Short-term physiological and biometrical responses of Lepidium sativum seedlings exposed to PET-made microplastics and acid rain.

Abstract Plastics enter in terrestrial natural system primarily by agricultural purposes, while acid rain is the result of anthropogenic activities. The synergistic effects of microplastics and acid rain on plant growth are not known. In this study, different sizes of polyethylene terephthalate (PET) and acid rain are tested on Lepidium sativum, in two separate experimental sets. In the first one we treated plants only with PET, in the second one we used PET and acid rain together. In both experimentations we analyzed: i) plant biometrical parameters (shoot height, leaf number, percentage inhibition of seed germination, fresh biomass), and ii) oxidative stress responses (hydrogen peroxide; ascorbic acid and glutathione). Results carried out from our experiments highlighted that different sizes of polyethylene terephthalate are able to affect plant growth and physiological responses, with or without acid rain supplied during acute toxicity (6 days). Short description This study showed that different sizes of PET microplastics affect physiological and biometrical responses of Lepidum sativum seedlings, with or without acid rain; roots and leaves responded differently

Intra-Laboratory Calibration Exercise for Quantification of Microplastic Particles in Fine-Grained Sediment Samples: Special Focus on the Influence of User Experience

An intra-laboratory calibration to quantify microplastic in fine-grained marine sediments was performed with two objectives: (a) to determine the recovery rate of self-produced microplastics characterized by a size ranging from 220 µm to 5 mm and differing in color (pink, orange, gray, yellow, silver), shape (fragments, filaments, spheres, films), and chemical composition (polystyrene, polyethylene, polyvinyl chloride, acrylonitrile-butadiene-styrene, polypropylene, poly(methyl methacrylate)) artificially introduced into real samples; and (b) to analyze whether operator experience can be a key factor in the quality of the results. To answer this question, the same protocol was assigned to an experienced and an inexperienced operator. The results of this comparison are detailed in terms of root mean square and percent error. Possible strategies to increase the recovery rate are presented, and an ad hoc category, namely “glitter”, was created to adjust the results with respect to this unique type of microplastic usually ignored and excluded from the analysi

Understanding Marine Biodegradation of Bio-Based Oligoesters and Plasticizers

The study reports the enzymatic synthesis of bio-based oligoesters and chemo-enzymatic processes for obtaining epoxidized bioplasticizers and biolubricants starting from cardoon seed oil. All of the molecules had MW below 1000 g mol-1 and were analyzed in terms of marine biodegradation. The data shed light on the effects of the chemical structure, chemical bond lability, thermal behavior, and water solubility on biodegradation. Moreover, the analysis of the biodegradation of the building blocks that constituted the different bio-based products allowed us to distinguish between different chemical and physicochemical factors. These hints are of major importance for the rational eco-design of new benign bio-based products. Overall, the high lability of ester bonds was confirmed, along with the negligible effect of the presence of epoxy rings on triglyceride structures. The biodegradation data clearly indicated that the monomers/building blocks undergo a much slower process of abiotic or biotic transformations, potentially leading to accumulation. Therefore, the simple analysis of the erosion, hydrolysis, or visual/chemical disappearance of the chemical products or plastic is not sufficient, but ecotoxicity studies on the effects of such small molecules are of major importance. The use of natural feedstocks, such as vegetable seed oils and their derivatives, allows the minimization of these risks, because microorganisms have evolved enzymes and metabolic pathways for processing such natural molecules

Lethal and Sublethal Responses of Hydropsyche pellucidula (Insecta, Trichoptera) to Commercial Polypropylene Microplastics after Different Preconditioning Treatments

