NURAY ALTINTAŞ, MUSTAFA ÖZTATLICI, NAZMİYE ALTINTAŞ, Aysegul UNVER, Aslan SAKARYA

Molecular analysis of cattle Isolates of Echinococcus granulosus in Manisa province of Turkey

Echinococcus granulosus insanda ve birçok evcil hayvanda kistik ekinokokkozise (KE) neden olan etkendir ve hala dünyada ve Türkiyede en önemli sağlık problemlerinden biridir. Metasestodlarla infeksiyon şiddetli hastalıklara ve yüksek ekonomik kayıplara neden olur. Bazı Echinococcus suşları, suşların epidemiyolojik ve biyolojik karakteristiklerine dayanarak tanımlanmaktadır. Çalışmamızda sığırlardan elde edilen toplam 18 örnek incelenmiştir. Örnekler 2010-2012 yılları arasında Manisa merkez mezbahasından elde edilmiştir. Total genomik DNA (gDNA) üretici firmanın talimatları doğrultusunda protoskoleks ve kistik germinal membranlardan RTA-DNA İzolasyon Kiti kullanılarak (Gebze/Kocaeli, Türkiye) izole edilmiştir. Bu çalışmanın amacı, Türkiyede Manisa ilindeki sığırlardan elde edilen E. granulosus izolatlarının Polimeraz Zincir Reaksiyonu (PZR) ile moleküler karakterizasyonunun elde edilmesidir. PZRdan sonra, izolatların genetik karakteristiklerini araştırmak için mitokondrial sitokrom c oksidaz alt ünite 1 (CO1) ve nikotinamid adenin dinükleotit dehidrogenaz alt ünite 1 (NAD1) genleri deoksiribonükleik asit dizileme ile ABI Prism Genetik Analizör 3100 cihazıyla çalışıldı. Çalışmamızın sonucu olarak, tüm (18) sığır izolatları E. granulosus sensu stricto (G1-G3 kompleksi) olarak teşhis edildi. Bu çalışma Manisa ilindeki sığırlardan elde edilen Echinococcus izolatlarının ilk moleküler genotiplendirme çalışmasıdır.

Anahtar Kelimeler:

genler, genetik analizler, genom, genotip, helmintoz, parasitoz, DNA, Echinococcus granulosus, NAD, Türkiye, sığır, sığır hastalıkları, ekinokokozis, teknikler

Manisa ilinde Echinococcus granulosusun sığır İzolatlarının moleküler analizi

Echinococcus granulosus is the causative agent of cystic echinococcosis (CE) in humans and many domestic animals, and still one of the most important global health problem in the world and in Turkey. Infection with metacestode causes severe illness and high economic losses. Several strains of Echinococcus have been identified based on the epidemiological and biological characteristics of strains. In this study, a total of 18 individual hydatid cyst samples from cattle were examined. They were obtained from central slaughterhouse in the province of Manisa/Turkey between 2010-2012. The total genomic DNA (gDNA) was extracted using RTA-DNA Isolation Kit (Gebze/Kocaeli, Turkey) according to manufacturer instructions from protoscoleces and cystic germinal membranes. The aim of this study was to provide molecular characterization of E. granulosus isolates which were obtained from cattles by using polymerase chain reaction (PCR) in Manisa province of Turkey. After PCR, to investigate the genetic characteristics of isolates, deoxyribonucleic acid sequencing of the mitochondrial cytochrome c oxidase subunit 1 (CO1) and nicotinamide adenine dinucleotide dehydrogenase subunit 1 (NAD1) genes were performed with ABI Prism Genetic Analyzer 3100 instrument. As a result of our study, all (18) cattle isolates were detected as E. granulosus sensu stricto (G1-G3 complex). This is the first molecular study report genotyping of Echinococcus isolates from cattle in Manisa province.

Keywords:

genes, genetic analysis, genomes, genotypes, helminthoses, parasitoses, DNA, Echinococcus granulosus, NAD, Turkey, cattle, cattle diseases, echinococcosis, techniques,

