A one-step multiplex real-time RT-PCR assay for rapid and simultaneous detection of human norovirus genogroup I, II and IV.

ABSTRACT

A one-step multiplex real-time reverse transcription-PCR (RT-PCR) assay was developed for one-tube and simultaneous detection of three genogroups of human norovirus, genogroup I, II and IV (GI, GII and GIV). The specificity and sensitivity of the assay were evaluated and 50 samples were tested by using this assay. The results showed that the multiplex assay had high sensitivity and specificity. The amplification efficiencies of the assay were 91.3%, 90.1%, 88.9% and the detection limits were up to 16.9, 6.3, 43.0 copies/reaction respectively for norovirus GI, GII and GIV detection. No cross-reaction with the other examined RNA viruses was observed, and the qualitative analysis of samples showed that the multiplex assay had a good consistency with its corresponding monoplex assays for the detection of norovirus GI, GII and GIV (Kappa values were 0.848, 0.876 and 0.812 respectively).