Erythrocytic and Nuclear Abnormalities in Natrix sp. from the Biga Stream (Çanakkale, Türkiye)

Yıl 2024, , 30 - 36, 30.06.2024

Nurşen Çördük , Begüm Boran , Bengi Baycan , İbrahim Uysal

https://doi.org/10.31594/commagene.1461253

Öz

The Grass Snake (Natrix natrix) and the Dice Snake (Natrix tessellata) are potential bioindicator water snake species in certain aquatic habitats. The aim of this study is to evaluate the frequency of erythrocytic abnormalities, immature erythrocyte index, and nuclear abnormalities in Natrix natrix and Natrix tessellata from Biga Stream in Çanakkale, Türkiye. Erythrocytic abnormalities and nuclear abnormalities such as micronuclei, lobbed nuclei, notched nuclei, blebbed nuclei, and binucleate cells were observed in peripheral blood smears for each sample. According to our results, total erythrocytic abnormalities in N. natrix (3.01±0.55%) were higher than in N. tessellata (2.52±0.32%). Similarly, the frequency of total nuclear abnormalities in N. natrix (13.40±2.7%) was higher than in N. tessellata (12.68±5.81%) but there was no statistically significant difference in total erythrocytic and nuclear abnormalities. However, student's t-test revealed that there was a statistically significant difference between the two species only in terms of immature erythrocyte index (p: 0.008) which was higher in N. natrix. Due to the results, it was suggested that N. natrix may exhibit more physiological responses to envirenmental stress factors.

Anahtar Kelimeler

Grass snake, dice snake, micronucleus test, ecotoxicology

Etik Beyan

The study was conducted with the permission of Çanakkale Onsekiz Mart University, Animal Experiments Local Ethics Committee (Permit no. 2017/04-16).

Biga Çayı (Çanakkale, Türkiye)’nda Dağılış Gösteren Natrix sp.’lerin Eritrositik ve Nüklear Anormallikleri

Öz

Yarı Sucul Yılan (Natrix natrix) ve Su Yılanı (Natrix tessellata), belirli sucul habitatlarda potansiyel biyoindikatör olan yılan türleridir. Bu çalışmanın amacı, Biga Çayı’nda (Çanakkale, Türkiye) bulunan Natrix natrix ve Natrix tessellata türlerinin eritrositik anormallik frekansı, olgunlaşmamış eritrosit indeksi ve nüklear anormallik frekansının belirlenmesidir. Her bir örnek için periferik kan yaymalarında, eritrositik anormallikler ve mikronükleus, loblu nükleus, çentikli nükleus, tomurcuklu nükleus, binükleat hücre gibi nüklear anormallikler gözlenmiştir. Sonuçlarımıza göre, N. natrix’teki toplam eritrositik anormallikler (%3.01±0.55) N. tessellata’dan (%2.52±0.32) daha yüksektir. Aynı şekilde N. natrix’teki toplam nüklear anormallik frekansının (%13.40±2.7) da N. tessellata’dan (%12.68±5.81) daha yüksek olduğu tespit edilmiştir fakat toplam eritrositik ve nüklear anormallikler bakımından istatistiksel olarak anlamlı bir fark bulunmamıştır. Ancak, student t testi ile iki tür arasında sadece olgunlaşmamış eritrosit indeksi (p: 0.008) açısından istatistiksel olarak anlamlı farklılık olduğu, N. natrix’de daha yüksek olduğu ortaya konmuştur. Bu sonuçlar doğrultusunda N. natrix’in çevresel stres faktörlerine karşı daha fazla fizyolojik tepkiler gösterebileceği düşünülmektedir.

