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10217904BB.Pdf T.C. ORDU ÜNİVERSİTESİ FEN BİLİMLERİ ENSTİTÜSÜ MELET IRMAĞI SU, SEDİMENT VE BAZI BALIK TÜRLERİNDE AĞIR METAL BİRİKİMİ VE GENOTOKSİK ETKİLERİNİN ARAŞTIRILMASI SEDA KONTAŞ DOKTORA TEZİ MOLEKÜLER BİYOLOJİ VE GENETİK ANABİLİM DALI ORDU 2018 T.C. ORDU ÜNİVERSİTESİ FEN BİLİMLERİ ENSTİTÜSÜ MOLEKÜLER BİYOLOJİ VE GENETİK ANABİLİM DALI MELET IRMAĞI SU, SEDİMENT VE BAZI BALIK TÜRLERİNDE AĞIR METAL BİRİKİMİ VE GENOTOKSİK ETKİLERİNİN ARAŞTIRILMASI SEDA KONTAŞ DOKTORA TEZİ ORDU 2018 ÖZET MELET IRMAĞI SU, SEDİMENT VE BAZI BALIK TÜRLERİNDE AĞIR METAL BİRİKİMİ VE GENOTOKSİK ETKİLERİNİN ARAŞTIRILMASI SEDA KONTAŞ ORDU ÜNİVERSİTESİ FEN BİLİMLERİ ENSTİTÜSÜ MOLEKÜLER BİYOLOJİ VE GENETİK ANABİLİM DALI DOKTORA TEZİ 256 SAYFA (TEZ DANIŞMANI: PROF. DR. DERYA BOSTANCI) Melet Irmağı Orta ve Doğu Karadeniz bölümleri arasında doğal bir sınır oluşturmaktadır. Jeolojik konumundan, maden yataklarınca zengin bir bölgede oluşundan, evsel atıklardan ve fındık tarımınında kullanılan pestisitlerden kaynaklı ağır metaller, ırmağı kirlilik açısından tehdit etmektedir. Bu çalışmada, Melet Irmağı üzerinde belirlenen Mahmudiye, Kıranyağmur, Kocaali ve nehir ağzı istasyonlarından, ilkbahar, yaz, sonbahar ve kış mevsimlerinde toplanan su, sediment ve balık örneklerinin kas, solungaç ve karaciğerlerindeki bazı elementlerin (Al, Ni, As, Cr, Cd, Pb, Mn, Fe, Co, Cu, Zn) konsantrasyonları belirlenmiştir. İstasyonların su ve sediment numunelerindeki element konsantrasyonlarının genotoksik potansiyellerinin belirlenmesinde, yakalanan balıkların eritrosit hücreleri kullanılarak, mikronukleus (MN) testi ve comet analizi yapılmıştır. İstasyonlardan alınan su örneklerinde yaz mevsiminde, arsenik elementinin Mahmudiye (As: 22.223 µg/L) ve nehir ağzı (As: 20.951 µg/L) istasyonunda; sonbahar mevsiminde alüminyumun, nehir ağzı (Al: 1332.788 µg/L) istasyonunda; demirin, Kocaali (Fe: 451.372 µg/L) ve nehir ağzı (Fe: 3091.824 µg/L) istasyonlarında; arseniğin ise Mahmudiye (As: 27.230 µg/L), Kıranyağmur (As: 20.398 µg/L), Kocaali (As: 26.908 µg/L) ve nehir ağzı (As: 37.140 µg/L) istasyonlarında yüksek miktarda olduğu tespit edilmiştir. Kış mevsiminde su içeriğinde, çinko Kocaali (Zn: 255.580 µg/L) istasyonunda ve arsenik Kıranyağmur (As: 20.035 µg/L) ve nehir ağzı (As: 20.664 µg/L) istasyonlarında su kalite kriterlerinde belirtilen değerlerinden daha yüksektir. İstasyonlara ait sediment örneklerinde çinko, kadmiyum ve kurşunun bazı mevsim ve istasyonlarda yer kabuğu değerlerinden yüksek olduğu saptanmıştır. Yaz mevsiminde çinko ve kadmiyum sırasıyla Kocaali istasyonunda (Zn: 99.38 µg/g ve Cd: 0.46 µg/g), sonbaharda kadmiyum Kocaali (Cd: 0.47 µg/g) ve nehir ağzı (Cd: 0.31 µg/g) istasyonlarında, kurşun ise Kocaali (Pb: 22.46 µg/g) istasyonunda limit değerlerin üzerinde belirlenmiştir. Balıkların kas dokusunda sadece kurşun miktarının tüm istasyonlarda bazı mevsimlerde yüksek olduğu görülmektedir. Mahmudiye istasyonundan yakalanan Capoeta banarescui bireylerinin kas dokusunda belirlenen element seviyeleri incelendiğinde, kurşun miktarlarının sonbahar (1.53 mg/kg) ve kış (0.49 mg/kg) mevsimlerinde kas dokuda yüksek olduğu tespit edilmiştir. Kıranyağmur istasyonundan yakalanan Vimba