title: Analytical performance determination and clinical validation of the novel Roche RealTime Ready Influenza A/H1N1 Detection Set.
abstract: The emergence of a novel pandemic human strain of influenza A (H1N1/09) virus in April 2009 has demonstrated the need for well-validated diagnostic tests that are broadly applicable, rapid, sensitive, and specific. The analytical performance and clinical validity of results generated with the novel Roche RealTime Ready Influenza A/H1N1 Detection Set using the LightCycler 2.0 instrument were characterized. Analytical performance was assessed by processing respiratory samples spiked with H1N1/09 and seasonal influenza A virus, a set of seasonal influenza A virus subtypes, and samples containing common viral and bacterial respiratory pathogens. The clinical validity of results was assessed in comparison to other assays by analyzing 359 specimens at three clinical sites and one reference laboratory. Direct sequencing was used to resolve samples with discrepant results. The assay detected virus concentrations down to <50 RNA copies per reverse transcription (RT)-quantitative PCR (qPCR). Various influenza A virus subtypes were covered. The analytical specificity was 100%. High clinical validity was demonstrated by the 99% positive agreement between seasonal influenza A viruses, 98% positive agreement between H1N1/09 viruses, and 88% agreement between negative results. The analytical sensitivity was compared to those of three other RT-qPCR assays and was found to be equivalent. The novel Roche RealTime Ready Influenza A/H1N1 Detection Set can be utilized on the widely used LightCycler platform. We demonstrate its usefulness for the rapid detection and surveillance of pandemic H1N1/09 influenza A virus infections.
authors.affiliation: Institute of Medical Microbiology and Hygiene, University of Regensburg, Franz Josef Strauss Allee 11, 93053 Regensburg, Germany. juergen.wenzel@klinik.uni-regensburg.de
authors.forename: Jürgen J
authors.initials: JJ
authors.lastname: Wenzel
authors.forename: Marcus
authors.initials: M
authors.lastname: Panning
authors.forename: Karen L
authors.initials: KL
authors.lastname: Kaul
authors.forename: Kathy A
authors.initials: KA
authors.lastname: Mangold
authors.forename: Paula A
authors.initials: PA
authors.lastname: Revell
authors.forename: Ruth Ann
authors.initials: RA
authors.lastname: Luna
authors.forename: Héctor
authors.initials: H
authors.lastname: Zepeda
authors.forename: Lizbeth
authors.initials: L
authors.lastname: Perea
authors.forename: Joel A
authors.initials: JA
authors.lastname: Vazquez-Perez
authors.forename: Steve
authors.initials: S
authors.lastname: Young
authors.forename: Bojana
authors.initials: B
authors.lastname: Rodic-Polic
authors.forename: Markus
authors.initials: M
authors.lastname: Eickmann
authors.forename: Christian
authors.initials: C
authors.lastname: Drosten
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Jilg
authors.forename: Udo
authors.initials: U
authors.lastname: Reischl
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002648:Child; D002675:Child, Preschool; D005260:Female; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D053118:Influenza A Virus, H1N1 Subtype / Q000302:isolation & purification*; D007251:Influenza, Human / Q000175:diagnosis* / Q000821:virology*; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D012680:Sensitivity and Specificity; D014773:Virology / Q000379:methods*; D055815:Young Adult