Corbicula fluminea gene expression modulated by CeO2 nanomaterials and salinity

  • Vanessa Koehlé-DivoEmail author
  • Sandrine Pain-Devin
  • Carole Bertrand
  • Simon Devin
  • Catherine Mouneyrac
  • Laure Giambérini
  • Bénédicte Sohm
Research Article


Cerium dioxide nanomaterials (CeO2 NMs) are used in different fields and incorporated in daily products. Several studies highlighted their effects on organism physiology, although molecular studies remain scarce. NM behavior is strongly dependent on the environment but few data are available using complex exposure media, raising the question of its environmental impacts. The aim of the present work was to assess the toxic potential of three CeO2 NMs in Corbicula fluminea at a molecular level by RT-qPCR under a more realistic scenario of exposure, in a multistress context at two different salinities (1.5 and 15 psu). C. fluminea was exposed for 28 days to pulses of the three selected NMs (reference, manufactured, and aged manufactured). In bivalves, the gills and digestive gland are two key organs used for ecotoxicological studies. The expression change of 12 genes was measured in control organisms after 28 days in both organs, allowing us to clearly separate the responses for both organs and salinities. As gills come in contact with the environment first, we monitored gene the expression at intermediate time points (7, 14, and 21 days) for this organ in order to highlight clams responses to NM and salinity. Two genes (Se-GPx, MnSOD) had a salinity-dependent level of expression. HSP70, Se-GPx, and Trxr mRNAs presented significant changes in their expressions in the presence of NM. This study was completed using an integrated statistical approach. The exposed organisms differed more from control at field salinity than those exposed to hyper-saline conditions. At 15 psu, salinity pressure seems to cause the first molecular impact. At 1.5 psu, gene expression patterns allowed the effect of each NM to separate clearly. These results confirmed the usefulness of gene expression studies. Moreover, we highlighted the necessity to assess the environmental toxicity of the different forms of manufactured NM.


Corbicula fluminea Gene expression CeO2 pristine and aged nanomaterials Mesocosm Salinity 


Funding information

Financial supports were provided by the French National Agency (ANR-3-CESA-0014/NANOSALT project) and CPER Lorraine-ZAM (Contrat Projet Etat Région Lorraine, Zone Atelier Moselle). This work is a contribution to the Labex Ressources 21 (ANR- 10-LABX-21-01, Strategic metal resources of the 21st century). The authors gratefully acknowledge CNRS for funding the iCEINT International Consortium for the Environmental Implications of NanoTechnology. KOEHLE-DIVO Vanessa received financial support for salary from the French Research Ministry. Sharon Kruger is gratefully acknowledged for her English corrections.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

11356_2019_4927_MOESM1_ESM.docx (78 kb)
ESM 1 (DOCX 78 kb)


  1. An MI, Choi CY (2010) Activity of antioxidant enzymes and physiological responses in ark shell, Scapharca broughtonii, exposed to thermal and osmotic stress: effects on hemolymph and biochemical parameters. Comp Biochem Physiol B: Biochem Mol Biol 155:34–42. CrossRefGoogle Scholar
  2. Auffan M, Rose J, Orsiere T, De Meo M, Thill A, Zeyons O, Proux O, Masion A, Chaurand P, Spalla O, Botta A, Wiesner MR, Bottero J-Y (2009) CeO nanoparticles induce DNA damage towards human dermal fibroblasts in vitro. Nanotoxicology 3:161–171.
