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Brain damage and behavioural disorders in fish induced by plastic nanoparticles delivered through the food chain


Abstract and Figures

The tremendous increases in production of plastic materials has led to an accumulation of plastic pollution worldwide. Many studies have addressed the physical effects of large-sized plastics on organisms, whereas few have focused on plastic nanoparticles, despite their distinct chemical, physical and mechanical properties. Hence our understanding of their effects on ecosystem function, behaviour and metabolism of organisms remains elusive. Here we demonstrate that plastic nanoparticles reduce survival of aquatic zooplankton and penetrate the blood-to-brain barrier in fish and cause behavioural disorders. Hence, for the first time, we uncover direct interactions between plastic nanoparticles and brain tissue, which is the likely mechanism behind the observed behavioural disorders in the top consumer. In a broader perspective, our findings demonstrate that plastic nanoparticles are transferred up through a food chain, enter the brain of the top consumer and affect its behaviour, thereby severely disrupting the function of natural ecosystems.
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Scientific RepoRts | 7: 11452 | DOI:10.1038/s41598-017-10813-0
Brain damage and behavioural
disorders in sh induced by plastic
nanoparticles delivered through the
food chain
Karin Mattsson1,2, Elyse V. Johnson3, Anders Malmendal1, Sara Linse
1,2, Lars-Anders
Hansson2,4 & Tommy Cedervall1,2
The tremendous increases in production of plastic materials has led to an accumulation of plastic
pollution worldwide. Many studies have addressed the physical eects of large-sized plastics on
organisms, whereas few have focused on plastic nanoparticles, despite their distinct chemical, physical
and mechanical properties. Hence our understanding of their eects on ecosystem function, behaviour
and metabolism of organisms remains elusive. Here we demonstrate that plastic nanoparticles reduce
survival of aquatic zooplankton and penetrate the blood-to-brain barrier in sh and cause behavioural
disorders. Hence, for the rst time, we uncover direct interactions between plastic nanoparticles
and brain tissue, which is the likely mechanism behind the observed behavioural disorders in the top
consumer. In a broader perspective, our ndings demonstrate that plastic nanoparticles are transferred
up through a food chain, enter the brain of the top consumer and aect its behaviour, thereby severely
disrupting the function of natural ecosystems.
e production of plastic material has increased tremendously during the last decades1, 2, and about 10% of the
plastics produced annually end up in the oceans3 through sewage treatment plants, waste handling or aerial
deposition4, constituting 60–80% of the total marine debris5. Plastic debris has been shown to aect over 660
marine species6 through entanglement and ingestion, and is thus a severe and potent pollutant in aquatic envi-
ronments6. Once in the aquatic environment, plastic material breaks up into smaller pieces through the action
of sunlight, waves, living organisms in the water and by the water itself 7, 8. Eventually plastic material is broken
down to nanoparticles912, which may be an even more potent threat since plastic nanoparticles are able to pass
through biological barriers13, penetrate tissues14 and accumulate in organs15 and aect behaviour and metabolism
of organisms16, 17. ose eects are generally not due to the toxicity of the material per se, but rather a result of the
physical features of the nanoparticles. Hence, the particle size plays a pivotal role for their biological impact18 as
size aects the curvature and provide a large surface area18, 19. Smaller particles are generally more toxic than the
corresponding bulk material at the same mass concentration2022, and the mobility, biological fate and bioavaila-
bility depend on size, shape, charge and other nanoparticle properties23, 24.
e freshwater invertebrate Daphnia magna can ingest nano- and micro-sized (20 nm to 70 μm) particles from
water21, 2527, are commonly used in toxicity studies28 and has a pivotal role in many food chains16, 17, 29. Previous
work has shown that the uptake rate depends on particles size25, 30, 31 and charge32. For example, Daphnia magna
were shown to have a lower uptake rate of 20 nm than 1000 nm polystyrene particles, although when compared at
equivalent surface area, the uptake was equal or higher for the small particles. Furthermore, indirect intake rate
via algal food was higher than direct intake from water30. e precise manner in which particle size, charge and
surface area aect the intake and biological impact of nanoparticles is, however, still unknown.
Here we report novel ndings on how plastic nanoparticles strongly aect an aquatic food chain from the
zooplankter Daphnia magna to the top consumer, the freshwater sh, Crucian carp (Carassius carassius), which is
common in anthropogenically aected waters. We exposed Daphnia magna to a range of polymeric nanoparticles
1Department of Biochemistry and Structural Biology, Lund University, P.O. Box 124, SE-221 00, Lund, Sweden.
2NanoLund, Lund University, SE-221 00, Lund, Sweden. 3CytoViva, Inc. 570 Devall Drive, Auburn, AL, 36832, USA.
4Department of Biology/Aquatic Ecology, Lund University, SE-223 62, Lund, Sweden. Correspondence and requests
for materials should be addressed to K.M. (email:
Received: 9 June 2017
Accepted: 14 August 2017
Published: xx xx xxxx
Scientific RepoRts | 7: 11452 | DOI:10.1038/s41598-017-10813-0
directly in water or via algae (Scendesmus sp.). We show that positively charged amino modied polystyrene nan-
oparticles aect both Daphnia and top consumer (sh) in a size dependent manner. We also show that the nano-
particles were transferred through a three-level food chain from algae through zooplankton to sh, which showed
behavioural disorders. Moreover, those behavioural disorders depended on the size of the nanoparticles and anal-
yses by hyperspectral microscopy showed that the plastic nanoparticles were present in the sh brains. Hence, we
here, for the rst time, demonstrate the mechanistic chain from uptake of nanoplastic particles by algae, through
transport up the food chain and, nally, eects on the brain physiology and behaviour of top consumer (sh). On
a broader scale such eects are likely to considerably aect natural ecosystems, since top predators have a crucial
impact on lower trophic levels and ecosystem functions33.