vimba bireylerinin kas dokusunda belirlenen kurşun miktarları yaz (1.53 mg/kg) ve sonbahar (0.53 mg/kg) mevsimlerinde; Kocaali istasyonundaki Alburnus chalcoides bireyleri için yaz (1.13 mg/kg), sonbahar (0.45 mg/kg) ve kış (0.51 mg/kg) mevsimlerinde limit değerlerin üzerindedir. Nehir ağzı istasyonundaki A. chalcoides bireyleri için kurşun miktarı ilkbahar (0.60 mg/kg), sonbahar (0.47 mg/kg) ve kış (0.93 mg/kg) mevsimlerinde kas dokuda yüksek bulunmuştur. Mikronukleus (MN), herhangi bir olumsuz çevre etkisi olmadığı durumlarda hücre çekirdeğinde meydana gelmesi beklenmeyen bir oluşumdur. Balık eritrositlerinde gözlemlenen mikronükleus oluşumları, hücrede kromozomal bir hasarın olduğunu II belirtmektedir. Yapılan değerlendirmelerde, mikronukleus oluşumu, Melet Irmağı balıklarında dört mevsimde de farklı frekanslarda bulunmuştur. Dört istasyonda da balıklarda en yüksek MN frekansı yaz mevsiminde (Mahmudiye: 2.20, Kıranyağmur: 5.20, Kocaali: 5.40, nehir ağzı: 9.60) ve en düşük MN frekansı ise kış mevsiminde (Mahmudiye: 0.40, Kıranyağmur: 1.10, Kocaali: 0.80, nehir ağzı: 1.90) tespit edilmiştir. Ayrıca, comet analizi kullanılarak balıkların eritrosit hücrelerinin çekirdeğindeki DNA zincirinde meydana gelen kırılmaların da tespiti gerçekleştirilmiştir. Bu çalışmada, eritrosit hücreleri üzerinde kuyruk boyu, kuyruk yoğunluğu, baş yoğunluğu, kuyruktaki % DNA, baştaki % DNA ve kuyruk momenti parametrelerinin ölçümleri yapılmıştır. Kuyruk boyu değeri, A. chalcoides’de en fazla yaz (29.11 µm) ve sırasıyla ilkbahar (28.97 µm), sonbahar (28.68 µm) ve kış (28.35 µm) mevsimlerinde nehir ağzında ölçülmüştür. Aynı istasyonda kuyruktaki % DNA değerleri sırasıyla 40.22 (yaz), 39.33 (ilkbahar), 39.06 (sonbahar) ve 38.01 (kış); kuyruk momenti ise 0.531 (yaz), 0.528 (ilkbahar), 0.516 (sonbahar), 0.498 (kış) olarak belirlenmiştir. C. banarescui’da kuyruktaki % DNA’nın en düşük değeri kış mevsiminde 11.51 olarak belirlenirken, ilkbahar, yaz ve sonbahar mevsimleri için sırasıyla 29.91, 34.04 ve 29.40 olarak hesaplanmıştır. Baştaki % DNA değerleri ise büyükten küçüğe sırasıyla kışın 88.49, sonbaharda 70.60, ilkbaharda 70.09 ve yazın 65.96 olarak hesaplanmıştır. Kuyruktaki % DNA ve kuyruk boyu değerlerinin yaz mevsiminde en yüksek değerde, kış mevsiminde ise en düşük değerde olduğu belirlenmiştir. DNA hasarının miktarını gösteren kuyruk boyu (27.75 µm), kuyruktaki % DNA (39.02) ve kuyruk momenti (0.470) değerleri Kocaali istasyonu bireylerinde yaz mevsiminde diğer mevsimlere göre en yüksek değerlerdedir. Melet Irmağı’nın genotoksik potansiyeli daha önce hiç bir çalışmada değerlendirilmemiştir. Yapılan mikronükleus testi ve comet analizi, ırmakta konsantrasyonu belirlenen elementlerin yanı sıra, ırmağın jeolojik konumu, maden yataklarınca zengin bir bölgede oluşu ve fındık tarımının yapıldığı arazilerde kullanılan pestisitler gibi diğer etkenlerden dolayı da, ırmağın genotoksik bir potansiyele sahip olduğunu göstermektedir. Bu çalışma, Ordu ilinde bulunan Melet Irmağı’nda yayılış gösteren bazı balık türlerinde yapılan ilk genotoksisite çalışmasıdır. Gerçekleştirilen bu çalışmada, hem Melet Irmağı hem de Karadeniz ve tüm Türkiye’deki akarsu sistemleri için genotoksisite çalışmalarına ışık tutacak veriler