  3. Auffan M, Tella M, Liu W, Pariat A, Cabié M, Borschneck D, Angeletti B, Landrot G, Mouneyrac C, Giambérini L, Rose J (2017) Structural and physical–chemical behavior of a CeO2 nanoparticle based diesel additive during combustion and environmental release. Environ Sci: Nano 4:1974–1980. Google Scholar
  4. Baker TJ, Tyler CR, Galloway TS (2014) Impacts of metal and metal oxide nanoparticles on marine organisms. Environ Pollut 186:257–271. CrossRefGoogle Scholar
  5. Balbi T, Smerilli A, Fabbri R, Ciacci C, Montagna M, Grasselli E, Brunelli A, Pojana G, Marcomini A, Gallo G, Canesi L (2014) Co-exposure to n-TiO2 and Cd2+ results in interactive effects on biomarker responses but not in increased toxicity in the marine bivalve M. galloprovincialis. Sci Total Environ 493:355–364. CrossRefGoogle Scholar
  6. Banni M, Sforzini S, Balbi T, Corsi I, Viarengo A, Canesi L (2016) Combined effects of n-TiO2 and 2,3,7,8-TCDD in Mytilus galloprovincialis digestive gland: a transcriptomic and immunohistochemical study. Environ Res 145:135–144. CrossRefGoogle Scholar
  7. Barmo C, Ciacci C, Canonico B, Fabbri R, Cortese K, Balbi T, Marcomini A, Pojana G, Gallo G, Canesi L (2013) In vivo effects of n-TiO2 on digestive gland and immune function of the marine bivalve Mytilus galloprovincialis. Aquat Toxicol 132–133:9–18. CrossRefGoogle Scholar
  8. Bertrand C, Zalouk-Vergnoux A, Giambérini L, Poirier L, Devin S, Labille J, Perrein-Ettajani H, Pagnout C, Châtel A, Levard C, Auffan M, Mouneyrac C (2016) The influence of salinity on the fate and behavior of silver standardized nanomaterial and toxicity effects in the estuarine bivalve Scrobicularia plana: Salinity influences Ag NM-300K behavior and toxicity in clam. Environ Toxicol Chem 35:2550–2561.
  9. Bertrand C, Devin S, Mouneyrac C, Giambérini L (2017) Eco-physiological responses to salinity changes across the freshwater-marine continuum on two euryhaline bivalves: Corbicula fluminea and Scrobicularia plana. Ecol Indic 74:334–342. CrossRefGoogle Scholar
  10. Bigot A, Doyen P, Vasseur P, Rodius F (2009) Metallothionein coding sequence identification and seasonal mRNA expression of detoxification genes in the bivalve Corbicula fluminea. Ecotoxicol Environ Saf 72:382–387. CrossRefGoogle Scholar
  11. Bigot A, Minguez L, Giambérini L, Rodius F (2011) Early defense responses in the freshwater bivalve Corbicula fluminea exposed to copper and cadmium: transcriptional and histochemical studies. Environ Toxicol 26:623–632. CrossRefGoogle Scholar
  12. Bottero J-Y, Auffan M, Borschnek D, Chaurand P, Labille J, Levard C, Masion A, Tella M, Rose J, Wiesner MR (2015) Nanotechnology, global development in the frame of environmental risk forecasting. A necessity of interdisciplinary researches. Compt Rendus Geosci 347:35–42.
  13. Boutet I, Tanguy A, Rousseau S, Auffret M, Moraga D (2003) Molecular identification and expression of heat shock cognate 70 (hsc70) and heat shock protein 70 (hsp70) genes in the Pacific oyster Crassostrea gigas. Cell Stress Chaperones 8:76–85Google Scholar
  14. Calzolai L, Ansorge W, Calabrese E, Denslow N, Part P, Lettieri T (2007) Transcriptomics and proteomics. Applications to ecotoxicology. Comp Biochem Physiol Part D Genomics Proteomics 2:245–249. CrossRefGoogle Scholar
  15. Canesi L, Corsi I (2016) Effects of nanomaterials on marine invertebrates. Sci Total Environ 565:933–940. CrossRefGoogle Scholar
