Genotoxic Response of Oreochromis niloticus Exposed to Tertiary Mixture of Pesticides

Faiza Ambreen, Mehmood Ahmed Husnain Hashmi, Sidra Abbas, Safina Kouser, Fariha Latif, Muhammad Javed


Background: At present the aquatic habitats of Pakistan become heavily polluted due to presence of heavy metals and pesticides. This research was carried out to check the percentage of DNA damage, Genetic Damage Index and Cumulative Tail Length of comets formed in the erythrocytes of Oreochromis niloticus following exposure to a tertiary mixture of pesticides (chlorpyrifos, endosulfan and bifenthrin) with the Comet assay.

Methods: Acute toxicity (96-hour LC50) of chlorpyrifos + endosulfan + bifenthrin mixture was determined for Oreochromis niloticus (180-day old), and then four sublethal concentrations (1/3rd, 1/4th, 1/5th , and 1/6th  of the LC50) were calculated. To control the possibility of temperature variation, fingerlings of O. niloticus were treated with four experimental pesticides concentrations used for duration of 90 days under constant conditions of laboratory (with negative and positive control). On day 14, 28, 42, 56, 70, and 84 fish peripheral blood cells were collected following exposure to assess DNA damage.

Results: DNA damage was observed to be statistically significant (p<0.05) throughout the exposure period due to the various test concentrations. In fish erythrocytes, a dose/concentration-dependent response was observed, with the greatest DNA damage occurring at 1/3rd of the LC50 exposure. Comparing DNA damage in Oreochromis niloticus peripheral blood erythrocytes across all sampling days revealed a continuous rise in the quantity of damaged DNA with increase in time of exposure.

Conclusion: Present investigation represented an unprecedented approach to study genotoxic effects of pesticides on fish. The widespread application of pesticides (chlorpyrifos, endosulfan, bifenthrin) in agriculture sector exerts adverse effects on various non-target organisms via trophic transfer that ultimately pose a serious threat for human beings. Current findings suggested minimized and sensible use of pesticides to avoid genetic threats to aquatic fauna and to maintain sustainable agriculture and aquaculture.

Keywords: Oreochromis niloticus; Endosulfan; Chlorpyrifos; DNA damage; Chronic exposure

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Cok I, Ulutas O, Okusluk O, Durmaz E, Demir N. Evaluation of DNA Damage in Common Carp (Cyprinus carpio L.) by Comet Assay for Determination of Possible Pollution in Lake Mogan (Ankara). Science World Journal, (2011); 11:1455-1461.

Dawood MA, Amer AA, Elbialy ZI, Gouda AH. Effects of including triticale on growth performance, digestive enzyme activity, and growth-related genes of Nile tilapia (Oreochromis niloticus). Aquaculture, (2020); 528: 568-768.

Yohannes YB, Ikenaka Y, Nakayama SM, Saengtienchai A. Organochlorine pesticides and heavy metals in fish from Lake Awassa, Ethiopia: Insights from stable isotope analysis. Chemosphere, (2013); 91(6): 857-863.

Brooks BW, Conkle JL. Commentary: Perspectives on aquaculture, urbanization and water quality. Comp. Biochem. Physiol. Part A: Environmental Toxicology and Pharmacology, (2019); 217: 1-4.

Nie X, Lan C, Wei T, Yang Y. Distribution of polychlorinated biphenyls in the water, sediment and fish from the Pearl River estuary, China. Marine Pollution Bulletin, (2005) ;50 (5): 537-546.

Binelli A, Provini A. POPs in edible clams from different Italian and European markets and possible human health risk. Marine Pollution Bulletin, (2003); 46(7): 879-886.

Kumari A, Sinha RK, Gopal K. Quantitative estimation of DDT residues in some freshwater fishes of River Ganga at Patna, Bihar. Environment and Ecology, (2001); 19: 396-399.

Huang Y, Xiao L, Li F, Xiao M, Lin D, Long X, Wu Z. Microbial Degradation of Pesticide Residues and an Emphasis on the Degradation of Cypermethrin and 3-phenoxy Benzoic Acid: A Review. Molecules, (2018); 23 (9): 2313

Jayaraj R, Megha P, Sreedev P. Review Article. Organochlorine pesticides, their toxic effects on living organisms and their fate in the environment. Interdisciplinary Toxicology, (2016); 9(3-4): 90-100.

Cosgrove S, Jefferson B, Jarvis P. Pesticide removal from drinking water sources by adsorption: a review. Environmental Technology Review, (2019); 8(1): 1-24.

Ali I, Alharbi O, Alothman Z, Al-Mohaimeed AM, Alwarthan A. Modeling of fenuron pesticide adsorption on CNTs for mechanistic insight and removal in water. Environmental Research, (2019); 170: 389-397.

Costa CS, Ronco AE, Trudeau VL, Marino D. Tadpoles of the horned frog Ceratophrys ornata exhibit high sensitivity to chlorpyrifos for conventional ecotoxicological and novel bioacoustic variables. Environmental Pollution, (2018); 235: 938-947.

