Molecular survey of Anaplasma phagocytophilum in small ruminants from Şanlıurfa

Öz

This study was performed to determine the presence and prevalence of Anaplasma phagocytophilum in sheep and goats in the Şanlıurfa region. Whole blood samples were taken from a total of 208 animals (134 sheep and 74 goats) in the Şanlıurfa region. Polymerase chain reaction (PCR) using primers specific to the A. phagocytophilum 16S SSU rRNA gene of the bacterium was applied to the blood samples. The amplified DNA fragments in PCR positive samples were sequenced in order to confirm their identity. PCR amplification indicated the presence of A. phagocytophilum in 9 animals (4.33 %). A. phagocytophilum was present in 5.97 % (8/134) of sheep and 1.35 % (1/74) of goats. The nucleotide sequence of a sample selected to represent the PCR positive samples was determined. The resulting DNA sequence was compared to other DNA sequences available in the GenBank using BLAST analysis and placed into GenBank under accession number as KJ183079. The presence of A. phagocytophilum in the sheep and goats in the Şanlıurfa region was demonstrated for the first time using PCR and sequence analyses and epidemiologic data regarding the pathogen were also obtained.

Anahtar Kelimeler

• Anaplasma phagocytophilum, goat, PCR, sheep, Şanlıurfa

Şanlıurfa yöresinde koyun ve keçilerde Anaplasma phagocytophilum’un moleküler yöntemlerle araştırılması

Öz

Bu araştırma, Şanlıurfa yöresinde koyun ve keçilerde Anaplasma phagocytophilum’un varlığı ve yaygınlığının belirlenmesi amacıyla yapılmıştır. Bu amaçla, Şanlıurfa yöresinde 134 koyun ve 74 keçi olmak üzere toplam 208 hayvandan tam kan örneği alınmıştır. Bu örneklerden total genomic DNA’lar izole edilmiş, elde edilen DNA örnekleri Anaplasma phagocytophilum’un varlığı yönünden 16S SSU rRNA genine spesifik primerler kullanılarak, polimeraz zincir reaksiyonu (PZR)’na tabi tutulmuştur. Polimeraz zincir reaksiyonu sonucunda pozitif olduğu belirlenen örnekleri temsilen seçilen bir örneğin 16S SSU rRNA geninin DNA dizilimi belirlenmiştir. İncelenen 208 koyun ve keçiye ait kan örneğinin 9 (% 4.33)’unda A. phagocytophilum’un varlığını gösteren PZR amplifikasyon ürünü elde edilmiştir. A. phagocytophilum pozitiflik oranının koyunlarda % 5.97 (8/134), keçilerde % 1.35 (1/74) olduğu saptanmıştır. Pozitif örnekleri temsilen seçilen bir örneğinin DNA dizilimi belirlenmiş, bu dizilim BLAST analiz yoluyla GenBankasındaki diğer dizilimlerle karşılaştırıldıktan sonra GenBanka KJ183079 accession numarası ile kayıt edilmiştir. Bu çalışma ile Şanlıurfa yöresindeki koyun ve keçilerde PZR ve sekans analizi ile A. phagocytohilium varlığı ilk defa ortaya konmuş ve etkene ait epidemiyolojik veriler de ilk olarak elde edilmiştir