  16. Canesi L, Ciacci C, Fabbri R, Marcomini A, Pojana G, Gallo G (2012) Bivalve molluscs as a unique target group for nanoparticle toxicity. Mar Environ Res 76:16–21. CrossRefGoogle Scholar
  17. Canesi L, Frenzilli G, Balbi T, Bernardeschi M, Ciacci C, Corsolini S, Della Torre C, Fabbri R, Faleri C, Focardi S, Guidi P, Kočan A, Marcomini A, Mariottini M, Nigro M, Pozo-Gallardo K, Rocco L, Scarcelli V, Smerilli A, Corsi I (2014) Interactive effects of n-TiO2 and 2,3,7,8-TCDD on the marine bivalve Mytilus galloprovincialis. Aquat Toxicol 153:53–65. CrossRefGoogle Scholar
  18. Carregosa V, Velez C, Soares AMVM, Figueira E, Freitas R (2014) Physiological and biochemical responses of three Veneridae clams exposed to salinity changes. Comp Biochem Physiol B: Biochem Mol Biol 177–178(1–9):1–9. CrossRefGoogle Scholar
  19. Cerqueira CCC, Fernandes MN (2002) Gill tissue recovery after copper exposure and blood parameter responses in the tropical fish Prochilodus scrofa. Ecotoxicol Environ Saf 52:83–91. CrossRefGoogle Scholar
  20. Châtel A, Lièvre C, Barrick A, Bruneau M, Mouneyrac C (2018) Transcriptomic approach: a promising tool for rapid screening nanomaterial-mediated toxicity in the marine bivalve Mytilus edulis —application to copper oxide nanoparticles. Comp Biochem Physiol, Part C: Toxicol Pharmacol 205:26–33. Google Scholar
  21. Chen J, Patil S, Seal S, McGinnis JF (2006) Rare earth nanoparticles prevent retinal degeneration induced by intracellular peroxides. Nat Nanotechnol 1:142–150CrossRefGoogle Scholar
  22. Chen H, Zha J, Yuan L, Wang Z (2015) Effects of fluoxetine on behavior, antioxidant enzyme systems, and multixenobiotic resistance in the Asian clam Corbicula fluminea. Chemosphere 119:856–862. CrossRefGoogle Scholar
  23. Cheng G, Guo W, Han L, Chen E, Kong L, Wang L, Ai W, Song N, Li H, Chen H (2013) Cerium oxide nanoparticles induce cytotoxicity in human hepatoma SMMC-7721 cells via oxidative stress and the activation of MAPK signaling pathways. Toxicol in Vitro 27:1082–1088. CrossRefGoogle Scholar
  24. Ciofani G, Genchi GG, Mazzolai B, Mattoli V (2014) Transcriptional profile of genes involved in oxidative stress and antioxidant defense in PC12 cells following treatment with cerium oxide nanoparticles. Biochim Biophys Acta Gen Subj 1840:495–506. CrossRefGoogle Scholar
  25. Cossu C, Doyotte A, Jacquin MC, Babut M, Exinger A, Vasseur P (1997) Glutathione reductase, selenium-dependent glutathione peroxidase, glutathione levels, and lipid peroxidation in freshwater bivalves, Unio tumidus, as biomarkers of aquatic contamination in field studies. Ecotoxicol Environ Saf 38:122–131Google Scholar
  26. D’Agata A, Fasulo S, Dallas LJ, Fisher AS, Maisano M, Readman JW, Jha AN (2014) Enhanced toxicity of ‘bulk’ titanium dioxide compared to ‘fresh’ and ‘aged’ nano-TiO2 in marine mussels (Mytilus galloprovincialis). Nanotoxicology 8:549–558. CrossRefGoogle Scholar
  27. Dale JG, Cox SS, Vance ME, Marr LC, Hochella MF (2017) Transformation of cerium oxide nanoparticles from a diesel fuel additive during combustion in a diesel engine. Environ Sci Technol 51:1973–1980. CrossRefGoogle Scholar
  28. Das M, Patil S, Bhargava N, Kang J-F, Riedel LM, Seal S, Hickman JJ (2007) Auto-catalytic ceria nanoparticles offer neuroprotection to adult rat spinal cord neurons. Biomaterials 28:1918–1925. CrossRefGoogle Scholar
  29. Devoille L, Revel M, Batana C, Feltin N, Giambérini L, Châtel A, Mouneyrac C (2018) Combined influence of oxygenation and salinity on aggregation kinetics of the silver reference nanomaterial NM-300K. Environ Toxicol Chem 37:1918–1925. CrossRefGoogle Scholar