Anahtar Kelimeler

Yarı sucul yılan, su yılanı, mikronükleus testi, ekotoksikoloji

Kaynakça

• Akbulut, M., Odabaşı, S.S., Odabaşı, D.A., & Çelik, E.Ş. (2006). Çanakkale İli'nin Önemli İçsuları ve Kirletici Kaynakları. Su Ürünleri Dergisi, 23(1), 9-15.
• Araki, T., Matsuhiko, H., & Saruta, T. (2006). Annion-exchange blocker enhances cytoplasmic vacuole formation and cell death in serum-deprived mouse kidney epithelial cells in mice. Cell Biology International, 30, 93–100. https://doi.org/10.1016/j.cellbi.2005.10.020
• Arıkan, H., & Çiçek, K. (2010). Morphology of peripheral blood cells from various species of Turkish herpetofauna. Acta Herpetologica, 5, 179–198.
• Ballard, B.M., & Cheek, R. (2003). Exotic animal medicine for the veterinary technician. Wiley and Sons / Blackwell Publishing, New York.
• Baran, İ., Ilgaz, Ç., Avcı, A., Kumlutaş, Y., & Olgun, K. (2012). Türkiye Amfibi ve Sürüngenleri. Tübitak Popüler Bilim Kitapları, 207, Semih Matbaacılık, Ankara. 204 p.
• Barni, S., Boncompagni, E., Grosso, A., Bertone, V., Freitas, I., Fasola, M., & Fenoglio, C. (2007). Evaluation of Rana snk esculenta blood cell response to chemical stressors in the environment during the larval and adult phases. Aquatic Toxicology, 81(1), 45-54. https://doi.org/10.1016/j.aquatox.2006.10.012
• Baycan, B., Boran, B., Gül, Ç., & Tosunoğlu, M. (2022). Clinical Hematology of the Nose-Horned Viper, Vipera ammodytes (Linnaeus 1758). Reptiles & Amphibians, 29(1), 461-469. https://doi.org/10.17161/randa.v29i1.16650
• Bolognesi, C., & Hayashi, M. (2011). Micronucleus assay in aquatic animals. Mutagenesis, 26, 205-213. https://doi.org/10.1093/mutage/geq073
• Campbell, T.W. (2006). Clinical pathology of reptiles, pp 453–470. In: D.R. Mader (ed.), Reptile Medicine and Surgery. Saunders, Philadelphia, PA.
• Canfield, P.J. (1998). Comparative cell morphology in the peripheral blood film from exotic and native animals. Australian Veterinary Journal, 76(12), 793–800. https://doi.org/10.1111/j.1751-0813.1998.tb12328.x
• Carrasco, K., Tilbury, K.L., & Myers, M.S. (1990). Assessment of the piscine micronucleus test as an in situ biological indicator of chemical contaminant effects. Canadian Journal of Fisheries and Aquatic Sciences, 47, 2123–2136. https://doi.org/10.1139/f90-237
• Erdoğan, D., & Tosunoğlu, M. (2017). Plasma biochemical parameters of Natrix natrix (Linnaeus, 1758; Squamata: Natricidae) Population in Çanakkale. Russian Journal of Herpetology, 24(1). https://doi.org/10.30906/1026-2296-2017-24-1-35-40
• Ergene, S., Cavas, T., Celik, A., Koleli, N., Kaya, F., & Karahan, A. (2007). Monitoring of nuclear abnormalities in peripheral erythrocytes of three fish species from the Goksu Delta (Turkey): genotoxic damage in relation to water pollution. Ecotoxicology, 16, 385-391. https://doi.org/10.1007/s10646-007-0142-4
• Fenech, M. (2000). The in vitro micronucleus technique. Mutation Research, 455, 81–95. https://doi.org/10.1016/S0027-5107(00)00065-8
• Frenzilli, G., Nigro, M., & Lyons, B.P. (2009). The Comet assay for the evaluation of genotoxic impact in aquatic environments. Mutation Research/Reviews in Mutation Research, 681(1), 80-92. https://doi.org/10.1016/j.mrrev.2008.03.001
• Guilherme, S., Válega, M., Pereira, M.E., Santos, M.A., & Pacheco, M. (2008). Erythrocytic nuclear abnormalities in wild and caged fish (Liza aurata) along an environmental mercury contamination gradient. Ecotoxicology and Environmental Safety, 70(3), 411-421. https://doi.org/10.1016/j.ecoenv.2007.08.016
• Gül, Ç., Hacıoğlu-Doğru, N., & Tosunoğlu, M. (2018). Impact of Some Hematological and Microbiological Factors on two Natrix Species in the Biga Stream (Çanakkale, Turkey). American Journal of Innovative Research and Applied Sciences, 6(4), 203-209.