Anahtar Kelimeler

• Anaplasma phagocytophilum, koyun, keçi, PZR, Şanlıurfa

Kaynakça

1. Dumler, J.S. (2005). Anaplasma and Ehrlichia Infection. Ann NY Acad Sci, 1063, 361-373.
2. Woldehiwet, Z. (2010). The natural history of Anaplasma phagocytophilum. Vet Parasitol, 167, 108-122.
3. McQuiston, J.H., Paddock, C.D., Holman, R.C., Childs, J.E. (1999). Human ehrlichiosis in the United States. Emerg Infect Dis, 5, 635-642.
4. Lotric-Furlan, S., Petrovec, M., Zupanc, T.A., Nicholson, W.L., Sumner, J.W., Childs, J.E., Strle, F. (1998). Human granulocytic ehrlichiosis in Europe: clinical and laboratory findings for four patients from Slovenia. Clin Infect Dis, 27, 424-428.
5. Dumler, J.S., Barbet, A.F., Bekker, C.P., Dasch, G.A., Palmer, G.H., Ray, S.C., Rikihisa, Y., Rurangirwa, F.R. (2001). Reorganization of genera in the families Rickettsiaceae and Anaplasmataceae in the order Rickettsiales: unification of some species of Ehrlichia with Anaplasma, Cowdria with Ehrlichia and Ehrlichia with Neorickettsia, descriptions of six new species combinations and designation of Ehrlichia equi and 'HGE agent' as subjective synonyms of Ehrlichia phagocytophila. Int J Syst Evol Micr, 51, 2145-2165
6. Dumler, J.S., Madigan, J.E., Pusterla, N., Bakken, J.S. (2007). Ehrlichiosis in humans epidemiology, clinical presentation, diagnosis and treatment. Clin Infect Dis, 45(1), 45-51.
7. Alessandra, T., Caracappa, S. (2012). Tick-borne diseases in sheep and goats : Clinical and diagnostic aspects. Small Ruminant Res, 1065, 6-11.
8. Karatepe, M., Karatepe, B. (2013). Anaplasmosis. Parazit Hastalıkları, Edr., İnci, A., Köroğlu, K., Karaer, Z., Eren, H., Yukarı, B.A., Dumanlı, N., Aydın, L., Yıldırım, A. 812-815, Türkiye Parazitoloji Derneği, İzmir.