  30. Doyen P, Vasseur P, Rodius F (2006) Identification, sequencing and expression of selenium-dependent glutathione peroxidase transcript in the freshwater bivalve Unio tumidus exposed to Aroclor 1254. Comp Biochem Physiol, Part C: Toxicol Pharmacol 144:122–129. Google Scholar
  31. Doyen P, Bigot A, Vasseur P, Rodius F (2008) Molecular cloning and expression study of pi-class glutathione S-transferase (pi-GST) and selenium-dependent glutathione peroxidase (Se-GPx) transcripts in the freshwater bivalve Dreissena polymorpha. Comp Biochem Physiol, Part C: Toxicol Pharmacol 147:69–77. Google Scholar
  32. Eom H-J, Choi J (2009) Oxidative stress of CeO2 nanoparticles via p38-Nrf-2 signaling pathway in human bronchial epithelial cell, Beas-2B. Toxicol Lett 187:77–83. CrossRefGoogle Scholar
  33. Ferreira-Rodríguez N, Pardo I (2016) An experimental approach to assess Corbicula fluminea (Müller, 1774) resistance to osmotic stress in estuarine habitats. Estuar Coast Shelf Sci 176:110–116. CrossRefGoogle Scholar
  34. Garaud M, Trapp J, Devin S, Cossu-Leguille C, Pain-Devin S, Felten V, Giamberini L (2015) Multibiomarker assessment of cerium dioxide nanoparticle (nCeO ) sublethal effects on two freshwater invertebrates, Dreissena polymorpha and Gammarus roeseli. Aquat Toxicol 158:63–74.
  35. Garaud M, Auffan M, Devin S, Felten V, Pagnout C, Pain-Devin S, Proux O, Rodius F, Sohm B, Giamberini L (2016) Integrated assessment of ceria nanoparticle impacts on the freshwater bivalve Dreissena polymorpha. Nanotoxicology 10:935–944. CrossRefGoogle Scholar
  36. Gomes T, Pereira CG, Cardoso C, Bebianno MJ (2013) Differential protein expression in mussels Mytilus galloprovincialis exposed to nano and ionic Ag. Aquat Toxicol 136–137:79–90. CrossRefGoogle Scholar
  37. Guan X, Shi W, Zha S, Rong J, Su W, Liu G (2018) Neurotoxic impact of acute TiO2 nanoparticle exposure on a benthic marine bivalve mollusk, Tegillarca granosa. Aquat Toxicol 200:241–246. CrossRefGoogle Scholar
  38. Guo R, Pan L, Ji R (2017) A multi-biomarker approach in scallop Chlamys farreri to assess the impact of contaminants in Qingdao coastal area of China. Ecotoxicol Environ Saf 142:399–409.
  39. Hoecke KV, Quik JT, Mankiewicz-Boczek J, Schamphelaere KAD, Elsaesser A, der Meeren PV, Barnes C, McKerr G, Howard CV, Meent DVD (2009) Fate and effects of CeO2 nanoparticles in aquatic ecotoxicity tests. Environ Sci Technol 43:4537–4546CrossRefGoogle Scholar
  40. Hussain S, Garantziotis S (2013) Interplay between apoptotic and autophagy pathways after exposure to cerium dioxide nanoparticles in human monocytes. Autophagy 9:101–103. CrossRefGoogle Scholar
  41. Keller AA, Wang H, Zhou D, Lenihan HS, Cherr G, Cardinale BJ, Miller R, Ji Z (2010) Stability and aggregation of metal oxide nanoparticles in natural aqueous matrices. Environ Sci Technol 44:1962–1967Google Scholar
  42. Kholodkevich SV, Kuznetsova TV, Trusevich VV, Kurakin AS, Ivanov AV (2009) Peculiarities of valve movement and of cardiac activity of the bivalve mollusc Mytilus galloprovincialis at various stress actions. J Evol Biochem Physiol 45:524–526. CrossRefGoogle Scholar
  43. Klaper R, Arndt D, Bozich J, Dominguez G (2014) Molecular interactions of nanomaterials and organisms: defining biomarkers for toxicity and high-throughput screening using traditional and next-generation sequencing approaches. Analyst 139:882–895. CrossRefGoogle Scholar
