Microplastics (MPs) and microfibers (MFs) represent environmental challenges threatening marine ecosystems due to their persistence and ubiquitous distribution in abiotic and biotic matrices. MFs have been often included in MPs, however these can be either of synthetic (e.g. polyamide and polyester) or natural origin (linen, cotton, and jute), and recent studies demonstrated the latter being the most abundant in the field environment. The great variety of shapes, polymers and sizes of MPs and MFs may influence their behavior in the marine environment, as well as their fate and effects in marine organisms. Furthermore, marine species are subjected to multiple stressors deriving from climate-change related factors and a growing number of anthropogenic disturbances: these emerging contaminants are therefore of concern even for the possibility to enhance the susceptibility of organisms toward other co-occurring stressors. This thesis aimed to (i) determine the biological effects of MFs, elucidating differences between synthetic- and natural-polymers and evaluating if these can increase the vulnerability of organisms toward a second stress, and (ii) understand if different dimensional classes of MPs have diverse biological effects, providing a relationship between size and toxicity. With such purposes, two experiments were carried out with the Mediterranean mussel Mytilus galloprovincialis. In the first experiment organisms were exposed for 14 days to polyester (PEST), polyamide (PA) or cotton (CO) microfibers at a concentration of 50 MFs L-1; after the exposure, organisms were allowed to recover for 7 days in clean seawater under two thermal regimes, one at the same temperature used throughout the exposure (recovery, 23°C) and the other simulating a heatwave scenario, choosing thermal stress (+4°C) as a second stress. Neuro-immune and genotoxic effects, antioxidant defenses, oxidative damages and changes in lipid metabolism were evaluated in mussels’ tissues at the end of each phase-condition. In the second experiment, M. galloprovincialis were exposed for 10 days to 1000 polyethylene-MPs L-1 of the following size classes: XS (20 - 50 µm), S (50 - 100 µm), M (100 - 250 µm), L (250 - 500 µm), XL (500 - 1000 µm). At the end of the exposure, mussels haemolymph and gills were collected to evaluate neuro-immune alterations and assess possible size-effect relationships. Exposure to MFs caused disturbance of the immune system and a higher demand of protection toward oxidative insult, particularly evident in synthetic-MFs exposed organisms; these effects were paralleled by the onset of carry-over alterations after 7 days of recovery, mainly affecting cholinergic enzymes, lipid peroxidation and metabolism. At the same time, however, organisms that had been exposed to MFs, independently on their origin, showed a higher susceptibility of the immune system to thermal stress. Concerning the effect of size, a reduction of haemocytes lysosomal membrane stability and an induction of AchE in the gills were observed in organisms exposed to 50 - 250 µm particles, while an increased number of nuclear alterations were detected in organisms exposed to largest size particles. Overall findings suggest that shape, polymeric-composition and size may deeply influence the effects of MPs in marine organisms, confirming the need to consider such intrinsic characteristics when assessing risks deriving from such emerging contaminants.
Le microplastiche (MPs) e le microfibre (MFs) rappresentano minacce ambientali per gli ecosistemi marini a causa della loro persistenza e distribuzione ubiquitaria nelle matrici abiotiche e biotiche. Le MFs sono spesso considerate parte delle MPs, tuttavia studi recenti hanno dimostrato che queste possono essere sia di origine sintetica (ad esempio poliammide e poliestere) che naturale (lino, cotone e iuta) e studi recenti hanno dimostrato che quest'ultima rappresenta la frazione più abbondante in ambiente. La grande varietà di forme, polimeri e dimensioni di MPs e MFs può influenzare il loro comportamento nell'ambiente marino, nonché il loro destino e gli effetti negli organismi marini. Inoltre, le specie marine sono soggette a molteplici stress derivanti da fattori legati ai cambiamenti climatici e da un numero crescente di disturbi antropogenici: questi contaminanti emergenti destano quindi preoccupazione anche per la possibilità che la loro presenza influenzi la suscettibilità degli organismi ad altri fattori di stress concomitanti. Questa tesi ha avuto i seguenti obiettivi: (i) determinare gli effetti biologici delle MF, chiarendo le differenze tra polimeri sintetici e naturali e valutando se questi possono aumentare la vulnerabilità degli organismi verso un secondo stress, e (ii) capire se diverse classi dimensionali di MPs hanno diversi effetti biologici, fornendo una relazione tra dimensione e tossicità. A tal fine sono stati condotti due esperimenti con il mitilo mediterraneo Mytilus galloprovincialis. Nel primo esperimento gli organismi sono stati esposti per 14 giorni a microfibre di poliestere (PEST), poliammide (PA) o cotone (CO) ad una concentrazione di 50 MF L-1; dopo l'esposizione, gli organismi sono stati lasciati per 7 giorni in acqua di mare pulita in due regimi termici, uno alla stessa temperatura utilizzata durante l'esposizione (recovery, 23°C) e l'altro simulando uno scenario di heatwave, scegliendo lo stress termico (+4 °C) come secondo stress. Alla fine di ogni fase sperimentale sono stati valutati gli effetti