Eects on Daphnia magna. Out of the tested nanoparticles of dierent size and charge (Table1) only ami-
no-modied positively charged polystyrene nanoparticles with a diameter of 52 nm aected Daphnia, whereas
larger particles of the same material (120–330 nm) had no eect on the animals and we therefore focused on this
particle size in our study. For the 52 nm particles, the toxicity was strongly dependent on the particle concentra-
tion. Up to a concentration of 0.025 g/L all Daphnia were still alive aer 24 hours, and above 0.075 g/L all were
dead within 13 h (Fig.1). Comparing the toxicity of dierently sized amino-modied polystyrene nanoparticles,
where either the mass, surface area or the number of particles were the same, revealed that size was the only
important factor for toxicity. To rule out a potential batch dependent toxicity of 52 nm amino modied polysty-
rene particles, the same type of particles but in a size of 53 nm, 57 nm as well as 58 nm were also tested.
Equipped with the insight obtained from Daphnia magna, we next set out to study the eects of plastic nan-
oparticles (53 nm and 180 nm) on the entire food chain and if the eects of nanoparticles are transferred to the
sh in a food chain starting from algae (Fig.2). Analyses of sh feeding times – the time it took for each group
(aquarium) to consume 50% of the provided Daphnia – showed that the sh receiving 53 nm particles ate more
slowly than the control sh. Contrary to our expectations, sh fed with 180 nm particles were the fastest feeders
(Fig.3A; p < 0.030 ANOVA). Furthermore, detailed analyses of the hunting behaviour showed that the sh fed
with 53 nm particles swam a longer distance to eat 50% of the provided zooplankton (Fig.3B, p < 0.02 ANOVA),
and explored less space within each aquarium (Figs3C and S1). ey also had a signicantly lower activity(px/s),
in contrast to the sh fed 180 nm particles which instead showed a higher activity than controls (Fig.3D; p < 0.001
ANOVA). In natural systems, a slower feeding rate combined with a longer swimming distance before successful
feeding likely leads to suboptimal energy use and a more pronounced exposure to predation. Collectively, these
consequences point to considerable eects on tness and thereby on ecosystem function for sh exposed to plas-
tic nanoparticles of about 50 nm size.
Particle Diameter size (nm) Concentration (g/L) Surface charge
PS-NH2 Amino-modied 52, 53, 57, 58, 120, 180, 330 0.005, 0.010, 0.025, 0.050, 0.075, 0.10, 0.15 positive
PS-COOH Carboxylate 261, 601, 92, 160, 190, 220 0.025, 0.050, 0.075, 0.10, 0.2, 0.4 negative
PS-OSO3H Sulfonated 25, 200 0.025, 0.050, 0.075, 0.10, 0.2, 0.4 negative
PMMA 68, 140 0.025, 0.050, 0.075, 0.10, 0.2, 0.4 negative
Table 1. Characteristics for particles tested for toxicity to Daphnia magna, including particle name, diameter
and concentration. Only the PS-NH2 amino modied particles was found to be toxic to Daphnia and we
therefore performed our sh study using this type of particles. 1Two dierent batches were tested.
Figure 1. Mortality aer exposure. Number of alive Daphnia magna 0–24 h aer exposure to dierent
concentrations (0.025–0.150 g/L) of 52 nm amino modied polystyrene nanoparticles. e gure shows that
Daphnia mortality rates depend on the concentration of the particles, n = 10 for each concentration.
Scientific RepoRts | 7: 11452 | DOI:10.1038/s41598-017-10813-0
Figure 2. Food chain. Food chain from algae-zooplankton-sh, nanoparticles (53 nm mass (dark blue), 53 nm
surface area (light blue) and 180 nm (red)).
Figure 3. Top consumer response. Fish exposed to amino modied polystyrene nanoparticles of dierent
sizes delivered through a food chain of algae and zooplankton behave dierently. (A) sh feeding time (s) aer
exposure to dierent sizes of nanoparticles, p < 0.030 with ANOVA post hoc between 53 nm and 180 nm, (B)
swimming distance during feeding time, p < 0.018 with ANOVA post hoc between 53 nm and 180 nm, (C) sh
exploration within each aquarium during the rst 120 s of the feeding time (D) mean activity (px/s) during the
rst 120 s of the feeding time, p < 0.001 with ANOVA post hoc between all groups. Data are shown for control
sh that were not exposed to nanoparticles (n = 6 aquaria), 180 nm nanoparticles (n = 6 aquaria), and 53 nm
particles with the same mass concentration as 180 nm particles (n = 5 aquaria). All graphs show the mean ± SE.
e graphs shown that sh fed with 53 nm nanoparticles had a longer feeding time, that they swam a longer
distance to feed and that they explored less area of the aquaria, as well as that they had a lower activity compared
to sh receiving 180 nm particles and control sh. Moreover, sh fed with 180 nm particles were the fastest
feeders and had the highest activity.
Scientific RepoRts | 7: 11452 | DOI:10.1038/s41598-017-10813-0
e observed behavioural changes in the sh suggest that their brains were aected by the particles. To conrm
this, we explored the unique spectral features of tracking polystyrene using a CytoViva Hyperspectral Imaging
System to obtain a wavelength spectrum from each pixel of the light scattered from sh brains. Polystyrene was
detected in the brains from all analysed sh fed with polystyrene nanoparticles, whereas no polystyrene was
detected in brains from control sh (Figs4B and S2).
We also found that sh exposed to nanoparticles had a higher weight loss and less water in their brains than
control sh (Fig.S3). Moreover, the microscope imaging shows that the gyri in the cerebral lobes were larger in
sh exposed to 53 nm particles (Fig.4A; p < 0.025 ANOVA), suggesting also morphological eects from those
particles. Hence, the morphological changes in the brains of the 53-nm-sized nanoparticle-fed sh suggest that
sh brains were directly aected by the plastic nanoparticles and that the eects depended on particle size. Taken
together, our results suggest that some of the plastic nanoparticles fed to sh through a food web end up in thesh
brains. Furthermore, we unravel a mechanistic link between behavioural disorders and the incorporation of nan-
oparticles in brain tissue.
e amount of plastics in the world’s water bodies is rapidly increasing and this material degrades in size over
time and will eventually break down into plastic nanoparticles. Due to their small size, they easily enter the basis
of natural food chains, although it is unclear how these particles aect aquatic ecosystems. We show here that
52 nm positively charged amino modied polystyrene nanoparticles are toxic to Daphnia and that sh feeding
on Daphnia containing plastic nanoparticles change their behaviour in terms of activity, feeding time and the
distance they need to swim to consume their provided food. Furthermore, the behavioural changes depend on
the size of the particles. However, sh receiving 180 nm particles were dierently aected as they were the fastest
feeders and had the highest activity. In nature, the particles likely become aggregated with biological or inorganic
material, but we here show that the nano-size eect remains aer passing through the Daphnia digestive system.