Microplastics (MPs) pose biological and chemical hazards in aquatic and terrestrial food webs across the globe. Research on microplastic contamination has long focused on marine ecosystems, whereas the toxicological impact on freshwater organisms is still little explored. In this study, the lethal and sublethal response of the freshwater macroinvertebrate Hydropsyche pellucidula exposed to polypropylene MPs after different pre-conditioning treatments was assessed. Field samples were collected in a riverine system (Vipacco river; northeast Italy) to assess the characteristics of the MPs in the aquatic environment Both water and sediment were contaminated by MPs (3.73 ± 2.11 items m−3 per min and 3.33 ± 4.16 items dm−3, respectively). The chemical MPs composition included polystyrene, polyethylene terephthalate, polyurethane, polyamide, polypropylene, and polyethylene. Polypropylene (PP), although not the most abundant polymer recorded in the study area, was preferred over the other types according to its abundance in freshwater and H. pellucidula feeding behavior. A housing test was performed to recreate the natural conditions of larvae sampled for a reliable response to the ecotoxicological tests. The microplastics underwent either preconditioning with Vipacco River water (PP-river) and surfactant Triton X-100 (PP-sf) or no pre-treatment (PP). Submersion of microplastics in 10 µg L−1 of surfactant solution for 24 h was sufficient to induce consistent spectral changes and modify the chemical profile of the plastic surface. Mortality rate differed according to treatment: PP and PP-river > positive control > PP-sf > negative control. Integrated biomarker response (IBRv2) and analysis of oxidative stress biomarker levels showed a greater response of superoxide dismutase and lipid peroxidation (malondialdehyde) in larvae treated with PP conditioned in surfactant. Our findings enhance knowledge on the toxicity of PP and conditioning phases on H. pellucidula larvae.This research received no external funding.Peer reviewe

Lethal and Sublethal Responses of Hydropsyche pellucidula (Insecta, Trichoptera) to Commercial Polypropylene Microplastics after Different Preconditioning Treatments

Microplastics (MPs) pose biological and chemical hazards in aquatic and terrestrial food webs across the globe. Research on microplastic contamination has long focused on marine ecosystems, whereas the toxicological impact on freshwater organisms is still little explored. In this study, the lethal and sublethal response of the freshwater macroinvertebrate Hydropsyche pellucidula exposed to polypropylene MPs after different pre-conditioning treatments was assessed. Field samples were collected in a riverine system (Vipacco river; northeast Italy) to assess the characteristics of the MPs in the aquatic environment Both water and sediment were contaminated by MPs (3.73 ± 2.11 items m−3 per min and 3.33 ± 4.16 items dm−3, respectively). The chemical MPs composition included polystyrene, polyethylene terephthalate, polyurethane, polyamide, polypropylene, and polyethylene. Polypropylene (PP), although not the most abundant polymer recorded in the study area, was preferred over the other types according to its abundance in freshwater and H. pellucidula feeding behavior. A housing test was performed to recreate the natural conditions of larvae sampled for a reliable response to the ecotoxicological tests. The microplastics underwent either preconditioning with Vipacco River water (PP-river) and surfactant Triton X-100 (PP-sf) or no pre-treatment (PP). Submersion of microplastics in 10 µg L−1 of surfactant solution for 24 h was sufficient to induce consistent spectral changes and modify the chemical profile of the plastic surface. Mortality rate differed according to treatment: PP and PP-river > positive control > PP-sf > negative control. Integrated biomarker response (IBRv2) and analysis of oxidative stress biomarker levels showed a greater response of superoxide dismutase and lipid peroxidation (malondialdehyde) in larvae treated with PP conditioned in surfactant. Our findings enhance knowledge on the toxicity of PP and conditioning phases on H. pellucidula larvae

Effetti tossicologici di Micro- e Nano-plastiche in diversi organismi marini modello

