Microplastics in oysters (Crassostrea gigas) and water at the Bahía Blanca Estuary (Southwestern Atlantic): An emerging issue of global concern

Regional Studies in Marine Science 32 (2019) 100829

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Regional Studies in Marine Science journal homepage: www.elsevier.com/locate/rsma

Microplastics in oysters (Crassostrea gigas) and water at the Bahía Blanca Estuary (Southwestern Atlantic): An emerging issue of global concern ∗

Melisa D. Fernández Severini a , , Diana M. Villagran a , Natalia S. Buzzi a,b , G. Chatelain Sartor a a b

Instituto Argentino de Oceanografía (IADO), Universidad Nacional del Sur (UNS)-CONICET, Bahía Blanca, Argentina Departamento de Biología, Bioquímica y Farmacia, Universidad Nacional del Sur (UNS), Bahía Blanca, Argentina

article

info

Article history: Received 13 August 2018 Received in revised form 26 August 2019 Accepted 8 September 2019 Available online 12 September 2019 Keywords: Plastics Fibers Environmental pollution Crassostrea gigas

a b s t r a c t Detection of microplastics (MPs) in biotic and abiotic matrices is relevant to evaluate how marine ecosystem’s exposure to these pollutants is of emerging environmental concern and at risk of loss of functionality and biodiversity. The presence of MPs was studied for the first time in the gut of benthic oysters (Crassostrea gigas) and in the water column in a eutrophic estuary under high anthropogenic pressure, in the southwestern Atlantic. Significant abundances of small plastic debris were found at all the sampling stations- mainly fibers, fragments, pellets, and beads. MPs were categorized and counted according to type, color, and size. Microfibers presented the highest percentage of abundance in the water column (98% with Van Dorn bottles and 72.73 % with a 60 µm plankton net) as well as in oysters (91%). In water collected with Van Dorn bottles, the total MP concentrations ranged from 5900 to 782,000 particles/m3 and from 42.6 to 113.6 particles/m3 in samples collected with a 60 µm plankton net. The widespread presence of fibers in all the assessed components could be related to the intense harbor activities in the area, such as the use of ropes for the mooring of boats and from fishing nets, as well as from domestic and industrial effluents. The presence of MPs in both the pelagic and benthic realms may imply risk for the animals that inhabit the estuary, and for human wellbeing, with respect to the potential transfer of MPs through the food web, affecting the provisioning of ecosystem services. © 2019 Elsevier B.V. All rights reserved.

1. Introduction Humans have been using plastics indiscriminately since their discovery in 1950s causing a substantial increase in the volume of debris of various sizes added to the ocean over the past 60 years (GESAMP, 2015; Plastics Europe, 2015). The term ‘‘plastic’’ is generally used to define a sub-category of polymers, which are very large molecules of high average weight due to their long-chain like molecular architecture (GESAMP, 2015). Another characteristic when considering these polymers as plastic material is that they soften when heated. In general, plastic debris termed microplastics (MPs) are small pieces of plastics (<5 mm) that may enter the ocean as such or may result from the fragmentation of larger pieces through weathering processes by UV radiation, physical break down, or through biological activity (Andrady et al., 1996; NOAA, 2015). Primary microplastics are those plastics manufactured as granules, pellets, and abrasive ∗ Corresponding author. E-mail address: [email protected] (M.D. Fernández Severini). https://doi.org/10.1016/j.rsma.2019.100829 2352-4855/© 2019 Elsevier B.V. All rights reserved.

microspheres, for a number of purposes like industrial abrasives or for cosmetic products that enter the environment directly. In contrast, secondary microplastics are produced through the environmental degradation of larger-sized pieces (Rillig, 2012). They are deliberately discarded, or unintentionally lost directly into the sea, or transported into the marine environment from land by rivers, drainage, sewage systems, or wind. The largest plastic producers are the sectors of packaging (39%) and construction (21%), followed by transportation, agriculture, household goods, and electronics (Pinto, 2012). Moreover, 80% of plastics in the ocean come from land, and beach litter is included within that 80%. However, beach litter can also be ocean-based litter as a consequence of the fishing industry, recreational activities and marine traffic. In this sense, about 18% of the marine plastic debris found in the ocean is attributed to the fishing industry (Andrady, 2011). There are many polymers of plastics but six classes are the most produced globally: polyethylene (PE, high and low density), polypropylene (PP), polyvinyl chloride (PVC), polystyrene (PS, including expanded EPS), polyurethane (PUR) and polyethylene terephthalate (PET).

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The ingestion of MPs by organisms implies a risk to their fitness, as this may cause weight loss (Besseling et al., 2013), decline in energy reserves and a lowering of the immune system (Wright et al., 2013), and toxicity from persistent organic pollutants (POPs) and metals that adsorb onto plastic surfaces in the environment (Andrady, 2011; Brennecke et al., 2016). Moreover, microplastics affect the antioxidant capacity and cause DNA damage, neurotoxicity and oxidative damage (Ribeiro et al., 2017). In particular, fibers are considered to be one of the most predominant types of MPs with global distribution, reaching remote areas such as polar glaciers (Obbard et al., 2014; Di Mauro et al., 2017; Salvador Cesa et al., 2017). These fibers are recorded in sediments from beaches and subtidal zones, surface and subsubsurface waters, as well as deep waters (Salvador Cesa et al., 2017), and therefore they are accidentally ingested by several marine organisms like fish, shrimp, bivalves and plankton (Di Mauro et al., 2017). Also, synthetic fibers were scored with a 7/10 degree priority as a MP source (Verschoor et al., 2014). Examples of this group of MPs are textile fibers, ropes, fishing lines and nets, and sanitary products along with others commonly found in sewage discharges. In recent years, studies have focused on sewage treatment plants as they are the intermediaries of fibers and other types of MPs from land into aquatic ecosystems. The United Nations Environment Programme (UNEP) has identified plastic pollution as a critical problem, analogous to that of climate change. Therefore, it is imperative that researchers are able to accurately isolate, identify and enumerate microplastic debris consumed by or entangled with biota (Lusher et al., 2017). The need for rapid, accurate assessment of the levels of microplastic in wild populations is essential for determining baseline levels of contamination, and assessing the risk of exposure of microplastics to organisms and ecosystems. The Bahía Blanca Estuary, located in Argentina along the SW Atlantic Ocean, represents a case study of contamination by MPs, because both sewage and industrial discharges are leading sources of these emergent pollutants into the estuarine environment. Intense port and maritime activities, fishing and tourism are also common in this estuary. Alongshore, cities have reached a population of ca. 370,000 inhabitants (Biancalana et al., 2019). The first records of MPs in the Bahía Blanca Estuary were in 2004, in samples collected with plankton nets (40 and 200 µm mesh size), where predominantly primary pellets and fibers were frequently observed (Fernández Severini unpublished data). Here, we aim to detect, quantify and classify the types of MPs that were found in water samples and in oysters (Crassostrea gigas) from the Bahía Blanca Estuary. Moreover, distribution of MPs related to industrial and domestic discharges are also assessed to track the origin and fate of these pollutants. 1.1. Study area The Bahía Blanca Estuary (38◦ 44′ –39◦ 27′ S; 61◦ 45′ –62◦ 30′ W) is regarded as one of the most important ports in the Southwestern Atlantic, characterized by strong human influence (Biancalana et al., 2019; Fernández Severini et al., 2018) with numerous chemical and petrochemical industries, turning it into one of the largest petrochemical centers in South America (Fig. 1). Among them, a factory that produces PVC and another that produces PE of various types may become one of the main potential sources of MPs, mainly primary microplastics. Hence, the estuary is affected by the contribution of exogenous substances from wastewater and industrial discharges, or leaked from wastelands, agrochemicals. Also, significant maritime traffic and port activities take place in this estuary that is considered to be the only deepwater port in Argentina (45 ft) (Fernández Severini et al., 2018). For this study sampling sites located along the estuary were selected according to their proximity to sewage and industrial discharges.