For example, Ward et al.34 exposed the blue mussel Mytilus edulis and the oyster Crassostrea virginica to poly-
styrene nanoparticles, aggregated nanoparticles and micro-particles and found a higher ingestion rate for the
aggregated nanoparticles34. Wegner et al.35 exposed the mussel Mytilus edulis to polystyrene nanoparticles both
as nano-sized particles and as aggregated polystyrene nanoparticles. ey found a reduced ltering rate and an
increased production of pseudofeces35. In this context, our results point to an acute need for a deeper understand-
ing of the size-dependent toxicity eects of nanoparticles when released into nature. How these particles aect
organisms higher up in the food web, such as sh, as well as how they aect birds and mammals are unclear. In
Figure 4. Brain eects. (A) Measured gyri size (px2) of sh brains aer exposure to dierent sizes of polystyrene
nanoparticles, including 53 nm (n = 7), 180 nm (n = 11) and control (n = 15) which was not exposed to any
nanoparticles. (B) number of detected pixels corresponding to polystyrene particles in homogenized sh brain
samples, (n = 3 for each treatment). Fish receiving 53 nm nanoparticles had signicantly larger gyri size than
the 180 nm group, p < 0.021 ANOVA with Turkey’s post hoc. Polystyrene was detected in all brains from sh
exposed to nanoparticles, whereas no polystyrene was detected in the control group.
Scientific RepoRts | 7: 11452 | DOI:10.1038/s41598-017-10813-0
2015, the estimated amount of plastics being released into the ocean was between 4.8 and 12.7 million tons, with a
steady increase the coming years36. Eventually this plastic will degrade in size and reach the nanometer size range.
Here we demonstrate how plastic nanoparticles are transported up the food chain and are detected in brain
tissue of the sh top consumer whereas no polystyrene were detected in the control group. Moreover, we also
here report macroscopic changes in the brain structure and water content in sh that have received plastic nano-
particles. By using hyperspectral microscopy, we were able to detect polystyrene particles in sh brain tissue and
thereby we have, for the rst time, demonstrated that the plastics nanoparticles can be transported across the
blood-brain barrier in sh. Moreover, this result suggests a mechanistic link between the observed behavioural
changes and the presence of plastic nanoparticles in the brain tissue. In the present study, we observed changes in
the brain which may have been caused by specic interactions between the plastics and the brain tissue, although
we cannot rule out that other organs may also be aected. Our study lasted for two months, but during the rst
half of the experiment we observed no changes in behaviour of the nanoparticle fed sh, suggesting that sh are
aected by the particles that are accumulated in the sh. In nature, the Daphnia and sh are likely exposed to low
concentrations of plastic nanoparticles during their whole life-time, which allows accumulation processes to act
for a much longer time period than in our study, since sh, such as crucian carp, may live for more than 10 years37.
However, our results also imply that eects on biota from plastic nanoplastics are dependent on both concentra-
tion and size of the particles, which opens up for manufacturers to adjust production of nanoparticles to sizes that
are less hazardous to organism metabolism and thereby ecosystem function.
e main conclusion from our study is that plastic nanoparticles are transferred through three tropic levels,
suggesting that they are likely to be transferred even further up the food web to ultimately reach humans, the
top-level consumer. Hence, in a broader perspective, our results may have implications for human wellbeing,
although such consequences of the accelerating disposal rate of plastics is yet not well recognized or understood.
Materials and Methods
Testing for suitable particles with Daphnia magna. Polystyrene particles with dierent surface mod-
ications, charges, sizes (25 nm to 330 nm) and at a range of concentrations (0.005 g/L to 0.150 g/L) were tested
for toxicity towards Daphnia magna. On day 1, 100 ml algae or water were added to each bottle together with
particles with dierent concentrations except for the control bottle, which received only water (Table1). e
bottles were then shaken for 2 minutes, and the algae were allowed to ingest the particles for 24 hours. On day 2,
900 ml water was added to the algae/water together with 10 adult Daphnia magna with an approximate size of 3
mm. e number of dead Daphnia was counted every hour for 24 hours and the bottles were then gently stirred
to distribute the algae or water evenly. To rule out a potential batch dependent toxicity of 52 nm amino modied
polystyrene particles, the same type of particles but in a size of 53 nm, 57 nm as well as 58 nm were also tested.
Nanoparticle preparation and characterization. Positively charged amino-modified polystyrene
(PAO2N) particles with diameters of 52 nm, 53 nm, 57 nm, 58 nm, 120 nm, 180 nm and 330 nm were purchased
from Bang laboratories (Fisher, IN, USA). e particles were dialyzed with fresh tap water for 24 hours. e
particle size was measured, to ensure that particles size remained constant during the experiment, with Dynamic
Light Scattering (DLS) both before and aer dialysis, as well as one week aer the dialysis. No change in particle
size was recorded during the study. We chose to not use surface labelled particles since this may aect the surface
chemistry. Moreover, it is unknown how passage through the digestive systems of Daphnia and especially sh
might aect the labelling and vice versa.
