Skip to main content
SpringerLink
Log in

Chlorpyrifos levels within permitted limits induce nuclear abnormalities and DNA damage in the erythrocytes of the common carp

  • Research Article
  • Published:
Environmental Science and Pollution Research Aims and scope Submit manuscript

Abstract

The organophosphate pesticide chlorpyrifos (CPF) is defined as a priority pollutant in surface freshwaters according to Directive 2013/39/EU of the European Parliament. The focus of this study was to assess the potential cytotoxic and genotoxic effects of permissible CPF levels on juvenile forms of the common carp. We found that low-level CPF exposure did not induce elevated levels of micronuclei, but significantly increased the frequency of total nuclear abnormalities (NAs) proportional to dose and time; notched, blebbed, lobed and eight-shaped nuclei, nuclear buds, nuclear bridges and binucleated cells were all detected. Decreased frequencies of polychromatic erythrocytes (PCEs) and DNA damage detected by comet assay were also observed, confirming the cytotoxic and genotoxic effects of CPF. Altogether, these data (1) demonstrate that CPF is toxic even at permissible levels, possessing considerable genotoxic and cytotoxic potential in peripheral erythrocytes of exposed fish and (2) validate the assessment of NAs, PCEs and comet assay performance as sensitive biomarkers for the early detection of CPF pollution. These findings can be applied to guide environmental risk assessment and biomonitoring programs.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  • Ali D, Nagpure NS, Kumar S, Kumar R, Kushwaha B, Lakra WS (2009) Assessment of genotoxic and mutagenic effects of chlorpyrifos in freshwater fish Channa punctatus (Bloch) using micronucleus assay and alkaline single-cell gel electrophoresis. Food Chem Toxicol 47:650–656

    CAS  Google Scholar 

  • Altun S, Özdemir S, Arslan H (2017) Histopathological effects, responses of oxidative stress, inflammation, apoptosis biomarkers and alteration of gene expressions related to apoptosis, oxidative stress, and reproductive system in chlorpyrifos-exposed common carp (Cyprinus carpio L.). Environ Pollut 230:432–443

    CAS  Google Scholar 

  • Ambreen F, Javed M (2018) Pesticide mixture induced DNA damage in peripheral blood erythrocytes of freshwater fish, Oreochromis niloticus. Pak Jf Zool 50(1). https://doi.org/10.17582/journal.pjz/2018.50.1.339.346

  • Anbumani S, Mohankumar MN (2011) Nuclear and cytoplasmic abnormalities in the fish Catla catla (Hamilton) exposed to chemicals and ionizing radiation. Res J Environ Sci 5(12):867–877

    CAS  Google Scholar 

  • APHA (2005) Standard methods for examination of water and wastewater, 21st Ed. – American public health association, Washington, DC, USA

  • Ayllon F, Garcia-Vazquez E (2000) Induction of micronuclei and other nuclear abnormalities in European minnow Phoxinus phoxinus and mollie Poecilia latipinna: an assessment of the fish micronucleus test. Mutat Res Genet Toxicol Environ Mutagen 467(2):177–186

    CAS  Google Scholar 

  • Banks KE, Hunter DH, Wachal DJ (2005) Chlorpyrifos in surface waters before and after a federally mandated ban. Environ Int 31:351–356

    CAS  Google Scholar 

  • Baršienė J, Butrimavičienė L, Grygiel W, Lang T, Michailovas A, Jackūnas T (2014) Environmental genotoxicity and cytotoxicity in flounder (Platichthys flesus), herring (Clupea harengus) and Atlantic cod (Gadus morhua) from chemical munitions dumping zones in the southern Baltic Sea. Mar Еnviron Res 96:56–67

    Google Scholar 

  • Baskaran S, Kookana RS, Naidu R (1999) Degradation of bifenthrin, chlorpyrifos and imidacloprid in soil and bedding materials at termiticidal application rates. Pestic Sci 55:1222–1228

    CAS  Google Scholar 

  • Bhatnagar A, Yadav AS, Cheema N (2016) Genotoxic effects of chlorpyrifos in freshwater fish Cirrhinus mrigala using micronucleus assay. Advances in Biology. https://doi.org/10.1155/2016/9276963

    Google Scholar 

  • Brock TC, Arts GH, Maltby L, Van den Brink PJ (2006) Aquatic risks of pesticides, ecological protection goals, and common aims in European Union legislation. Integr Environ Assess Manag 2(4):20–46