2. Materials and methods Water samples were taken at 6 sampling sites (PR: Puerto Rosales, CV: Canal Vieja, W: White, PG: Puerto Galván, M: Maldonado, and PC: Puerto Cuatreros) along the estuary in the first half of 2018 (Fig. 1). The sampling stations are located over the Main Navigation Channel of the estuary. Puerto Rosales is in the middle of the estuary. This area receives the poorly treated sewage from Punta Alta (∼60 thousand inhabitants). Its main operational activity is by boats and pontoons that support tugboat tasks in moored monobuoys, artisanal fishing boats, tourism and sport boats. Sometimes the port works with the naval workshop in repairing boats. The CV site receives untreated sewage from the city of Bahía Blanca (∼300,000 inhabitants), and PG and W have very diverse operational activities. They are commercial harbors with a constant shipping traffic; this area is considered a highly industrialized core of the estuary since prominent industries have been established there: oil, plastic polymers, by-product derivative refineries, and small-scale commercial fishing. Next to them, M receives water input from the Maldonado stream, running through farmlands and across the city of Bahía Blanca. Finally, PC is located in the innermost part of the estuary, a small recreational/fishing harbor in the vicinity of low urbanized/rural lands. Previous registries of microplastics in some plankton samples in 2004 (Fernández Severini unpublished data) were at stations PC, PG and W. Two samples of surface water were collected at each station: (1) One using a 60 µm plankton net and (2) the other one with a Van Dorn Bottle of 2 L volume. One replica was collected for each type of sample. The net was towed from a dock in the upper 50 cm of the water surface with a Hydro-Bios flowmeter for the duration of 10 min during low tide (Di Mauro et al., 2017). The volume of water filtered by the net was calculated with the flowmeter placed slightly off-center in the mouth of the net. The end of the net was folded and tied shut to avoid using a plastic (PVC) cod end. The samples were washed from the net and then transferred into glass jars and fixed with 4% formol. Bulk seawater samples were collected with a 2 L Van Dorn Bottle at approximately 1 m depth from the surface and then transferred into glass jars. Thereafter, at the laboratory, 5 aliquots of 300 ml bulk water samples (Van Dorn Bottle) were vacuum filtered on low using Glass Fiber (GF) Membrane Filters of 0.22 µm pore size and a 47 mm diameter. After filtration, the filters were placed in aluminum packets stored in wooden paper bags until analysis. Precautions were taken to avoid contamination from airborne fibers or other unwanted particles (Di Mauro et al., 2017) while processing the samples. All the material used for sample processing was made of glass. Also the glass Petri dishes used with the filters were covered with glass watches. The filters as well as the samples from the net were examined under a stereoscopic microscope (Nikon SMZ 1500) and an optical microscope (Nikon Eclipse 80i) with precaution and always covered to prevent contamination. Furthermore, identical glass Petri dishes used for control purposes were placed next to the ones with the sample to recover particles from the air, and they were checked for MPs under the stereomicroscope, but no type of airborne contamination was seen. Microplastics were counted, sized and photographed with a digital camera attached to the microscopes (Nikon). Five aliquots of 1 ml of water samples collected with the plankton net were examined for MPs using a SedgwickRafter chamber with a Pasteur pipette and the above mentioned microscopes, following the same protocol for MP identification as in bulk water. Also, glassware, metal containers and instruments were used during the sample processing, whenever possible, to avoid contamination of the samples.

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Fig. 1. Geographic location of the sampling sites at the Bahía Blanca Estuary (BBE). PR: Puerto Rosales, CV: Canal Vieja, W: White, PG: Puerto Galván, M: Maldonado and PC: Puerto Cuatreros.

In addition, 17 organisms of the oyster Crassostrea gigas were randomly collected at Puerto Galván station, where oysters presented the highest abundances in the estuary. Each oyster was analyzed twice, one for each type of digestion. Analysis of the gut content in Crassostrea gigas oysters was carried out considering two types of digestions: one with 20 ml of 22.5 M HNO3 , digested overnight (12 h) at room temperature, followed by 30 min of boiling (100 ◦ C) according to Claessens et al. (2013). The resulting mixture was then diluted 1:10 with warm deionized water, before filtration and microscopical observation. The other digestion, was based on H2 O2 30% at 55 ◦ C for seven days and then all samples were observed under the microscope, identified, sized and counted (Nuelle et al., 2014). Again, precautions to avoid contamination were taken while processing the microplastic samples during the analysis procedures and no type of airborne contamination was observed. 3. Results and discussion Several types of microplastics (MPs) of varying shapes, colors, and sizes were found in the bottle and net samples in the water column from the Bahía Blanca Estuary. Fibers, fragments, pellets, and beads were recorded (Figs. 2 and 3) and most of them were probably of secondary origin (from fragmented plastic debris that had been exposed to the marine environment for a long time). Additionally, small plastic debris were found at all of the sampling stations. Fig. 4 shows the variation of the total MP abundance (particles m−3 ) and the percentage (%) of each type of item per site in both the net (Fig. 4a, c) and bottle samples (Fig. 4b, d). One way-ANOVA was conducted to evaluate if the sampling sites displayed different abundance of plastics. Two ANOVA were conducted, one for the samples collected with Van Dorn bottles and the other for the net samples. After the statistical analysis, neither detected differences in abundance between the sampling sites (p = 0.99 in both cases, Tables 1, 2). This means that similar concentrations of MPs were found at all the stations regardless of their location. Therefore, despite the fact that industrial and urban discharges are localized (Biancalana et al., 2012), the extensive

Table 1 Results of the One-way ANOVA analysis for MPs sampled with Van Dorn bottles at different sampling sites at the BBE. Variability

Sum of squares

df

Quadratic average

F

Sig.

Between groups Inside groups Total

1 436 198 977 4.00E+15 4.00E+15

5 24 29

28 723 979.3 1.67E+14

0.002

0.99

Table 2 Results of the One-way ANOVA analysis for MPs sampled with plankton net at different sampling sites at the BBE. Variability

Sum of squares

df

Quadratic average

F

Sig.

