title: Clinical evaluation of real-time PCR assays for rapid diagnosis of SARS coronavirus during outbreak and post-epidemic periods.
abstract: BACKGROUND
  The protocols of WHO network laboratories facilitated development of rapid diagnosis for SARS coronavirus (CoV) using reverse transcription (RT)-PCR assays. However, several reports have shown that conventional and real-time PCR assays were very specific for SARS CoV but lack sensitivity depending on the assay, specimen, and time course of disease.
  
  
  OBJECTIVE
  To evaluate an automatic nucleic acid extraction system and two standardized real-time PCR assays for rapid diagnosis of SARS CoV during outbreak and post-epidemic periods in Hong Kong.
  
  
  METHODS
  Specimens from clinically suspected SARS patients collected during outbreak and post-epidemic periods were tested by an automatic nucleic acid extraction system followed by our first generation conventional RT-PCR and two standardized real-time PCR assays (Artus GmbH, Germany and Roche Diagnostics, Germany). Paired serum samples were assayed for increasing titer against SARS CoV.
  
  
  RESULTS
  In the SARS epidemic, Artus and Roche PCR assays exhibited sensitivities of 87% and 85% for respiratory specimens (n = 64), 91% and 88% for stool (n = 44), and 82% for urine (n = 29). A specificity of 100% was exhibited by both PCR assays except Artus attained only a 92% specificity for stool. For post-epidemic period, no SARS CoV was identified among 56 respiratory specimens by all PCR assays. Inhibitors to PCR assays were detected at an average rate of 7-8% among 202 clinical specimens.
  
  
  CONCLUSIONS
  This study highlights the high throughput and performance of automatic RNA extraction in coordination with standardized real-time PCR assays suitable for large-scale routine diagnosis in case of future SARS epidemic. As no SARS CoV was detected among specimens collected during post-epidemic period, the positive predictive value of real-time PCR assays for detection of SARS CoV during low epidemic requires further evaluation.
authors.affiliation: Department of Microbiology, The University of Hong Kong, University Pathology Building, Queen Mary Hospital Compound, Pokfulam, Hong Kong SAR, PR China. wcyam@hku.hk
authors.forename: W C
authors.initials: WC
authors.lastname: Yam
authors.forename: K H
authors.initials: KH
authors.lastname: Chan
authors.forename: K H
authors.initials: KH
authors.lastname: Chow
authors.forename: L L M
authors.initials: LL
authors.lastname: Poon
authors.forename: H Y
authors.initials: HY
authors.lastname: Lam
authors.forename: K Y
authors.initials: KY
authors.lastname: Yuen
authors.forename: W H
authors.initials: WH
authors.lastname: Seto
authors.forename: J S M
authors.initials: JS
authors.lastname: Peiris
mesh_terms: D004196:Disease Outbreaks; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011933:Reagent Kits, Diagnostic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D045169:Severe Acute Respiratory Syndrome / Q000175:diagnosis* / Q000453:epidemiology* / Q000821:virology; D013997:Time Factors
chemical_list: D011933:Reagent Kits, Diagnostic