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Particles in the oceans: Implication for a safe marine environment
Marine Environmental Research 111 (2015) 1e4
Contents lists available at ScienceDirect
Marine Environmental Research journal homepage: www.elsevier.com/locate/marenvrev
Editorial
Particles in the oceans: Implication for a safe marine environment a b s t r a c t Strategies and technologies for the ecosafety assessment and design of engineered particles entering the marine environment are urgently needed. As the application of nanoparticles in science and technology grows, the need to understand their impact on the marine environment becomes increasingly important. This Editorial introduces a Special Issue on the topic of a sustainable and safety use of nanoparticles for protecting, recovering and supporting the oceans’ environment and consequently human health. The issue focus on the impact of micro/nano-plastics and metallic nanoparticles on marine organisms, as well as some methodological aspects associated to the eco/toxicity and analytical approaches for in deep physico-chemical characterization of nanoparticles in marine waters and sediment media. Important and urgent topics are addressed in the field of nano-ecosafety in order to assess more precisely both exposure routes and environmental hazards of nanoparticles in the ocean. Ecotoxicological and toxicological data, obtained using a wide variety of organisms representative of different trophic levels and biological organization, from whole animals to macromolecules, will be useful for a better definition of cleaner and safer nanoparticles. Efforts in developing a broad understanding of target species, expected results, benchmarks and timelines, will be of primary importance.
1. Scope The scientific literature on the occurrence, fate, behaviour and toxicity of particles in the marine environment has increased exponentially in the last years. However, the production and release into the marine environment of new nano products is growing at high rate and represent a major challenge for the scientific community. Thus, for example, graphene (GFN) represents a new carbonaceous nanomaterial, rapidly developed, but information about its environmental behaviour and toxicity on marine organisms is limited (Hu et al., 2015) and it can be considered in an early stage (Zhao et al., 2014). Additionally, difficulties to develop standardized protocols have been pointed out because ecotoxicity tests for dissolved substances (e.g. EPA and OECD) could not be adequate for nanoparticles (NPs) where toxicity is not only related to the mass or particulate number concentration but also to the aggregation degree and resulting particle size distribution (Kühnel and Nickel, 2014; Parker et al., 2014). Handy et al. (2012) summarized specific issues for toxicity test of nanoparticles. The accent on the need to investigate the hazards posed by NPs on marine environments, looking at model species and using molecular approaches has been introduced recently (Matranga and Corsi, 2012). This also introduced concepts related to the behaviour of a given NP in sea water, thus affecting its characteristics, bioavailability and toxicity (Corsi et al., 2014). The collection of papers of this Special Issue (SI) is to stimulate a comprehensive discussion on the effects of nanoparticles and http://dx.doi.org/10.1016/j.marenvres.2015.10.001 0141-1136/© 2015 Published by Elsevier Ltd.
micro- and nano-plastics in the marine environment and enlarge the marine ecotoxicology community working on such emerging issues at both European and International levels (USA, Japan, China, others). Ecotoxicological and toxicological data provided by the several papers included in the present Special Issue might be useful to achieve a certain level of knowledge on the interactions of nanoparticles in the sea water media which will be useful to categorize or classify safer and/or cleaner NPs for the environment, and consequently for human health. As far as scientific contributions on micro-plastics, they address important challenges, such as the need for: i) validation and harmonisation of analytical methods, ii) identification and quantification of micro-plastics in wildlife and iii) more detailed ecotoxicological studies on their potential impact on marine organisms. Most of the articles included herewith relate to the effects of several metallic NPs on various marine organisms, belonging to different trophic levels, targeting different organs, tissues, cells and macromolecules. Although probably not fully comprehensive, this SI is an attempt to fill important gaps in assessing the environmental hazards posed by NPs in the marine environment, and to stress the need for further investigations on the risks posed by both micro- and nano-plastics. 2. Outline A
number
of
manuscripts
contributed
by
renowned
2
Editorial / Marine Environmental Research 111 (2015) 1e4
international experts in the field were received in response to the invitation of Julian Blasco. In addition, we included a selected collection of manuscripts consequent to oral presentations given at the 2nd Marine NanoEcosafety Workshop, which was held at the Area della Ricerca CNR in Palermo, Italy, in November 2014 (http://bimat2014.azuleon.org/welcome_w.php), co-organized by Ilaria Corsi and Valeria Matranga. In the above, participated invited experts from Italy, Spain, Portugal, France, UK, Belgium, Netherlands, USA, Australia and Japan. The presentations given by acknowledged experts were related to NPs interaction, fate, behaviour and toxicity in the marine environment, as well as the impact of micro- and nano-plastics. After an independent peer-reviewing process, eighteen papers were accepted for publication in this SI that contains relevant contributions on the following topics: i) micro- and nano-plastics (1e4), ii) silver nanoparticles (5e7), iii) copper oxide nanoparticles (8e12), iv) titanium dioxide nanoparticles (13e16), and v) methodological aspects related to toxicity and analytical approaches (17e18). The first paper is a review by Van Cauwenberghe et al. (pp. 5e17) titled “Micro-plastics in sediments: a review of techniques, occurrence and effects” that well introduces the topic of microplastics and their fate into the marine environment. The review summarizes the data presented in more than 100 articles, appeared over the last 50 years, with the aim of evaluating the current microplastic extraction techniques, discuss their occurrence and worldwide distribution in sediments and assess their potential adverse effects on marine organisms. The main issues outlined are the urgent need for standardized protocols to be used for the extraction and detection of micro-plastics in environmental samples including biota, unified reporting units and a more realistic assessment of noxious effects. In the paper by Avio et al. (pp. 18e26) titled: “Experimental development of a new protocol for extraction and characterization of micro-plastics in fish tissues: first observation in commercial species from Adriatic Sea” the authors provide a new protocol allowing good extraction yields of micro-plastics from fish tissues (recovery from 78% to 98%, depending on the polymer size). Authors compare the efficacy of some existing approaches and document the presence and distribution of micro-plastics in various fish species collected along the Adriatic Sea (North-Eastern Mediterranean Sea). The paper by Gago et al. (pp. 27e33) titled: “First observation on neustonic plastics in waters off NW Spain (spring 2013 and 2014)” summarizes the presence and distribution of plastic particles in sea waters off the NW Spanish Atlantic coast, as resulting from a field sampling program run in 2013. Plastic particles of various types were found in 95% of the investigated sites, both as micro- and meso-particles. The authors stressed the urgent need for the harmonization of protocols for the determination (size, type and quantity) of plastic particles to ensure coherence in data implementation, at both European and International levels. In the paper by Canesi et al. (pp. 34e40) titled: “Evidence for immunomodulation and apoptotic processes induced by cationic polystyrene nanoparticles in the hemocytes of the marine bivalve Mytilus”, are reported the first data on cytotoxicity of nano-sized (40 nm) plastics in mussel's hemocytes, using amino-polystyrene nanoparticles as a model. The authors reported that the mechanism of action is similar to the one observed in mammalian cells and suggest that in marine invertebrates the immune system can represent a significant target for polystyrene NPs, which are also particularly abundant among plastic debris. Such findings stress again the need for further research on specific mechanisms of toxicity and cellular uptake of both micro and nano-sized plastics in marine organisms for risk assessment purposes.
