Development of a Real-Time Quantitative PCR Based on a TaqMan-MGB Probe for the Rapid Detection of Theileria haneyi

文献类型： 外文期刊

第一作者： Zhou, Bingqian

作者： Zhou, Bingqian;Yang, Guangpu;Hu, Zhe;Chen, Kewei;Guo, Wei;Wang, Xiaojun;Du, Cheng

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关键词： Theileria haneyi; real-time quantitative PCR; nested PCR

期刊名称：MICROORGANISMS （ 影响因子：4.5； 五年影响因子：4.8 ）

ISSN：

年卷期： 2023 年 11 卷 11 期

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收录情况： SCI

摘要： Equine piroplasmosis (EP) is a parasitic disease caused by Theileria equi (T. equi), Babesia caballi (B. caballi) and Theileria haneyi (T. haneyi). This disease is considered to be reportable by the World Organization for Animal Health (WOAH). Real-time quantitative PCR (qPCR) is regarded as a straightforward, rapid and sensitive diagnostic method to detect pathogens. However, qPCR has not been employed in the various epidemiological investigations of T. haneyi. In this study, we developed a new qPCR method to detect T. haneyi based on the chr1sco (chromosome 1 single-copy open reading frame (ORF)) gene, which has no detectable orthologs in T. equi or B. caballi. A TaqMan MGB probe was used in the development of the qPCR assay. A plasmid containing the chr1sco gene was constructed and used to establish the standard curves. The novel qPCR technique demonstrated great specificity for detecting additional frequent equine infectious pathogens and sensitivity for detecting diluted standard plasmids. This qPCR was further validated by comparison with an optimized nested PCR (nPCR) assay in the analysis of 96 clinical samples. The agreement between the nPCR assay and the established qPCR assay was 85.42%. The newly established method could contribute to the accurate diagnosis of T. haneyi infections in horses.

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