title: Development of a Recombinase Polymerase Amplification Assay for Detection of Epidemic Human Noroviruses.
abstract: Human norovirus is a leading cause of viral gastroenteritis worldwide. Rapid detection could facilitate control, however widespread point-of-care testing is infrequently done due to the lack of robust and portable methods. Recombinase polymerase amplification (RPA) is a novel isothermal method which rapidly amplifies and detects nucleic acids using a simple device in near real-time. An RT-RPA assay targeting a recent epidemic human norovirus strain (GII.4 New Orleans) was developed and evaluated in this study. The assay successfully detected purified norovirus RNA from multiple patient outbreak isolates and had a limit of detection of 3.40 ± 0.20 log10genomic copies (LGC), which is comparable to most other reported isothermal norovirus amplification methods. The assay also detected norovirus in directly boiled stool, and displayed better resistance to inhibitors than a commonly used RT-qPCR assay. The assay was specific, as it did not amplify genomes from 9 non-related enteric viruses and bacteria. The assay detected norovirus in some samples in as little as 6 min, and the entire detection process can be performed in less than 30 min. The reported RT-RPA method shows promise for sensitive point-of-care detection of epidemic human norovirus, and is the fastest human norovirus amplification method to date.
authors.affiliation: Department of Food, Bioprocessing and Nutrition Sciences, North Carolina State University, 315 Schaub Hall, 400 Dan Allen Drive, Raleigh, North Carolina 27695, United States of America.
authors.forename: Matthew D
authors.initials: MD
authors.lastname: Moore
authors.affiliation: Department of Food, Bioprocessing and Nutrition Sciences, North Carolina State University, 315 Schaub Hall, 400 Dan Allen Drive, Raleigh, North Carolina 27695, United States of America.
authors.forename: Lee-Ann
authors.initials: LA
authors.lastname: Jaykus
mesh_terms: D017250:Caliciviridae Infections / Q000175:diagnosis* / Q000235:genetics; D058872:Epidemics / Q000517:prevention & control*; D006801:Humans; D029322:Norovirus / Q000235:genetics / Q000302:isolation & purification*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D045522:Recombinases / Q000032:analysis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*
chemical_list: D045522:Recombinases