Between groups Inside groups Total

152.388 15984.11 16136.48

5 24 29

30.478 666.005

0.046

0.99

internal circulation and mixing of the water at the estuary and high residence time (Perillo et al., 2001), result in the MPs being distributed and accumulated throughout the internal/middle zone of the estuary in a homogeneous manner. This implies a high risk of impact for the biota because MPs are widespread along the estuary. In regards to the size of the MPs in the water samples, they varied from 0.17 to 5 mm. The mean size and the standard error of the different types of MPs were: fibers 2.3 ± 0.87 mm, fragments 1.60 ± 0.97 mm, pellets 0.3 ± 0.2 mm and beads 0.24 ± 0.1 mm. The fibers significantly outnumbered the plastic particles; they showed the highest percentage of abundance (72.7–98.5%) in 100% of the water samples (bottle and net). Moreover, the fibers presented different sizes and colors (Fig. 2a, b, c, f and Fig. 3a, b, c, e, f, g) and density from 113.6 to 7.8 × 105 fibers/m3 in the net and bottle samples, respectively. The most common color was blue; under the microscope these fibers presented different characteristics regarding lattice and thickness (Fig. 3). On the other hand, the white fibers had a different structure compared to the blue ones and possibly originated from a different source. However, these differences do not necessarily indicate a different

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Fig. 2. Examples of the microplastic particles found in water samples: fibers (a,b,c,d and f); irregular fragments (c, d and e). MPs in Van Dorn bottle: a,b,c. MPs in plankton net: d,e. MPs in C.gigas: f.

type of polymer. To confirm the identity of the particles found and to determine the type of plastic, i.e. the polymer composition, a Fourier Transformed Infrared Spectroscopy in attenuated total reflectance mode (FTIR-ATR) is needed, but unfortunately in this study we had operative challenges. According to Lusher et al. (2017) a large amount of non-synthetic organic fibers (i.e. not plastic) are present in aquatic environments, and in this work the authors discussed the mistakes that can be made by confusing organic fibers with plastic. However, several measurements were taken to try to minimize this error as much as possible. Comparing our results with the work of Di Mauro et al. (2017) the blue fibers (Fig. 3d, e, g) and white fibers (Fig. 3f) in the BBE seem to be very similar to those plastic ones found by these authors in the shelf waters in the northern Gulf of Mexico, taking into account the thickness, structure, and shape. The fibers found by Di Mauro

et al. (2017) were confirmed to be plastic after evaluation with Fourier Transform Infra-Red (FTIR) microscopy and pre-treatment with hydrofluoric acid. Additionally, we compared the results of the present study with the images of plastic fibers reported by Lares et al. (2018) in different stages of a Wastewaters treatment plant in Finland, and the fibers were very similar to those found in the BBE. Also, these authors found that 96.3% of the microplastic fibers were polyester fibers being equivalent to 79.1% of the total amount of MPs collected from all of the samples. As in the BBE, fibers are predominantly seen in the study of Lares et al. (2018) with the same tendency—predominance of microplastic fibers over microplastic particles, and as also reported in the literature for wastewater effluents (Mason et al., 2016; Michielssen et al., 2016; Leslie et al., 2017; Gies et al., 2018) and the environment (Browne et al., 2011). Moreover, three rules were applied in order

M.D. Fernández Severini, D.M. Villagran, N.S. Buzzi et al. / Regional Studies in Marine Science 32 (2019) 100829

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Fig. 3. Fibers (a,b,c,e,f, and g); pellet (d); bead (h). MPs in Van Dorn bottle: a,b,c,d,e. MPs in plankton net: h. MPs in C.gigas: f,g.

to discriminate between plastic and non-plastic as described by Hidalgo-Ruz et al. (2012): (1) the object should not have any cellular or organic structures, (2) the fibers should be equally thick throughout the entire length, (3) and the color of the particles should be clear and homogeneous throughout. After considering these rules, all the fibers found in the BBE presented the characteristics of plastic fibers. It is worth mentioning that a subsample of the fibers was isolated and subjected to the effects of acid nitric for 24 hs in glass Petri dishes to check the plastic composition. After the acid digestion these fibers were still present in the Petri

dishes so therefore we considered them as plastic. Finally, when possible, the fiber was taken with a clamp and heated with a hot needle as proposed by Lusher et al. (2017). Moreover, regarding the type of fibers, 60% of world fibers that are consumed are synthetic fibers, among which polyester, polyamide, acrylic, polypropylene and polyolefin are the most common (FAO-ICAC, 2013). Also, PP, one of the most common compositions in fibers, are less dense than seawater, and therefore they are concentrated in surface waters (Takana and Takana, 2016). Moreover, Andrady (2011) affirms that, globally, almost

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Fig. 4. Variation of the total MPs abundances (particles m−3 ) and the percentage of each type of item per site in both net samples (Fig. 4a, c) and bottle samples (Fig. 4b, d).

all fishing activities use synthetic materials, such as polyamide and polyolefin, and some of the most common objects lost or discarded are constituted of textile fibers, or include ropes, lines, and fishing nets (Cole et al., 2011). In the study area, fishing activities, as well as maritime traffic, have a significant presence so part of the plastic fibers in the BBE may account from these activities. On the other hand, Lusher et al. (2013) recorded fibers in 68.3% of plastics in fish from the English Channel, identified as rayon, polyamide, or polyester. They also found that rayon is commonly found in clothing, furnishings, female hygiene products, and diapers, hence high levels could be the result of sewage discharges. Items made of rayon can disintegrate rapidly (Park et al., 2004) which could explain their abundance. In the study of Browne et al. (2011), the authors found that polyester and acrylic fibers used in clothing resembled those found in habitats that receive sewage-discharges and sewage-effluent itself and they demonstrated that clothing can release more than 1900 fibers at each wash. So sites in close proximity to sewage discharge may contain proportions of fibers resembling the proportions used in synthetic clothing. This is in agreement with the findings of the present study, because some of the sampling stations (CL, W, M, and PR) are located a few meters away from sewage

discharges, or nearby (PG and PC). The station CL is 1000 m from the industrial discharge (Industrial Collector Channel) and 300 m from the domestic one, where there is a primary treatment plant for domestic effluents (First Basin). W station is 2300 m from this latter effluent. Also, M station is located at 1700 m from another treatment plant of domestic effluents called Third Basin, as well as the PC station that is 900 m from this point. On the other hand, the PG station is 900 m from the industrial discharge (Industrial collector channel) and PR that is 3500 m from the domestic effluents of the city of Punta Alta. All these effluents are ultimately discharged into the estuary and therefore could contribute to the MPs pool. The wastewater treatment plant of domestic effluents called First Basin is often operating improperly; when the filters are working inefficiently they could increase the levels of fibers and other MPs in the estuary. At this plant, the primary filtering treatment is carried out through two rotating systems of sieves that remove part of the solid material. After a primary treatment the waters are discharged into the estuary. However, seeing as though this primary treatment does not often operate properly, raw sewage is actually being poured directly into the estuary most of the time. Moreover, during the sampling in this study, the primary treatment at the First Basin

M.D. Fernández Severini, D.M. Villagran, N.S. Buzzi et al. / Regional Studies in Marine Science 32 (2019) 100829

Fig. 5. Fragment of red plastic (1.6 mm) found in Crassostrea gigas. Table 3 Results of the ‘‘t’’ student test to compare digestion protocols (Nitric acid and hydrogen peroxide ) of MPs in gut content of C. gigas. t

df

Sig.

Mean diff.

