A novel HRM assay for differentiating classical strains and highly pathogenic strains of type 2 porcine reproductive and respiratory syndrome virus

文献类型： 外文期刊

第一作者： Sun, Junying

作者： Sun, Junying;Liu, Zhicheng;Zhang, Chunhong;Shen, Haiyan;Zhang, Jianfeng;Bingga, Gali;Guo, Pengju;Sun, Junying;Liu, Zhicheng;Zhang, Chunhong;Shen, Haiyan;Zhang, Jianfeng;Sun, Junying;Liu, Zhicheng;Zhang, Chunhong;Shen, Haiyan;Zhang, Jianfeng

作者机构：

关键词： PRRSV; Classical trains; Highly pathogenic strains; PCR; High resolution melting curve analysis; Unlabeled probe

期刊名称：MOLECULAR AND CELLULAR PROBES （ 影响因子：2.365； 五年影响因子：2.386 ）

ISSN： 0890-8508

年卷期： 2018 年 39 卷

页码：

收录情况： SCI

摘要： Differentiation of classical strains and highly pathogenic strains of porcine reproductive and respiratory syndrome virus is crucial for effective vaccination programs and epidemiological studies. We used nested PCR and high resolution melting curve analysis with unlabeled probe to distinguish between the classical and the highly pathogenic strains of this virus. Two sets of primers and a 20 bp unlabeled probe were designed from the NSP3 gene. The unlabeled probe included two mutations specific for the classical and highly pathogenic strains of the virus. An additional primer set from the NSP2 gene of the highly pathogenic vaccine strain JXA1-R was used to detect its exclusive single nucleotide polymorphism. We tested 107 clinical samples, 21 clinical samples were positive for PRRSV (consistent with conventional PCR assay), among them four were positive for the classical strain with the remainder 17 for the highly pathogenic strain. Around 10 C difference between probe melting temperatures showed the high discriminatory power of this method. Among highly pathogenic positive samples, three samples were determined as positive for JXA1-R vaccine-related strain with a 95% genotype confidence percentage. All these genotyping results using the high resolution melting curve assay were confirmed with DNA sequencing. This unlabeled probe method provides an alternative means to differentiate the classical strains from the highly pathogenic porcine reproductive and respiratory syndrome virus strains rapidly and accurately.

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