Fish experiment. Two sizes of particles were chosen for the sh experiment, one with the size of 53 nm that was
shown to aect the Daphnia and one larger, 180 nm, that did not show any toxicity towards Daphnia. e particle
size were conrmed with DLS and measured 56 nm (PI: 27%) and 174 nm (PI: 18%) in the water used during the
experiment. Twenty-four aquaria with three sh in each were divided into four groups. e rst group (the 180 nm
group) received 180 nm particles at a concentration of 0.1 g/L. e second group received the same mass concentra-
tion (0.1 g/L) of 53 nm particles (the 53 nm mass group). e third group also received 53 nm particles, but at a lower
concentration corresponding to the same surface area as the group receiving 180 nm particles (the 53 nm surface
area group, concentration 0.029 g/L). e results from this treatment are, for clarity, presented in Supplementary
material, (TableS1). e fourth group, the control group, did not receive any nanoparticles. All sh were measured
and weight before the experiment started. e study was performed under the permission from the Malmö/Lund
Ethical committee (D nr 14 13–12) and was performed according to the current laws in Sweden.
Food chain. Algae (Scenedesmus sp.) with a diameter of approximately 25 μm were cultivated in aquaria. On
day 1, 500 mL algae with a concentration of 450
were mixed with water and particles to a total volume of 1 L
in four dierent test bottles (except for the control bottle, which received only water). Aer 24 hours Daphnia
magna (20 Daphnia/sh) were added to the algae medium. Aer 2 hours, the Daphnia were collected on a net
with a mesh size of 150
and washed two times with 150 mL water. Each sh (Crucian carp, Carassius carassius)
was then served 20 Daphnia, i.e. 60 Daphnia per aquarium.
We replicated this natural food chain such that the sh eventually ingested, via algae and Daphnia, the same
type of amino-modied polystyrene nanoparticles as used for the Daphnia toxicity with diameters of 53 nm and
180 nm (Fig.2). To distinguish between size and mass eects, two concentrations of the 53 nm particles were
used, one that corresponded to the same surface area and one that corresponded to the same mass as the 180 nm
particles. e three groups: 180 nm, 53 nm surface area (TableS1) and 53 nm mass were studied together with the
control group, that did not receive any nanoparticles. Sixty Daphnia individuals were introduced as food to each
sh aquarium every third day for a period of 67 days.
Scientific RepoRts | 7: 11452 | DOI:10.1038/s41598-017-10813-0
Video analysis. On day 62, we monitored the hunting behaviour of the sh by video recording each aquar-
ium separately during 2 minutes before the sh received food and 10 minutes aer. Since the smaller particles
were toxic to the Daphnia and thereby possibly aected their interaction with the sh, all groups of sh were on
the 62nd day fed with Daphnia that had not received any nanoparticles. Each sh position was registered each sec-
ond during the whole tracking period using the soware ImageJ. e feeding time – the time it took for the sh to
consume 50% of the provided food (Daphnia) – was registered. An ANOVA post hoc was used to test dierences
between treatments.
Brain analysis. On day 64, all sh were collected and anaesthetized using benzocaine. ey were measured
and weighed before the neck was cut and the brain was extracted. All samples were stored at 80 °C. e brain
was weighed and an image was recorded with Olympus SZX7 microscope with an Innity 1 camera and then
freeze-dried and weighed again before it was homogenized in PBS buer. e area of two gyri in all brain images
was measured in pixels2 using ImageJ and further calculated with Matlab. Finally, three brains from each group
were analysed with CytoViva hyperspectral microscope. is microscope was equipped with an enhanced dark-
eld illuminator and visible-near infrared (400–1000 nm) hyperspectral imaging components. e homogenized
brain samples were imaged under 60x magnication. Each image captured one pixel line at a time using an
automated stage. ese pixel lines were compiled to form hyperspectral images, also known as datacubes, which
contain spatial and spectral data for each pixel. For each image of exposed brain that was acquired, a spectral
library corresponding to polystyrene was created. is was accomplished by gathering several regions of interest
from each exposed brain image and ltering the spectra associated with those regions against 3 negative control
images (homogenized brain with no polystyrene). Any spectra that matched spectra in the negative controls were
eliminated from the polystyrene spectral libraries. en, the polystyrene was spectrally mapped and identied in
the exposed brain images using the Spectral Angle Mapper algorithm.
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We would like to thank the Centre for Environmental and Climate Research (CEC), the NanoLund at Lund
University, e Swedish Research Council and Mistra.
Author Contributions
K.M., A.M., S.L., L.A.H. and T.C. designed the study. K.M. and E.V.J. performed the experiments. K.M., E.V.J. and
T.C. preformed the analysis, and all authors wrote the manuscript.
Additional Information
Supplementary information accompanies this paper at doi:10.1038/s41598-017-10813-0
Competing Interests: e authors declare that they have no competing interests.
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... Later, Volkheimer showed that 5-110 µm particles of polyvinyl chloride appeared in the blood 2 h after ingestion by dogs, and smaller particles were translocated more rapidly than larger ones (Volkheimer, 1975). In fish, the translocation of plastic particles across the intestine has been implied by the discovery of particles in various tissues of several species (Collard et al., 2017;Elizalde-Velázquez et al., 2020;Jovanović et al., 2018;Kim et al., 2020;Mattsson et al., 2017;Zeytin et al., 2020), although the evidence is not indisputable. For example, the presence of two microplastic (MP) particles of 39 and 90 μm was reported in the livers of wild anchovy, Engraulis encrasicolus (Collard et al., 2017), but the precise location of the particles in or near blood cells could not be accurately inferred due to inadequate tissue preparation. ...
... Experimental studies have also suggested transfer of 200-600 μm MPs from the digestive tract to the liver in grey mullet, Mugil cephalus, and the presence of polystyrene (PS) particles (ranging from 214 to 288 μm) was observed in the livers of a few individual seabream, Sparus aurata, 45 days after their addition to the diet (Jovanović et al., 2018). In Crucian carp, Carassius carassius, fed food containing 53 nm and 180 nm polystyrene nanoplastic (PS-NPs) particles, the particles were detected in the brain by hyperspectral imaging (Mattsson et al., 2017). Similarly, in fathead minnow, Pimephales promelas, 60-70 nm PS-NPs were found in liver and kidney 48 h after their addition to the diet (Elizalde-Velázquez et al., 2020). ...
