Standardized Positive Controls for Detection of Norovirus by Reverse Transcription PCR

Background:
Norovirus is one of the most common causes of nonbacterial gastroenteritis in humans. Rapid spread by contaminated food and person-to-person transmission through the fecal-oral route are characteristics of norovirus and result in high morbidity in vulnerable patient populations. Therefore, detection of norovirus is a major public health concern. Currently, the most common method for detecting and differentiating among noroviruses in clinical and environmental samples is reverse transcription PCR (RT-PCR). Standardized positive controls used in RT-PCR assays to detect norovirus are designed to overcome the problem of false-negative results due to PCR inhibitors and suboptimal reaction conditions.

Results:
In the current study, four types of RNA transcripts were produced from plasmids: norovirus GI-5 and GII-4 capsid regions with human rotavirus (HRV) fragment insertions, and norovirus GI-6 and GII-4 capsid regions with hepatitis A virus (HAV) fragment insertions. These size-distinguishable products were used as positive controls under the RT-PCR assay conditions used to detect NoV in stool and groundwater samples. Their reliability and reproducibility was confirmed by multiple sets of experiments.

The article is published in the Virology Journal and is free to access.