• Gül, Ç., Tosunoğlu, M., Bilgin, H., Baycan, B., Boran, B., & Kurtul, D. (2019). Determination of the effect of body size on the hematological parameters in the species of Natrix tessellata (Tessellated water snake) distributed in Çanakkale (Turkey), I. International Symposium on Biodiversity Research, Çanakkale, Türkiye, 2-4 Mayıs 2019, pp. 317.
• Haskins, D.L., Brown, M.K., Bringolf, R.B., & Tuberville, T.D. (2021). Brown watersnakes (Nerodia taxispilota) as bioindicators of mercury contamination in a riverine system. Science of The Total Environment, 755, 142545. https://doi.org/10.1016/j.scitotenv.2020.142545.
• Heddle, J.A., & Countryman, R.I. (1976). The production of micronuclei from chromosome aberration in irradiated cultures of human lymphocytes. Mutation Research/Fundamental and Molecular Mechanism of Mutagenesis, 41, 321-332. https://doi.org/10.1016/0027-5107(76)90105-6
• Hojati, V., Faghiri, A., & Shiravi, A. (2012). Diet of the grass snake, Natrix natrix (Linnaeus, 1758) (Serpentes: Colubridae), in northern Iran. Zoology in the Middle East, 55(1), 132-134. https://doi.org/10.1080/09397140.2012.10648929
• IUCN. (2023). The IUCN Red List of Threatened Species. Version 2022-2. https://www.iucnredlist.org
• Josende, M. E., Tozetti, A. M., Alalan, M. T., Filho, V.M., Ximenez, S., Junior, F.M.R., & Martins, S.E. (2015). Genotoxic evaluation in two amphibian species from Brazilian subtropical wetlands. Ecological Indicators, 49, 83–87. https://doi.org/10.1016/j.ecolind.2014.10.007
• Koleva, V., Kornilev, Y., Telenchev, I., Lukanov, S., Hristova, B., & Natchev, N. (2017). Salt tolerance’s toll: prolonged exposure to saline water inflicts damage to the blood cells of dice snakes (Natrix tessellata). Web Ecology, 17(1), 1-7. https://doi.org/10.5194/we-17-1-2017
• Kornilev, Y. V., Popgeorgiev, G., Plachiyski, D., Dyugmedzhiev, A., Mladenov, V., Andonov, K., ... & Naumov, B. Distribution of the grass snake (Natrix natrix) and dice snake (N. tessellata) in Bulgaria. (2023). Historia Naturalis Bulgarica, 45(9), 239-254. https://doi.org/10.48027/hnb.45.093
• Kousar, S., & Javed, M. (2015). Studies on Induction of Nuclear Abnormalities in Peripheral Blood Erythrocytes of Fish Exposed to Copper. Turkish Journal of Fisheries and Aquatic Sciences, 15, 879-886. https://doi.org/10.4194/1303-2712-v15_4_11
• Lettoof, D.C., Cornelis, J., Jolly, C.J., Aubret, F., Gagnon, M.M., Hyndman, T.H., ... & Bateman, P.W. (2022). Metal (loid) pollution, not urbanisation nor parasites predicts low body condition in a wetland bioindicator snake. Environmental Pollution, 295, 118674. https://doi.org/10.1016/j.envpol.2021.118674
• Lisičić, D., Đikić, D., Benković, V., Knežević, A.H., Oršolić, N., & Tadić, Z. (2013). Biochemical and hematological profiles of a wild population of the nosehorned viper Vipera ammodytes (Serpentes: Viperidae) during autumn, with a morphological assessment of blood cells. Zoological Studies, 52, 1–9. https://doi.org/10.1186/s40409-017-0098-7.
• Manolis, S., Webb, G., & Britton, A. (2002). Crocodilians and Other Reptiles: Bioindicators of Pollution.
• Marcussi, S., Santos, P.R.S., Menaldo, D., Silveira L., Santos-Filho, N., Mazzi, M., Silva, S., Stábeli, R., Greggi Antunes, L., & Soares, A. (2011). Evaluation of the genotoxicity of Crotalus durissus terrificus snake venom and its isolated toxins on human lymphocytes. Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis, 724(1-2), 59-63. https://doi.org/10.1016/j.mrgentox.2011.06.004
• Minissi, S., Ciccotti, E., & Rizzoni, M. (1996). Micronucleus test in erythrocytes of Barbus plebejus (Teleostei, Pisces) from two natural environments: a bioassay for the in situ detection of mutagens in freshwater. Mutation Research, 367, 245–251. https://doi.org/10.1016/S0165-1218(96)90084-1.