Plastic pollution in marine ecosystems constitutes an important threat to marine life. For vertebrates, macro/microplastics can obstruct and/or transit into the airways and digestive tract whereas nanoplastics (NPs; < 1000 nm) have been observed in non-digestive tissues such as the liver and brain. Whether NPs cross the intestinal epithelium to gain access to the blood and internal organs remains controversial, however. Here, we show directly NP translocation across the intestinal barrier of a fish, the European seabass, Dicentrarchus labrax, ex vivo. The luminal side of median and distal segments of intestine were exposed to fluorescent polystyrene NPs (PS-NPs) of 50 nm diameter. PS-NPs that translocated to the serosal side were then detected quantitatively by fluorimetry, and qualitatively by scanning electron microscopy (SEM) and pyrolysis coupled to gas chromatography and high-resolution mass spectrometry (Py-GC-HRMS). Fluorescence intensity on the serosal side increased 15–90 min after PS-NP addition into the luminal side, suggesting that PS-NPs crossed the intestinal barrier; this was confirmed by both SEM and Py-GC-HRMS. This study thus evidenced conclusively that NPs beads translocate across the intestinal epithelium in this marine vertebrate.
... 27 It has previously been shown that 50 nm PS-NH 2 are toxic to D. magna after 24 h exposure, whereas 200 nm PS-NH 2 , and 60 nm and 200 nm PS-COOH are not acutely toxic. 28 More recently the importance of the surface charge on PS nanoparticles 29 and the size of PS-NH 2 (ref. 30) has been studied in detail. ...
... These proteins are mainly related to the epithelium and intracellular structures and processes. Even though the latter nanoparticles are shown to be non-toxic after acute exposure, 28 there were a couple of proteins (serine protease and vitellogenin-1), that bound to all nanoparticles, which partly could explain the toxicity of 62 nm PS-COOH after a long-term exposure. 47 We found that neither 53 nm nor 200 nm PS-NH 2 bound to lipids, this could be due to the electrostatic repulsion between the positively charged parts of the lipid head and the positive group on the particle surface. ...
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Toxic and non-toxic polystyrene particles bind different proteins during filtration by zooplankton.
... This can increase their reactivity with inorganic and organic compounds, including biomolecules and other contaminants [34,35] ( Figure 1). It also enhances NP ingestion and absorption by aquatic organisms, as evidenced by the buildup of NPs in various organs, such as the gills, brain, heart, liver, yolk sac, gonads, and digestive organs of vertebrate and invertebrate species [36][37][38], especially at size values below 100 nm (Figure 1). The presence of NPs in several tissues of exposed organisms suggests that their small size increases the likelihood of tissue translocation and systemic distribution, with endocytosis, transcytosis, paracellular diffusion between tight junctions, and uptake through enterocyte barriers all playing important roles [39,40] ( Figure 1). ...
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Plastic production began in the early 1900s and it has transformed our way of life. Despite the many advantages of plastics, a massive amount of plastic waste is generated each year, threatening the environment and human health. Because of their pervasiveness and potential for health consequences, small plastic residues produced by the breakdown of larger particles have recently received considerable attention. Plastic particles at the nanometer scale (nanoplastics) are more easily absorbed, ingested, or inhaled and translocated to other tissues and organs than larger particles. Nanoplastics can also be transferred through the food web and between generations, have an influence on cellular function and physiology, and increase infections and disease susceptibility. This review will focus on current research on the toxicity of nanoplastics to aquatic species, taking into account their interactive effects with complex environmental mixtures and multiple stressors. It intends to summarize the cellular and molecular effects of nanoplastics on aquatic species; discuss the carrier effect of nanoplastics in the presence of single or complex environmental pollutants, pathogens, and weathering/aging processes; and include environmental stressors, such as temperature, salinity, pH, organic matter, and food availability, as factors influencing nanoplastic toxicity. Microplastics studies were also included in the discussion when the data with NPs were limited. Finally, this review will address knowledge gaps and critical questions in plastics’ ecotoxicity to contribute to future research in the field.
... Plastic litter of micro and nano-size may impact the base of the ocean food chain, causing potential damage to the entire trophic chain, including humans [11][12][13][14]. Although the effects of plastics in aquatic systems are widely documented [15,16], the potential impact of NPs on humans is still unclear [17] as, to date, there are still limited studies assessing the Nanomaterials 2022, 12,1947 2 of 21 potential toxic effects and biological interactions of NPs in mammalian systems and mainly performed using polystyrene nanobeads [18][19][20][21][22][23]. ...
Sub-micrometer particles derived from the fragmentation of plastics in the environment can enter the food chain and reach humans, posing significant health risks. To date, there is a lack of adequate toxicological assessment of the effects of nanoplastics (NPs) in mammalian systems, particularly in humans. In this work, we evaluated the potential toxic effects of three different NPs in vitro: two NPs obtained by laser ablation (polycarbonate (PC) and polyethylene terephthalate (PET1)) and one (PET2) produced by nanoprecipitation. The physicochemical characterization of the NPs showed a smaller size, a larger size distribution, and a higher degree of surface oxidation for the particles produced by laser ablation. Toxicological evaluation performed on human cell line models (HePG2 and Caco-2) showed a higher toxic effect for the particles synthesized by laser ablation, with PC more toxic than PET. Interestingly, on differentiated Caco-2 cells, a conventional intestinal barrier model, none of the NPs produced toxic effects. This work wants to contribute to increase knowledge on the potential risks posed by NPs.
... Revealing the toxicological consequences of plastic pollution on fish especially their early life-history stage is of great significance. Previous studies showed that microplastics and nanoplastics can impair the locomotion, energy reserves or other physiological functions of fish adults (Mattsson et al., 2015;Mattsson et al., 2017;Yin et al., 2018;Yin et al., 2019). In the present study, our results indicated that the 75nm PS-NPs impacted energy metabolism and several important molecular pathways, including protein digestion, response to damaged protein, gonad development and muscle contraction, in the larval channel catfish Ietalurus punetaus. ...