  44. Koehlé-Divo V, Cossu-Leguille C, Pain-Devin S, Simonin C, Bertrand C, Sohm B, Mouneyrac C, Devin S, Giambérini L (2018) Genotoxicity and physiological effects of CeO2 NPs on a freshwater bivalve (Corbicula fluminea). Aquat Toxicol 198:141–148. CrossRefGoogle Scholar
  45. Li J, Schiavo S, Xiangli D, Rametta G, Miglietta ML, Oliviero M, Changwen W, Manzo S (2018) Early ecotoxic effects of ZnO nanoparticle chronic exposure in Mytilus galloprovincialis revealed by transcription of apoptosis and antioxidant-related genes. Ecotoxicology 27:369–384. CrossRefGoogle Scholar
  46. Lin W, Huang Y, Zhou X-D, Ma Y (2006) Toxicity of cerium oxide nanoparticles in human lung cancer cells. Int J Toxicol 25:451–457. CrossRefGoogle Scholar
  47. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 25:402–408. CrossRefGoogle Scholar
  48. Luo L, Ke C, Guo X, Shi B, Huang M (2014) Metal accumulation and differentially expressed proteins in gill of oyster (Crassostrea hongkongensis) exposed to long-term heavy metal-contaminated estuary. Fish Shellfish Immunol 38:318–329. CrossRefGoogle Scholar
  49. Metian M, Warnau M, Cosson RP, Oberhänsli F, Bustamante P (2008) Bioaccumulation and detoxification processes of Hg in the king scallop Pecten maximus: field and laboratory investigations. Aquat Toxicol 90:204–213. CrossRefGoogle Scholar
  50. Morimoto Y, Izumi H, Yoshiura Y, Tomonaga T, Oyabu T, Myojo T, Kawai K, Yatera K, Shimada M, Kubo M, Yamamoto K, Kitajima S, Kuroda E, Kawaguchi K, Sasaki T (2015) Pulmonary toxicity of well-dispersed cerium oxide nanoparticles following intratracheal instillation and inhalation. J Nanopart Res 17:442. CrossRefGoogle Scholar
  51. Navarro A, Faria M, Barata C, Piña B (2011) Transcriptional response of stress genes to metal exposure in zebra mussel larvae and adults. Environ Pollut 159:100–107. CrossRefGoogle Scholar
  52. Niu J, Azfer A, Rogers L, Wang X, Kolattukudy P (2007) Cardioprotective effects of cerium oxide nanoparticles in a transgenic murine model of cardiomyopathy. Cardiovasc Res 73:549–559CrossRefGoogle Scholar
  53. OECD (2010) List of manufactured nanomaterials and list of endpoints for phase one of the sponsorship programme for the testing of manufactured nanomaterials: revision. Series on the Safety of Manufactured Nanomaterials No, In, p 27Google Scholar
  54. Park B, Martin P, Harris C, Guest R, Whittingham A, Jenkinson P, Handley J (2007) Initial in vitro screening approach to investigate the potential health and environmental hazards of Envirox™ – a nanoparticulate cerium oxide diesel fuel additive. Part Fibre Toxicol 4:12. CrossRefGoogle Scholar
  55. Park E-J, Choi J, Park Y-K, Park K (2008) Oxidative stress induced by cerium oxide nanoparticles in cultured BEAS-2B cells. Toxicology 245:90–100. CrossRefGoogle Scholar
  56. Philipp EER, Wessels W, Gruber H, Strahl J, Wagner AE, Ernst IMA, Rimbach G, Kraemer L, Schreiber S, Abele D, Rosenstiel P (2012) Gene expression and physiological changes of different populations of the long-lived bivalve Arctica islandica under low oxygen conditions. PLoS One 7:e44621. CrossRefGoogle Scholar
  57. Piña B, Casado M, Quirós L (2007) Analysis of gene expression as a new tool in ecotoxicology and environmental monitoring. TrAC Trends Anal Chem 26:1145–1154. CrossRefGoogle Scholar
  58. R Core Team (2014) R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria URL