Typical error of the difference

3.354

4

0.028

5

1.49

treatment plant was not working. Accordingly, the authors of this study have personally observed in this area, plastic remains of all sizes of waste discharge, as well as fabrics, agglomerates of soap powder and even female hygiene products of domestic origin. In addition, industrial discharges may also contribute to the MPs pool. As mentioned previously here, two plants of PVC and PP are also located at this estuary, and may possibly be dumping their waste into the estuary. The above mentioned treatment plant called Third Basin also presents some deficiencies with the primary treatment which could also have negative effects due to its contribution of MPs in the estuary. Domestic washings of textiles produce fiber emissions, which eventually end up in the BBE as a consequence of these deficiencies in the WWTP. In this sense, Browne et al. (2011) detected that the individual washing of a textile article can spread >1900 microplastics. Once in the environment, fibers can reach concentrations of up to thousands of particles per cubic meter, and so they are available for ingestion by a broad range of species, such as C. gigas in the present study. Other sewage of industrial origin is also dumped into an industrial waste collector channel, which then discharges into the BBE waters. Moreover, some streams that flow into the estuary located near the sampling points (900– 5000 m) receive industrial and domestic effluents which could also be potential sources of microplastics. Therefore these results are a warning sign because these wastewater treatment plants (WWTPs) may be acting as routes of microplastics (MPs) into the environment instead of being retention plants. For example, in C. gigas fibers (4.2 fibers/ind.), plastic fragments (2 fragments/ind.; Fig. 5), and beads (0.5 item/ind.; Fig. 3h) have been detected in both protocols of MP digestion. However, statistical differences were detected between these protocols considering the number of plastic items (‘‘t’’ Student test: p = 0.028, n = 3 for each treatment, Table 3). Diverse digestion methods are described in the literature for MP isolation (e.g. Vandermeersch et al., 2015; Lusher et al., 2017) and the chosen digestion type can also have an influence on the quantification of microplastics. In the present study, gut samples of oysters digested with 30% H2O2 at 55 ◦ C for seven days, presented incomplete digestion and lower MP abundance (2 item/ind.), with many tissue and lipid residues that obstructed the detection of microplastics. On the other hand,

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samples in 22.5 M HNO3 presented a more complete digestion, with a cleaner sample and higher abundance (7 item/ind.). The results of the digestion with 30% H2 O2 at 55 ◦ C for seven days were similar to that reported by Nuelle et al. (2014) where it was demonstrated that in the treatment with 35% H2 O2 , only 25% of the biological material is removed after an H2 O2 exposure period of 7 days. Moreover, in the review of Lusher et al. (2017) the authors mentioned that nitric acid (HNO3 ), like the one employed in this study, is a strong oxidizing mineral acid, capable of molecular cleavage and rapid dissolution of biogenic material. When tested against hydrochloric acid (HCl), hydrogen peroxide (H2 O2 ) and sodium hydroxide (NaOH), HNO3 showed the highest digestion efficacies, with >98% weight loss of biological tissue. In the present study, acid digestion with nitric acid also seemed to be the most appropriate method for C. gigas, however some small tissue remnants were observed after acid digestion. Therefore, in order to fully remove these residues, further studies with C. gigas in the BBE will consider perchloric acid (HClO4 ) like De Witte et al. (2014) who proposed a mixture of 65% HNO3 and 68% perchloric acid (HClO4 ) in a 4:1 v/v ratio (500 ml acid to 100 g tissue) overnight at room temperature, followed by 10 min boiling. However, this method with perchloric acid overnight followed by 10 min boiling, could result in fiber degradation (Lusher et al., 2017), so samples will be exposed to fewer hours with this acid and only at room temperature. For this purpose, tests with oyster samples, as well as fibers with perchloric acid for a different numbers of hours, will be carried out to determine the minimum amount of hours necessary to digest the sample without damaging the fibers that the oysters may contain. Moreover, Claessens et al. (2013) and the International Council for the Exploration of the Sea (ICES) advice group (2015) mentioned the potential negative effect of nitric acid and perchloric acid on fibers and determined that nylon fibers are possibly destroyed. However, other types of synthetic fibers, like polyethylene (fishing nets or dolly rope) and polypropylene fibers, which are also found in great abundance in the marine environment, should withstand this type of digestion. In the present study, only nitric acid was used, not perchloric acid, which is stronger, therefore fibers may have resisted this acid attack. Another type of digestion is alkaline hydrolysis, in which NaOH or KOH are used to digest biological material. However, Cole et al. (2014) showed that alkaline treatment causes physical damage and discoloration of microplastics. On the other hand, Dehaut et al. (2016) have tested several protocols and they suggested the modification of Rochman et al. (2015) (10% KOH, 24 h, 60 ◦ C) as the most suitable protocol in terms of the absence of substantial degradation of the plastic polymers and good tissue degradation. Finally, enzymatic methods (Cole et al., 2014) are difficult methodologies to implement and have some digestion deficiencies (Dehaut et al., 2016). As can be seen, there is currently no single method of digestion of tissues associated with plastics, so depending on which protocol is chosen the quantification of MPs varies. Moreover, the characterization and quantification methods, such as sample size, contamination measures, inclusion of procedural blanks, digestion method, pore size filters, visualization method and assessment, blank correction, also vary (Hidalgo-Ruz et al., 2012; Vandermeersch et al., 2015; Dehaut et al., 2016). Under this scenario, further research is necessary to determine the optimal digestion assay, as well as a harmonized characterization for a better and more accurate quantification of MPs in pollution studies. In this particular case of fibers found in C. gigas, this type of plastic was the most important in terms of abundance and size (Fig. 3f, g) as in the water samples. Statistical analysis also shows that no differences were detected in the percentage of the types of MPs in oysters or those in a plankton net (Table 4) as well as in bottle samples (Table 5). Numerous studies show the prevalence of microfibers

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in marine biota. In mussels, for example Karlsson et al. (2017) found that MP concentrations were approximately a thousandfold higher compared to those in surface water and sediment, from the same location. In addition, the authors found that 50% of the microplastics found in mussels were fibers with a higher average length than the other particles. Another study on MP ingestion in bivalves in China found that microfibers accounted for more than half of the items present (Li et al., 2015). In our study, a higher percentage of fibers was recorded but their lengths were also greater than the other MPs. These results indicate that C. gigas in the BBE effectively incorporate MPs in the same way as detected in mussels and oysters worldwide (De Witte et al., 2014; Leslie et al., 2017; Karlsson et al., 2017; Jauregui, 2017). The novelty of these findings is that these results correspond to the first records of MPs ingested by the exotic and invasive oyster C. gigas in the Bahia Blanca Estuary and also in Argentina. Even though the composition of fibers in both water samples (bottle and net) and oysters is unknown in the present study, most of the fibers reported worldwide are of PP and so the fibers in the BBE could be composed of this polymer. As previously mentioned, among the major polymers only PE and PP are less dense than seawater, and therefore they are predominant in surface waters (Takana and Takana, 2016). In the case of C. gigas sampled in this study, they were attached to dock pilings, where ships load and unload merchandise; hence fibers in the water could be present as a consequence of these port activities and so part of ingested plastics is due the water filtered by the oysters . Since oysters filter water to feed on plankton and suspended particulates, a large proportion of the fibers found in the stomach content are due precisely to this water. Oysters capture plankton by opening their shells and filter water through their gills to extract food from the water. While they are feeding, the oysters are cleaning the water which is a secondary function of their feeding processes. The size of material, typically less than five (5) millimeters like MPs, that passes through their filters (gills) is what they will consume. Thus, the water they filter is the source of fibers. As long as the material is the same size of their food, they will eat it regardless if it has no nutritional value. Also, zooplankton in the water column, ingested by the oysters may be another source of MPs because they also present MPs inside their body, which were accidentally incorporated. In addition, there is an industrial waste collector channel near this site at 900 m, a discharge that may also contribute to the MP pool in the water. Also, some small tributaries located at 10 km receive domestic (from the Third basin WWTP) and industrial effluents that could also be a source of the MPs found in the oysters. The other WWTP, called First Basin, is also located at 10 km and due to the lack of the primary treatment previously mentioned, may also be a source of MPs, especially fibers. Regarding the colors of the MPs in all the samples, most of the MPs found in the present study were colored or white (Figs. 2, 3 and 5) and this deserves special attention because colors cause microplastics to resemble the natural food that is probably ingested by the organisms (Andrady, 2011; Di Mauro et al., 2017). Also, fibers for example can be very small and become available to zooplankton as a hazardous food item (Cole et al., 2011). Alternatively, organisms at higher trophic levels than the plankton, such as oysters, may display fibers or other MPs from eating plankton that had consumed MPs accidentally (Fendall and Sewell, 2009). Therefore, part of MPs in the oyster C. gigas at the estuary may be from zooplankton ingestion. Mytilus edulis for example can ingest MPs via the inhalant siphon and then filter them out through the gills and transport them to the labial palps for digestion or rejection. However, it was shown that microspheres were also translocated in the circulatory system of these organisms (Cole et al., 2011). Au et al. (2015) found negative effects of fibers in an amphipod crustacean and Savoca (2018), in an interesting article, discussing the