    Google Scholar 

  • Buschini A, Martino A, Gustavino B, Monfrinotti M, Poli P, Rossi C, Santoro M, Dörr AJM, Rizzoni M (2004) Comet assay and micronucleus test in circulating erythrocytes of Cyprinus carpio specimens exposed in situ to lake waters treated with disinfectants for potabilization. Mutat Res Genet Toxicol Environ Mutagen 557(2):119–129

    CAS  Google Scholar 

  • Carrasco KR, Tilbury KL, Myers MS (1990) Assessment of the piscine micronucleus test as an in situ biological indicator of chemical contaminant effects. Can J Fish Aquat Sci 47(11):2123–2136

    CAS  Google Scholar 

  • Çavaş T (2008) In vivo genotoxicity of mercury chloride and lead acetate: micronucleus test on acridine orange stained fish cells. Food Chem Toxicol 46(1):352–358

    Google Scholar 

  • Çavaş T (2011) In vivo genotoxicity evaluation of atrazine and atrazine-based herbicide on fish Carassius auratus using the micronucleus test and the comet assay. Food Chem Toxicol 49:1431–1435

    Google Scholar 

  • Çavaş T, Ergene-Gözükara S (2003) Micronuclei, nuclear lesions and interphase silver-stained nucleolar organizer regions (AgNORs) as cyto-genotoxicity indicators in Oreochromis niloticus exposed to textile mill effluent. Mutat Res Genet Toxicol Environ Mutagen 538(1–2):81–91

    Google Scholar 

  • Çavaş T, Ergene-Gözükara S (2005) Induction of micronuclei and nuclear abnormalities in Oreochromis niloticus following exposure to petroleum refinery and chromium processing plant effluents. Aquat Toxicol 74:264–271

    Google Scholar 

  • Çavaş T, Garanko NN, Arkhipchuk VV (2005) Induction of micronuclei and binuclei in blood, gill and liver cells of fishes subchronically exposed to cadmium chloride and copper sulphate. Food Chem Toxicol 43(4):569–574

    Google Scholar 

  • Çavaş T, Könen S (2007) Detection of cytogenetic and DNA damage in peripheral erythrocytes of goldfish (Carassius auratus) exposed to a glyphosate formulation using the micronucleus test and the comet assay. Mutagenesis 22(4):263–268

    Google Scholar 

  • Chen D, Zhang Z, Yao H, Liang Y, Xing H, Xu S (2015) Effects of atrazine and chlorpyrifos on oxidative stress-induced autophagy in the immune organs of common carp (Cyprinus carpio L.). Fish Shellfish Immun 44(1):12–20

    CAS  Google Scholar 

  • Collins AR, Oscoz AA, Brunborg G, Gaivao I, Giovannelli L, Kruszewski M, Smith CC, Štětina R (2008) The comet assay: topical issues. Mutagenesis 23(3):143–151

    CAS  Google Scholar 

  • da Silva Souza T, Fontanetti CS, (2006) Micronucleus test and observation of nuclear alterations in erythrocytes of Nile tilapia exposed to waters affected by refinery effluent. Mutat Res Genet Toxicol Environ Mutagen 605 (1-2):87-93

  • De Silva PMCS, Samayawardhena LA (2005) Effects of chlorpyrifos on reproductive performances of guppy (Poecilia reticulata). Chemosphere 58(9):1293–1299

    Google Scholar 

  • Deb N, Das S (2013) Chlorpyrifos toxicity in fish: a review. Curr world environ 8(1):77–84. https://doi.org/10.12944/CWE.8.1.17

    Article  CAS  Google Scholar 

  • Directive 2010/63/EU of the European parliament and of the Council of 22 September 2010 on the protection of animals used for scientific purposes. https://eur-lex.europa.eu/legal-content/EN/TXT/?uri=celex%3A32010L0063

  • Directive 2013/39/EU of the European Parliament and of the Council of 12 August 2013 amending Directives 2000/60/EC and 2008/105/EC as regards priority substances in the field of water policy. https://eur-lex.europa.eu/LexUriServ/LexUriServ.do?uri=OJ:L:2013:226:0001:0017:EN:PDF

  • Du Preez LH, Jansen van Rensburg PJ, Jooste AM, Carr JA, Giesy JP, Gross TS, Kendall PJ, Smith EE, Van Der Kraak G, Solomon KR (2005) Seasonal exposures to triazine and other pesticides insurface waters in the Western Highveld corn-production region in South Africa. Environ Pollut 135:131–141

    Google Scholar 

  • DWAF Department of Water Affairs and Forestry (1996): South African water quality 570 guidelines. – Pretoria, South Africa

  • Fenech M (2007) Cytokinesis-block micronucleus cytome assay. Nat Protoc 2(5):1084

    CAS  Google Scholar 

  • Fenech M, Kirsch-Volders M, Natarajan AT, Surralles J, Crott JW, Parry J, Norppa H, Eastmond DA, Tucker JD, Thomas P (2011) Molecular mechanisms of micronucleus, nucleoplasmic bridge and nuclear bud formation in mammalian and human cells. Mutagenesis 26(1):125–132

    CAS  Google Scholar 

  • Furnus GNA, Caffetti JD, García EM, Benítez MF, Pastori MC, Fenocchio AS (2014) Baseline micronuclei and nuclear abnormalities frequencies in native fishes from the Paraná River (Argentina). Braz J Biol 74(1):217–221

    CAS  Google Scholar 

  • Giesy JP, Solomon KA (2014) Ecological risk assessment for chlorpyrifos in terrestrial and aquatic systems in North America. Springer

  • Ismail M, Khan QM, Ali R, Ali T, Mobeen A (2014) Evaluation of the genotoxicity of chlorpyrifos in common indus valley toad, Bufo stomaticus using alkaline single-cell gel electrophoresis (Comet) assay. Agric Sci 5(04):376. http://www.scirp.org/journal/as. https://doi.org/10.4236/as.2014

    Article  CAS  Google Scholar 

  • Ismail M, Ali R, Shahid M, Khan MA, Zubair M, Ali T, Mahmood Khan Q (2018) Genotoxic and hematological effects of chlorpyrifos exposure on freshwater fish Labeo rohita. Drug Chem Toxicol 41(1):22–26

    CAS  Google Scholar 

  • Kumar SP (2012) Micronucleus assay: a sensitive indicator for aquatic pollution. Int J Res Biosci 1(2):32–37

    Google Scholar 

  • Li X, Liu L, Zhang Y, Fang Q, Li Y, Li Y (2013) Toxic effects of chlorpyrifos on lysozyme activities, the contents of complement C3 and IgM, and IgM and complement C3 expressions in common carp (Cyprinus carpio L.). Chemosphere 93(2):428–433

    CAS  Google Scholar 

  • Li D, Huang Q, Lu M, Zhang L, Yang Z, Zong M, Tao L (2015) The organophosphate insecticide chlorpyrifos confers its genotoxic effects by inducing DNA damage and cell apoptosis. Chemosphere 135:387–393

    CAS  Google Scholar 

  • Mitkovska V, Dimitrov H, Chassovnikarova T (2017) In vivo genotoxicity and cytotoxicity assessment of allowable concentrations of nickel and lead: comet assay and nuclear abnormalities in acridine orange stained erythrocytes of common carp (Cyprinus carpio L.). Acta Zool Bulgar S8:47–56

    Google Scholar 

  • Muranli FDG, Güner U (2011) Induction of micronuclei and nuclear abnormalities in erythrocytes of mosquito fish (Gambusia affinis) following exposure to the pyrethroid insecticide lambda-cyhalothrin. Mutat Res Genet Toxicol Environ Mutagen 726(2):104–108

    CAS  Google Scholar 

  • OECD Test No. 203 (1992) Fish, acute toxicity test. Organisation for Economic Cooperation and Development, Paris

  • Palma P, Palma VL, Fernandes RM, Bohn A, Soares AMVM, Barbosa IR (2009) Embryo-toxic effects of environmental concentrations of chlorpyrifos on the crustacean Daphnia magna. Еcotoxicol Environ Saf 72:1714–1718

    CAS  Google Scholar 

  • Papadakis EN, Tsaboula A, Kotopoulou A, Kintzikoglou K, Vryzas Z, Papadopoulou-Mourkidou E (2015a) Pesticides in the surface waters of Lake Vistonis Basin, Greece: occurrence and environmental risk assessment. Sci Total Environ 536:793–802

    CAS  Google Scholar 

  • Papadakis EN, Vryzas Z, Kotopoulou A, Kintzikoglou K, Makris KC, Papadopoulou-Mourkidou (2015b) A pesticide monitoring survey in rivers and lakes of northern Greece and its human and ecotoxicological risk assessment. Ecotoxicol Environ Saf 116:1–9