Questo progetto di dottorato si sviluppa su due linee di ricerca, ciascuna delle quali focalizzata su una particolare frazione del marine litter: le MICROplastiche (MP, < 5 mm) e le NANOplastiche (NP, < 1\ub5m). In riferimento alla prima linea di ricerca, nel primo lavoro pubblicato e interamente riportato nella tesi, sono state investigate le possibili interazioni fra MP di PET (polietilentereftalato) ed altri fattori potenzialmente capaci di influenzarne la tossicit\ue0, quali il pH dell\u2019acqua di mare (Acido = 7.5 vs. Standard = 8.0) e la disponibilit\ue0 di cibo (presenza/assenza), testando la tossicit\ue0 sia degli eluati che delle sospensioni di MP di diversa dimensione (5-3,000 \ub5m) in un test multispecie composto da Vibrio fischeri, Phaeodactylum tricornutum e Paracentrotus lividus. Le analisi sono state poi, ulteriormente estese a giovanili di pesce pagliaccio Amphiprion ocellaris (esposti per 3 giorni a sospensioni di MP in presenza di cibo). La risposta biologica si \ue8 basata sulla misura dell\u2019attivit\ue0 enzimatica di singoli biomarker legati allo stress ossidativo (CAT; GST) e sulla capacit\ue0 totale di scavenger ossidoradicale per mezzo del saggio TOSCA. Dai risultati ottenuti nella prima linea di ricerca, e alla luce delle concentrazioni testate, risulta una bassa tossicit\ue0 legata alle MP di PET. In particolare: i) le risposte sono state specie-specifiche; ii) gli eluati si sono dimostrati generalmente pi\uf9 tossici delle sospensioni; iii) la taglia pi\uf9 piccola delle MP non sempre ha causato risposte biologiche pi\uf9 evidenti; iv) il pH e le variazioni di cibo sono in grado di modulare le risposte ecotossicologiche degli echinodermi secondo meccanismi non dimostrabili allo stato attuale delle conoscenze; v) non \ue8 stata evidenziata una risposta concentrazione-dipendente nei giovanili di pesce pagliaccio, infatti la concentrazione pi\uf9 bassa di MP ha indotto effetti 1.87 volte superiori rispetto alla concentrazione pi\uf9 alta. Riguardo la seconda linea di ricerca, nella review pubblicata e interamente riportata nella tesi, viene indagata la conoscenza scientifica dell\u2019inquinamento da NP, dalla degradazione dei polimeri alla loro presenza in campioni di acqua marina, dedicando particolare attenzione agli aspetti riguardanti gli effetti ecotossicologici. Nel secondo lavoro riportato nella tesi, viene descritto l\u2019esperimento di 7 giorni in cui giovanili di pesce pagliaccio sono stati esposti a NP di polistirene (PS-COOH). Le possibili implicazioni biologiche vengono indagate per mezzo di un approccio multidisciplanare basato sulla risposta biochimica connessa allo stress ossidativo in aggiunta a quella molecolare basata su tecniche di Next Generation Sequencing quali il sequenziamento del trascrittoma codificante (RNA-seq). Da quanto appreso durante la preparazione della review, le NP sembrerebbero essere un contaminante dagli effetti allarmanti, tuttavia, molte domande (tra tutte, la quantificazione del fenomeno) rimangono irrisolte e dunque risulta prematuro prendere una posizione sul rischio reale che le NP eserciterebbero sulla biodiversit\ue0 e sul funzionamento degli ecosistemi naturali. Il test realizzato su giovanili di pesce pagliaccio ha voluto colmare parzialmente la scarsit\ue0 di conoscenze sugli effetti delle NP nei pesci marini ed ha evidenziato come: i) la CAT, la GST, la GR e il saggio TOSC idrossilico e perossilico, non siano stati significativamente alterati; ii) l\u2019analisi integrata delle risposte biologiche (indice IBRv2) riporti una riduzione globale dello stato di salute dei pesci esposti alle NP dimostrando come tale indice possa rappresentare un approccio rapido, semplice ed informativo; iii) la concentrazione pi\uf9 bassa sia in grado di suscitare risposte tossicogenomiche maggiori rispetto alle concentrazioni medie ed elevate; iv) i geni con un cambiamento pi\uf9 elevato nei livelli trascrizionali siano stati quelli coinvolti nell'infezione virale e nella risposta infiammatoria.This PhD project was developed on two parallel lines of research, each of which concentrated on a specific fraction of the marine litter: the MICROplastic (MP, < 5 mm) and the NANOplastic (NP, < 1\ub5m). With reference to the first line of research, the first full paper reported in this thesis is an investigation of the possible interactions between PET (polyethylene terephthalate) MP and some other factors potentially able to influence MP toxicity, namely water pH (Ac=7.5 and St=8.0) and food variations (presence/absence). The toxicity of leachates and suspensions of different size (5-3,000 \ub5m) of PET MPs was evaluated in a multispecies test composed by Vibrio fischeri, Phaeodactylum tricornutum and Paracentrotus lividus. The analyses were subsequently extended also to marine teleosts: juveniles of Amphiprion ocellaris were investigated in a 3-days water-exposure to suspensions of PET MPs under feeding conditions. Both single enzymes related to oxidative stress response (CAT; GST) and total scavenger capacity (TOSCA assay) were tested. According to the final results obtained, considering the MP concentrations analyzed in these studies, a low toxicity is attributable to PET MPs and, in particular: i) responses were species-specific; ii) leachates demonstrated to be more toxic than suspensions of MPs; iii) smaller sizes of plastics not always corresponded to stronger biological responses; iv) pH and food variations modulated the ecotoxicological responses of echinoderms according to mechanisms not demonstrable in the current state of knowledge; v) a concentration-dependent response was not evident in clownfish juveniles in which the lower concentration of MPs induced effects 1.87 times greater than the higher concentration. With regard to the second line of research, the first full paper reported in this thesis explores the scientific knowledge on the topic, from polymer degradation to occurrence in marine water, passing across laboratory evidence of nanoplastic formation. The works examined were classified on the basis of the aspects covered, with particular attention being paid to those relating to ecotoxicological effects, and suggestions were also made for future studies. In the second full paper reported in this thesis, a 7-days exposure to 100 nm PS-COOH nanoplastics in juveniles of clownfish explores the possible biological implications adopting a multidisciplinary approach spanning from biochemical responses related to oxidative stress, to gene expression analysis by the Next Generation Sequencing (NGS) of the whole transcriptome (RNA-seq technique). As we learn from the review, the framework on nanoplastics toxicity seems to be alarming. However, many knowledge gaps (quantification of the phenomenon first) still exist and therefore it is difficult to take a position about the risk they pose to the biodiversity and functioning of natural ecosystems. The experiment performed on clownfish juveniles detected a minor response in oxidative stress and contributed in understanding the cascade of molecular events that are activated by the exposure to PS-NPs in the clownfish juveniles. Specifically: i) sensitive biomarkers such as CAT, GST, GR and TOSC Assay (hydroxylic and peroxylic) were not significantly altered; ii) the integrated analysis of biomarker responses reported a reduction in overall health status of fishes demonstrating how the IBRv2 index may represent a quick, useful and informative approach; iii) RNA-seq analysis highlighted significant dysregulation of immune genes especially in the low condition which elicited higher toxicogenomic response than medium and high concentrations; iv) remarkably, the genes showing higher change in transcriptional levels were predominantly factors involved in viral infection and inflammatory response, and to less extent in ciliogenesis, energy metabolism, olfactory and reproductive functions