9. Leah, A.C. (2014). Ehrlichiosis and related infections. Vet Clin N Am-Small, 33, 863-884.
10. Aktas, M. (2014). A survey of ixodid tick species and molecular identification of tick-borne pathogenns. Vet Parasitol, 200 (3-4), 276-283.
11. Altay, K., Dumanli, N., Aktas, M., Ozübek, S. (2014). Survey of Anaplasma infections in small ruminants from East part of Turkey. Kafkas Univ Vet Fak Derg, 20(1), 1-4.
12. d’Oliveria, C., van der Weide, M., Habela, M.A., Jacquiet, P., Jongejan, F. (1995). Detection of Theileria annulata in blood by PCR. J Clin Microbiol, 33, 2665-2669.
13. Dumanli, N., Aktas, M., Cetinkaya, B., Cakmak, A., Koroglu, E., Saki, C.E., Erdogmus, Z., Nalbantoglu, S., Ongör, H., Simsek, S., Karahan, M., Altay, K. (2005). Prevalence and distribution of tropical theileriosis in eastern Turkey. Vet Parasitol, 127(1), 15-21.
14. Hoşgör, M, Bilgiç, H.B, Bakırcı, S, Ünlü, A.H., Karagenç, T., Eren, H. (2015). Aydın yöresinde sığırlarda ve kenelerde Anaplasma/Ehrlichia türlerinin belirlenmesi. Türkiye Parazitol Derg, 39, 291-298.
15. Kawahara, M., Rikihisa, Y., Lin, Q., Isogai, E., Tahara, K., Itagaki, A., Hiramitsu, Y., Talima, T. (2006). Novel genetic variants of Anaplasma phagocytophilum, Anaplasma bovis, Anaplasma centrale, and a novel Ehrlichia sp. in wild deer and ticks on two major islands in Japan. Appl Environ Microb, 72, 1102-1109.
16. Derdakova, M., Stefancikova, A., Spitalska, E., Tarageova, V., Kostalova, T., Hrklova, G., Kybicova, K., Schanilec, P., Majlathova, V., Varady, M., Petko, B. (2011). Emergence and genetic variability of Anaplasma species in small ruminants and ticks from Central Europe. Vet Microbiol, 153, 293-298.
17. Gökçe, H.I., Genç, O., Akça, A., Vatansever, Z., Unver, A., Erdoğan, H.M. (2008). Molecular and serological evidence of Anaplasma phagocytophilum infection farm animals in the Black Sea Region of Turkey. Acta Vet Hung, 56(3), 281-292.
18. Grova, L., Olesen, I., Steinshamn, H., Stuen, S. (2011). Prevalence of Anaplasma phagocytophilum infection and effect on lamb growth. Acta Vet Scand, 53, 30-36.
19. Lillini, E., Macri, G., Proietti, G., Scapulla, M. (2006). New findings on anaplasmosis caused by infection with Anaplasma phagocytophilum. Ann NY Acad Sci, 1081, 360-370.
20. Stuen, S., Bergström, K. (2001). Serological investigation of granulocytic Ehrlichia infection in sheep in Norway. Acta Vet Scand,42, 331-338.
21. Zhan, L., Cao, W.C., Jiang, J.F., Zhang, X.A., Wu, X.M., Zhang, W.Y., Liu, W., Zuo, S.Q., Cao, Z.W., Yang, H., Richardus, J.H., Habbema, J.D. (2010). Anaplasma phagocytophilum in livestoc k and small rodents. Vet Microbiol, 144, 405-408.
22. Aktas, M., Vatansever, Z., Altay, K., Aydin, M.F., Dumanli, N. (2010). Molecular evidence for Anaplasma phagocytophilum in Ixodes ricinus from Turkey. T Roy Soc Trop Med H, 104: 10-15.
23. Selçuk, Ö., Alver, O., Çatık, S., Aydın, L., Şenlik, B. (2015). Determination of diagnostic value of cELISA for the diagnosis of Anaplasma in clinically suspected ruminants. Kafkas Univ Vet Fak Derg, 21 (5), 691-695.
24. Bekker, C.P., de Vos, S., Taoufik, A., Sparagano, O.A., Jongejan, F. (2002). Simultaneus detection of Anaplasma and Ehrlichia species in ruminants and detection of Ehrlichia ruminantum in Amblyomma variegatum ticks by reverse line blot hybridization. Vet Mic, 89(2-3), 223-228.
25. Schouls, L.M., Van de Pol, I., Rijpkema, S.G., Schot, C.S. (1999). Detection and identification of Ehrlichia, Borrelia burgdorferi sensu lato, and Bartonella species in Dutch Ixodes ricinus ticks. J Clin Microbiol, 37(7), 2215-2222.
26. Pfaffle, M., Littwin, N., Muders, S.V., Petney, T.V. (2013). The ecology of tick-borne diseases. Int J Parasitol, 43(12-13), 1059-1077.
27. Aydın, L., Bakırcı, S. (2007). Geographical distribution of ticks in Turkey. Parasitol Res, 101, 163-166.
28. Dumanlı, N., Altay, K., Aydın, M.F. (2012). Türkiye’de sığır, koyun ve keçilerde belirlenen kene türleri. Türkiye Klinikleri Vet Bilim Derg, 3(2), 67-72.
29. Karaer, Z., Yukarı, B.A., Aydın, L. (1997). Türkiye keneleri ve vektörlükleri. Parazitolojide Artropod Hastalıkları ve Vektörler, Edr., Özcel, M.A., Daldal N., 363-433, Türkiye Parazitoloji Derneği, İzmir.
30. Ogden, N.H., Casey, A.N.J., French, N.P., Woldehiwet, Z. (2002). Review of studies on the transmission of Anaplasma phagocytophilum from sheep: implications for the force of infection in endemic cycles. Exp and Appl Acarol, 28(1-4), 195-202.
31. Richter, P., Kimsey, R., Madigan, J., Barlough, J.E., Dumler, J.S., Brooks, D.L. (1996). Ixodes pacificus (Acari: Ixodiddae) as vector of Ehrlichia equi (Rickettsiales: Ehrlichieae). J Med Entomol, 33, 1–5.
32. Aktas, M., Altay, K., Dumanli, N. (2011). Molecular detection and identification of Anaplasma and Ehrlichia species in cattle from Turkey. Ticks Tick Borne Dis, 2, 62-66.
33. Stuen, S. (2013). Tick-borne infections in small ruminants in northern Europe. Small Ruminant Res, 110, 142-144.
34. Öter, K., Çetinkaya, H., Vuruşaner, C., Toparlak, M., Ergünay, K. (2015). Molecular detection and typing of Anaplasma species in small ruminants in Thrace region of Turkey. Kafkas Univ Vet Fak Derg, 22 (1), 133-138.