数字农科院2.0

Novel real-time quantitative RT-PCR for detection of PRRSV-1 and PRRSV-2 strains circulating in China: A study based on whole-genome characteristics and epidemiological features

文献类型： 外文期刊

作者： Bangjun Gong; Lirun Xiang; Jinhao Li; Chao Li ; Jiahao Shi ;Zhenyang Guo ;Siyu Zhang ;Zixuan Feng ;Haonan Kang ;Xueli Zhang ;Ziyu Song ;Chaoliang Leng ;Qian Wang; Jinmei Peng; Guohui Zhou; Kuan Zhao; Yan-Dong Tang; Hongyuan Zhao; Xuehui Cai; Hu Xu; Zhi-Jun Tian; Hongliang Zhang

关键词： PRRSV;Genomic variation; Real-time RT-qPCR; Differential diagnosis; Detection and quantification

期刊名称： VETERINARY JOURNAL

ISSN： 1090-0233

年卷期： 2025 年

页码：

收录情况： SCIE（2025版）

摘要： Recent studies have shown significant changes in the whole-genome characteristics of both PRRSV-1 and PRRSV2 in China, which may compromise the accuracy of previous detection methods. Herein, based on the current whole-genome characteristics and a comprehensive sequence database of Chinese PRRSV-1 and PRRSV-2, we identified conserved and specific genes for both viruses and designed corresponding primers and probes. After systematic optimization and evaluation, we developed a duplex real-time RT-qPCR for simultaneously identi fying and differentiating PRRSV-1 and PRRSV-2, along with two simplex quantitative RT-PCRs for viral load measurement. To assess the performance of the duplex real-time RT-qPCR, a comprehensive sample repository was established to refine its positive criterion, evaluate its specificity and inclusivity, and compare its detection capacity with that of three commercial kits. The evaluation demonstrated that a positive result for both PRRSV-1 and PRRSV-2 channels was defined as an S-shaped amplification curve with a Ct value ≤ 36.0, while a suspected positive result was characterized by an S-shaped amplification curve with a Ct value greater than 36.0 but not exceeding 37.5. Samples exhibiting a Ct value exceeding 37.5 were classified as negative. Moreover, the method successfully detected diverse PRRSV-1 and PRRSV-2 strains, accurately differentiated between the two types of viruses, and exhibited higher inclusivity for PRRSV-1 and better differentiation capability between PRRSV-1 and PRRSV-2 than three commercial kits. For the two simplex quantitative RT-PCRs, validation using sera from pigs infected with multiple PRRSV-1 and PRRSV-2 strains showed that no viral RNA was detected at 0 dpi, while the expected viral load was successfully quantified between 3 and 21 dpi. Overall, the three developed methods can serve as effective tools for the detection, discrimination, and quantification of PRRSV-1 and PRRSV-2 strains circulating in China.

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