conclusione, questa tesi fornisce nuove informazioni sui rischi ecotossicologici delle MP per gli organismi marini oltre che un’importante linea di base sul livello di contaminazione da MP nel biota Mediterraneo.Microplastics (MPs) are a growing but still unexplored environmental concern for marine organisms. Although several species can ingest MPs, a clear evidence of their accumulation pathways, trophic transfer and adverse effects is still lacking. In this thesis, the potential role of MPs as vectors of chemical pollutants was initially evaluated in virgin macroplastics, beached macro- and microplastics; laboratory experiments further characterized adsorbing kinetics of pyrene and cadmium on polyethylene (PE) and polystyrene (PS) MPs, and their capability to transfer adsorbed pyrene to mussels Mytilus galloprovincialis. In these organisms, several molecular, biochemical and cellular effects were analyzed in term of immunological responses, lysosomal alterations, peroxisomal proliferation, antioxidant and neurotoxic effects, genotoxicity and gene expression profile. A new protocol for extraction and FT-IR characterization of MPs in marine organisms was optimized and applied to several species of Mediterranean fish and invertebrates. Load of PAHs was higher in weathered and micron-sized particles, and an efficient adsorption of chemicals was confirmed with a time- and dose-dependent trend for both PE and PS. Histological analyses revealed occurrence of ingested MPs in haemolymph, gills and digestive tissues of mussels. Adsorbed PAHs were desorbed from MPs and bioavailable for mussels that showed many altered several molecular and cellular biomarkers. The developed extraction protocol allowed to demonstrate the presence of MPs in the stomach and, for the first time, in liver of exposed Mugil cephalus. Field studies highlighted the occurrence of MPs in 38% of analyzed wild organisms, mostly represented by fragments and lines, while PE, PS and nylon were the dominant polymers. In conclusion, this thesis provides new insights on the ecotoxicological risks of MPs for marine organisms and an important baseline for assessing the level of MPs contamination in Mediterranean biot

Neustonic microplastics in the Southern Adriatic Sea. Preliminary results.

Neustonic micro-plastic abundance and polymeric composition were determined after a cruise conducted in the Southern Adriatic Sea between May 9th and 17th 2013. Plankton samples were collected using a Neuston net (200 µm mesh size) which sampled the first 50 cm of the sea surface at a speed of ~2 kts for 5-6 minutes. Samples were then stored in ethanol 70% and in the laboratory micro-plastics were hand-picked using a dissecting stereomicroscope, counted, weighed and split into 7 different size classes. On a subset of collected particles (> 0.7mm) FT-IR analyses were performed to characterize the polymeric composition of the items. All 29 surface tows contained plastic particles of various typologies (e.g. filaments, fragments, thin plastic films), colours and sizes. A total of 5940 plastic particles were collected during the survey, the vast majority of which were hard plastic fragments (78.5%) or synthetic fibers and filaments (19.2%). Most particles were white (27.8%), transparent (22.5%) or black/grey (21.4%). 98.2% of all the particles were < 5 mm and plastic abundance markedly increased with decreasing size (i.e. 52.8% of all the particles were smaller than 0.5 mm), indicating very high fragmentation rates. Overall, an average concentration of 1.05 ± 1.13 particles/m2 and 442.88 ± 1145.96 g/km2 was observed throughout the study area, with micro-plastic densities ranging from 0.10 particles/m2 to a maximum of 4.86 particles/m2. FT-IR analyses indicated polyethylene as the predominant polymer (41%), followed by polyester and paint (12%), polypropylene (10%), polystyrene and polyimide (5%), polyammide (3%), paraffin (4%) and 1% bioplastic (i.e. polycaprolactone). In addition, 7% of the items were characterized as non-plastic materials (i.e. minerals, cellulose and cotton fabric), suggesting a potential bias when visually sorting for micro-plastics. On the whole, very high levels of plastic pollution have been found in our study area. Despite any clear geographical pattern in plastic distribution was identified, the conspicuous spatial heterogeneity in plastic abundances and polymeric compositions seem to confirm the existence of multiple pollution sources insisting on the Adriatic Sea

Major Role of Surrounding Environment in Shaping Biofilm Community Composition on Marine Plastic Debris

Plastic debris in aquatic environments is colonized by microbes, yet factors influencing biofilm development and composition on plastics remain poorly understood. Here, we explored the microbial assemblages associated with different types of plastic debris collected from two coastal sites in the Mediterranean Sea. All plastic samples were heavily colonized by prokaryotes, with abundances up to 1.9 × 107 cells/cm2. Microbial assemblages on plastics significantly differed between the two geographic areas but not between polymer types, suggesting a major role of the environment as source for the plastisphere composition. Nevertheless, plastic communities differed from those in the surrounding seawater and sediments, indicating a further selection of microbial taxa on the plastic substrates. The presence of potential pathogens on the plastic surface reflected the levels of microbial pollution in the surrounding environment, regardless of the polymer type, and confirmed the role of plastics as carriers for pathogenic microorganisms across the coastal ocean, deserving further investigations

Experimental development of a new protocol for extraction and characterization of microplastics in fish tissues: First observations in commercial species from Adriatic Sea

The presence of microplastics in the marine environment has raised scientific interest during the last decade. Several organisms can ingest microplastics with potentially adverse effects on the digestive tract, respiratory system and locomotory appendages. However, a clear evidence of tissue accumulation and transfer of such microparticles in wild organisms is still lacking, partially hampered by technical difficulties in isolation and characterization protocols from biological samples. In this work, we compared the efficacy of some existing approaches and we optimized a new protocol allowing an extraction yield of microplastics from fish tissues ranging between 78% and 98%, depending on the polymer size. FT-IR analyses confirmed that the extraction procedure did not affect the particles characteristics. The method was further validated on the fish mullet, Mugil cephalus, exposed under laboratory conditions to polystyrene and polyethylene; the particles were isolated and quantified in stomach and liver, and their presence in the hepatic tissue was confirmed also by histological analyses. A preliminary characterization revealed the presence and distribution of microplastics in various fish species collected along the Adriatic Sea. FT-IR analyses indicated polyethylene as the predominant polymer (65%) in the stomach of fish. The overall results confirmed the newly developed method as a reliable approach to detect and quantify microplastics in the marine biota

