title: Detection of circulating norovirus genotypes: hitting a moving target.
abstract: BACKGROUND
  Although national surveillance programs are in place to monitor norovirus epidemiology, the emergence of new strains and the genetic diversity among genotypes can be challenging for clinical laboratories. This study evaluated the analytical and clinical performance characteristics of one real-time RT-PCR and two end-point RT-PCRs commonly used in microbiology laboratories.
  
  
  METHODS
  Lower limit of detection (LoD) was determined using 10-fold dilutions of noroviruses belonging to different genotypes. The clinical performance of the real-time and end-point RT-PCRs was assessed in parallel using nucleic acids extracted from 186 stool specimens.
  
  
  RESULTS
  The real-time RT-PCR was highly sensitive and specific for the detection of norovirus genotypes that are currently circulating in Canada. In contrast, the two end-point RT-PCRs displayed poor analytical sensitivity or complete failure to detect certain norovirus genotypes, which was correlated to sequence mismatches in the primer-binding sites. In an attempt to improve norovirus detection with the end-point RT-PCRs, both assays were processed concurrently and detection from either assay was considered a positive result. Concurrent testing resulted in only a modest increase in clinical sensitivity (75.0%) compared to each assay alone (62.5% and 71.9%). However, the false positivity rate increased from 1.98% and 3.36% for the assays alone to 5.47% with concurrent testing.
  
  
  CONCLUSIONS
  This study emphasizes the benefits of a real-time method and provides support for routine surveillance to monitor norovirus epidemiology and ongoing proficiency testing to ensure detection of circulating norovirus genotypes.
authors.forename: Brenda-Lee
authors.initials: BL
authors.lastname: Rooney
authors.forename: Janice
authors.initials: J
authors.lastname: Pettipas
authors.forename: Elsie
authors.initials: E
authors.lastname: Grudeski
authors.forename: Oksana
authors.initials: O
authors.lastname: Mykytczuk
authors.forename: Xiao-Li
authors.initials: XL
authors.lastname: Pang
authors.forename: Tim F
authors.initials: TF
authors.lastname: Booth
authors.forename: Todd F
authors.initials: TF
authors.lastname: Hatchette
authors.affiliation: Dalhousie University, Halifax, Nova Scotia, Canada. Jason.leblanc@cdha.nshealth.ca.
authors.forename: Jason J
authors.initials: JJ
authors.lastname: LeBlanc
mesh_terms: D017250:Caliciviridae Infections / Q000175:diagnosis* / Q000821:virology*; D005759:Gastroenteritis / Q000175:diagnosis* / Q000821:virology*; D005838:Genotype; D006801:Humans; D029322:Norovirus / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity