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Development of Taqman-Based Real-Time PCR Assays for Diagnostic Detection of Babesia bovis and Babesia bigemina

Two TaqMan-based real-time polymerase chain reaction (PCR) assays devised for the detection of two bovine Babesia parasites, Babesia bovis and B. bigemina, were evaluated for their diagnostic utility using cultured parasites and 92 field bovine blood samples collected from cattle living in Brazil. The real-time PCR assays were compared with previously established nested-PCR assays. The detection limits of both B. bovis- and B. bigemina–real-time PCR assays were identical at the value of 2.5 parasites/µL of the infected blood. When 92 field bovine blood samples were tested using the real-time assays, B. bovis-positive signals were observed in 30 samples among 31 B. bovis-positive blood samples in the nested-PCR assay (96.9% sensitivity and 100% specificity), whereas the B. bigemina–real-time PCR assay could detect the parasite from all of 45 B. bigemina-positive blood samples in the nested-PCR assay (100% sensitivity and specificity). The real-time assays using the TaqMan-system can therefore be practically implemented in the epidemiologic survey for bovine babesiosis. Further studies will be necessary to investigate the clinical value of this technique, especially for the quantitative detections of the parasites.

Received May 7, 2007. Accepted for publication July 5, 2007.

Acknowledgments: This research was supported by Grants-in-Aid for Scientific Research from the Japan Society for the Promotion of Science (JSPS), the Program of Promotion of Basic Research Activities for Innovative Biosciences (PROBRAIN), and The 21^st Century COE Program (A-1), Ministry of Education, Culture, Sports, Science, and Technology, Japan.

^* Address correspondence to Ikuo Igarashi, National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-cho, Obihiro, Hokkaido 080-8555, Japan. E-mail: igarcpmi{at}obihiro.ac.jp

Authors’ addresses: Chulmin Kim, Hiroshi Iseki, Maria Shirley Herbas, Naoaki Yokoyama, Hiroshi Suzuki, Xuenan Xuan, Kozo Fujisaki, and Ikuo Igarashi, National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-cho, Obihiro, Hokkaido 080-8555, Japan, Tel.: +81-155-49-5641, Fax: +81-155-49-5643, E-mail: igarcpmi{at}obihiro.ac.jp.

Reprint requests: Ikuo Igarashi, National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-cho, Obihiro, Hokkaido 080-8555, Japan. Telephone: +81-155-49-5641. Fax: +81-155-49-5643, E-mail: igarcpmi{at}obihiro.ac.jp.