Rapid and Visual Detection of Type 2 Porcine Reproductive and Respiratory Syndrome Virus by Real-Time Fluorescence-Based Reverse Transcription Recombinase-Aided Amplification
Wenlong Xia,
Yao Chen,
Xue Ding,
Xiaoming Liu,
Huipeng Lu,
Changming Guo,
Hua Zhang,
Zhijun Wu,
Jing Huang,
Zhongjun Fan,
Shupei Yu,
Huaichang Sun,
Shanyuan Zhu,
Zhi Wu
Affiliations
Wenlong Xia
Jiangsu Key Laboratory for Bioresources of Saline Soils, School of Ocean and Biological Engineering, Yancheng Teachers University, Yancheng 224007, China
Yao Chen
Jiangsu Key Laboratory for Bioresources of Saline Soils, School of Ocean and Biological Engineering, Yancheng Teachers University, Yancheng 224007, China
Xue Ding
Jiangsu Key Laboratory for Bioresources of Saline Soils, School of Ocean and Biological Engineering, Yancheng Teachers University, Yancheng 224007, China
Xiaoming Liu
College of Veterinary Medicine, Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Diseases and Zoonoses, Yangzhou University, Yangzhou 225009, China
Huipeng Lu
Jiangsu Key Laboratory for High-Tech Research and Development of Veterinary Biopharmaceuticals, Jiangsu Agri-Animal Husbandry Vocational College, Taizhou 225300, China
Changming Guo
Jiangsu Key Laboratory for High-Tech Research and Development of Veterinary Biopharmaceuticals, Jiangsu Agri-Animal Husbandry Vocational College, Taizhou 225300, China
Hua Zhang
School of Pharmacy, Yancheng Teachers University, Yancheng 224007, China
Zhijun Wu
School of Pharmacy, Yancheng Teachers University, Yancheng 224007, China
Jing Huang
Jiangsu Key Laboratory for Bioresources of Saline Soils, School of Ocean and Biological Engineering, Yancheng Teachers University, Yancheng 224007, China
Zhongjun Fan
Jiangsu Key Laboratory for Bioresources of Saline Soils, School of Ocean and Biological Engineering, Yancheng Teachers University, Yancheng 224007, China
Shupei Yu
Yancheng Animal Husbandry and Veterinary Station, Yancheng 224001, China
Huaichang Sun
College of Veterinary Medicine, Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Diseases and Zoonoses, Yangzhou University, Yangzhou 225009, China
Shanyuan Zhu
Jiangsu Key Laboratory for High-Tech Research and Development of Veterinary Biopharmaceuticals, Jiangsu Agri-Animal Husbandry Vocational College, Taizhou 225300, China
Zhi Wu
Jiangsu Key Laboratory for High-Tech Research and Development of Veterinary Biopharmaceuticals, Jiangsu Agri-Animal Husbandry Vocational College, Taizhou 225300, China
Porcine reproductive and respiratory syndrome (PRRS) is one of the most important diseases that has brought significant economic losses to the swine industry worldwide. Rapid and accurate PRRS virus (PRRSV) detection is one of the key factors for PRRS prevention and control. This study developed a real-time fluorescence-based reverse transcription recombinase-aided amplification (RF-RT-RAA) method for type 2 PRRSV (PRRSV-2) detection. The RF-RT-RAA assay could be performed at 42 °C for 20 min with the optimal primers and a probe. RF-RT-RAA results could be monitored using real-time fluorescence read-out or visually observed with the naked eye using a portable blue light transilluminator. The method had a strong specificity; no cross-reaction was identified with the detected common swine viruses. Moreover, the technique yielded high sensitivity with the lowest detection limit of 101 copies/μL and exhibited good repeatability and reproductively with the coefficients of variation (CV) less than 10%. Eighty-seven clinical samples were tested using RF-RT-RAA and a commercial PRRSV-2 RT-qPCR detection kit. The coincidence rate was 100% between RF-RT-RAA (real-time fluorescence read-out) and RT-qPCR, and 97.7% between RF-RT-RAA (visually observed) and RT-qPCR. The RF-RT-RAA assay provides a new method for rapid and visual detection of PRRSV-2.
