Parasitology Research

, Volume 108, Issue 4, pp 837–843 | Cite as

Identification of different Theileria species (Theileria lestoquardi, Theileria ovis, and Theileria annulata) in naturally infected sheep using nested PCR–RFLP

  • Mahdieh Zaeemi
  • Hamidreza Haddadzadeh
  • Parvaneh Khazraiinia
  • Bahram Kazemi
  • M. Bandehpour
Original Paper


Ovine theileriosis is an important hemoprotozoal disease of sheep and goats in tropical and subtropical regions that leads to economic losses in these animals. A nested PCR–restriction fragment length polymorphism (RFLP) was carried out to identification Theileria species in sheep in some area in western half of Iran (Sari, Rasht, Urmia, Ilam, and Ahvaz). Two hundred and fifty blood samples were taken from sheep during tick activating season (summer of 2008). Microscopic examination revealed that 9.2% (23/250) sheep were infected by Theileria spp. piroplasms. Parasitemia ranged from 0.011% to 0.015%. In nested PCR assessment of DNA samples, 32.8% (82/250) sheep were positive. The negative samples were confirmed by amplifying of ovine beta-actin gene as an internal control. The differentiation of Theileria species was based on RFLP patterns using three restriction enzymes: HpaII, Rsa1, and Bsh 1285I. Out of 82 positive samples, 54.8% (45/82) and 40.2% (33/82) were positive for Theileria lestoquardi and Theileria ovis respectively. Mixed infection was detected in 4.8% (4/82) cases. Based on their PCR product digestion pattern with HpaII (1178, 900, 278, and 106 bp), it seemed to be mixture of Theileria annulata and T. lestoquardi. The presence of T. annulata was supported by sequence analysis. This is the first report of naturally infected sheep with T. annulata in Iran. Geographical distribution of Theileria species in sheep is shown according to the result of microscopy and nested PCR and RFLP data.


Restriction Fragment Length Polymorphism Blood Smear Mixed Infection Mean Annual Temperature Restriction Fragment Length Polymorphism Pattern 



This study was supported financially by the Tehran University. We acknowledged also the Center of Excellence of Pathobiology, the Center of Excellence on Veterinary Research on Iranian Indigenous Animals, and the Iranian Research Center for Ticks and Tick-borne Diseases (IRCTTD) for their support.


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Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Mahdieh Zaeemi
    • 1
  • Hamidreza Haddadzadeh
    • 2
  • Parvaneh Khazraiinia
    • 1
  • Bahram Kazemi
    • 3
  • M. Bandehpour
    • 3
  1. 1.Department of Clinical Science, Faculty of Veterinary MedicineUniversity of TehranTehranIran
  2. 2.Department of Parasitology, Faculty of Veterinary MedicineUniversity of TehranTehranIran
  3. 3.Cellular and Molecular Biology Research Center, Department of Parasitology, School of MedicineShaheed Beheshti University of Medical SciencesTehranIran

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