Abstract
BackgroundHuman noroviruses are one of the main causes of foodborne illnesses and represent a serious public health concern. Rapid and sensitive assays for human norovirus detection are undoubtedly necessary for clinical diagnosis, especially in regions without more sophisticated equipment.MethodThe rapid reverse transcription recombinase-aided amplification (RT-RAA) is a fast, robust and isothermal nucleic acid detection method based on enzyme reaction. This method can complete the sample detection at 39 °C in 30 min. In this study, we successfully established a rapid reverse transcription recombinase-aided amplification (RT-RAA) assay for the detection of human norovirus GII.4 and applied this assay to clinical samples, as well as comparison with commercial reverse transcription real-time fluorescence quantitative PCR (RT-qPCR).ResultsAt 95% probability, the detection sensitivity of RT-RAA was 3.425 log10 genomic copies (LGC)/reaction. Moreover, no cross-reaction was observed with other norovirus genogroups and other common foodborne viruses. Stool samples were examined by RT-RAA and reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR). Compared of RT-qPCR, kappa values for human norovirus detection with RT-RAA were 0.894 (p < 0.001), indicating that both assays were in agreement.ConclusionThis RT-RAA assay provides a rapid, specific, and sensitive assay for human norovirus detection and is suitable for clinical testing.
Highlights
Human noroviruses are one of the main causes of foodborne illnesses and represent a serious public health concern
Stool samples were examined by reverse transcription recombinase-aided amplification (RT-recombinase-aided amplification (RAA)) and reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR)
Compared of real-time fluorescence quantitative PCR (RT-qPCR), kappa values for human norovirus detection with RT-RAA were 0.894 (p < 0.001), indicating that both assays were in agreement
Summary
Human noroviruses are one of the main causes of foodborne illnesses and represent a serious public health concern. Rapid and sensitive assays for human norovirus detection are undoubtedly necessary for clinical diagnosis, especially in regions without more sophisticated equipment. It been noted that NoV infects people of all ages, but is more severe in children and the elderly, especially in low-income countries, leading to over 200,000 deaths every year [3]. Traditional detection methods for NoV, such as immunoassays and electron microscopy, are not suitable for rapid diagnosis because most of them are generally time-consuming and laborious. Common nucleic molecular diagnostic assays for NoV detection with high specificity and sensitivity, such as fluorescence-based real-time PCR and nested PCR, have been used in disease diagnosis [4, 5]. Expensive equipment and the need for trained technicians make these traditional detection methods restrictive and impractical for rapid and convenient detection in low-income regions
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