The effects on marine organisms caused by Ag-NPs is introduced by the paper titled “Effect of silver nanoparticles on marine organisms belonging to different trophic levels” by Gambardella et al. (pp. 41e49), that investigates the potential toxicity on species belonging to four ecologically-relevant phyla, such as algae, cnidarians, crustaceans and echinoderms. Authors measured algal growth, ephyra jellyfish immobilization and frequency of pulsations, crustacean's mortality and swimming behavior and sea urchin sperm motility. They found that although differences in sensitivity among different organisms were observed, an effect of Ag-NPs towards all the selected species was found, suggesting that Ag-NPs affect differently organisms belonging to various trophic levels within the marine ecosystem. A scale of species sensitivity to Ag-NPs was proposed based on results obtained. The paper by Buric et al., (pp. 50e59) titled “Effect of silver nanoparticles on Mediterranean sea urchin embryonal development is species specific and depends on moment of first exposure” focuses on the species-specific effects of low concentrations (from 100 down to 1 mg/L 1) of Ag-NPs on embryonic development. Authors found that the three major sea urchin species present in the Mediterranean sea, namely Arbacia lixula, Paracentrotus lividus and Sphaerechinus granularis, differ in their sensitivity to Ag-NPs. The most sensitive species is A. lixula whose embryos show an impaired development at the lowest Ag-NPs concentrations (1e10 mg/L 1) tested. It follows S. granularis, with an effective Ag-NP concentration range of 10e50 mg/L 1, and last P. lividus (50e100 mg/L 1). This suggests that given its sensitivity, A. lixula may be the appropriate choice for NPs toxicity testing. The stage at which embryos were exposed to Ag-NPs was also important. Authors tested the effects on developmental impairments at four stages: eggs, 4 cells, blastula and gastrula, demonstrating that the earlier embryos were exposed, the harsher were the effects. Therefore, it is also important for future applications of the sea urchin embryo development test for NPs toxicity to specify the stage at which embryos are exposed. A comprehensive review article titled “Toxicity of silver and gold nanoparticles on marine microalgae” by Moreno-Garrido et al., (pp. 60e73) is centered on the fate and toxicity of silver, gold, and goldsilver alloy NPs on microalgae. They emphasize that the current literature describes a wide variation of results on toxicity, mainly due to the different methodologies used in the bioassays involving microalgae. The toxicity EC50 values can span in the case of Ag-NPs from 0.005 to 83.3 mM, representing five orders of magnitude. Authors point out that there is an urgent need for the development of standardised bioassays before the comparison of toxicity results can be made. This is particularly important in view of establishing predictive models on a wide database of toxicity responses and physico-chemical characterisation of nanoparticles. The review article by Lopes Rocha et al. (pp. 74e88) titled: “Ecotoxicological impact of engineered nanomaterials in bivalve molluscs: an overview” summarizes the information about NPs behaviour and fate in the aquatic environment, and their ecotoxicological impact on marine and freshwater bivalves. The paper reinforces the notion that bivalves are key the model species and digestive glands the target tissue. On the other hand, standard protocols for NPs ecotoxicological testing need to be developed. In the article by Thit et al. (pp. 89e98) titled “Influence of copper oxide nanoparticle shape on bioaccumulation, cellular internalization and effects in the estuarine sediment-dwelling polychaete, Nereis diversicolor” the worms were exposed to sediment spiked with CuO-NPs with different sizes and shapes, or aquaeous Cu in a range of concentrations (7e140 mg Cu/g d.w.). Worms accumulated Cu-NPs in a concentration-dependent manner, although TEM images were not conclusive about the internalization as particles in the worm tissues. Cu-NPs forms and concentrations seem to be the main drivers of their bioaccumulation and adverse effects.
