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Detection and Genogrouping of Noroviruses from Children's Stools By Taqman One-step RT-PCR
Published on: July 22, 2012
Diagnosing norovirus-associated infectious intestinal disease using viral load
Gemma Phillips1, Ben Lopman, Clarence C Tam
1Department of Gastrointestinal, Emerging and Zoonotic Infections, Health Protection Agency Centre for Infections, 61 Colindale Avenue, London, UK. gemma.phillips@hpa.org.uk
BMC Infectious Diseases
|May 16, 2009
Summary
Determining norovirus as the cause of infectious intestinal disease (IID) is challenging due to asymptomatic positives. This study used viral load via real-time RT-PCR and ROC analysis to establish a cut-off, improving norovirus diagnosis in IID cases.
Area of Science:
- Virology
- Infectious Diseases
- Molecular Diagnostics
Background:
- Reverse transcription-polymerase chain reaction (RT-PCR) is standard for norovirus diagnosis.
- Asymptomatic norovirus shedding complicates attributing illness to the virus in infectious intestinal disease (IID) cases.
- Accurate pathogen identification is crucial for clinical management and epidemiological studies of IID.
Purpose of the Study:
- To investigate the utility of fecal viral load in determining if norovirus is the causative agent of IID.
- To establish a reliable method for differentiating norovirus infection from asymptomatic shedding in IID patients.
Main Methods:
- Real-time RT-PCR was employed to quantify norovirus genogroup II viral load in fecal samples from IID cases and healthy controls.
- Cycle threshold (Ct) values served as a proxy for viral load.
- Receiver-operating characteristic (ROC) analysis was utilized to identify an optimal Ct value cut-off for diagnosing norovirus-associated IID.
Main Results:
- An optimal Ct value cut-off of 31 was identified using ROC analysis for attributing IID to norovirus.
- This cut-off remained consistent whether using healthy controls or culture-confirmed bacterial pathogen cases as the negative reference group.
- The reference negative group can be sourced from specimens routinely processed in clinical virology laboratories.
Conclusions:
- Receiver-operating characteristic (ROC) analysis effectively identifies a cut-off for norovirus real-time RT-PCR assays, aiding in the diagnosis of IID.
- Routine clinical virology laboratory specimens can be used to establish appropriate assay cut-offs.
- Laboratories can implement this method to define in-house cut-offs, enhancing diagnostic accuracy for clinicians and public health.
