In the present study, the SYBR green I-based duplex quantitative polymerase chain reaction (qPCR) was developed for simultaneous detection of classical swine fever virus (CSFV) and porcine circovirus 3 (PCV3). The assay was used to detect both CSFV and PCV3 in one sample by their distinct melting temperatures (melting peaks at 87°C for CSFV and 81.5 °C for PCV3), and no specific fluorescence signals were detected for other non-targeted porcine pathogens. The assay had a high degree of linearity (R2 > 0.998) with the detection limits of 23 copies/μL for CSFV and 36 copies/μL for PCV3, and exhibited high repeatability and reproducibility with a low coefficient of variation below 2.0% in both intra- and inter-assay. In this study, 130 clinical samples collected from sick pigs in the field were tested by this assay with the positive rates of 9.23% (12/130) for CSFV and 21.54% (28/130) for PCV3 respectively, and the positive rate of CSFV and PCV3 co-infection was 6.92% (9/130). Our results showed that the developed method was a reliable diagnostic tool to monitor and survey CSFV, PCV3 and CSFV/PCV3 co-infection in the field.

中文翻译：

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基于SYBR Green I的双工实时荧光定量PCR同时检测经典猪瘟病毒和猪圆环病毒3。

在本研究中，开发了基于SYBR green I的双重定量聚合酶链反应（qPCR），用于同时检测经典猪瘟病毒（CSFV）和猪圆环病毒3（PCV3）。该测定法通过其独特的解链温度（CSFV在87°C和PCV3在81.5°C的解链峰）检测一个样品中的CSFV和PCV3，并且未检测到其他非靶向猪病原体的特异性荧光信号。该测定法具有高度的线性度（R2> 0.998），CSFV的检出限为23拷贝/μL，PCV3的检出限为36拷贝/μL，在两种内标法中均显示出低于2.0％的低变异系数的高重复性和重现性。 -批间。在这项研究中，通过此测定法测试了从现场生病猪收集的130个临床样品，阳性率为9。CSFV的感染率为23％（12/130），PCV3的感染率为21.54％（28/130），CSFV和PCV3合并感染的阳性率为6.92％（9/130）。我们的结果表明，所开发的方法是在现场监测和调查CSFV，PCV3和CSFV / PCV3共感染的可靠诊断工具。