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the highest concern is centered on the extreme accumulation level found in sediments collected in the estuarine station, which is certainly attributed to the untreated sewage and stormwater discharges occur-ring nearby.
While the intensification of urban runoff and river discharges, typical of the winter season due to rainfall, ended up fueling the abundance of MPs at the surface, this was not observed in sediments.
The absence of such seasonal influence in the accumulation of MPs in the seabed may be linked to the increase of the Sado river current velocity in winter, delaying the sinking process of MPs. Despite such dissimilarities regarding temporal distribution patterns, both studies indicate a higher accumulation in the station closer to Setúbal city and suggest that the pool of MPs occurring in the remaining stations result from the weak, but continuous, exportation from the Sado estuary. In fact, the expected gradual seaward decrease in MPs abundance, at this south faced coastal area, was not observed. Instead, while in seawater surface there is a nearshore retention effect potentially occurring due to the shelter provided by Arrábida mountain chain against the prevailing north and north-west winds, in sediments there is an abrupt decrease of MPs abundance at the ocean exposed stations.
The categorization of MPs found in both matrices suggests a consistent predominance of frag-ments at the most polluted station: inside the estuary. However, while at the water surface, both foam and film types were also quite represented, being a pattern also observed in the remaining stations; at the estuarine sediments, the fragment type largely prevailed in contrast to the other types, and, in marine sediments, fibers were the most abundant type of MPs. Whereas the preponderance of secondary MPs (fragments, foams and films) at the surface may be attributed to the breakdown of larger objects, as a potential source; the predominance of fragments in estuarine sediments, i.e., close to their potential sources, was majorly linked to their lower surface area to volume ratio in comparison to fibers, that allows a faster sinking process.
Another aspect to be noted, regarding MPs vertical distribution inside the estuary, concerns to their size. Although both studies do not share the same exact size classes, their comparison suggest that, at the surface, MPs are bigger, being the 1–2 mm the predominant size class, while in sediments most MPs belong to the 0.250–0.500 mm size class. Although such size difference might be strongly related to the sampling methods applied, the smaller MPs in sediments could be also explained by granulometric characteristics of sediments and by the fragmentation process occurring during the settlement of parti-cles. Further sampling taking place in our study area should consider the collection of surface samples with a smaller mesh size to guarantee the quantification of smaller MPs which remained underestimated.
At the end, this baseline data also intends to be use as a reference dataset in both future moni-toring and experimental studies. For example, the MP:neuston, MP:ichthyoplankton and MP:meiofauna ratios provided in chapter 2. (D. Rodrigues et al., 2020) and chapter 3. (D. Rodrigues et al., 2022) may be useful in further ecotoxicological research when aiming at selecting environmental realistic exposure
conditions. Moreover, the analysis of MPs composition and distribution patterns in subtidal compart-ments has also enabled the evaluation of the use of a fish species as a bioindicator of MPs pollution in this coastal area (chapter 4.).
In fact, the concurrent collection of fish specimens and environmental samples (water and sed-iments) enabled a proper comparison which has been rarely performed. Though the selected species, Boops boops (Linnaeus, 1758), has been frequently suggested as a potential bioindicator in the Medi-terranean, our study shows a weak match between MPs ingested and those available in this species feeding grounds (most similarities with sediments). Such match mainly relies in the predominance of fibers among the MPs ingested and, on the polymers identified, which were denser than seawater. The lack of a stronger coincident tendency could be explained by the observed decrease in food intake along the sampling campaigns, which consequently relates to this fish species reproduction season. Therefore, in order to properly conclude about this species suitability as bioindicator in Portuguese waters, further studies should be conducted during the spring/summer months.
Lastly, this thesis last aim was to contribute to the understanding of the effects of MPs ingestion (directly and by trophic transfer) in fish larvae, a critical and fragile life stage, which also faces other ecotoxicological risks, as being exposed to pollutants (as nonylphenol; NP) through dermal uptake and prey ingestion. Although the analysis of water samples from the aquaria indicated a considerably smaller concentration of NP in water than the nominal one (5 µg/l), thus approaching levels which are not con-sidered as toxic, our findings suggest that under a realistic exposure, namely to more than one type of MPs, to different polymers, to an environmentally relevant MP:prey ratio and to a relevant concentration of NP in water, there are no significant effects occurring in mortality and biomarkers response to report.
Nevertheless, fish larvae simultaneously exposed to NP and MPs presented the lowest levels of both antioxidant and detoxification enzymes (catalase and glutathione S-transferase) and the highest of vitel-logenin. It should be also noticed that growth was only affected in larvae exposed to water borne nonylphenol, suggesting that the exposure to MPs may contribute to remove NP from their tissues and consequently play a detoxification role. Further studies are thus necessary to understand if the activity of such biomarkers would occur differently if the exposure was longer or if the measured concentration of NP was in fact closer to 5 µg/l.
To conclude, besides the immediate contribution of this thesis for the scientific community working on microplastics pollution, I believe that it consists of a critical foundation for further studies that aim to monitor such important study area. Moreover, findings here reported mirror the importance of sharing the knowledge with all sectors of society, which I intended to assure in all the opportunities which crossed my PhD path. Investing in environmental education is critical and should be faced as a solution for microplastics pollution.