Presence of microplastics in benthic and epibenthic organisms: Influence of habitat, feeding mode and trophic level

The exponential production and use of plastics has generated increasing environmental release over the past decades, and microplastics (MPs) have been reported across all the oceans. Field studies have documented the occurrence of MPs in several species, but important knowledge gaps still remain. In the present study, we characterized the distribution of MPs in ten sediment-dwelling and epibenthic species representative of different habitat, feeding modes and trophic levels within the inner Oslofjord (Oslo, Norway), an area subjected to moderate anthropogenic pressures. Analyzed species included fish, bivalves, echinoderms, crustaceans and polychaetes. MPs were present in all the species with a frequency up to 65% of positive individuals for some species. In most cases, 1 or 2 MPs were found per individual, but some organisms contained up to 7 particles. A total of 8 polymer typologies were identified, with PE and PP being the most common according to our extraction protocol. MP sizes ranged from 41 µm to lines as long as 9 mm. Our results indicate that occurrence of MPs in analysed biota is not influenced by organism habitat or trophic level, while characteristics and typology of polymers might be significantly affected by feeding mode of organisms

Microplastics pollution after the removal of the Costa Concordia wreck: First evidences from a biomonitoring case study

Microplastics (MPs) represent a matter of growing concern for the marine environment. Their ingestion has been documented in several species worldwide, but the impact of specific anthropogenic activities remains largely unexplored. In this study, MPs were characterized in different benthic fish sampled after 2.5 years of huge engineering operations for the parbuckling project on the Costa Concordia wreck at Giglio Island. Fish collected in proximity of the wreck showed a high ingestion of microplastics compared to both fish from a control area and values reported worldwide. Also the elevated percentage of nylon, polypropylene lines and the presence of polystyrene are quite unusual for marine organisms sampled in natural field conditions, thus supporting the possible relationship of ingested microplastics with maritime operations during wreck removal. On the other hand, the use of transplanted mussels revealed a lower frequency of ingested MPs, and did not discriminate differences between the wreck and the control area. Some variations were observed in terms of typology and size of particles between surface- and bottom-caged mussels highlighting the influence of a different distribution of MPs along the water column. In conclusion, this study demonstrated that MPs pollution in the area of Costa Concordia was more evident on benthonic environment than on seawater column, providing novel insights on the possibility of using appropriate sentinel organisms for monitoring specific anthropogenic sources of MPs pollution in the marine environment

Occurrence of Microplastics in Commercial Seafood under the Perspective of the Human Food Chain. A Review

The occurrence of microplastics in the marine ecosystem and aquatic organisms, their trophic transfer along the food web, and the identification of seafood species as suitable indicators have become a research priority. Despite the high quantity of research in this field, a comparison between the available data and an appropriate risk assessment remains difficult. In this perspective, as an innovative approach, the association of the feeding strategies of commercial seafood and the microplastic level was considered. Further research to assess the occurrence of microplastics in the marine food web, the long-term effects on animals and humans, and the health implications is needed

Pollutants bioavailability and toxicological risk from microplastics to marine mussels

Microplastics represent a growing environmental concern for the oceans due to their potential of adsorbing chemical pollutants, thus representing a still unexplored source of exposure for aquatic organisms. In this study polyethylene (PE) and polystyrene (PS) microplastics were shown to adsorb pyrene with a time and dose-dependent relationship. Results also indicated a marked capability of contaminated microplastics to transfer this model PAH to exposed mussels Mytilus galloprovincialis; tissue localization of microplastics occurred in haemolymph, gills and especially digestive tissues where a marked accumulation of pyrene was also observed. Cellular effects included alterations of immunological responses, lysosomal compartment, peroxisomal proliferation, antioxidant system, neurotoxic effects, onset of genotoxicity; changes in gene expression profile was also demonstrated through a new DNA microarray platform. The study provided the evidence that microplastics adsorb PAHs, emphasizing an elevated bioavailability of these chemicals after the ingestion, and the toxicological implications due to responsiveness of several molecular and cellular pathways to microplastics

Benthic Crustacean Digestion Can Modulate the Environmental Fate of Microplastics in the Deep Sea

Microplastics (MPs) are ubiquitous contaminants of the marine environment, and the deep seafloor is their ultimate sink compartment. Manipulative and field experiments provided evidence of the ingestion of MPs by deep-sea fauna, but knowledge of MPs' fate once ingested still remains scant. We provide evidence of MP partial retention and fragmentation mediated by digestion activity of a Norwegian langoustine, a good bioindicator for MP contamination of the deep sea. We report here that MPs in the intestines were more abundant and significantly smaller (up to 1 order of magnitude in surface) than those in the stomachs. Our results show that the stomach can act as a size-bottleneck for ingested MPs, enhancing the retention of larger particles within the stomach and promoting fragmentation into smaller plastic debris, which is then released in the intestine. Our results provide evidence that the langoustine is responsible for the fragmentation of MPs already accumulated in sediments through its scavenging activity and digestion. These findings highlight the existence of a new peculiar kind of "secondary" MPs, introduced in the environment by biological activities, which could represent a significant pathway of plastic degradation in a secluded and stable environment such as the deep sea