title: Comparison of real-time RT-PCR assays for hepatitis E virus RNA detection.
abstract: BACKGROUND
  Hepatitis E virus (HEV) is an increasing cause of acute viral hepatitis in developed countries. Serological testing alone may fail to diagnose acute infection, especially in immunocompromised patients, which justifies the use of molecular assays for diagnosis. Few studies have compared accuracy of HEV RNA detection assays.
  
  
  OBJECTIVE
  The performances of five real-time PCR procedures for HEV RNA detection were compared.
  
  
  METHODS
  First, RNA quantification of hepatitis E diluted standards of 3a and 3b genotypes were performed. Secondly, forty-seven clinical samples of patients with known acute HEV infection were tested using five hepatitis E RNA detection methods of assigned letters A, B, C, D and E.
  
  
  RESULTS
  Standards of HEV 3a genotype were detected in 100% of replicates with 2500 UI/ml of sensitivity by using A, B and C assays. Standards of HEV 3b genotype were more accurately detected with a sensitivity of 25 UI/ml in 100% of replicates using C assay and were detected in 100% of replicates with 2500 UI/ml of sensitivity by using A, B and E assays. Overall, B assay detected all of 250 UI/ml dilution and occasionally the 25 UI/ml dilution on both subtypes. The detection rates of clinical samples were 100%, 100% 97%, 97% and 83% for the respective A, B, C, D and E assay. Assays A and B were well correlated, independently of the subtype. However, discrepancies were observed when these techniques were compared to C, D and E assays according to the different subtypes.
  
  
  CONCLUSIONS
  A and B assays appear reliable for HEV RNA detection. These assays target the ORF2/3 overlapping region, described as more conserved than ORF2.
authors.affiliation: AP-HP, Hôpital Paul Brousse, Centre National de Référence des hépatites A et E, Villejuif 94804, France.
authors.forename: Camelia
authors.initials: C
authors.lastname: Mokhtari
authors.forename: Eric
authors.initials: E
authors.lastname: Marchadier
authors.forename: Stephanie
authors.initials: S
authors.lastname: Haïm-Boukobza
authors.forename: Asma
authors.initials: A
authors.lastname: Jeblaoui
authors.forename: Sophie
authors.initials: S
authors.lastname: Tessé
authors.forename: Jeanine
authors.initials: J
authors.lastname: Savary
authors.forename: Anne Marie
authors.initials: AM
authors.lastname: Roque-Afonso
mesh_terms: D016751:Hepatitis E / Q000175:diagnosis* / Q000821:virology; D016752:Hepatitis E virus / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D014773:Virology / Q000379:methods*
chemical_list: D012367:RNA, Viral
keywords: HEV diagnosis; HEV genotypes; Real-time RT-PCR assay