Editorial / Marine Environmental Research 111 (2015) 1e4
In the paper by Dai et al., (pp. 99e106) titled “Influence of copper oxide nanoparticle form and shape on toxicity and bioaccumulation in the deposit feeder, Capitella teleta” the CuO-NPs characteristics in terms of particle form and shape are considered in relation to their toxicity and bioaccumulation. The exposure route was spiked sediment and the target species, the deposit feeder C. teleta. All forms and shapes of CuO-NPs were toxic to the species, but the effects were evident during the depuration period only. Particle shape affected the bioaccumulation process and current risk assessment approaches should be refined to be sufficiently safe for the organism and the environment. Ruiz et al., (pp. 107e120) in their paper titled “Short-term effects on antioxidant enzymes and long-term genotoxic and carcinogenic potential of CuO nanoparticles compared to bulk CuO and ionic Cu in mussels Mytilus galloprovincialis” examine the shorteterm and long-term effects of CuO-NPs in comparison with bulk CuO and ionic Cu, looking also at the genotoxic and carcinogenic potential in the long-term exposures. Also in this case, accumulation was dependent on the metal form (bulk or ionic) and NPs were found preferentially in lysosomes of the digestive cells. CuO-NPs induced the activity of antioxidant enzymes (SOD and catalase) and also induced genotoxic effects, as determined by micronuclei frequency, histological analysis and mitotic division, although they dissapeared during the post-exposure period. The manuscript by Maisano et al. (pp. 121e127) titled “Developmental abnormalities and neurotoxicological effects of CuO-NPs on the black sea urchin A. lixula by embryotoxicity assay”analyses the embryotoxicity of CuO-NPs by measuring the abnormalities occurring early in development, as well as neurotoxic effects measured by AchE activity. CuO-NPs with a nominal size of 12 nm were found to aggregate in artificial seawater (80e200 nm, observed by Field Emission Scanning Electron Microscopy-FESEM) and embryos were able to internalise them. The CuO-NPs impact on development and larval morphology was recorded. Alteration in neurotransmission pathway was related to observed abnormalities. As a result, A. lixula is proposed as a valid model species for ecotoxicological assays and biomonitoring programmes. The review by Canesi et al., (pp. 128e134) titled: “Interactive effects of nanoparticles with other contaminants in aquatic organisms: friend or foe?” summarizes toxicity data obtained on carbon based NPs and n-TiO2, as examples of widespread NPs in aquatic organisms. In addition, complex and unexpected interactive effects of n-TiO2 combined with common marine pollutants, such as dioxin and cadmium, are discussed based on the available data on the model marine bivalve Mytilus and in comparison with freshwater species. Overall, the authors conclude that interactive effects of NPs with other contaminants do not necessarily lead to increased toxicity or harmful effects in aquatic organisms. Johnson et al. (pp. 135e143) in their paper titled “Cellular responses of eastern oysters, Crassostrea virginica, to titanium dioxide nanoparticles” examine the potential toxicity of TiO2-NPs to oysters; their combination with natural sunlight was also considered. Bio-reactivity and toxicity was reported in hepatopancreas exposed to levels corresponding to those predicted in the environment. Brief low-intensity light exposures may potentiate the TiO2-NPs toxicity, but it was not related to ROS production, although lysosomal function was impaired. In vitro studies show the usefulness as screening tool for toxicity in oysters and the lysosomal destabilization as biomarker of nanoparticle bio-reactivity. In the paper by Rocco et al. (pp. 144e148) titled: “Genomic and chromosomal damage in the marine mussel M. galloprovincialis: effects of the combined exposure to titanium dioxide nanoparticles and cadmium” the interactive effects in terms of genotoxicity are reported in the gills and digestive glands of a marine mussel species. Exposure to n-TiO2 was responsible for a significant increase
3
of micro-nucleated cell frequency in gill tissue, while no chromosomal damage was observed after CdCl2 exposure or after the combined exposure to both substances (n-TiO2 and CdCl2). The authors confirmed that the interaction does not necessarily lead to increased harmful effects, but can be still tissue- and speciesspecific and strongly depends on the type of NP and the chemical compounds tested. In the Morelli et al., paper (pp. 149e157) titled “The response of Phaeodactylum tricornutum to quantum dot exposure: acclimation and changes in protein expression” authors examine the effect of exposure to CdSe/ZnS quantum dots (QDs) in the diatom marine model, P. tricornutum, using different physiological, biochemical and molecular approaches. Time-dependent physico-chemical transformations of QDs are reported. Transcriptional expression of selected stress responsive genes (hsp101, cat and gsr2) showed their up-regulation in the QD-exposed algae. A comparison of the proteomes of exposed and un-exposed cells highlighted a large number of differentially expressed proteins. Molecular pathways involved in cellular response to NPs remains still an open question in marine species. Last, but not least, two papers on methodological aspects associated to toxicity and analytical approaches close this SI. In particular, the paper by Hu et al., (pp. 158e161) titled “Neutral red retention time assay in determination of toxicity of nanoparticles” describes an improvement of the neutral red retention time (NRRT) assay, which detects the lysosomal membrane stability, applied to mussels hemocytes. The new assay is revisited to be performed in microtitre plates and tested on a panel of metal and metal oxide NPs (2 ppm), namely: copper, chromium, cobalt, gold and titanium. Author showed that copper, chromium, cobalt NPs are toxic by this assay, while gold and titanium NPs are not. nio et al., (pp. 162e169) titled “Assessing silThe paper by Anto ver nanoparticles behaviour in artificial seawater by mean of AF4 and spICP-MS” provides an experimental methodology for the analysis of the fate and behaviour of Ag-NPs in sea water. By combining different characterization techniques, i.e. asymmetric flow field-flow fractionation, single particle ICP-MS, UVeVis, authors determined in detail the agglomeration process of Ag-NPs in artificial seawater. In particular, they showed that Ag-NPs form agglomerates quite quickly in marine waters, while they are practically stable in de-ionized water. The presence of dissolved organic matter stabilizes Ag-NPs in sea water and might change the interaction with living organisms. 