Journey into the Local Market in Search of &ldquo;Glitter&rdquo; Microparticles: Mini Product Investigation and Relative Chemical-Physical Characterization

There is one type of primary microplastic, glitter, which has not received the same attention as microbeads. The general thought associates the use of glitter with extraordinary and rare events, in reality, its presence in everyday life is much more ubiquitous than one might think. The main objective of this preliminary study was to create a mini product survey with the aim of analyzing the distribution and composition of glitter in the local market of the city of Trieste (Italy), thus creating a starting point for larger future studies. The analysis of a real context served as a snapshot for the description of the phenomenon, from which useful information was obtained: glitter is everywhere, cheap, disperses easily in the environment, is made of different materials, often has sharp-edged shapes, and mainly affects sensitive audiences, such as children. It is time to put glitter in the spotlight

Journey into the Local Market in Search of “Glitter” Microparticles: Mini Product Investigation and Relative Chemical-Physical Characterization

There is one type of primary microplastic, glitter, which has not received the same attention as microbeads. The general thought associates the use of glitter with extraordinary and rare events, in reality, its presence in everyday life is much more ubiquitous than one might think. The main objective of this preliminary study was to create a mini product survey with the aim of analyzing the distribution and composition of glitter in the local market of the city of Trieste (Italy), thus creating a starting point for larger future studies. The analysis of a real context served as a snapshot for the description of the phenomenon, from which useful information was obtained: glitter is everywhere, cheap, disperses easily in the environment, is made of different materials, often has sharp-edged shapes, and mainly affects sensitive audiences, such as children. It is time to put glitter in the spotlight