Detection and quantification of human and bovine novoviruses by a TaqMan RT-PCR assay with a control for inhibition.

Noroviruses are single-stranded RNA viruses belonging to the family Caliciviridae. They are a major cause
of epidemic and sporadic gastroenteritis in humans and calves. Reverse transcription-polymerase chain
reaction (RT-PCR) has become the ‘‘gold standard’’ for detection of noroviruses in faecal and environmental
samples. However, false negative results due to co-concentration of RT-PCR inhibitors are
a continuous concern. A TaqMan real-time RT-PCR assay making use of a foreign internal RNA control and
a RNA standard was developed. Very interestingly, this method is capable of detecting human
noroviruses belonging to genogroups I and II, and bovine noroviruses belonging to genogroup III.
Inhibitors were removed efficiently by 1/10 dilution of the sample or addition of bovine serum albumin
to the RT-PCR mix. This assay was validated with human and bovine stool samples previously tested for
norovirus by conventional RT-PCR. The ability to detect norovirus in stool samples that were negative by
conventional RT-PCR assay demonstrate the higher sensitivity of the TaqMan assay compared to the
conventional RT-PCR assay. This real-time RT-PCR assay allows the detection of both human and bovine
noroviruses, avoids false negative results and is able to quantify the level of norovirus contamination.