3. Concluding remarks The production of new NPs has increased in the last years, although the knowledge about their occurrence, fate, behaviour and toxicity in the marine environments is rather limited. Thus, it appears clear that more work is still needed in order to fully assess the impact of particles (nano and micro-sized) into the marine environment and more efforts should be spent to standardize protocols for testing NPs in exposure media. The detailed characterizations of NPs before and during toxicity assays, jointly innovative analytical techniques for measuring their levels in environmental media as sediment and sea water and overall good bench practices are essential in order to assure a comparison of results obtained. Such challenges are in agreement with European Commission work programme 2016e2017 and included in the upcoming calls for Nanotechnologies, Advanced Materials, Biotechnology and Production which will be launched in mid October 2015 (https://ec.europa.eu/ programmes/horizon2020/en/draft-work-programmes-2016-17). Validation and harmonisation of methods, identification and quantification of micro-plastics and more detailed ecotoxicological studies on their potential impact on marine organisms are three
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Editorial / Marine Environmental Research 111 (2015) 1e4
main themes listed as scientific priorities of the last JPI Ocean call on microplastics from which 4 projects (7.4 milion euro budget) have been funded all around Europe and will start from December 2015 (http://www.jpi-oceans.eu/news-events/news/results-%E2% 82%AC75-million-call-microplastics-published). All authors of papers appearing in this SI emphasize that there is an urgent need for the development of standardised procedures and consistent protocols in order to reach a consensus in the Scientific Community working on the nanosafety of particles and give the possibility to authorities to compare results coming from different laboratories in the world. The safeguard of the oceans health is nowadays one of the key issues that the EU Marine Board is giving attention to, in view of the interconnections between oceans and human health (Depledge et al., 2013, Moore et al., 2013; Fleming et al., 2014). Acknowledgements The Guest Editors (JB, IC and VM) would like to thank the Scientific Committee and all the participants in the 2nd Marine NanoEcosafety Workshop (MANET-2014), held under the aegis of the Nano Safety Cluster (http://www.nanosafetycluster.eu/), who contributed with their insightful thoughts and stimulating discussions to the success of the meeting, putting the basis for the materialization of this special issue (for a comprehensive review, see also the MANET 2014 book of Abstracts at: http://bimat2014.azuleon. org/welcome_w.php). The MANET-2014 would not have been possible without the financial support of the National Research Council of Italy, the University of Siena. The Azuleon Meetings organising secretariat, as well as the local and technical support of Mr M. Biondo and A. Pensato are also sincerely acknowledged. Guest Editors express their warm appreciation to the Editors of Marine Environmental Research and Elsevier for supporting this initiative and to the anonymous reviewers who by their appropriate and bright comments and suggestions contributed in increasing the quality of papers contained in this issue. We are also grateful to the project CTM2012-38720-C03-03, funded by the Spanish Ministry of Economy and Competitiveness to JB; the Italian Ministry of Research PRIN2009FHHP2W to IC; the CNR Flagship Project POM-FBdQ 2013-2014 to VM.
Marcomini, A., Sabbioni, E., Matranga, V., 2014. Common strategies and technologies for the ecosafety assessment and design of nanomaterials entering the marine environment. ACS Nano 8, 9694e9709. Depledge, M.H., Harvey, A., Brownlee, C., Frost, M., Moore, M.N., Fleming, L.E., 2013. Changing views of the interconnections between oceans and human health in Europe. Microbiol. Ecol. 65, 852e859. Fleming, L.E., McDonough, N., Austen, M., Mee, L., Moore, M., Hess, P., Depledge, M.H., White, M., Philippart, K., Bradbrook, P., Smalley, A., 2014. Oceans and Human Health: A rising tide of challenges and opportunities for Europe. Mar. Envir. Res. 99, 16e19. Handy, R.D., Cornelis, G., Fernandes, T., Tsyusko, O., Decho, A., Sabo-Attwood, T., Metcalfe, C., Steevens, J.A., Klaine, S.J., Koelmans, A.A., Horne, N., 2012. Ecotoxicity test methods for engineered nanomaterials: practical experiences and recommendations from the bench. Environ. Toxicol. Chem. 31 (1), 15e31. Hu, Xiangang, Ouyang, Shaohu, Mu, Li, An, Jing, Zhou, Qixing, 2015. Effects of graphene oxide and oxidized carbon nanotubes on the cellular division, microstructure, uptake, oxidative stress, and metabolic profiles. Env. Sci. Technol. 49, 10825e10833. https://ec.europa.eu/programmes/horizon2020/en/draftwork-programmes-2016-17. http://www.jpi-oceans.eu/news-events/news/ results-%E2%82%AC75-million-call-microplastics-published. Kühnel, D., Nickel, C., 2014. The OECD expert meeting on ecotoxicology and environmental fate d Towards the development of improved OECD guidelines for the testing of nanomaterials. Sci. Total Environ. 472, 347e353. Matranga, V., Corsi, I., 2012. Toxic effects of engineered nanoparticles in the marine environment: model organisms and molecular approaches. Mar. Environ. Res. 76, 32e40. Moore, M., Depledge, M.H., Fleming, L.E., Hess, P., Less, D., Leonard, P., Madsen, L., Owen, R., Pirlet, H., Seys, J., Vasconcelos, V., Viarengo, A., 2013. Oceans and human health (OHH): a European perspective from the marine board of the european science foundation (Marine Board-ESF). Microbiol. Ecol. 65, 889e900. http://dx.doi.org/10.1007/s00248-013-0204-5. Parker, S., Woodhall, J., Ma, G., Veinot, J.C., Cresser, M., Boxall, A.B.A., 2014. Regulatory eotoxicity testing of engineered nanoparticles: are the resuts relevant to the natural environment? Nanotoxicology 8 (5), 583e592. Zhao, Jian, Wang, Zhenyu, Jason, C., White, Xing, Baoshan, 2014. Graphene in the aquatic environment: adsorption, dispersion, toxicity and transformation. Environ. Sci. Technol. 48, 9995e10009.
Julian Blasco* Institute of Marine Sciences of Andalusia (ICMAN-CSIC), Campus Río San Pedro, 11510 Puerto Real, Cadiz, Spain Ilaria Corsi Department of Physical, Earth and Environmental Sciences, University of Siena, Siena 53100, Italy Valeria Matranga CNR-Institute of Biomedicine and Molecular Immunology “Alberto Monroy”, Palermo 90146, Italy *
References Corsi, I., Cherr, G.N., Lenihan, H.S., Labille, J., Hassellov, M., Canesi, L., Dondero, F., Frenzilli, G., Hristozov, D., Puntes, V., Della Torre, C., Pinsino, A., Libralato, G.,
Corresponding author.