• Morris, R.G., Hargreaves, A.D., Duvall, E., & Wyllie, A.H. (1984). Hormoneinduced cell death. 2. Surface changes in thymocytes undergoing apoptosis. American Journal of Pathology, 115, 426–436
• Mower, D.A., Peckham, D.W., Illera, V.A., Fishbaugh, J.K., Stunz, L.L., & Ashman, R.F. (1994). Decreased membrane phospholipids packing and decreased cell size precede DNA cleavage in mature mouse B cell apoptosis. Journal of Immunology, 152, 4832–4842. https://doi.org/10.4049/jimmunol.152.10.4832
• Napierska, D., Barŝienė, J., Mulkiewicz, E., Podolska, M., & Rybakovas, A. (2009). Biomarker responses in flounder Platichthys flesus from the Polish coastal area of the Baltic Sea and applications in biomonitoring. Ecotoxicology, 18, 846–859. https://doi.org/10.1007/s10646-009-0328-z
• Nascimento, F.F., Pereira, L.G., Geise, L., Bezerra, A.M., & D'Andrea, P.S. (2011). Colonization process of the Brazilian common vesper mouse, Calomys expulsus (Cricetidae, Sigmodontinae): a biogeographic hypothesis. Journal of Heredity, 102, 260–268. https://doi.org/10.1093/jhered/esr012
• O’Connor, J.S., & Dewling, R.T. (1986). Indices of marine degradation: their utility. Environmental Management, 10, 335–343. https://doi.org/10.1007/BF01867257
• Obiakor, M.O., Okonkwo, J.C., Nnabude, P.C., & Ezeonyejiaku, C.D. (2012). Eco-genotoxicology: Micronucleus assay in fish erythrocytes as in situ aquatic pollution biomarker: a review. Journal of Animal Science Advances, 2, 123-133.
• Oliveira-Júnior, A.A., Tavares-Dias, M., & Marcon, J.L. (2009). Biochemical and hematological reference ranges for Amazon freshwater turtle, Podocnemis expansa (Reptilia: Pelomedusidae), with morphologic assessment of bloodcells. Research in Veterinary Science, 86, 146–151. https://doi.org/10.1016/j. rvsc.2008.05.015.
• Pacheco, M., & Santos, M.A. (2002). Biotransformation, genotoxic, and histopathological effects of environmental contaminants in European eel (Anguilla anguilla L.). Ecotoxicology and Environmental Safety, 53, 331–347. https://doi.org/10.1016/s0147-6513(02)00017-9.
• Paoletti, M. G., & Sommaggio, D. (1996). Biodiversity indicators for sustainability. Assessment of rural landscapes. Bioindicator Systems for Soil Pollution, NATO ASI Series 16, 123-140.
• Peltzer, P.M., Lajmanovich, R.C., Attademo, A.M., Junges, C.M., Cabagna-Zenklusen, M.C., Repetti, M.R., Sigrist, M.E., & Beldomenico, H. (2013). Effect of exposure to contaminated pond sediments on survival, development, and enzyme and blood biomarkers in veined treefrog (Trachycephalus typhonius) tadpoles. Ecotoxicology Environmental Safety, 98, 142-151. https://doi.org/10.1016/j.ecoenv.2013.09.010
• Pollo, F.E., Grenat, P.R., Otero, M.A., Babini, S., Salas, N.E., & Martino, A.L. (2019). Evaluation in situ of genotoxic and cytotoxic response in the diploid/polyploid complex Odontophrynus (Anura: Odontophrynidae) inhabiting agroecosystems. Chemosphere, 216, 306-312. https://doi.org/10.1016/j.chemosphere.2018.10.149
• Salakij, C., Salakij, J., Suthunmapinunta, P., & Chanhome, L. (2002). Hematology, morphology and ultrastructure of blood cells and blood parasites from puff-faced watersnakes (Homalopsis buccata). Kasetsart Journal (Nat Sci), 36, 35–43.
• Saquib, Q., Al-Khedhairy, A.A., Siddiqui, M.A., Abou-Tarboush, F.M., Azam, A., & Musarrat, J. (2012). Titanium dioxide nanoparticles induced cytotoxicity, oxidative stress and DNA damage in human amnion epithelial (WISH) cells. Toxicology in Vitro, 26(2), 351-361. https://doi.org/10.1016/j.tiv.2011.12.011
