Xiuli Sun , Huanhuan Lu , Yanqing Tie , Mengchuan Zhao , Ruiqing Zhang , Zhenlu Sun , Guohao Fan , Fengyu Li , Fengyu Tian , Yaxin Hu , Mengyi Zhang , Xinxin Shen , Xuejun Ma , Zhishan Feng
{"title":"A one-step reverse-transcription recombinase aided PCR assay for the rapid and sensitive detection of human enteroviruses","authors":"Xiuli Sun , Huanhuan Lu , Yanqing Tie , Mengchuan Zhao , Ruiqing Zhang , Zhenlu Sun , Guohao Fan , Fengyu Li , Fengyu Tian , Yaxin Hu , Mengyi Zhang , Xinxin Shen , Xuejun Ma , Zhishan Feng","doi":"10.1016/j.bsheal.2023.03.002","DOIUrl":null,"url":null,"abstract":"<div><p>Human enteroviruses (HEVs) include many different types that cause a wide range of diseases, and an effective method of genus-level identification has therefore significant clinical implications. However, quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR), the gold-standard method, still has shortfalls in diagnostic sensitivity and timeliness. Here we established a one-step real-time reverse-transcription recombinase-aided PCR assay (RT-RAP) to detect HEV fragment within an hour. The RT-RAP assay showed a detection limit of 5 copies/μL using recombinant plasmids and was extensively verified using 15 HEV strains. Among 15 types of HEV (species A-C), the sensitivity of RT-RAP was approximately 2–8 folds lower than that of the qRT-PCR in 9 types, and no-cross reaction with other viruses was observed. RT-RAP was further applied to analyze CSF and fecal specimens; the clinical performance demonstrated that the RT-RAP and the commercial qRT-PCR kit provided consistent results. These results indicated that RT-RAP assay may be a promising approach for rapid and sensitive detection of HEV.</p></div>","PeriodicalId":36178,"journal":{"name":"Biosafety and Health","volume":null,"pages":null},"PeriodicalIF":3.5000,"publicationDate":"2023-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Biosafety and Health","FirstCategoryId":"3","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S2590053623000253","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"PUBLIC, ENVIRONMENTAL & OCCUPATIONAL HEALTH","Score":null,"Total":0}
引用次数: 0
Abstract
Human enteroviruses (HEVs) include many different types that cause a wide range of diseases, and an effective method of genus-level identification has therefore significant clinical implications. However, quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR), the gold-standard method, still has shortfalls in diagnostic sensitivity and timeliness. Here we established a one-step real-time reverse-transcription recombinase-aided PCR assay (RT-RAP) to detect HEV fragment within an hour. The RT-RAP assay showed a detection limit of 5 copies/μL using recombinant plasmids and was extensively verified using 15 HEV strains. Among 15 types of HEV (species A-C), the sensitivity of RT-RAP was approximately 2–8 folds lower than that of the qRT-PCR in 9 types, and no-cross reaction with other viruses was observed. RT-RAP was further applied to analyze CSF and fecal specimens; the clinical performance demonstrated that the RT-RAP and the commercial qRT-PCR kit provided consistent results. These results indicated that RT-RAP assay may be a promising approach for rapid and sensitive detection of HEV.