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Microplastics (nanoplastics) pollution has been a major ecological issue threatening global aquatic ecosystems. However, knowledge of the adverse effects of nanoplastics and the effects on freshwater ecosystems is still limited. To understand the impacts of nanoplastics on freshwater ecosystems, it is essential to reveal the physiological changes caused by nanoplastics in freshwater organisms, especially at their early life-history stages. In the present study, the larval channel catfish Ietalurus punetaus were exposed to gradient concentrations (0, 5, 10, 25, and 50 mg/L) of 75-nm polystyrene nanoplastics (PS-NPs) for 24 h or 48 h, and changes in contents of energy metabolites, metabolic enzyme activities and transcriptome were assessed. The results showed that glucose and triglyceride contents increased after 24 h of exposure to 10 or 25 mg/L of PS-NPs but decreased with increased concentrations or prolonged exposure duration. Activities of most metabolic enzymes analyzed decreased in the larvae after 48 h of exposure, especially in 25 or 50 mg/L of PS-NPs. These suggested that PS-NPs caused huge energy consumption and disturbed the energy metabolism in larval fish. Transcriptomic analysis showed that 48 h of exposure to 50 mg/L PS-NPs affected the expression of genes involved in protein digestion and induced response of proteasomes or heat shock proteins in the larval I. punetaus. The genes involved in peroxisome proliferator-activated receptors (PPAR) pathway and biosynthesis of amino acids were activated after the exposure. PS-NPs also depressed the expression of the genes involved in gonad development or muscle contraction in the larval I. punetaus. Overall, acute exposure to 75-nm PS-NPs disrupted the energy metabolism by consuming the energy reserves, and affected a series of molecular pathways which may further affect the development and survival of fish. This study provided the information about adverse effects of nanoplastics on the fish larvae and revealed the molecular pathways for the potential adverse outcomes.
Our oceans and seas have been polluted with plastics for nearly 60 years. The increase in plastic consumption all over the world, the possibility of plastics remaining in the environment for hundreds of years without decomposing, the decomposition of plastics into smaller pieces, the detection of organisms at all levels of the marine food chain, and the possibility of human exposure to microplastics through food increase the awareness on this issue day by day. With the introduction of microplastics and nanoplastics, scientists have started to work on this pollution in water, especially since 2010. The common view is that the impact of this type of pollution on the environment will increase and harm living things.
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Microplastic pollution is one of the leading global conservation issues. Microplastics generate various physical and toxicological effects on wildlife, which have received significant scientific and conservation-based attention. However, most research to date has been conducted in aquatic environments, while terrestrial ecosystems are still underrepresented. Moreover, there is a need for model species with the help of which we could compare the scale of microplastic pollution in different landscapes on a continental scale. In this study we investigated the potential of thrushes, a group of songbirds with an exceptionally terrestrial lifestyle and wide distribution range, to be used as a proxy of microplastic contamination in terrestrial ecosystems. Hence, we analysed the gastrointestinal tracts of Common Blackbirds Turdus merula and Song Thrushes Turdus philomelos in search of microplastics (over 100 μm in size) and assessed whether their contamination differed regarding the age of the birds and the time of year. We used birds that had died as a result of collision with anthropogenic infrastructure, which were sampled during wildlife monitoring of anthropogenic infrastructures and citizen science projects. We found that all the analysed individuals contained microplastics in their gastrointestinal tracts, mostly consisting of transparent fibers below 1mm in size. However, we found no seasonal or age-related differences in microplastic ingestion in either species. Slight discrepancies in microplastic ingestion observed between the studied species may be related to differences in their foraging strategies and habitat utilization. While our results show an ubiquity of microplastics in terrestrial environments, they also indicate that thrushes may be used as indicators of microplastic pollution in terrestrial ecosystems.
With the widespread presence of plastic wastes, knowledge about the potential environmental risks and bioavailability of micro- or nanoplastics fragmented from large analogs is of utmost importance. As the particle size matters in mediating endocytic mechanism and particle internalization, we first studied the effects of polystyrene microparticles (PS-MPs, 1 μm) and polystyrene nanoparticles (PS-NPs, 100 nm) of two different sizes at varying concentrations of 5, 25 and 75 μg/mL on the mouse hippocampal neuronal HT22 cells. The in vitro study showed efficient cellular uptake of PS-MPs and PS-NPs of both sizes. The adverse effects of cellular metabolic activity as reflective of excess Reactive Oxygen Species (ROS) and cell cycle S phase arresting were observed especially at the greater concentration of smaller-sized PS particles, consequently leading to mild cytotoxicity. We further evaluated the dynamic particle-cell interaction with a continuous supply of PS particles using a microfluidic device. By recapitulating the in vivo mechanical microenvironments while allowing homogeneous distribution of PS particles, the dynamic exposure to PS particles of both sizes under flowing conditions resulted in much lesser viability of neural cells than the traditional static exposure. As the flowing dynamics may avoid the gravitational settling of particles and allow more efficient cellular uptake, the size distribution, together with the exposure configurations, contributed significantly to the determination of the PS particle cytotoxicity. The on-chip investigation and a better understanding of particle translocation mechanisms would offer very much to the risk assessment of PS particles on human health.
Plastics are a group of synthetic materials made of organic polymers and some additives with special characteristics. Plastics have become part of our daily life due to their many applications and uses. However, inappropriately managed plastic waste has raised concern regarding their ecotoxicological and human health risks in the long term. Due to the non-biodegradable nature of plastics, their waste may take several thousands of years to partially degrade in natural environments. Plastic fragments/particles can be very minute in size and are mistaken easily for prey or food by aquatic organisms (e.g., invertebrates, fishes). The surface properties of plastic particles, including large surface area, functional groups, surface topography, point zero charge, influence the sorption of various contaminants, including heavy metals, oil spills, PAHs, PCBs and DDT. Despite the fact that the number of studies on the biological effects of plastic particles on biota and humans has been increasing in recent years, studies on mixtures of plastics and other chemical contaminants in the aquatic environment are still limited. This review aims to gather information about the main characteristics of plastic particles that allow different types of contaminants to adsorb on their surfaces, the consequences of this adsorption, and the interactions of plastic particles with aquatic biota. Additionally, some missing links and potential solutions are presented to boost more research on this topic and achieve a holistic view on the effects of micro- and nanoplastics to biological systems in aquatic environments. It is urgent to implement measures to deal with plastic pollution that include improving waste management, monitoring key plastic particles, their hotspots, and developing their assessment techniques, using alternative products, determining concentrations of micro- and nanoplastics and the contaminants in freshwater and marine food-species consumed by humans, applying clean-up and remediation strategies, and biodegradation strategies.