    CAS  Google Scholar 

  • Pastor S, Creus A, Parrón T, Cebulska-Wasilewska A, Siffel C, Piperakis S, Mercos (2003) Biomonitoring of four European populations occupationally exposed to pesticides: use of micronuclei as biomarkers. Mutagenesis 18(3):249–258

    CAS  Google Scholar 

  • Patetsini E, Dimitriadis BK, Kaloyianni M (2013) Biomarkers in marine mussels, Mytilus galloprovincialis, exposed to environmentally relevant levels of the pesticides, chloropyrifos and penoxsulam. Aquat Toxicol 126:338–345

    CAS  Google Scholar 

  • Polard T, Jean S, Merlina G, Laplanche C, Pinelli E, Gauthier L (2011) Giemsa versus acridine orange staining in the fish micronucleus assay and validation for use in water quality monitoring. Ecotoxicol Environ Saf 74(1):144–149

    CAS  Google Scholar 

  • Rey Vázquez G, Lo Nostro F (2014) Changes in Hematological Parameters of Cichlasoma dimerus (Teleostei, Perciformes) Exposed to Sublethal Concentrations of 4-tert-Octylphenol. Arch Environ Con Tox 66 (3):463-469

    Google Scholar 

  • Ruiz de Arcaute C, Pérez-Iglesias JM, Nikoloff N, Natale GS, Soloneski S, Larramendy L (2014) Genotoxicity evaluation of the insecticide imidacloprid on circulating blood cells of Montevideo tree frog Hypsiboas pulchellus tadpoles (Anura, Hylidae) by comet and micronucleus bioassays. Ecol Indic 45:632-639

    CAS  Google Scholar 

  • Ruiz de Arcaute C, Soloneski S, Larramendy ML (2016) Toxic and genotoxic effects of the 2, 4-dichlorophenoxyacetic acid (2, 4-D)-based herbicide on the Neotropical fish Cnesterodon decemmaculatus. Ecotoxicol Environ Saf 128:222–229

    CAS  Google Scholar 

  • Singh NP, McCoy MT, Tice RR, Schneider EL (1988) A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res 175 (1):184-191

    CAS  Google Scholar 

  • Sunanda M, Rao JCS, Neelima P, Rao KG, Simhachalam G (2016) Effects of chlorpyrifos (an organophosphate pesticide) in fish. Int J Pharm Sci Rev Res 39(1):299–305

    CAS  Google Scholar 

  • Suzuki Y, Nagae Y, Li J, Sabaka H, Mazowa K, Takahashi A, Shimuzu H (1989) The micronucleus test and erythropoiesis: effects of erythropoietin and a mutagen on the ratio of polychromatic to normochromatic erythrocytes (P/N ratio). Mutagenesis 4:420–424

    CAS  Google Scholar 

  • Shimizu N, Itoh N, Utiyama H, Wahl MG (1998) Selective entrapment of extrachromosomally amplified DNA by nuclear budding and micronucleation during S phase. J Cell Biol 140(6):1307–1320

    CAS  Google Scholar 

  • Shimizu N, Shimuara T, Tanaka T (2000) Selective elimination of acentric double minutes from cancer cells through the extrusion of micronuclei. Mutat Res 448:81–90

    CAS  Google Scholar 

  • Ueda T, Hayashi M, Ohtsuka Y, Nakamura T, Kobayashi J, Sofuni T (1992) A preliminary study of the micronucleus test by acridine orange fluorescent staining compared with chromosomal aberration test using fish erythropoietic and embryonic cells. Water Sci Technol 25(11):235–240

    CAS  Google Scholar 

  • Udroiu I (2006) The micronucleus test in piscine erythrocytes. Aquat Toxicol 79(2):201–204

    CAS  Google Scholar 

  • Vera-Candioti J, Soloneski S, Larramendy ML (2013) Single-cell gel electrophoresis assay in the ten spotted live-bearer fish, Cnesterodon decemmaculatus (Jenyns, 1842), as bioassay for agrochemical-induced genotoxicity. Ecotoxicol Environ Saf 98:368–373

    CAS  Google Scholar 

  • Vera-Candioti J, Soloneski S, Larramendy ML (2014) Chlorpyrifos-based insecticides induced genotoxic and cytotoxic effects in the ten spotted live-bearer fish, Cnesterodon decemmaculatus (Jenyns, 1842). Environ Toxicol 29(12):1390–1398

