Establishment of a Real-Time Reverse Transcription Recombinase-Aided Isothermal Amplification (qRT-RAA) Assay for the Rapid Detection of Bovine Respiratory Syncytial Virus

Abstract

Background: Bovine respiratory syncytial virus (BRSV) is a significant cause of bovine respiratory disease, resulting in significant losses to the cattle industry. For rapid detection of BRSV, a real-time recombinase-aided isothermal amplification assay (qRT-RAA) based on the F gene of BRSV was developed in this study. Results: The developed qRT-RAA assay showed good exponential amplification of the target fragment in 20 min at a constant temperature of 39 °C. And this assay displayed a high specificity for BRSV, without cross-reactions with Infectious Bovine Rhinotracheitis Virus (IBRV), Bovine Parainfluenza Virus Type 3 (BPIV3), Bovine Viral Diarrhea Virus (BVDV), and Bovine Coronavirus (BCoV). With the standard RNA of BRSV serving as a template, the limit of detection for qRT-RAA was 102 copies/μL. We examined ninety-seven clinical samples from cattle with respiratory disease using this method and determined a positive rate of 7.2% (7/97), consistent with results using the classical PCR method reported previously. Conclusions: A qRT-RAA assay for BRSV detection was established in this study. The method is specific and sensitive and can be completed within 20 min at 39 °C. These works demonstrate that the generated qRT-RAA assay is an effective diagnostic tool for rapidly detecting BRSV in resource-limited settings, which may be applied for the clinical detection of BRSV.