Detection of norovirus quantificationally by realtime RT-PCR and its evaluation

Objective:To establish a TaqMan-based realtime fluorescent quantitative RT-PCR assay for detection and quantitation of Norovirus GⅠand GⅡ.Methods:The reaction conditions and systems were optimized so that the assay for detection the Norovirus by Realtime RT-PCR was establish.The sensitivity,specificity,repetition were evaluated.Laboratory diagnoses of some clinical specimens of this methods were also conducted.Results:The TaqMan assay can detect as few as 1 copy/μl virus.It showed a high linear dynamic range of quantitaion between 108～101 copy/μl.There was no cross reaction with Rotavirus,Adenovirus,Astrovirus,Enterovirus.Also There was no cross reaction between Norovirus GⅠand GⅡ.The standard deviation(SD) of intra-assay and extra-assay was 0.10～0.59.And the coefficient of variation(CV) was 0.43%～2.84%.181 specimens were detected by this assay,31 of which were positive for Norovirus.10 positive specimens were selected to conduct the sequencing analysis and it was confirmed that all of them were Norovirus.Conclusion:The established TaqMan quantitative realtime RT-PCR is a specific and sensitive method for rapid detection of Norovirus.It can distinguish the Norovirus GⅠand GⅡ,which are the most important genotypes that infected to human beings.This method can be used for early laboratory diagnosis of Norovirus.