    CAS  Google Scholar 

  • Wang X, Xing H, Li X, Xu S, Wang X (2011) Effects of atrazine and chlorpyrifos on the mRNA levels of IL-1 and IFN-γ2b in immune organs of common carp. Fish Shellfish Immunol 31:126–133

    CAS  Google Scholar 

  • Wang C, Zhang Z, Yao H, Zhao F, Wang L, Wang X, Xing H, Xu S (2014) Effects of atrazine and chlorpyrifos on DNA methylation in the liver, kidney and gill of the common carp (Cyprinus carpio L.). Ecotoxicol Environ Saf 108:142–151

    CAS  Google Scholar 

  • Wild D (1975) Mutagenicity studies on organophosphorus insecticides. Mutat Res 32:133–150

    CAS  Google Scholar 

  • Xing H, Li S, Wang Z, Gao X, Xu S, Wang X (2012a) Histopathological changes and antioxidant response in brain and kidney of common carp exposed to atrazine and chlorpyrifos. Chemosphere 88:377–383

    CAS  Google Scholar 

  • Xing H, Li S, Wang Z, Gao X, Xu S, Wang X (2012b) Oxidative stress response and histopathological changes due to atrazine and chlorpyrifos exposure in common carp. Pestic Biochem Physiol 103:74–80

    CAS  Google Scholar 

  • Xing H, Wang X, Sun G, Gao X, Xu S, Wang X (2012c) Effects of atrazine and chlorpyrifos on activity and transcription of glutathione S-transferase in common carp (Cyprinus carpio L.). Environ Toxicol Pharmacol 33(2):233–244

    CAS  Google Scholar 

  • Xing H, Wang Z, Gao X, Chen D, Wang L, Li S, Xu S (2015) Atrazine and chlorpyrifos exposure induces liver autophagic response in common carp. Ecotoxicol Environ Saf 113:52–58

    CAS  Google Scholar 

  • Yancheva V, Mollov I, Georgieva E, Stoyanova S, Tsvetanova V, Velcheva I (2017) Ex situ effects of chlorpyrifos on the lysosomal membrane stability and respiration rate in Zebra mussel (Dreissena polymorpha Pallas, 1771). Acta Zool Bulg S8:85–90

    Google Scholar 

  • Yancheva V, Velcheva I, Georgieva E, Mollov I, Stoyanova S (2019) Chlorpyrifos induced changes on the physiology of common carp (Cyprinus carpio Linnaeus, 1785): a laboratory exposure study. Appl Ecol Environ Res 17(2):5139–5157

    Google Scholar 

  • Yin X, Zhu G, Li XB, Liu S (2009) Genotoxicity evaluation of chlorpyrifos to amphibian chinese toad (amphibian: Anura) by comet assay and micronucleus test. Mutat Res Genet Toxicol Environ Mutagen 680:2–6

    CAS  Google Scholar 

  • Zhang GW, Wang JR, Zhan CR, Hu X (2010) Study of influences of chlorpyrifos on potential damage of DNA. J Nanchang Univ (Eng Technol) 32:10–13 (in Chinese)

    Google Scholar 

  • Zhang X, Shen Y, Yu X, Liu X (2012) Dissipation of chlorpyrifos and residue analysis in rice, soil and water under paddy field conditions. Ecotoxicol Environ Saf 78:276–280

    CAS  Google Scholar 

Download references

Funding

This study was supported by the National Program “Young Researchers and Postdocs, 2018” financed by the Ministry of Education and Science, Bulgaria.

Author information

Authors and Affiliations

  1. Department of Zoology, Faculty of Biology, University of Plovdiv Paisii Hilendarski, 24 Tzar Asen Street, 4000, Plovdiv, Bulgaria

    Vesela Mitkovska & Tsenka Chassovnikarova

  2. Institute of Biodiversity and Ecosystem Research, Bulgarian Academy of Sciences, 1 Tsar Osvoboditel Blvd, 1000, Sofia, Bulgaria

    Tsenka Chassovnikarova

Authors
  1. Vesela Mitkovska
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Tsenka Chassovnikarova
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Tsenka Chassovnikarova.

Additional information

Responsible Editor: Thomas Braunbeck

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Mitkovska, V., Chassovnikarova, T. Chlorpyrifos levels within permitted limits induce nuclear abnormalities and DNA damage in the erythrocytes of the common carp. Environ Sci Pollut Res 27, 7166–7176 (2020). https://doi.org/10.1007/s11356-019-07408-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11356-019-07408-9

Keywords

Search

Navigation