Contents lists available at ScienceDirect
Marine Environmental Research journal homepage: www.elsevier.com/locate/marenvrev
Editorial
Particles in the oceans: Implication for a safe marine environment a b s t r a c t Strategies and technologies for the ecosafety assessment and design of engineered particles entering the marine environment are urgently needed. As the application of nanoparticles in science and technology grows, the need to understand their impact on the marine environment becomes increasingly important. This Editorial introduces a Special Issue on the topic of a sustainable and safety use of nanoparticles for protecting, recovering and supporting the oceans’ environment and consequently human health. The issue focus on the impact of micro/nano-plastics and metallic nanoparticles on marine organisms, as well as some methodological aspects associated to the eco/toxicity and analytical approaches for in deep physico-chemical characterization of nanoparticles in marine waters and sediment media. Important and urgent topics are addressed in the field of nano-ecosafety in order to assess more precisely both exposure routes and environmental hazards of nanoparticles in the ocean. Ecotoxicological and toxicological data, obtained using a wide variety of organisms representative of different trophic levels and biological organization, from whole animals to macromolecules, will be useful for a better definition of cleaner and safer nanoparticles. Efforts in developing a broad understanding of target species, expected results, benchmarks and timelines, will be of primary importance.
1. Scope The scientific literature on the occurrence, fate, behaviour and toxicity of particles in the marine environment has increased exponentially in the last years. However, the production and release into the marine environment of new nano products is growing at high rate and represent a major challenge for the scientific community. Thus, for example, graphene (GFN) represents a new carbonaceous nanomaterial, rapidly developed, but information about its environmental behaviour and toxicity on marine organisms is limited (Hu et al., 2015) and it can be considered in an early stage (Zhao et al., 2014). Additionally, difficulties to develop standardized protocols have been pointed out because ecotoxicity tests for dissolved substances (e.g. EPA and OECD) could not be adequate for nanoparticles (NPs) where toxicity is not only related to the mass or particulate number concentration but also to the aggregation degree and resulting particle size distribution (Kühnel and Nickel, 2014; Parker et al., 2014). Handy et al. (2012) summarized specific issues for toxicity test of nanoparticles. The accent on the need to investigate the hazards posed by NPs on marine environments, looking at model species and using molecular approaches has been introduced recently (Matranga and Corsi, 2012). This also introduced concepts related to the behaviour of a given NP in sea water, thus affecting its characteristics, bioavailability and toxicity (Corsi et al., 2014). The collection of papers of this Special Issue (SI) is to stimulate a comprehensive discussion on the effects of nanoparticles and http://dx.doi.org/10.1016/j.marenvres.2015.10.001 0141-1136/© 2015 Published by Elsevier Ltd.
micro- and nano-plastics in the marine environment and enlarge the marine ecotoxicology community working on such emerging issues at both European and International levels (USA, Japan, China, others). Ecotoxicological and toxicological data provided by the several papers included in the present Special Issue might be useful to achieve a certain level of knowledge on the interactions of nanoparticles in the sea water media which will be useful to categorize or classify safer and/or cleaner NPs for the environment, and consequently for human health. As far as scientific contributions on micro-plastics, they address important challenges, such as the need for: i) validation and harmonisation of analytical methods, ii) identification and quantification of micro-plastics in wildlife and iii) more detailed ecotoxicological studies on their potential impact on marine organisms. Most of the articles included herewith relate to the effects of several metallic NPs on various marine organisms, belonging to different trophic levels, targeting different organs, tissues, cells and macromolecules. Although probably not fully comprehensive, this SI is an attempt to fill important gaps in assessing the environmental hazards posed by NPs in the marine environment, and to stress the need for further investigations on the risks posed by both micro- and nano-plastics. 2. Outline A
number
of
manuscripts
contributed
by
renowned
2
Editorial / Marine Environmental Research 111 (2015) 1e4
international experts in the field were received in response to the invitation of Julian Blasco. In addition, we included a selected collection of manuscripts consequent to oral presentations given at the 2nd Marine NanoEcosafety Workshop, which was held at the Area della Ricerca CNR in Palermo, Italy, in November 2014 (http://bimat2014.azuleon.org/welcome_w.php), co-organized by Ilaria Corsi and Valeria Matranga. In the above, participated invited experts from Italy, Spain, Portugal, France, UK, Belgium, Netherlands, USA, Australia and Japan. The presentations given by acknowledged experts were related to NPs interaction, fate, behaviour and toxicity in the marine environment, as well as the impact of micro- and nano-plastics. After an independent peer-reviewing process, eighteen papers were accepted for publication in this SI that contains relevant contributions on the following topics: i) micro- and nano-plastics (1e4), ii) silver nanoparticles (5e7), iii) copper oxide nanoparticles (8e12), iv) titanium dioxide nanoparticles (13e16), and v) methodological aspects related to toxicity and analytical approaches (17e18). The first paper is a review by Van Cauwenberghe et al. (pp. 5e17) titled “Micro-plastics in sediments: a review of techniques, occurrence and effects” that well introduces the topic of microplastics and their fate into the marine environment. The review summarizes the data presented in more than 100 articles, appeared over the last 50 years, with the aim of evaluating the current microplastic extraction techniques, discuss their occurrence and worldwide distribution in sediments and assess their potential adverse effects on marine organisms. The main issues outlined are the urgent need for standardized protocols to be used for the extraction and detection of micro-plastics in environmental samples including biota, unified reporting units and a more realistic assessment of noxious effects. In the paper by Avio et al. (pp. 18e26) titled: “Experimental development of a new protocol for extraction and characterization of micro-plastics in fish tissues: first observation in commercial species from Adriatic Sea” the authors provide a new protocol allowing good extraction yields of micro-plastics from fish tissues (recovery from 78% to 98%, depending on the polymer size). Authors compare the efficacy of some existing approaches and document the presence and distribution of micro-plastics in various fish species collected along the Adriatic Sea (North-Eastern Mediterranean Sea). The paper by Gago et al. (pp. 27e33) titled: “First observation on neustonic plastics in waters off NW Spain (spring 2013 and 2014)” summarizes the presence and distribution of plastic particles in sea waters off the NW Spanish Atlantic coast, as resulting from a field sampling program run in 2013. Plastic particles of various types were found in 95% of the investigated sites, both as micro- and meso-particles. The authors stressed the urgent need for the harmonization of protocols for the determination (size, type and quantity) of plastic particles to ensure coherence in data implementation, at both European and International levels. In the paper by Canesi et al. (pp. 34e40) titled: “Evidence for immunomodulation and apoptotic processes induced by cationic polystyrene nanoparticles in the hemocytes of the marine bivalve Mytilus”, are reported the first data on cytotoxicity of nano-sized (40 nm) plastics in mussel's hemocytes, using amino-polystyrene nanoparticles as a model. The authors reported that the mechanism of action is similar to the one observed in mammalian cells and suggest that in marine invertebrates the immune system can represent a significant target for polystyrene NPs, which are also particularly abundant among plastic debris. Such findings stress again the need for further research on specific mechanisms of toxicity and cellular uptake of both micro and nano-sized plastics in marine organisms for risk assessment purposes.