Table 4 Results of the ‘‘t’’ student test to compare percentage of the types of MPs in oysters and those in plankton net. t

df

Sig.

Mean Diff.

Typical error of the difference

0.000

6

0.99

0

25.45

Table 5 Results of the ‘‘t’’ student test to compare percentage of the types of MPs in oysters and those in bottle samples. t

df

Sig.

Mean Diff.

Typical error of the difference

0.000

6

0.99

0

26.43

mistaken notion that all seabirds consume plastic simply because of its appearance. This notion is incongruous with decades of research demonstrating that Procellariiformes are highly reliant on odor cues for foraging. In addition, some species never ingest plastic, whereas others seem unable to avoid it. Seabirds possess an acute olfactory sense and are also severely affected by the ingestion of plastic. In particular, marine predators—including certain species of fish, seabirds and sea turtles—track relative concentrations of DMS (dimethyl sulfide) over the open ocean in order to find productive regions to forage. DMS, is an odoriferous algal-derived molecule. Some Procellariiform seabirds (burrownesting petrels and shearwaters) are likely to follow an odor trail of DMS, to locate nutrient-rich zooplankton (such as krill). When phytoplankton are consumed by zooplankton, DMS is released in mass from the depredated phytoplankton. Savoca et al. (2017) found that those species that use DMS for foraging were almost six times more likely to consume plastic than those that are not attracted to DMS. Moreover, they found that every sample of ocean-weathered (i.e. biofouled) plastic emitted DMS at concentrations six orders of magnitude above the detection threshold for Procellariiforms and three orders of magnitude above background DMS in the environment. Furthermore, there was no DMS signature on clean plastics in the study of Savoca et al. (2017). The distinct DMS signature associated with marine plastic debris originates from the algal biofilm that rapidly coats plastic at sea. In conclusion Savoca (2018) found that marine plastics may falsely amplify an olfactory signal that certain species associate with foraging opportunities and suggested that plastic debris may be more confusing and appetizing to marine organisms than previously thought possible. This behavior could also extend to the oysters at the BBE. Small microorganisms like plankton can grow onto plastic and utilize it as a platform for transportation as well. These slimly looking substances will release dimethyl sulfide (DMS) into the water as it is being eaten and/or crushed and DMS can behave as a cue to organisms like C. gigas that food is in the area. These oysters could be ‘‘attracted’’ to plankton that emit a DMS type odor, and thus increase the water filtration rate. Therefore, plankton can give a false positive that the plastic they are eating is food. Since they have no true way of distinguishing between actual foods and plastic, oysters can be easily deceived. However, the potential mechanism by which oysters can detect DMS and thus increase the water filtration rate must be tested. In the present study, fibers accounted for between 72.73% and 98.54% of items found in the study area. High percentages of fibers were also recorded in other areas, which indicate that this item is commonly found in oceans around the world. For example, in the review of Rochman et al. (2015) the authors found that anthropogenic debris recovered from fish in the USA were primarily fibers. In the Río de la Plata Estuary, a coastal ecosystem also located in the SW Atlantic, Pazos et al. (2017) studied the presence of MPs in the gut contents of coastal freshwater fish

Table 6 Informative data of MPs abundances around the world found in different types of samples. Type of microplastic

Type of sample

Abundance

Tunisian coast

Total microplastics

Sediments

141.20 ± 25.98–461.25 ± 29.74 items kg−1 d.w.

Golf of Mexico

Fibers Bead Irregular Total microplastics

Bongo and Neuston water samples

0.2–11.9 E4 m−3 (µ = 3.86 ± 1.36 particles m−3 ) 0.2–1.9 m−3 (µ = 0.76 ± 0.23 particles m−3 ) 0.8–16.9 m−3 (µ = 6.73 ± 2.03 particles m−3 ) 5.1–23.7 m−3 (µ = 11.3 ± 3.26 particles m−3 )

Fibers Bead Irregular Total microplastics

Niskin × 10000

5.9–11.9 (µ = 7.4 ± 1.8 particles m−3 ) 0.4–1.9 (µ = 0.8 ± 0.4 particles m−3 ) 0.8–3.5 (µ = 2 ± 0.6 particles m−3 ) 6–15.7 (µ = 10.2 ± 2.1 particles m−3 ) g −1 g −1 g −1 g −1

w.w. w.w. w.w. w.w.

Reference Abidli et al. (2018)

1.0 ml of 40%–45% (∼22.6 M) HF at room temperature for 24 h

Di Mauro et al. (2017)

Nitric acid and perchloric acid

Griet et al. (2015)

Italy Spain

Fibers Particles Particles Fibers

Mussel Mytilus galloprovincialis

0.29 0.06 0.05 0.15

USA

Fibers

Pacific oyster Crassostrea gigas

0–2 (µ = 0.6 ± 0.9 pieces per animal)

10% KOH solution

Norway

Fiber, film, fragment

Surface waters Subsurface waters

0–1.31 m−3 (µ = 0.34 ± 0.31 particles m−3 ) 0–11.5 m−3 (µ = 2.68 ± 2.95 particles m−3 )

10% KOH solution

Atlantic and Indian Ocean and Mediterranean Sea

Fiber

Deep sea sediments

1.4–40 piece 50 ml−1 (µ = 13.4 ± 3.5)

Concentrated NaCl solution and filtering with three sequential extractions

Woodall et al. (2014)

Yangtze River estuary (Pacific Ocean)

Total microplastics

Surface water

µ = 4137.3 ± 2461.5 particles m−3

30% H2 O2 and saturated zinc chloride solution

Zhao et al. (2014)

Geoje Island, South Korea

Total microplastics

English Channel

Bead, fiber, planar fragment, granular

Subsurface waters

0.24–0.35 item m−3 (µ = 0.26 item m−3 )

Enzymatic (Proteinase-K) digestion

Cole et al. (2014)

Atlantic Ocean

Bead, fiber, foam, fragment

Subsurface waters

0–22.5 m−3 (µ = 2.46 ± 2.43 particles m−3 )

Filtered samples with glass microfiber paper (GF/C)

Lusher et al. (2014)

Pacific Ocean

Fiber fragment

Subsurface waters

8.51–9180 m−3 (µ = 2080 ± 2190 particles m−3 )

Acid digested at 80–90 C for 3 h using concentrated HCl

Desforges et al. (2014)

Belgian coastal zone

Total microplastics

Harbor sediments Sublittoral sediments

66.9 ± 7.9–390.7 ± 32.6 particles kg−1 d.w. 71.5 ± 6.4–115.8 ± 13.3 particles kg−1 d.w.

