Gong Ling, Wang Xiaowen, Li Zhu, Huang Guichuan, Zhang Wei, Nie Jin, Wu Chunyan, Liu Daishun
The First Clinical Medical College, Jinan University, Guangzhou, China.
Department of Respiratory Medicine, The Third Affiliated Hospital of Zunyi Medical University, The First People's Hospital of Zunyi, Zunyi, China.
Front Microbiol. 2022 Mar 31;13:819931. doi: 10.3389/fmicb.2022.819931. eCollection 2022.
Respiratory syncytial virus (RSV) is a common virus that causes respiratory infection, especially severe respiratory infection in infants and young children, the elderly people over 65 years old, and people with weak immunity. Currently, RSV infection has no effective vaccine and antiviral treatment. The number of deaths due to RSV infection increases every year. Moreover, RSV A infection occurs in a large number and has severe clinical symptoms and complications than RSV B infection. Therefore, the development of a simple, rapid, and inexpensive detection method with high amplification efficiency, high sensitivity, and specificity is very important for the diagnosis of RSV A or RSV B infection, which can help in the early clinical medication and prevent the progress of the disease. Therefore, we developed an integrated trinity test with an RPA-CRISPR/Cas12a-fluorescence (termed IT-RAISE) assay system to detect RSV A or RSV B. The characteristic of the IT-RAISE system is that after target recognition, the reporter single-stranded DNA (ssDNA) is cleaved by Cas12a that is activated by different crRNAs to detect the generated fluorescent signal. This method is simple and helps in adding all reagents rapidly. It is a high-sensitive method that can detect 1.38 × 10 copies/μl of the target sequences, and it can distinguish RSV A or RSV B infection within 37 min. In addition, clinical specimens were detected for IT-RAISE system. It was found that the sensitivity and specificity of RSV A were 73.08 and 90%, respectively, and those of RSV B were 42.86 and 93.33%, respectively. The cost of ONE specimen for IT-RAISE system was approximately $ 2.6 (excluding rapid RNA extraction and reverse transcription costs). IT-RAISE system has good clinical application prospects for detecting RSV A or RSV B infection; it is a simple, rapid, and inexpensive method with high amplification efficiency, high sensitivity, and high specificity. The IT-RAISE system might also detect other viral or bacterial infections.
呼吸道合胞病毒（RSV）是一种常见病毒，可引起呼吸道感染，尤其是婴幼儿、65岁以上老年人以及免疫力低下人群的严重呼吸道感染。目前，RSV感染尚无有效的疫苗和抗病毒治疗方法。因RSV感染导致的死亡人数逐年增加。此外，RSV A感染数量众多，且与RSV B感染相比，具有更严重的临床症状和并发症。因此，开发一种简单、快速且廉价的检测方法，具有高扩增效率、高灵敏度和特异性，对于RSV A或RSV B感染的诊断非常重要，这有助于早期临床用药并防止疾病进展。因此，我们开发了一种集成三位一体检测方法，即RPA-CRISPR/Cas12a-荧光（称为IT-RAISE）检测系统，用于检测RSV A或RSV B。IT-RAISE系统的特点是在靶标识别后，报告单链DNA（ssDNA）被不同crRNAs激活的Cas1分子切割，以检测产生的荧光信号。该方法简单，有助于快速添加所有试剂。它是一种高灵敏度方法，可检测1.38×10拷贝/微升的靶标序列，并且能够在37分钟内区分RSV A或RSV B感染。此外，对临床标本进行了IT-RAISE系统检测。结果发现，RSV A的灵敏度和特异性分别为73.08%和90%，RSV B的灵敏度和特异性分别为42.86%和93.33%。IT-RAISE系统检测一个标本的成本约为2.6美元（不包括快速RNA提取和逆转录成本）。IT-RAISE系统在检测RSV A或RSV B感染方面具有良好的临床应用前景；它是一种简单、快速且廉价的方法，具有高扩增效率、高灵敏度和高特异性。IT-RAISE系统也可能检测其他病毒或细菌感染。