sunulmuştur. Melet Irmağı’nın genotoksisite değerlendirmesinin yapıldığı bu çalışmada, mikronukleus testi ve comet analizi sonuçları birbirini desteklemiştir. Anahtar Kelimeler: Ağır Metal, Comet Analizi, Cyprinidae, Çekirdek Anormallikleri, DNA Hasarı, Genotoksisite, Kirleticiler, Melet Irmağı, Mikronukleus, Toksisite. III ABSTRACT ACCUMULATION OF HEAVY METALS IN WATER, SEDIMENT AND SOME FISH SPECIES OF MELET RIVER AND INVESTIGATION OF GENOTOXIC EFFECT SEDA KONTAŞ ORDU UNIVERSITY INSTITUTE OF NATURAL AND APPLIED SCIENCES MOLECULAR BIOLOGY AND GENETIC DOCTORATE THESIS, 256 PAGES (SUPERVISOR: PROF. DR. DERYA BOSTANCI Melet River creates a natural border between the Central and Eastern Black Sea regions. Geological location of river, the formation of rich region of mineral deposits and heavy metals from domestic wastes and pesticides used in hazelnut agriculture threaten the river in terms of pollution. In this study, some elements concentrations (Al, Ni, As, Cr, Cd, Pb, Mn, Fe, Co, Cu, Zn) were determined in water, sediment and muscle, gill and liver of fish samples collected in spring, summer, autumn and winter seasons from the Mahmudiye, Kıranyağmur, Kocaali and river mouth stations determined on Melet River. Micronucleus (MN) test and comet analysis were performed using the erythrocyte cells of the captured fish to determine the genotoxic potentials of the element concentrations in water and sediment samples of the stations. The values of arsenic were high in Mahmudiye (As: 22.223 μg / L) and river mouth (As: 20.951 μg / L) in the summer; aluminium was high in river mouth (Al: 1332.788 µg/L); iron was high in Kocaali (Fe: 451.372 µg/L) and river mouth (Fe: 3091.824 µg/L) in the autumn and As was high for Mahmudiye (As: 27.230 µg/L), Kıranyağmur (As: 20.398 µg/L), Kocaali (As: 26.908 µg/L) and river mouth (As: 37.140 µg/L) stations in autumn for water samples from stations. Zn value was higher for Kocaali (Zn: 255.580 µg/L) station, As values were higher for Kıranyağmur (As: 20.035 µg/L) and river mouth (As: 20.664 µg/L) stations than the water quality criteria values in winter. In the sediment samples belonging to the stations, zinc, cadmium and lead were higher than earth's crust value at some seasons and stations. Zinc and cadmium at the Kocaali station (Zn: 99.38 µg/g ve Cd: 0.46 µg/g) in summer, cadmium at Kocaali (Cd: 0.47 µg/g) and River Mouth (Cd: 0.31 µg/g) stations, and also the lead at Kocaali station (Pb: 22.46 µg/g) in the autumn were determined above the limit values. It is observed that only the amount of lead in muscles of fish is high in all stations at some seasons. When the levels of elements determined in the muscle tissue of Capoeta banarescui individuals from Mahmudiye station were examined, it was determined that lead amounts were high in autumn (1.53 mg/kg) and winter (0.49 mg/kg) seasons. The lead amount in muscle tissue were above the limit values in the summer (1.53 mg/kg) and autumn (0.53 mg/kg) seasons for Vimba vimba individuals caught from the Kıranyağmur station and in the summer (1.13 mg/kg), autumn (0.45 mg/kg) and winter (0.51 mg/kg) seasons for Alburnus chalcoides individuals in Kocaali station.
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