Particles were separated by flotation

Plymouth (UK)

Fiber, fragment

Sandy sediments Estuarine sediments Subtidal sediments

<1 fibers 50 ml−1 <3 fibers 50 ml−1 <7 fibers 50 ml−1

Particles were separated by flotation

Portugal

particles particles particles particles

Digestion or other isolation method

µ = 211 ± 117 particles m−3

Rochman et al. (2015) Lusher et al. (2015)

Song et al. (2015)

Claessens et al. (2011)

M.D. Fernández Severini, D.M. Villagran, N.S. Buzzi et al. / Regional Studies in Marine Science 32 (2019) 100829

Area

Thompson et al. (2004)

9

10

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and they found that MPs were recorded in 100% of fish samples and that fibers represented 96% of the MPs, being the highest percentage documented in the bibliography according to these authors. High percentages of fibers were also reported by Boerger et al. (2010) (94%) and Lusher et al. (2013) (68.3%). Recently, Vermaire et al. (2017) also found that fibers were the most common form of plastic (70%–100%) and Salvador Cesa et al. (2017) affirm that fibers can reach concentrations of up to thousands of particles per cubic meter. Considering all this information, it is evident that fibers are one of the most important MP items worldwide. Hence, studies on fibers in the aquatic environments are very important because, as previously mentioned, this type of pollutant is commonly found in domestic discharges as well as from fishing and it may reach remote areas on earth. Unfortunately, to date, apart from the urgent call for a reduction in fiber emission, general textile sources, for example, are not known or are poorly explored (Salvador Cesa et al., 2017). On the other hand, the survey of the BBE coastal environment in the SW Atlantic showed that MPs were present in all sampling sites, an observation consistent with the ubiquitous nature of MPs in the marine environment. Although there were some variations between the sampling sites in terms of abundance (Fig. 4a,b) and composition, our results are comparable to those reported in other studies worldwide (e.g. Peng et al., 2017; Di Mauro et al., 2017; Salvador Cesa et al., 2017). For example a dominant number of fibers in sandy and sedimentary samples were reported in previous studies (Claessens et al., 2011; Thompson et al., 2004; Abidli et al., 2018), and Zhao et al. (2015) found fibers as the second most common type of plastic debris on sandy beaches, most of which was polypropylene. Also, Salvador Cesa et al. (2017), in a review of synthetic fibers like MPs in marine environments, described some instances when the abundance was similar to that recorded in the BBE in the water samples collected from Van Dorn bottles. Di Mauro et al. (2017) also found similar results of higher concentrations of MPs in Niskin bottles than in net samples, and the Niskin bottles collected smaller plastic particles than the nets which were primarily fibers. Moreover, the values recorded in this study were among the highest reported globally. Table 6 presents informative data of the above mentioned studies on MPs worldwide. However, it is worth mentioning that the comparison of concentrations of microplastics between different regions is complex due to the use of different sampling methods as well as the microplastic treatments to isolate them from the unwanted particles or tissues (Hidalgo-Ruz et al., 2012; Di Mauro et al., 2017; Lusher et al., 2017). Moreover, there is also a high degree of variability in the sampling nets used in studies that evaluated the concentration of pelagic microplastics (sea surface and water column). Overall, there is an urgent need to harmonize procedures for sampling, extraction, identification, assessment, as well as quality assurance, in order to mitigate airborne contamination. 4. Conclusions To our knowledge, this study represents the southernmost latitudinal record of MPs in South America, in water as well as in the oyster C. gigas, and it demonstrates the presence of microplastics in the BBE for the first time. They are of different types, colors, shapes and sizes with a high dominance of fibers. Also, these findings show the high availability or chance to consume fibers by organisms such as plankton, bivalves, crabs and fish, indicating potential negative effects on these organisms that inhabit the coastal environment of the BBE. Particularly, in the case of the oyster C. gigas in the BBE, the ingestion of microplastics was demonstrated through the analysis of gut contents. Acid digestion with HNO3 , as employed in the present study, was a good treatment that provided the confirmation of plastic particles

at low cost and high effectivity in both water samples and the gut contents of oysters. Also, the visual comparison with images of plastics in other studies was also a good complement. On the other hand, analytical methodologies for microplastics discrimination and determination including FTIR or Raman microspectroscopy (RMS) are important tools and a complement to the chemical treatment of the MPs samples. Therefore, a subset of previous acid confirmed plastics could then be analyzed with FTIR or RMS to determine the type of polymer. It is necessary to consider samples from both the plankton net of 60 µm mesh size as well as Varn Dorn bottles for MPs analysis at the BBE. Moreover, Van Dorn bottles are useful for fibers because they may be underestimated with plankton nets, however plankton nets are very useful in those areas where MP concentrations are low. This study was the first attempt to document MPs in the BBE over a short period of time and only covering the inner zone of the estuary. This study was a pilot-scale attempt to document microplastics in the BBE and the spatiotemporal bounds and sampling design were limited. These results, however, are essential for establishing ecologically relevant data, which finally provide a clear view of the quantity and types of plastic encountered by biota in the natural environment, such as the BBE. Microplastics appear to be present in the area at concentrations that are of a similar magnitude as those found in hotspots around the world. Untreated domestic and industrial effluents, located near the sampling points, would be the main source of MPs which together with the heavy marine traffic negatively affect the estuary. Future studies will be carried out by the authors of this work and will analyze the concentrations, characteristics and sources of MPs over the whole estuary as well as the sandy beach areas closely associated with the BBE. Additionally, more exhaustive samplings at the WWTPs will be incorporated. In particular, the analysis of fibers will be continued and their sources and pathways into the environment will be evaluated on account of their great abundance found in the estuary, as well as the urgent need to explore issues related to this kind of pollution. Also future research on other organisms, together with the environmental occurrence of MPs should be considered in order to determine the potential effects of MPs in the BBE and nearby coastal areas, such as the beaches at Monte Hermoso and Pehuen-Co, as well as to establish the interaction between organisms and MPs. References Abidli, S., Antunes, J.C., Ferreira, J.L., Lahbib, Y., Sobral, P., El Menif, N.T., 2018. Microplastics in sediments from the littoral zone of the north Tunisian coast (Mediterranean Sea). Estuar. Coast. Shelf Sci. 205, 1–9. Andrady, A.L., 2011. Microplastics in the marine environment. Mar. Pollut. Bull. 62 (8), 1596–1605. Andrady, A.L., Pegram, J.E., Searle, N.D., 1996. Wavelength sensitivity of enhanced photodegradable polyethylenes, ECO, and LDPE/MX. J. Appl. Polym. Sci. 62 (9), 1457–1463. Au, S.Y., Bruce, T.F., Bridges, W.C., Klaine, S.J., 2015. Responses of Hyalella azteca to acute and chronic microplastic exposures. Environ. Toxicol. Chem. 34 (11), 2564–2572. Besseling, E., Wegner, A., Foekema, E.M., van den Heuvel-Greve, M.J., Koelmans, A.A., 2013. Effects of microplastic on fitness and PCB bioaccumulation by the lugworm Arenicola marina (L.). Environ. Sci. Technol. 47 (1), 593–600. Biancalana, F., Fernandez-Severini, M.D., Villagran, D.M., Berasategui, A.A., Tartara, M.N., Spetter, C.V., Guinder, V., Marcovecchio, J.E., Lara, R.J., 2019. Assessment of chitin variation in seston of a temperate estuary (Bahía Blanca Argentina). Environ. Earth Sci. 78, 4. Biancalana, F., Menéndez, M.C., Berasategui, A.A., Fernández-Severini, M.D., Hoffmeyer, M.S., 2012. Sewage pollution effects on mesozooplankton structure in a shallow temperate estuary. Environ. Monit. Assess. 184 (6), 3901–3913. Boerger, C.M., Lattin, G.L., Moore, S.L., Moore, C.J., 2010. Plastic ingestion by planktivorous fishes in the North Pacific Central Gyre. Mar. Pollut. Bull. 60, 2275–2278.

