Rapid detection of norovirus genogroup II in clinical and environmental samples using recombinase polymerase amplification

Norovirus is the leading cause of acute gastroenteritis all over the world, and the most genotype that causes its epidemic is norovirus genogroup II (NoVs GII). Rapid detection of NoVs is important because it can facilitate timely diagnosis. In this study, we designed universal specific primers and an Exo probe to hybridize to all genetic clusters of NoVs GII based on the conserved region at the ORF1-ORF2 junction of the genome. For the first time, we established a rapid and reliable reverse transcription recombinase polymerase amplification (RT-RPA) method for the detection of NoVs GII within 20 min. This method can specifically amplify NoVs GII, and the detection limit was as low as 1.66 × 102 copies/μL. The method was validated in terms of LOD, accuracy, and specificity. We tested 55 real samples including foods, water, and feces. The results showed a sensitivity of 96% and specificity of 100% to NoVs GII. The whole procedure can be operated by a mobile suitcase laboratory, which is useful for resource-limited diagnostic laboratories. This novel real-time RT-RPA assay is an accurate tool for point-of-care testing of NoVs, providing practical support for norovirus-caused disease diagnosis and prevention.