neuro-immunitari e genotossici, le difese antiossidanti, i danni ossidativi e le variazioni del metabolismo lipidico nei tessuti dei mitili. Nel secondo esperimento, gli organismi sono stati esposti per 10 giorni a 1000 MPs di polietilene L-1 delle seguenti classi dimensionali: XS (20 - 50 µm), S (50 - 100 µm), M (100 - 250 µm), L (250 - 500 µm), XL (500 - 1000 µm). Al termine dell'esposizione l'emolinfa e le branchie dei mitili sono stati utilizzati per valutare possibili relazioni dimensione-effetto in termini di alterazioni neuro-immunitarie e di genotossicità. L'esposizione a MFs ha causato disturbi del sistema immunitario e una maggiore richiesta di protezione antiossidante, particolarmente evidente negli organismi esposti a MF sintetiche; questi effetti sono stati seguiti dall'insorgenza di alterazioni anche dopo 7 giorni di recovery, principalmente coinvolgendo enzimi colinergici, il metabolismo lipidico e accumulo di prodotti della perossidazione lipidica. Allo stesso tempo, gli organismi che erano stati esposti a MFs, indipendentemente dalla loro origine, hanno mostrato una maggiore suscettibilità del sistema immunitario allo stress termico. Per quanto riguarda l'effetto delle dimensioni, negli organismi esposti a particelle di 50-250 µm è stata osservata una riduzione della stabilità delle membrane lisosomiali degli emociti e un'induzione dell’AchE nelle branchie, mentre l’esposizione a particelle di dimensioni maggiori ha causato un aumento del numero di alterazioni nucleari. Nel complesso, i risultati suggeriscono che la forma, la composizione polimerica e le dimensioni possono influenzare profondamente gli effetti degli MPs negli organismi marini, confermando la necessità di considerare tali caratteristiche intrinseche nella valutazione dei rischi derivanti da tali contaminanti emergenti.
RISPOSTE BIOLOGICHE DI MITILI ESPOSTI A MICROPLASTICHE DI FORMA E TAGLIA DIFFERENTE.
MONGERA, FEDERICA
2020/2021
Abstract
Microplastics (MPs) and microfibers (MFs) represent environmental challenges threatening marine ecosystems due to their persistence and ubiquitous distribution in abiotic and biotic matrices. MFs have been often included in MPs, however these can be either of synthetic (e.g. polyamide and polyester) or natural origin (linen, cotton, and jute), and recent studies demonstrated the latter being the most abundant in the field environment. The great variety of shapes, polymers and sizes of MPs and MFs may influence their behavior in the marine environment, as well as their fate and effects in marine organisms. Furthermore, marine species are subjected to multiple stressors deriving from climate-change related factors and a growing number of anthropogenic disturbances: these emerging contaminants are therefore of concern even for the possibility to enhance the susceptibility of organisms toward other co-occurring stressors. This thesis aimed to (i) determine the biological effects of MFs, elucidating differences between synthetic- and natural-polymers and evaluating if these can increase the vulnerability of organisms toward a second stress, and (ii) understand if different dimensional classes of MPs have diverse biological effects, providing a relationship between size and toxicity. With such purposes, two experiments were carried out with the Mediterranean mussel Mytilus galloprovincialis. In the first experiment organisms were exposed for 14 days to polyester (PEST), polyamide (PA) or cotton (CO) microfibers at a concentration of 50 MFs L-1; after the exposure, organisms were allowed to recover for 7 days in clean seawater under two thermal regimes, one at the same temperature used throughout the exposure (recovery, 23°C) and the other simulating a heatwave scenario, choosing thermal stress (+4°C) as a second stress. Neuro-immune and genotoxic effects, antioxidant defenses, oxidative damages and changes in lipid metabolism were evaluated in mussels’ tissues at the end of each phase-condition. In the second experiment, M. galloprovincialis were exposed for 10 days to 1000 polyethylene-MPs L-1 of the following size classes: XS (20 - 50 µm), S (50 - 100 µm), M (100 - 250 µm), L (250 - 500 µm), XL (500 - 1000 µm). At the end of the exposure, mussels haemolymph and gills were collected to evaluate neuro-immune alterations and assess possible size-effect relationships. Exposure to MFs caused disturbance of the immune system and a higher demand of protection toward oxidative insult, particularly evident in synthetic-MFs exposed organisms; these effects were paralleled by the onset of carry-over alterations after 7 days of recovery, mainly affecting cholinergic enzymes, lipid peroxidation and metabolism. At the same time, however, organisms that had been exposed to MFs, independently on their origin, showed a higher susceptibility of the immune system to thermal stress. Concerning the effect of size, a reduction of haemocytes lysosomal membrane stability and an induction of AchE in the gills were observed in organisms exposed to 50 - 250 µm particles, while an increased number of nuclear alterations were detected in organisms exposed to largest size particles. Overall findings suggest that shape, polymeric-composition and size may deeply influence the effects of MPs in marine organisms, confirming the need to consider such intrinsic characteristics when assessing risks deriving from such emerging contaminants.| File | Dimensione | Formato | |
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https://hdl.handle.net/20.500.12075/8242