  59. Renault S, Baudrimont M, Mesmer-Dudons N, Gonzalez P, Mornet S, Brisson A (2008) Impacts of gold nanoparticle exposure on two freshwater species: a phytoplanktonic alga (Scenedesmus subspicatus) and a benthic bivalve (Corbicula fluminea). Gold Bull 41:116–126. CrossRefGoogle Scholar
  60. Ringwood AH, McCarthy M, Bates TC, Carroll DL (2010) The effects of silver nanoparticles on oyster embryos. Mar Environ Res 69:S49–S51. CrossRefGoogle Scholar
  61. Rocha TL, Gomes T, Sousa VS, Mestre NC, Bebianno MJ (2015) Ecotoxicological impact of engineered nanomaterials in bivalve molluscs: An overview. Mar Environ Res 111:74–88. CrossRefGoogle Scholar
  62. Ruiz P, Katsumiti A, Nieto JA, Bori J, Jimeno-Romero A, Reip P, Arostegui I, Orbea A, Cajaraville MP (2015) Short-term effects on antioxidant enzymes and long-term genotoxic and carcinogenic potential of CuO nanoparticles compared to bulk CuO and ionic copper in mussels Mytilus galloprovincialis. Mar Environ Res 111:107–120. CrossRefGoogle Scholar
  63. Shi W, Han Y, Guo C, Zhao X, Liu S, Su W, Zha S, Wang Y, Liu G (2017) Immunotoxicity of nanoparticle nTiO2 to a commercial marine bivalve species, Tegillarca granosa. Fish Shellfish Immunol 66:300–306. CrossRefGoogle Scholar
  64. Singh S (2016) Cerium oxide based nanozymes: redox phenomenon at biointerfaces. Biointerphases 11:04B202. CrossRefGoogle Scholar
  65. Singh C, Institute for Health and Consumer Protection (2014) Cerium Dioxide NM-211, NM-212, NM-213, characterisation and test item preparation JRC repository: NM-series of representative manufactured nanomaterials. Publications Office of the European Union, LuxembourgGoogle Scholar
  66. Tran D, Fournier E, Durrieu G, Massabuau J-C (2003) Copper detection in the Asiatic clam Corbicula fluminea: optimum valve closure response. Aquat Toxicol 65:317–327. CrossRefGoogle Scholar
  67. Volland M, Hampel M, Martos-Sitcha JA, Trombini C, Martínez-Rodríguez G, Blasco J (2015) Citrate gold nanoparticle exposure in the marine bivalve Ruditapes philippinarum: uptake, elimination and oxidative stress response. Environ Sci Pollut Res Int 22:17414–17424. CrossRefGoogle Scholar
  68. Volland M, Hampel M, Katsumiti A, Yeste MP, Gatica JM, Cajaraville M, Blasco J (2018) Synthesis methods influence characteristics, behaviour and toxicity of bare CuO NPs compared to bulk CuO and ionic cu after in vitro exposure of Ruditapes philippinarum hemocytes. Aquat Toxicol 199:285–295CrossRefGoogle Scholar
  69. Wang Y, Nowack B (2018a) Dynamic probabilistic material flow analysis of nano-SiO2, nano iron oxides, nano-CeO2, nano-Al2O3, and quantum dots in seven European regions. Environ Pollut 235:589–601. CrossRefGoogle Scholar
  70. Wang Y, Nowack B (2018b) Environmental risk assessment of engineered nano-SiO2, nano iron oxides, nano-CeO2, nano-Al2O3, and quantum dots: environmental risk assessment of engineered nanomaterials. Environ Toxicol Chem 37:1387–1395. CrossRefGoogle Scholar
  71. Wiesner MR, Bottero J-Y (2016) Environmental nanotechnology: applications and impacts of nanomaterials, 2nd edn. McGraw Hill, New York, p 141–172Google Scholar
  72. Zhang H, He X, Zhang Z, Zhang P, Li Y, Ma Y, Kuang Y, Zhao Y, Chai Z (2011) Nano-CeO2 exhibits adverse effects at environmental relevant concentrations. Environ Sci Technol 45:3725–3730. CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Université de Lorraine, CNRS, LIECMetzFrance
  2. 2.Laboratoire Mer, Molécules et Santé (MMS, EA2160)Université Catholique de l’OuestAngers Cedex 01France

Personalised recommendations