Rapid diagnosis is an important intervention in managing the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) outbreak. Real time reverse transcription polymerase chain reaction (RT-qPCR) remains the primary means for diagnosing the new virus strain but it is time consuming and costly. Recombinase polymerase amplification (RPA) is an isothermal amplification assay that does not require a PCR machine. It is an affordable, rapid, and simple assay. In this study, we developed and optimized a sensitive reverse transcription (RT)-RPA assay for the rapid detection of SARS-CoV-2 using SYBR Green I and/or lateral flow (LF) strip. The analytical sensitivity and specificity of the RT-RPA assay were tested by using 10-fold serial diluted synthetic RNA and genomic RNA of similar viruses, respectively. Clinical sensitivity and specificity of the RT-RPA assay were carried out using 78 positive and 35 negative nasopharyngeal samples. The detection limit of both RPA and RT-qPCR assays was 7.659 and 5 copies/μL RNA, respectively with no cross reactivity with other viruses. The clinical sensitivity and specificity of RT-RPA were 98% and 100%, respectively. Our study showed that RT-RPA represents a viable alternative to RT-qPCR for the detection of SARS-CoV-2, especially in areas with limited infrastructure.
【초록키워드】 severe acute respiratory syndrome coronavirus 2, viruses, SARS-CoV-2, coronavirus, Diagnosis, Intervention, severe acute respiratory syndrome Coronavirus, coronavirus 2, RNA, lateral flow, polymerase chain reaction, specificity, RT-qPCR, SYBR Green, PCR, Sensitivity and specificity, outbreak, clinical, Rapid, reverse transcription, isothermal amplification, RT-RPA, respiratory, recombinase polymerase amplification, Cross reactivity, analytical sensitivity, RPA, Detection limit, genomic RNA, real time, RT-qPCR assay, clinical sensitivity, acute respiratory syndrome coronavirus, RT-qPCR assays, Nasopharyngeal samples, Real time reverse transcription polymerase chain reaction, SYBR Green I, new virus, diagnosing, positive, polymerase chain, tested, carried, costly, 【제목키워드】 SARS-CoV-2, coronavirus, coronavirus 2, reverse transcription, respiratory, development, polymerase, Amplification assay,