• Scali, S. (2011). Ecological comparison of the Dice Snake (Natrix tessellata) and the Viperine Snake (Natrix maura) in northern Italy. Mertensiella, 18, 131-144.
• Sharma, R.C., & Rawat, J.S. (2009). Monitoring of aquatic macroinvertebrates as bioindicator for assessing the health of wetlands: a case study in the Central Himalayas, India. Ecological Indicators, 9, 118–128. https://doi.org/10.1016/j. ecolind.2008.02.004.
• Shimizu, N., Itoh, N., Utiyama, H., & Vahl, G.M. (1998). Selective entrapment of extrachromosomally amplified DNA by nuclear budding and micronucleation during the S-phase. Journal of Cell Biology, 140, 1307–1320. https://doi.org/10.1083/jcb.140.6.1307
• Siddig, A.A.H., Ellison, A.M., Ochs, A., Villar-Leeman, C., & Lau, M.K. (2016). How do ecologists select and use indicator species to monitor ecological change? Insights from 14 years of publication. Ecological Indicators, 60, 223–230. https://doi.org/10.1016/j.ecolind.2015.06.036.
• Speybroeck J., Beukema W., Bok B., Van Der Voort J., Velikov I. (2016). Field Guide to the Amphibians and Reptiles of Britain and Europe. Bloomsbury Publishing, London, 432 pp.
• Strunjak-Perovic, I., Lisicic, D., Coz-Rakovac, R., Topic Popovic, N., Jadan, M., Benkovic, V. & Tadic, Z. (2010). Evaluation of micronucleus and erythrocytic nuclear abnormalities in Balkan whip snake Hierophis gemonensis. Ecotoxicology, 19, 1460-1465. https://doi.org/10.1007/s10646-010-0531-y
• Šukalo G., Đorđević S., Gvozdenović S., Simović A., Anđelković M., Blagojević V., Tomović L. (2014). Intra- and inter-population variability of food preferences of two Natrix species on the Balkan peninsula. Herpetological Conservation and Biology, 9(1), 123−136.
• Thrall, A., Baker, D.C., Campbell, T.W., DeNicola, D., Feetman, M.J., Lassen, E.D., Rebar, A., & Weiser, G. (2004). Veterinary hematology and clinical chemistry. Blackwell Publishing. Philadephia, USA.
• Titenko-Holland, N., Windham, G., Kolachana, P., Reinisch, F., Parvatham, S., Osorio, A.M., & Smith, M.T. (1997). Genotoxicity of malathion in human lymphocytes assessed using the micronucleus assay in vitro and in vivo: a study of malathion-exposed workers. Mutation Research/Reviews in Mutation Research, 388(1), 85-95. https://doi.org/10.1016/S1383-5718(96)00140-4
• Tok, C. V., Tosunoğlu, M., Gül, Ç., Yiğini, B., Türkakın, M., Saruhan, G., & Kaya, S. (2006). Erythrocyte count and size in some colubrids (Reptilia: Ophidia) from Turkey. Russian Journal of Herpetology, 13(2), 97-100. https://doi.org/10.30906/1026-2296-2006-13-2-97-100
• Tok, C.V., Hayretdağ, S., Gürkan, M., & Yakın, B.Y. (2014). A preliminary study on micronuclei and nuclear abnormalities in the erythrocytes of some colubrid snakes from Turkey. Biharean Biologist, 8, 53–55.
• Tosunoğlu, M., Gül, Ç., Yılmaz, N., & Topyıldız, H. (2011). Hematological Reference Intervals of Some Snake Species in Turkey. Turkish Journal of Zoology, 35(2), 237-243. https://doi.org/10.3906/zoo-0905-9
• Valbona, A., Mihallaq, Q., & Evis, L. (2015). Grass snake, Natrix sp, as a potential bioindicator of freshwater ecosystem’s health. A case study in Erzeni River, Tirana, Albania. In International CAFE Conference, France.
• van der Horst, G., Walton, S., & Hofmeyr, M.D. (2013). Accurate automated quantitative imaging of tortoise erythrocytes using the NIS image analysis system. Biotechnic & Histochemistry, 88(5), 242-249. https://doi.org/10.3109/10520295.2013.765594
• Yermolenko, S.V., Gasso, V.Y., Hagut, A.M., Hasso, I.A., & Spirina, V.A. (2019). Bioaccumulation of heavy metals in the dice snake Natrix tessellata (Reptilia, Colubridae) from ecosystems of different anthropogenic loads in modern conditions of the steppe Dnieper area. Ecology and Noospherology, 30(2), 113-117. https://doi.org/10.15421/031919