M.D. Fernández Severini, D.M. Villagran, N.S. Buzzi et al. / Regional Studies in Marine Science 32 (2019) 100829 Brennecke, D., Duarte, B., Paiva, F., Caçador, I., Canning-Clode, J., 2016. Microplastics as vector for heavy metal contamination from the marine environment. Estuar. Coast. Shelf Sci. 178, 189–195. Browne, M.A., Crump, P., Niven, S.J., Teuten, E., Tonkin, A., Galloway, T., 2011. Accumulation of microplastic on shorelines worldwide: sources and sinks. Environ. Sci. Technol. 45 (21), 9175–9179. Claessens, M., De Meester, S., Van Landuyt, L., De Clerck, K., Janssen, C.R., 2011. Occurrence and distribution of microplastics in marine sediments along the Belgian coast. Mar. Pollut. Bull. 62 (10), 2199–2204. Claessens, M., Van Cauwenberghe, L., Vandegehuchte, M.B., Janssen, C.R., 2013. New techniques for the detection of microplastics in sediments and field collected organisms. Mar. Pollut. Bull. 70, 227–233. Cole, M., Lindeque, P., Halsband, C., Galloway, T.S., 2011. Microplastics as contaminants in the marine environment: A review. Mar. Pollut. Bull. 62, 2588–2597. Cole, M., Webb, H., Lindeque, P.K., Fileman, E.S., Halsband, C., Galloway, T.S., 2014. Isolation of microplastics in biota-rich seawater samples and marine organisms. Sci. Rep. 4, 4528. De Witte, B., Devriese, L., Bekaet, K., Hoffman, S., Vandermeersch, G., Cooreman, K., Robbens, J., 2014. Quality assessment of the blue mussel ( Mytilus edulis) comparison between commercial and wild types. Mar. Pollut. Bull. 85, 146–155. Dehaut, A., Cassone, A.L., Frère, L., Hermabessiere, L., Himber, C., Rinnert, E., Rivière, G., Lambert, C., Soudant, P., Huvet, A., Duflos, G., Paul-Pont, I., 2016. Microplastics in seafood: Benchmark protocol for their extraction and characterization. Environ. Pollut. 215, 223–233. Desforges, J., Galbraith, M., Dangerfield, N., Ross, 2014. Widespread distribution of microplastics in subsurface seawater in the NE Pacific Ocean. Mar. Pollut. Bull. 79, 94–99. Di Mauro, R., Kupchik, M.J., Benfield, M.C., 2017. Abundant plankton-sized microplastic particles in shelf waters of the northern Gulf of Mexico. Environ. Pollut. 230, 798–809. FAO-ICAC, 2013. Food and Agriculture Organization of the United Nations and International Cotton Advisory Committee, World Apparel Fiber Consumption Survey, Washington. (5 pp. (Retrieved from)). https://www.icac.org/cotton_info/publications/statistics/world-apparelsurvey/FAO-ICAC-Survey-2013-Update-and-2011-Text.pdf. Fendall, L.S., Sewell, M.A., 2009. Contributing to marine pollution by washing your face: Microplastics in facial cleansers. Mar. Pollut. Bull. 58, 1225–1228. Fernández Severini, M.D., Carbone, M.E., Villagran, D.M., Marcovecchio, J.E., 2018. Toxic metals in a highly urbanized industry-impacted estuary (Bahia Blanca Estuary, Argentina) spatio-temporal analysis based on GIS. Environ. Earth Sci. 77, 393–412. GESAMP, 2015. Sources, fate and effects of microplastics in the marine environment: a global assessment (Kershaw, P. J. ed.). In: (IMO/FAO/UNESCOIOC/UNIDO/WMO/IAEA/UN/UNEP/UNDP Joint Group of Experts on the Scientific Aspects of Marine Environmental Protection). Rep. Stud. GESAMP (90) 96 p. Gies, E.A., LeNoble, J.L., Noël, M., Etemadifar, A., Bishay, F., Hall, E.R., Ross, P.R., 2018. Retention of microplastics in a major secondary wastewater treatment plant in vancouver, Canada. Mar. Pollut. Bull. 133, 553–561. Griet, V., Van Cauwenberghe, L., Janssen, C.R., Marques, A., Granby, K., Fait, G., Kotterman, M.J.J., Diogène, J., Bekaert, K., Robbens, J., Devriese, L., 2015. A critical view on microplastic quantification in aquatic organisms. Environ. Res. 143 (2015), 46–55. Hidalgo-Ruz, V., Gutow, L., Thompson, R.C., Thiel, M., 2012. Microplastics in the marine environment: a review of the methods used for identification and quantification. Environ. Sci. Technol. 46, 3060–3075. ICES, 2015. OSPAR Request on Development of a Common Monitoring Protocol for Plastic Particles in Fish Stomachs and Selected Shellfish on the Basis of Existing Fish Disease Surveys. ICES Special Request Advice, book, 1, 6 pp. Jauregui, M.K., 2017. Microplastic Concentrations in Crassostrea gigas: Establishing a Baseline of Microplastic Contamination in Oregon’s Oyster Aquacultures. University Honors Theses. Paper 494. Karlsson, T.M., Vethaak, A.D., Almrothd, B.C., Ariese, F., van Velzen, M., Hassellöv, M., Leslie, H.A., 2017. Screening for microplastics in sediment, water, marine invertebrates and fish: Method development and microplastic accumulation. Mar. Pollut. Bull. 122, 403–408. Lares, M., Chaker Ncibi, M., Sillanpää, M., Sillanpää, M., 2018. Occurrence, identification and removal of microplastic particles and fibers in conventional activated sludge process and advanced MBR technology. Water Res. 133, 236–246. Leslie, H.A., Brandsma, S.H., van Velzen, M.J., Vethaak, A.D., 2017. Microplastics en route: field measurements in the dutch river delta and Amsterdam canals, wastewater treatment plants, North Sea sediments and biota. Environ. Int. 101, 133–142.