The effects on marine organisms caused by Ag-NPs is introduced by the paper titled “Effect of silver nanoparticles on marine organisms belonging to different trophic levels” by Gambardella et al. (pp. 41e49), that investigates the potential toxicity on species belonging to four ecologically-relevant phyla, such as algae, cnidarians, crustaceans and echinoderms. Authors measured algal growth, ephyra jellyfish immobilization and frequency of pulsations, crustacean's mortality and swimming behavior and sea urchin sperm motility. They found that although differences in sensitivity among different organisms were observed, an effect of Ag-NPs towards all the selected species was found, suggesting that Ag-NPs affect differently organisms belonging to various trophic levels within the marine ecosystem. A scale of species sensitivity to Ag-NPs was proposed based on results obtained. The paper by Buric et al., (pp. 50e59) titled “Effect of silver nanoparticles on Mediterranean sea urchin embryonal development is species specific and depends on moment of first exposure” focuses on the species-specific effects of low concentrations (from 100 down to 1 mg/L 1) of Ag-NPs on embryonic development. Authors found that the three major sea urchin species present in the Mediterranean sea, namely Arbacia lixula, Paracentrotus lividus and Sphaerechinus granularis, differ in their sensitivity to Ag-NPs. The most sensitive species is A. lixula whose embryos show an impaired development at the lowest Ag-NPs concentrations (1e10 mg/L 1) tested. It follows S. granularis, with an effective Ag-NP concentration range of 10e50 mg/L 1, and last P. lividus (50e100 mg/L 1). This suggests that given its sensitivity, A. lixula may be the appropriate choice for NPs toxicity testing. The stage at which embryos were exposed to Ag-NPs was also important. Authors tested the effects on developmental impairments at four stages: eggs, 4 cells, blastula and gastrula, demonstrating that the earlier embryos were exposed, the harsher were the effects. Therefore, it is also important for future applications of the sea urchin embryo development test for NPs toxicity to specify the stage at which embryos are exposed. A comprehensive review article titled “Toxicity of silver and gold nanoparticles on marine microalgae” by Moreno-Garrido et al., (pp. 60e73) is centered on the fate and toxicity of silver, gold, and goldsilver alloy NPs on microalgae. They emphasize that the current literature describes a wide variation of results on toxicity, mainly due to the different methodologies used in the bioassays involving microalgae. The toxicity EC50 values can span in the case of Ag-NPs from 0.005 to 83.3 mM, representing five orders of magnitude. Authors point out that there is an urgent need for the development of standardised bioassays before the comparison of toxicity results can be made. This is particularly important in view of establishing predictive models on a wide database of toxicity responses and physico-chemical characterisation of nanoparticles. The review article by Lopes Rocha et al. (pp. 74e88) titled: “Ecotoxicological impact of engineered nanomaterials in bivalve molluscs: an overview” summarizes the information about NPs behaviour and fate in the aquatic environment, and their ecotoxicological impact on marine and freshwater bivalves. The paper reinforces the notion that bivalves are key the model species and digestive glands the target tissue. On the other hand, standard protocols for NPs ecotoxicological testing need to be developed. In the article by Thit et al. (pp. 89e98) titled “Influence of copper oxide nanoparticle shape on bioaccumulation, cellular internalization and effects in the estuarine sediment-dwelling polychaete, Nereis diversicolor” the worms were exposed to sediment spiked with CuO-NPs with different sizes and shapes, or aquaeous Cu in a range of concentrations (7e140 mg Cu/g d.w.). Worms accumulated Cu-NPs in a concentration-dependent manner, although TEM images were not conclusive about the internalization as particles in the worm tissues. Cu-NPs forms and concentrations seem to be the main drivers of their bioaccumulation and adverse effects.
