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Multiplex PCR for diagnosis of Theileria uilenbergi, Theileria luwenshuni, and Theileria ovis in small ruminants

Parasitology Research volume 113, pages 527–531 (2014)Cite this article

Abstract

Infections with Theileria sp. may cause significant economic losses to the sheep industry. Species identification based on microscopic examination is difficult, and more suitable methods are required for the rapid detection and identification of Theileria sp, in clinical specimens. In this study, a multiplex polymerase chain reaction (mPCR) assay was developed to simultaneously identify three individual Theileria species in small ruminants. Three pairs of specific, sensitive primers were designed on the basis of the 5.8S ribosomal RNA gene (Theileria luwenshuni and Theileria ovis) and the 18S ribosomal RNA gene (Theileria uilenbergi) to generate target products of 303, 884, and 530 bp, respectively. Standard DNA for each of the three species was extracted from blood recovered from infected sheep, and a preliminary study was conducted on 56 sheep to verify the reliability of the system. Optimal PCR conditions, including primer concentration, annealing time, and the number of amplification cycles, were established. The assay sensitivity under these conditions was 10−3 % parasitemia, and its specificity was 100 %. The results of the study suggest that mPCR represents a simple, efficient test method as a practical alternative for the rapid detection and identification of Theileria species in small ruminants.

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Acknowledgments

This study was financially supported by the NSFC (no. 31272556, 30972182, 31072130, and 31001061), NBCITS.MOA (CARS-38), 973 Program (2010CB530206), State Key Laboratory of Veterinary Etiological Biology Project (SKLVEB2008ZZKT019), EPIZONE (FOOD-CT-2006-016236), ARBOZOONET (no. 211757), PIROVAC (KBBE-3-245145), and European Commission, Brussels, Belgium. The authors thank Edanz Editing for help with revision of this article.

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  1. State Key Laboratory of Veterinary Etiological Biology, Xujiaping 1, Lanzhou, Gansu, 730046, People’s Republic of China

    Xiao Zhang, Zhijie Liu, Jifei Yang, Ze Chen, Guiquan Guan, Qiaoyun Ren, Aihong Liu, Jianxun Luo, Hong Yin & Youquan Li

  2. Key Laboratory of Veterinary Parasitology of Gansu Province, Xujiaping 1, Lanzhou, Gansu, 730046, People’s Republic of China

    Xiao Zhang, Zhijie Liu, Jifei Yang, Ze Chen, Guiquan Guan, Qiaoyun Ren, Aihong Liu, Jianxun Luo, Hong Yin & Youquan Li

  3. Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Science, Xujiaping 1, Lanzhou, Gansu, 730046, People’s Republic of China

    Xiao Zhang, Zhijie Liu, Jifei Yang, Ze Chen, Guiquan Guan, Qiaoyun Ren, Aihong Liu, Jianxun Luo, Hong Yin & Youquan Li

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  1. Xiao Zhang
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  2. Zhijie Liu
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  9. Hong Yin
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Correspondence to Youquan Li.

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Zhang, X., Liu, Z., Yang, J. et al. Multiplex PCR for diagnosis of Theileria uilenbergi, Theileria luwenshuni, and Theileria ovis in small ruminants. Parasitol Res 113, 527–531 (2014). https://doi.org/10.1007/s00436-013-3684-9

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  • DOI: https://doi.org/10.1007/s00436-013-3684-9

Keywords

  • Polymerase Chain Reaction
  • Multiplex Polymerase Chain Reaction
  • Small Ruminant
  • Babesia
  • Prime Concentration