The distribution, accumulation, and transfer of micro(nano)plastics (MnPs) in animals has been a key task of ecological risk research. Understanding the translocation of MnPs may help extrapolate toxicity and health risks to organisms. Here, we briefly summarize the distribution of MnPs in the tissues and organs of aquatic and terrestrial animals. Then, we review and discuss three transfer modes of MnPs: translocation in tissues and organs, transfer by intergeneration, and movement through the food web. Additionally, factors affecting translocation are discussed. Together, these phenomena present urgent and important challenges for future research.
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Nanowires (NWs) have unique electrical and optical properties of value for many applications including lighting, sensing, and energy harnessing. Consumer products containing NWs increase the risk of NWs being released in the environment, especially into aquatic ecosystems through sewage systems. Daphnia magna is a common, cosmopolitan freshwater organism sensitive to toxicity tests and represents a likely entry point for nanoparticles into food webs of aquatic ecosystems. Here we have evaluated the effect of NW diameter on the gut penetrance of NWs in Daphnia magna. The animals were exposed to NWs of two diameters (40 and 80 nm) and similar length (3.6 and 3.8 µm, respectively) suspended in water. In order to locate the NWs in Daphnia, the NWs were designed to comprise one inherently fluorescent segment of gallium indium phosphide (GaInP) flanked by a gallium phosphide (GaP) segment. Daphnia mortality was assessed directly after 24 hours of exposure, 24 h after exposure and 7 days after exposure. Translocation of NWs across the intestinal epithelium was investigated using confocal fluorescence microscopy directly after 24 h of exposure and was observed in 89% of Daphnia exposed to 40 nm NWs and in 11% of Daphnia exposed to 80 nm NWs. A high degree of fragmentation was observed for NWs of both diameters after ingestion by the Daphnia, although 40 nm NWs were fragmented to a greater extent, which could possibly facilitate translocation across the intestinal epithelium. Our results show that the feeding behavior of animals may enhance the ability of NWs to penetrate biological barriers and that penetrance is governed by the NW diameter.
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The release of plastics into the environment has been identified as an important issue for some time. Recent publications have suggested that the degradation of plastic materials will result in the release of nano-sized plastic particles to the environment. Nanoparticle tracking analysis was applied to characterise the formation of nanoplastics during the degradation of a polystyrene (PS) disposable coffee cup lid. The results clearly show an increase in the formation of nanoplastics over time. After 56 days' exposure the concentration of nanoplastics in the PS sample was 1.26 × 108 particles/ml (average particles size 224 nm) compared to 0.41 × 108 particles/ml in the control.
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The use of nanoparticles in consumer products, like cosmetic, sunscreens and electrical devices has increased tremendously the last decade, despite insufficient knowledge about their effects on human health and ecosystem function. Moreover, the amount of plastic waste products entering natural ecosystems, such as oceans and lakes, is increasing and degradation of the disposed plastics produces smaller particles towards the nano scale. Therefore it is of outmost importance to gain knowledge about how plastic nanoparticles enter and affect living organisms. Here we have administered polystyrene nanoparticles to fish through an aquatic food chain and studied the effects on behavior and metabolism. We find severe effects on feeding and shoaling behavior as well as metabolism in the fish. Hence, we conclude that polystyrene nanoparticles have severe effects on both behavior and on metabolism in fish and that commonly used nano-sized particles may have considerable effects on natural systems and on ecosystem services derived from them.
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The amount of nano- and microplastic in the aquatic environment rises due to the industrial production of plastic and the degradation of plastic into smaller particles. Concerns have been raised about their incorporation into food webs. Little is known about the fate and effects of nanoplastic, especially for the freshwater environment. In this study, effects of nano polystyrene (Nano-PS) on the growth and photosynthesis of the green alga Scenedesmus obliquus and the growth, mortality, neonate production and malformations of the zooplankter Daphnia magna were assessed. Nano-PS reduced population growth and reduced chlorophyll concentrations in the algae. Exposed Daphnia showed a reduced body size and severe alterations in reproduction. Numbers and body size of neonates were lower, while the number of neonate malformations among neonates rose to 68% of the individuals. These effects of Nano-PS were observed between 0.22 and 103 mg Nano-PS/L. Malformations occurred from 30 mg Nano-PS/L onwards. Such plastic concentrations are much higher than presently reported for marine as well as freshwater, but may eventually occur in sediment pore waters. As far as we know, these results are the first to show that direct life history shifts in algae and Daphnia populations may occur as a result of exposure to nanoplastic.
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Limnologistas han venido estudiando patrones de la productividad primaria en los lagos por mas de 60 anos. En lo relativo a la entroficacion a puesto la atencion en el suministro de nutrientes como un regulador de la productividad del lago. Sin embargo la adicion de nutrientes no siempre explica la variacion de la productividad primaria de todos los lagos del mundo. Este articulo se refiere a algunos casos practicos
In this work, we present for the first time undeniable evidence of nano-plastic occurrence due to solar light degradation of marine micro-plastics under controlled and environmentally representative conditions. As observed during our recent expedition (Expedition 7th Continent), plastic pollution will be one of the most challenging ecological threats for the next generation. Up to now, all studies have focused on the environmental and the economic impact of millimeter scale plastics. These plastics can be visualized, collected and studied. We are not aware of any studies reporting the possibilities of nano-plastics in marine water. Here, we developed for the first time a new solar reactor equipped with a nanoparticle detector to investigate the possibility of the formation of nano-plastics from millimeter scale plastics. With this system, correlated with electronic microscopy observations, we identified for the first time the presence of plastics at the nano-scale in water due to UV degradation. Based on our observations large fractal nano-plastic particles (i.e., >100 nm) are produced by UV light after the initial formation of the smallest nano-plastic particles (i.e., <100 nm). These unprecedented results show the new and unprecedented potential hazards of plastic waste at the nanoscale, which had not been taken into account previously.