Quiroga LB, Sanabria EA, Fornes MW, Bustos DA, Tejedo M. Sublethal concentrations of chlorpyrifos induce changes in the thermal sensitivity and tolerance of anuran tadpoles in the toad Rhinella arenarum. Chemosphere, (2019); 219: 671-677.

Rutkoski C, Macagnan N, Folador A, Skovronski VJ, Doamaral AM. Leitemperger J, Hartmann MT. Morphological and biochemical traits and mortality in Physalaemus gracilis (Anura: Leptodactylidae) tadpoles exposed to the insecticide chlorpyrifos. Chemosphere, (2020); 250: 126-162.

Kaur M, Jindal R. Biomarker enzyme activities and ultrastructure of liver of the grass carp Ctenopharyngodon idella (Valenciennes, 1844) exposed to the organophosphate pesticide chlorpyrifos. Indian Journal of Fisheries, (2017); 64: 1-7.

Gopika CM, Sumi N, Chitra KC. Involvement of reactive oxygen species in the toxicity of acrylamide in muscle tissue of the fish, Oreochromis niloticus (Linnaeus, 1758). World Journal Pharmaceutical Research, (2018); 7(1): 1617-1628.

Sebastian, R, Raghavan SC. Induction of DNA damage and erroneous repair can explain genomic instability caused by endosulfan. Carcinogenesis, (2016); 37(10): 929-940.

Wei J, Zhang L, Ren L, Lihua R, Jin Z, Jianhui L, Junchao D, Yang Y, Yanbo L, Cheng P, Xianqing Z, Zhiwei S. Endosulfan induces cell dysfunction through cycle arrest resulting from DNA damage and DNA damage response signaling pathways. Science of Total Environment, (2017); 589: 97-106.

James T, Nenov MN, Tapia CM, Lecchi M, Koshy S, Green TA, Laezza F. Consequences of acute Na v 1.1 Exposure to deltamethrin. Neurotoxicology, (2017); 60: 150-160.

Okda ES, Abdel-Hamid MA, Hamdy AM. Immunological and genotoxic effects of occupational exposure to α-cypermethrin pesticide. International Journal of Occupational Medicine and Environmental Health, (2017); 1:30(4): 603.

Cunha FDS, Natanlino CS, Ruda FBS, Juliana OM, Marcia VSC, Fabricio TCA, Jose GSF, Paulo CFC, Alexendere NM, Rodrigo YF. Deltamethrin-induced nuclear erythrocyte alteration and damage to the gills and liver of Colossoma macropomum. Environmental Science and Pollution, (2018); 25(15): 15102-15110.

Paravani EV, Simoniello MF, Poletta GL, Casco VH. Cypermethrin induction of DNA damage oxidative stress in zebrafish gill cells. Ecotoxicology and Environmental Safety, (2019); 173: 1-7.

Ullah S, Li Z, Zuberi A, Alagawany M, Farang MR, Dadar M, Karthik K, Tiwari R, Dhama K, Iqbal HM. Biomarkers of pyrethroid toxicity in fish. Environmental Chemistry Letters, (2019); 17(2): 945-973.

Zhang H, Hong X, Yan S, Zha J, Qin J. Environmentally relevant concentrations of bifenthrin induce changes in behaviour, biomarkers, histological characteristics, and the transcriptome in Corbicula fluminea. Science of Total Environment, (2020); 728: 138-821.

Ullah S, Zuberi A, Alagawany M, Farag MR, Dadar M, Karthik K, Tiwari R, Dhama K, Iqbal HM. Cypermethrin induced toxicities in fish and adverse health outcomes: Its prevention and control measure adaptation. Journal of Environmental Management, (2018); 206: 863-871.

Siraj M, Khisroon M, Khan A, Zaidi F, Ullah ARG. Bio-monitoring of Tissue Accumulation and Genotoxic Effect of Heavy Metals in Cyprinus carpio from River Kabul Khyber Pakhtunkhwa Pakistan. Bulletin of Environmental Contamination and Toxicology, (2018); 100(3): 344-349.

Shah N, Ajmal K, Nazma HK, Muhammad K. Genotoxic Consequences in Common Grass Carp (Ctenopharyngodon idella Valenciennes, 1844) Exposed to Selected Toxic Metals. Biological Trace Element Research, (2021); 199(1): 305-314.

Cai J, Leung P. Short-term projection of global fish demand and supply gaps. FAO Fish. Technical Papers, (2017); 607: 1-128.

Frenzilli G, Vittoria S, Ilaria DB, Marco N, Lars F, Claudia B, Joachim S. DNA damage in eelpout (Zoarces viviparus) from Goteborg harbour. Mutation Research, (2004);

(1-2): 187-195.

Cavas T. In vivo genotoxicity of mercury chloride and lead acetate: Micronucleus test on acridine orange stained fish cells. Food and Chemical Toxicology, (2008); 46(1): 352-358.

Lourenco J, Pereira R, Goncalves F, Mendo S. Metal bioaccumulation, genotoxicity and gene expression in the European wood mouse (Apodemus sylvaticus) inhabiting an abandoned uranium mining area. Science of Total Environment, (2013); 443: 673-680.