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In the paper by Dai et al., (pp. 99e106) titled “Influence of copper oxide nanoparticle form and shape on toxicity and bioaccumulation in the deposit feeder, Capitella teleta” the CuO-NPs characteristics in terms of particle form and shape are considered in relation to their toxicity and bioaccumulation. The exposure route was spiked sediment and the target species, the deposit feeder C. teleta. All forms and shapes of CuO-NPs were toxic to the species, but the effects were evident during the depuration period only. Particle shape affected the bioaccumulation process and current risk assessment approaches should be refined to be sufficiently safe for the organism and the environment. Ruiz et al., (pp. 107e120) in their paper titled “Short-term effects on antioxidant enzymes and long-term genotoxic and carcinogenic potential of CuO nanoparticles compared to bulk CuO and ionic Cu in mussels Mytilus galloprovincialis” examine the shorteterm and long-term effects of CuO-NPs in comparison with bulk CuO and ionic Cu, looking also at the genotoxic and carcinogenic potential in the long-term exposures. Also in this case, accumulation was dependent on the metal form (bulk or ionic) and NPs were found preferentially in lysosomes of the digestive cells. CuO-NPs induced the activity of antioxidant enzymes (SOD and catalase) and also induced genotoxic effects, as determined by micronuclei frequency, histological analysis and mitotic division, although they dissapeared during the post-exposure period. The manuscript by Maisano et al. (pp. 121e127) titled “Developmental abnormalities and neurotoxicological effects of CuO-NPs on the black sea urchin A. lixula by embryotoxicity assay”analyses the embryotoxicity of CuO-NPs by measuring the abnormalities occurring early in development, as well as neurotoxic effects measured by AchE activity. CuO-NPs with a nominal size of 12 nm were found to aggregate in artificial seawater (80e200 nm, observed by Field Emission Scanning Electron Microscopy-FESEM) and embryos were able to internalise them. The CuO-NPs impact on development and larval morphology was recorded. Alteration in neurotransmission pathway was related to observed abnormalities. As a result, A. lixula is proposed as a valid model species for ecotoxicological assays and biomonitoring programmes. The review by Canesi et al., (pp. 128e134) titled: “Interactive effects of nanoparticles with other contaminants in aquatic organisms: friend or foe?” summarizes toxicity data obtained on carbon based NPs and n-TiO2, as examples of widespread NPs in aquatic organisms. In addition, complex and unexpected interactive effects of n-TiO2 combined with common marine pollutants, such as dioxin and cadmium, are discussed based on the available data on the model marine bivalve Mytilus and in comparison with freshwater species. Overall, the authors conclude that interactive effects of NPs with other contaminants do not necessarily lead to increased toxicity or harmful effects in aquatic organisms. Johnson et al. (pp. 135e143) in their paper titled “Cellular responses of eastern oysters, Crassostrea virginica, to titanium dioxide nanoparticles” examine the potential toxicity of TiO2-NPs to oysters; their combination with natural sunlight was also considered. Bio-reactivity and toxicity was reported in hepatopancreas exposed to levels corresponding to those predicted in the environment. Brief low-intensity light exposures may potentiate the TiO2-NPs toxicity, but it was not related to ROS production, although lysosomal function was impaired. In vitro studies show the usefulness as screening tool for toxicity in oysters and the lysosomal destabilization as biomarker of nanoparticle bio-reactivity. In the paper by Rocco et al. (pp. 144e148) titled: “Genomic and chromosomal damage in the marine mussel M. galloprovincialis: effects of the combined exposure to titanium dioxide nanoparticles and cadmium” the interactive effects in terms of genotoxicity are reported in the gills and digestive glands of a marine mussel species. Exposure to n-TiO2 was responsible for a significant increase
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of micro-nucleated cell frequency in gill tissue, while no chromosomal damage was observed after CdCl2 exposure or after the combined exposure to both substances (n-TiO2 and CdCl2). The authors confirmed that the interaction does not necessarily lead to increased harmful effects, but can be still tissue- and speciesspecific and strongly depends on the type of NP and the chemical compounds tested. In the Morelli et al., paper (pp. 149e157) titled “The response of Phaeodactylum tricornutum to quantum dot exposure: acclimation and changes in protein expression” authors examine the effect of exposure to CdSe/ZnS quantum dots (QDs) in the diatom marine model, P. tricornutum, using different physiological, biochemical and molecular approaches. Time-dependent physico-chemical transformations of QDs are reported. Transcriptional expression of selected stress responsive genes (hsp101, cat and gsr2) showed their up-regulation in the QD-exposed algae. A comparison of the proteomes of exposed and un-exposed cells highlighted a large number of differentially expressed proteins. Molecular pathways involved in cellular response to NPs remains still an open question in marine species. Last, but not least, two papers on methodological aspects associated to toxicity and analytical approaches close this SI. In particular, the paper by Hu et al., (pp. 158e161) titled “Neutral red retention time assay in determination of toxicity of nanoparticles” describes an improvement of the neutral red retention time (NRRT) assay, which detects the lysosomal membrane stability, applied to mussels hemocytes. The new assay is revisited to be performed in microtitre plates and tested on a panel of metal and metal oxide NPs (2 ppm), namely: copper, chromium, cobalt, gold and titanium. Author showed that copper, chromium, cobalt NPs are toxic by this assay, while gold and titanium NPs are not. nio et al., (pp. 162e169) titled “Assessing silThe paper by Anto ver nanoparticles behaviour in artificial seawater by mean of AF4 and spICP-MS” provides an experimental methodology for the analysis of the fate and behaviour of Ag-NPs in sea water. By combining different characterization techniques, i.e. asymmetric flow field-flow fractionation, single particle ICP-MS, UVeVis, authors determined in detail the agglomeration process of Ag-NPs in artificial seawater. In particular, they showed that Ag-NPs form agglomerates quite quickly in marine waters, while they are practically stable in de-ionized water. The presence of dissolved organic matter stabilizes Ag-NPs in sea water and might change the interaction with living organisms. 