Plastic debris in the marine environment is widely documented, but the quantity of plastic entering the ocean from waste generated on land is unknown. By linking worldwide data on solid waste, population density, and economic status, we estimated the mass of land-based plastic waste entering the ocean. We calculate that 275 million metric tons (MT) of plastic waste was generated in 192 coastal countries in 2010, with 4.8 to 12.7 million MT entering the ocean. Population size and the quality of waste management systems largely determine which countries contribute the greatest mass of uncaptured waste available to become plastic marine debris. Without waste management infrastructure improvements, the cumulative quantity of plastic waste available to enter the ocean from land is predicted to increase by an order of magnitude by 2025. Copyright © 2015, American Association for the Advancement of Science.
Nanoplastic debris, resulted from run-off and weathering breakdown of macro and microplastics, represent an emerging concern for marine ecosystems. The aim of the present study was to investigate disposition and toxicity of polystyrene nanoparticles (PS NPs) in early development of sea urchin embryos Paracentrotus lividus. NPs with two different surface charges where chosen, carboxylated (PS-COOH) and amine (PS-NH2) polystyrene, the latter being a less common variant, known to induce cell death in several in vitro cell systems. NPs stability in natural seawater (NSW) was measured while disposition and embryotoxicity were monitored within 48 h post-fertilization (hpf). Modulation of genes involved in cellular stress response (cas8, 14-3-3ε, p-38 MAPK, Abcb1, Abcc5) was investigated. PS-COOH form micro-aggregates (PDI > 0.4) in NSW, while PS-NH2 result better dispersed (89 nm ± 2 nm) initially, though they also aggregated partially with time. Their respectively anionic and cationic nature was confirmed by zeta potential measurements. No embryotoxicity was observed for PS-COOH up to 50 µg mL-1 while PS-NH2 caused severe developmental defects (EC50 3.85 µg mL-1 -24 hpf and EC50 2.61 µg mL-1 -48 hpf). PS-COOH accumulated inside embryo's digestive trait while PS-NH2 were more dispersed. Abcb1 gene resulted up-regulated at 48 hpf by PS-COOH while PS-NH2 induced cas8 gene at 24 hpf, suggesting an apoptotic pathway. In line with the results obtained with the same PS NPs in several human cell lines, also in sea urchin embryos, differences in surface charges and aggregation in sea water strongly affect their embryotoxicity.
The toxic effects of two differently sized ZnO nanopowders have been studied in Daphnia magna using advanced microscopy techniques. Five nanoZnO suspensions (0.1, 0.33, 1, 3.3 and 10 mg/L) were tested. The results of the 48-h acute toxicity tests performed with ZnO < 100 nm (bZnO) and ZnO < 50 nm (sZnO) showed slight effects, with EC50 values of 3.1 and 1.9 mg/L for bZnO and sZnO, respectively. Specimens exposed to 1 and 3.3 mg/L have been microscopically analysed and nanoparticles (NPs) from both concentrations have been found into midgut cells: i) in the microvilli; ii) in endocytic vesicles near the upper cell surface; iii) in some endosomes, as well as in mitochondria, in multivesicular and multilamellar bodies; iv) into the enterocytes' nuclei; v) free in the cytoplasm; vi) in the paracellular space between adjacent cells; vii) into the folded basal plasma membrane, and viii) in the gut muscolaris, suggesting that not only both nanoZnOs are able to interact with the plasmatic membrane of D. magna enterocytes, but also that they are capable to cross epithelial barriers. The ultrastructural changes increased with increasing concentrations and the worst morphological fields came from samples exposed to 3.3 mg/L of both nanoZnOs. Morphological effects were qualitatively similar between the two nanomaterials, but they appear to be much more frequent for sZnO NPs. Data from ICP-OES analyses demonstrated that the maximum Zn(++) concentration in our tested suspensions was 0.137 mg/L, which is well below the reported NOEC for the soluble Zinc. The corresponding Zn-salt exposures (0.1 mg/L Zn(++)) gave 0% of immobilized daphnids for both NPs suggesting that in our test medium nanoZnO toxicity is not driven by their solubilized ions. The large presence of NPs inside midgut cells after only 48-h exposure to nanoZnOs and their effects on the intestinal cells highlighted the toxic potential of these nanomaterials, also suggesting that studies on chronic effects are needed.
Abstract The developmental toxicity of nanostructured materials, as well as their impact on the biological barriers, represents a crucial aspect to be assessed in a nanosafety policy framework. Nanosized metal oxides have been demonstrated to affect Xenopus laevis embryonic development, with nZnO specifically targeting the digestive system. To study the mechanisms of the nZnOinduced intestinal lesions, we tested two different nominally sized ZnO nanoparticles (NPs) at effective concentrations. Advanced microscopy techniques and molecular markers analyses were applied in order to describe the NP-epithelial cell interactions and the mechanisms driving NP toxicity and translocation through the intestinal barrier. We attributed the toxicity to NP-induced cell oxidative damage, the small sized NPs being the more effective. This outcome is sustained by a marked increase in anti-oxidant genes' expression and high lipid peroxidation level in the enterocytes, where disarrangement of the cytoskeleton and cell junctions' integrity were evidenced. These events led to diffuse necrotic changes in the intestinal barrier, and trans- and paracellular NP permeation through the mucosa. The uptake routes, leading NPs to cross the intestinal barrier and reach secondary target tissues, have been documented. nZnOs embryotoxicity was confirmed to be crucially mediated by the NPs' reactivity rather than their dissolved ions. The ZnO NPs ability to overwhelm the intestinal barrier must be taken into high consideration for a future design of safer ZnO NPs.