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1. McManus DP: The molecular epidemiology of Echinococcus granulosus and cystic hydatid disease. Trans Roy Soc Med Hyg, 96, 151-157, 2002.
2. Eryıldız C, Sakru N: Molecular characterization of human and animal isolates of Echinococcus granulosus in the Thrace Region, Turkey. Balkan
3. nábel V, Altintas N, DAmelio S, Nakao M. Romig T, Yolasigmaz A, Gunes K, Turk M, Busi M, Hüttner M, evcová D, Ito A, Altintas N, Dubinský P: Cystic echinococcosis in Turkey: Genetic variability and first record of the pig strain (G7) in the country. Parasitol Res, 105, 145-154, 2009.
4. Altintas N: Parasitic zoonotic diseases in Turkey. Veterinaria Italiana, 44 (4): 633-646, 2008.
5. Altintas N: Past to present: Echinococcosis in Turkey. Acta Tropica, 85, 105-112, 2003.
6. Utuk AE, Simsek S, Koroglu E, McManus DP: Molecular genetic characterization of different isolates of Echinococcus granulosus in east and southeast reagions of Turkey. Acta Tropica, 107, 192-194, 2008.
7. Beyhan YE, Umur S: Molecular characterization and prevalence of cystic echinococcosis in slaughtered water buffaloes in Turkey. Vet Parasitol, 181, 174-179, 2011.
8. Utuk AE, Piskin FC, Dalkilic B: Molecular characterization of sheep isolates of Echinococcus granulosus in Kilis province. Kafkas Univ Vet Fak Derg, 18 (Suppl-A): A35-A38, 2012.
9. Yildiran FAB, Yildiz K, Cakir S, Gazyagci AN: The molecular characterization of Echinococcus granulosus isolates from sheep in Kırıkkale region. Kafkas Univ Vet Fak Derg, 16 (2): 245-250, 2010.
10. Ütük AE: Echinococcus granulosusun Doğu ve Güneydoğu Anadolu bölgesi izolatlarının moleküler ayrımı. Doktora Tezi, Fırat Üniv. Sağlık Bil. Enst., 2008.
11. Sharbatkhori M, Mirhendi H, Jex AR, Pangasa A, Campbell BE, Kia EB, Eshraghian MR, Harandi MF, Gasser RB: Genetic categorization of Echinococcus granulosus from humans and herbivorous hosts in Iran using an integrated mutation scanning-phylogenetic approach. Electrophoresis, 30, 2648-2655, 2009.
12. Acıöz M: Muş yöresinde köpeklerde PCR yöntemiyle, kesim hayvanlarında kesim takipleriyle ve insanlarda ELISA yöntemiyle Echinococcosis sıklığının araştırılması. Doktora Tezi, Cumhuriyet Üniv. Sağlık Bil. Enst., 2008.
13. Bowles J, Mc Manus DP: Molecular variation in Echinococcus. Acta Trop, 53, 291-305, 1993.
14. Bowles J, Blair D, McManus DP: Genetic variants within the genus Echinococcus identified by mitochondrial DNA sequencing. Mol Biochem Parasitol, 54, 165-174, 1992.
15. Mwambete KD, Ponce-Gordo F, Cuesta-Bandera C: Genetic identification and host range of the Spanish strains of Echinococcus granulosus. Acta Trop, 91, 87-93, 2004.
16. Schneider R, Gollackner B, Edel B, Schmid K, Wrba F, Tucek G, Walochnik J, Auer H: Development of a new PCR protocol for the detection of species and genotypes (strains) of Echinococcus in formalin-fixed, parafin-embedded tissues. Int J Parasitol, 38, 1065-1071, 2008.
17. Varcasia A, Canu S, Lightowlers MW, Scala A, Garippa G: Molecular characterization of Echinococcus granulosus strains in Sardinia. Parasitol Res, 98, 273-277, 2006.
18. Mrad S, Fiisetti D, Oudni M, Mekki M, Belguith M, Nouri A, Sayadi T, Lahmar S, Candolfi E, Azaiez R, Mezhoud H, Baba H: Molecular evidence of ovine (G1) and camel (G6) strains of Echinococcus granulosus in Tunisia and putative role of cattle in human contamination. Vet Parasitol, 129, 267-272, 2005.
19. Breyer I, Georgieva D, Kurdova R, Gottstein B: Echinococcus granulosus strain typing in Bulgaria: The G1 genotype is predominant in intermediate and definitive wild hosts. Parasitol Res, 93, 127-130, 2004.
20. Bağcı Ö, Vural G, Baca AÜ, Gauci C: Türkiyede koyunlarda cytochrom c oxidase subunit 1 gen analizi ile Echinococcus granulosus genotipinin belirlenmesi. TÜBİTAK Proje No: 104 V 132, 2007.
21. Simsek S, Balkaya I, Koroglu E: Epidemiological survey and molecular characterization of Echinococcus granulosus in cattle in an endemic area of eastern Turkey. Vet Parasitol, 172, 347-349, 2010.
22. Simsek S, Eroksuz Y: Occurrence and molecular characterization of Echinococcus granulosus in Turkish mouflon (Ovis gmelinii anatolica). Acta Tropica, 109, 167-169, 2009.
23. Vural G, Baca AU, Gauci CG, Bagci O, Gicik Y, Lightowlers MW: Variability in the Echinococcus granulosus cytochrome c oxidase 1 mitochondrial gene sequence from livestock in Turkey and a re-appraisal of the G1-3 genotype cluster. Vet Parasitol, 154, 347- 350, 2008.
24. Utuk AE, Simsek S: Molecular characterization of the horse isolate of Echinococcus granulosus in Turkey. J Helminthol, 1 of 4 doi:10.1017/S0022149X12000363 Cambridge University Press, 2012.