11

Li, J., Yang, D., Li, L., Jabeen, K., Shi, H., 2015. Microplastics in commercial bivalves from China. Environ. Pollut. 207, 190–195. Lusher, A., Burke, A., O’Connor, A., Officer, R., 2014. Microplastic pollution in the northeast atlantic ocean: validated and opportunistic sampling. Mar. Pollut. Bull. 88 (1–2), 325–333. Lusher, A.L., McHugh, M., Thompson, R.C., 2013. Occurrence of microplastics in the gastrointestinal tract of pelagic and demersal fish from the English Channel. Mar. Pollut. Bull. 67, 94–99. Lusher, A.L., Tirelli, V., O’Connor, I., Officer, R., 2015. Microplastic in Arctic polar waters: the first reported values of particles in surface and sub-surface samples. Sci. Report 5 (14947), 1–9. http://dx.doi.org/10.1038/srep14947. Lusher, A.L., Welden, N.A., Sobral, P., Cole, M., 2017. Sampling, isolating and identifying microplastics ingested by fish and invertebrates. Anal. Methods 9, 1346–1360. Mason, S.A., Garneau, D., Sutton, R., Chu, Y., Ehmann, K., Barnes, J., Fink, P., Papazissimos, D., Rogers, D.L., 2016. Microplastic pollution is widely detected in US municipal wastewater treatment plant effluent. Environ. Pollut. 218, 1045–1054. Michielssen, M.R., Michielssen, E.R., Ni, J., Duhaime, M.B., 2016. Fate of microplastics and other small anthropogenic litter (SAL) in wastewater treatment plants depends on unit processes employed. Environ. Sci. J. Integr. Environ. Res. Water Res. Technol. 2 (6), 1064–1073. NOAA, 2015. A NOAA MDP research project focuses on types and abundance of microplastics. Detecting microplastics in the marine environment. Available from: http://marinedebris.noaa.gov/research/detectingmicroplastics-marine-environment. Nuelle, M.T., Dekiff, J.H., Remy, D., Fries, E., 2014. A new analytical approach for monitoring microplastics in marine sediments. Environ. Pollut. 184, 161–169. Obbard, R., Sadri, S., Wong, Y.Q., Khitun, A.A., Baker, I., Thompson, R.C., 2014. Global warming releases microplastic legacy frozen in Artic Sea ice. Earth’s Future 2, 315–320. Park, C.H., Kang, Y.K., Im, S.S., 2004. Biodegradability of cellulose fabrics. J. Appl. Polym. Sci. 94 (1), 248–253. Pazos, R.S., Maiztegui, T., Colauttia, D.C., Paracampo, A.H., Gómez, N., 2017. Microplastics in gut contents of coastal freshwater fish from Río de la Plata estuary. Mar. Pollut. Bull. 122, 85–90. Peng, G., Zhu, B., Yang, D., Su, L., Shi, H., Li, D., 2017. Microplastics in sediments of the changjiang estuary, China.. Environ. Pollut. 225, 283–290. Perillo, G.M.E., Piccolo, M.C., Parodi, E.R., Freije, R.H., 2001. The Bahía Blanca Estuary ecosystem: a review. In: Seelinger, U., Lacerda, L., Kjerve, B. (Eds.), Coastal Marine Ecosystems of Latin America. Springer Verlag, Heidelberg, Germany, pp. 205–217. Pinto, J.C., 2012. Impactos ambientais causados pelos plásticos: uma discussão abrangente sobre os mitos e os dados científicos (2a ed.) Editora E-papers. Plastics Europe, 2015. Plastics-The Facts 2014/2015: An Analysis of European Plastics Production, Demand and Waste Data. Retrieved May, 2016, from. http://www.plasticseurope.org/documents/document/20150227150049final_ plastics_the_facts_2014_2015_260215.pdf. Ribeiro, F., Garcia, A.R., Pereira, B.P., Fonseca, M., Mestre, N.C., Fonseca, T.G., Ilharco, L.M., Bebianno, M.J., 2017. Microplastics effects in scrobicularia plana. Mar. Pollut. Bull. 122, 1–2. Rillig, M.C., 2012. Microplastic in terrestrial ecosystems and the soil?. Environ. Sci. Technol. 46, 6453–6454. Rochman, C.M., Tahir, A., Williams, S.L., Baxa, D.V., Lam, R., Miller, J.T., The, F.C., Werorilangi, S., The, S.J., 2015. Anthropogenic debris in seafood: Plastic debris and fibers from textiles in fish and bivalves sold for human consumption. Sci. Rep. 5, 14340. Salvador Cesa, F., Turra, A., Baruque-Ramos, J., 2017. Synthetic fibers as microplastics in the marine environment: A review from textile perspective with a focus on domestic washings. Sci. Total Environ. 598, 1116–1129. Savoca, M., 2018. The ecology of an olfactory trap. Science 362 (6417), 904–905. Savoca, M.S., Tyson, C.W., McGill, M., Slager, C.J., 2017. Odours from marine plastic debris induce food search behaviours in a forage fish. Proc. R. Soc. B 284, 20171000. Song, Y.K., Hong, S.H., Jang, M., Han, G.M., Shim, W.J., 2015. Occurrence and distribution of microplastics in the sea surface microlayer in Jinhae Bay, South Korea. Arch. Environ. Contam. Toxicol. 69 (3), 279–287. Takana, K., Takana, H., 2016. Microplastic fragments and microbeads in digestive tracts of planktivorous fish from urban coastal waters. Sci. Rep. 6, 34351. Thompson, R.C., Olsen, Y., Mitchell, R.P., Davis, A., Rowland, S.J., John, A.W.G., McGonigle, D., Russel, A.E., 2004. Lost at sea: where is all the plastic?. Science 304 (5672), 838. Vandermeersch, G., Van Cauwenberghe, L., Janssen, C.R., Marques, A., Granby, K., Fait, G., Kotterman Michiel, J.J., Diogène, J., Bekaert, K., Robbens, J., Devriese, L., 2015. A critical view on microplastic quantification in aquatic organisms. Environ. Res. 143 (Part B), 46–55.

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Vermaire, J.C., Pomeroy, C., Herczegh, S.M., Haggart, O., Murphy, M., 2017. Microplastic abundance and distribution in the open water and sediment of the Ottawa River, Canada, and its tributaries. FACETS 2, 301–314. Verschoor, A., Poorter, L., Roex, E., Bellert, B., 2014. Quick scan and prioritization of microplastic sources and emissions. RIVM Advisory Letter 250012001. National Institute for Public Health and Environment, Bilthoven (NL) :p. 41 (Retrieved from). http://www.rivm.nl/dsresource?objectid=b14d58ab-55794b17-aae2-5c5e1e21eb4f&type=org&disposition=inline. Woodall, L.C., Sanchez-Vidal, A., Canals, M., Paterson, G.L.J., Coppock, R., Sleight, V., Calafat, A., Rogers, A.D., Narayanaswamy, B.E., Thompson, R.C., 2014. The deep sea is a major sink for microplastic debris. R. Soc. Open Sci. 1 (140317), 1–8.

Wright, S.L., Rowe, D., Thompson, R.C., Galloway, T.S., 2013. Microplastic ingestion decreases energy reserves in marine worms. Curr. Biol. 23 (23), 1031–1033. Zhao, S., Zhu, L., Li, D., 2015. Characterization of small plastic debris on tourism beaches around the south China Sea. Reg. Stud. Mar. Sci. 1, 55–62. Zhao, S., Zhu, L., Wang, T., Daoii, L., 2014. Suspended microplastics in the surface water of the Yangtze Estuary system in China. Mar. Pollut. Bull. 86 (1–2), 562–568.