Shugart LR. DNA damage as biomarker of exposure. Ecotoxicology, (2000); 9(5): 329-340.

Khisroon M, Ajmal K, Maryam N, Naheed A, Sardar K, Shyed Basit R. Evaluation of DNA damage in lymphocytes of radiology personnel by comet assay. Journal of Occupational Health, (2015); 57(3): 268-274.

Azqueta A, Damian M, Elisa BR, Mirta M, Maria D, Gunnar B, Peter M, Andrew RC. Technical recommendations to perform the alkaline standard and enzyme-modified comet assay in human biomonitoring studies. Mutation Research Genetic Toxicology and Environmental Mutagens, (2019); 843: 24-32.

Siraj M, Muhammad K, Ajmal K, Farrah Z, Ahmad U, Ghani R. Bio-monitoring of Tissue Accumulation and Genotoxic Effect of Heavy Metals in Cyprinus carpio from River Kabul Khyber Pakhtunkhwa Pakistan. Bulletin of Environmental Contamination and Toxicology, (2018); 100(3): 344-349.

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

Jose S, Jayesh P, Mohandas A, Rosamma P, Singh ISB. Application of primary haemocyte culture of Penaeus monodon in the assessment of cytotoxicity and genotoxicity of heavy metals and pesticides. Marine Environmental Research, (2011); 71(3): 169-177.

Steel RGD, Torrie JH, Dinkely DA. Principles and procedures of statistics (3rd Ed.) McGraw Hill Book Co., Singapore. 1996.

Al-Mamun A. Pesticide Degradations, Research on Environmental Contamination, (2017); 87-102.

Amenyogbe E, Huang JS, Chen G. An overview of the pesticides’ impacts on fishes and humans. International Journal of Aquatic Biology, 2021; 9(1): 55-56.

Suliman, Ajmal K, Syed SAS, Naila G, Muhammad K, Muhammad Z. Toxicity evaluation of pesticide chlorpyrifos in male Japanese quails (Coturnix japonica). Environmental Science and Pollution Research, (2020); 27(20): 25353-25362.

Altinok I, Capkin E, Boran H. Mutagenic, genotoxic and enzyme inhibitory effects of carbosulfan in rainbow trout Oncorhynchus mykiss. Pesticide Biochemistry and Physiology, (2012); 102(1): 61-67.

Kapour D, Nagpure NS. Training on genotoxic assays in fishes. Kapour, D. and N.S. Nagpure (Eds). National Bureau of fish genetic resources. Dilknsha, Telibagh, India. (2005); 68.

Kousar S, Javed M. 2014. Assessment of DNA damage in peripheral blood erythrocytes of fish exposed to arsenic under laboratory conditions. International Journal of Current Microbiology and Applied Sciences, (2014); 3: 877-888.

Klobucar GIV, Stambuk A, Pavlica M, Peric MS, Kutuzovic B, Hylland K. Genotoxicity monitoring of freshwater environments using caged carp (Cyprinus carpio). Ecotoxicology, (2010); 19(1): 77-84.

Tope A, Rogers P. Evaluation of protective effects of sulforaphane on DNA damage caused by exposure to low levels of pesticide mixture using comet assay. Journal of Environmental Sciences and Health, (2009); 44(7): 657-662.

Liu Y, Zhang Y, Liu J, Huang D. The role of reactive oxygen species in the herbicide acetochlor-induced DNA damage on Bufo raddei tadpole liver. Aquatic Toxicology, (2006); 78(1): 21-26.

Guilherme S, Santos MA, Barroso C, Pacheco M. Differential genotoxicity of Roundup® formulation and its constituents in blood cells of fish (Anguilla anguilla): considerations on chemical interactions and DNA damaging mechanisms. Ecotoxicology, (2012); 21(5): 1381-1390.

Palanikumar L, Kumaraguru AK, Ramakritinan CM, Anand M. Toxicity, biochemical and clastogenic response of chlorpyrifos and carbendazim in milkfish Chanos chanos. International Journal of Environmental Sciences and Technology, (2014); 11(3): 765-774.

Jadhav TJ, Pawar SM. Organic pesticides effect on freshwater fish Barbus ticto. International Journal of Scientific and Engineering Research, (2018); 7(8): 1-2

Nwani C, Nagpure N, Kumar R, Kushwaha B, Lakra WS. DNA damage and oxidative stress modulatory effects of glyphosate-based herbicide in freshwater fish, Channa punctatus. Environmental Toxicology and Pharmacology, (2013); 36(2): 539-547.

Ilyas R. Effects of selected pesticides on growth and DNA damage in peripheral blood erythrocytes of fish. PhD. Thesis. Department of Zoology and Fisheries, University of Agriculture, Faisalabad, Pakistan. (2015); 1-329.

Ambreen F, Javed M. Pesticide Mixture Induced DNA Damage in Peripheral Blood Erythrocytes of Freshwater Fish, Oreochromis niloticus. Pakistan Journal of Zoology, (2018); 50(1): 339-346.



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