3. Concluding remarks The production of new NPs has increased in the last years, although the knowledge about their occurrence, fate, behaviour and toxicity in the marine environments is rather limited. Thus, it appears clear that more work is still needed in order to fully assess the impact of particles (nano and micro-sized) into the marine environment and more efforts should be spent to standardize protocols for testing NPs in exposure media. The detailed characterizations of NPs before and during toxicity assays, jointly innovative analytical techniques for measuring their levels in environmental media as sediment and sea water and overall good bench practices are essential in order to assure a comparison of results obtained. Such challenges are in agreement with European Commission work programme 2016e2017 and included in the upcoming calls for Nanotechnologies, Advanced Materials, Biotechnology and Production which will be launched in mid October 2015 (https://ec.europa.eu/ programmes/horizon2020/en/draft-work-programmes-2016-17). Validation and harmonisation of methods, identification and quantification of micro-plastics and more detailed ecotoxicological studies on their potential impact on marine organisms are three
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main themes listed as scientific priorities of the last JPI Ocean call on microplastics from which 4 projects (7.4 milion euro budget) have been funded all around Europe and will start from December 2015 (http://www.jpi-oceans.eu/news-events/news/results-%E2% 82%AC75-million-call-microplastics-published). All authors of papers appearing in this SI emphasize that there is an urgent need for the development of standardised procedures and consistent protocols in order to reach a consensus in the Scientific Community working on the nanosafety of particles and give the possibility to authorities to compare results coming from different laboratories in the world. The safeguard of the oceans health is nowadays one of the key issues that the EU Marine Board is giving attention to, in view of the interconnections between oceans and human health (Depledge et al., 2013, Moore et al., 2013; Fleming et al., 2014). Acknowledgements The Guest Editors (JB, IC and VM) would like to thank the Scientific Committee and all the participants in the 2nd Marine NanoEcosafety Workshop (MANET-2014), held under the aegis of the Nano Safety Cluster (http://www.nanosafetycluster.eu/), who contributed with their insightful thoughts and stimulating discussions to the success of the meeting, putting the basis for the materialization of this special issue (for a comprehensive review, see also the MANET 2014 book of Abstracts at: http://bimat2014.azuleon. org/welcome_w.php). The MANET-2014 would not have been possible without the financial support of the National Research Council of Italy, the University of Siena. The Azuleon Meetings organising secretariat, as well as the local and technical support of Mr M. Biondo and A. Pensato are also sincerely acknowledged. Guest Editors express their warm appreciation to the Editors of Marine Environmental Research and Elsevier for supporting this initiative and to the anonymous reviewers who by their appropriate and bright comments and suggestions contributed in increasing the quality of papers contained in this issue. We are also grateful to the project CTM2012-38720-C03-03, funded by the Spanish Ministry of Economy and Competitiveness to JB; the Italian Ministry of Research PRIN2009FHHP2W to IC; the CNR Flagship Project POM-FBdQ 2013-2014 to VM.
Marcomini, A., Sabbioni, E., Matranga, V., 2014. Common strategies and technologies for the ecosafety assessment and design of nanomaterials entering the marine environment. ACS Nano 8, 9694e9709. Depledge, M.H., Harvey, A., Brownlee, C., Frost, M., Moore, M.N., Fleming, L.E., 2013. Changing views of the interconnections between oceans and human health in Europe. Microbiol. Ecol. 65, 852e859. Fleming, L.E., McDonough, N., Austen, M., Mee, L., Moore, M., Hess, P., Depledge, M.H., White, M., Philippart, K., Bradbrook, P., Smalley, A., 2014. Oceans and Human Health: A rising tide of challenges and opportunities for Europe. Mar. Envir. Res. 99, 16e19. Handy, R.D., Cornelis, G., Fernandes, T., Tsyusko, O., Decho, A., Sabo-Attwood, T., Metcalfe, C., Steevens, J.A., Klaine, S.J., Koelmans, A.A., Horne, N., 2012. Ecotoxicity test methods for engineered nanomaterials: practical experiences and recommendations from the bench. Environ. Toxicol. Chem. 31 (1), 15e31. Hu, Xiangang, Ouyang, Shaohu, Mu, Li, An, Jing, Zhou, Qixing, 2015. Effects of graphene oxide and oxidized carbon nanotubes on the cellular division, microstructure, uptake, oxidative stress, and metabolic profiles. Env. Sci. Technol. 49, 10825e10833. https://ec.europa.eu/programmes/horizon2020/en/draftwork-programmes-2016-17. http://www.jpi-oceans.eu/news-events/news/ results-%E2%82%AC75-million-call-microplastics-published. Kühnel, D., Nickel, C., 2014. The OECD expert meeting on ecotoxicology and environmental fate d Towards the development of improved OECD guidelines for the testing of nanomaterials. Sci. Total Environ. 472, 347e353. Matranga, V., Corsi, I., 2012. Toxic effects of engineered nanoparticles in the marine environment: model organisms and molecular approaches. Mar. Environ. Res. 76, 32e40. Moore, M., Depledge, M.H., Fleming, L.E., Hess, P., Less, D., Leonard, P., Madsen, L., Owen, R., Pirlet, H., Seys, J., Vasconcelos, V., Viarengo, A., 2013. Oceans and human health (OHH): a European perspective from the marine board of the european science foundation (Marine Board-ESF). Microbiol. Ecol. 65, 889e900. http://dx.doi.org/10.1007/s00248-013-0204-5. Parker, S., Woodhall, J., Ma, G., Veinot, J.C., Cresser, M., Boxall, A.B.A., 2014. Regulatory eotoxicity testing of engineered nanoparticles: are the resuts relevant to the natural environment? Nanotoxicology 8 (5), 583e592. Zhao, Jian, Wang, Zhenyu, Jason, C., White, Xing, Baoshan, 2014. Graphene in the aquatic environment: adsorption, dispersion, toxicity and transformation. Environ. Sci. Technol. 48, 9995e10009.
Julian Blasco* Institute of Marine Sciences of Andalusia (ICMAN-CSIC), Campus Río San Pedro, 11510 Puerto Real, Cadiz, Spain Ilaria Corsi Department of Physical, Earth and Environmental Sciences, University of Siena, Siena 53100, Italy Valeria Matranga CNR-Institute of Biomedicine and Molecular Immunology “Alberto Monroy”, Palermo 90146, Italy *
References Corsi, I., Cherr, G.N., Lenihan, H.S., Labille, J., Hassellov, M., Canesi, L., Dondero, F., Frenzilli, G., Hristozov, D., Puntes, V., Della Torre, C., Pinsino, A., Libralato, G.,
Corresponding author.