title: Comparison of real-time multiplex human papillomavirus (HPV) PCR assays with the linear array HPV genotyping PCR assay and influence of DNA extraction method on HPV detection.
abstract: Real-time human papillomavirus (HPV) type-specific multiplex PCR assays were developed to detect HPV DNA in specimens collected for the efficacy determination of the quadrivalent HPV (type 6, 11, 16, and 18) L1 virus-like particle (VLP) vaccine (Gardasil). We evaluated the concordance between type-specific multiplex HPV PCR and the widely used, commercially available Roche Linear Array genotyping PCR assay. Female genital swab specimens were tested for the presence of L1, E6, and E7 sequences of HPV type 6 (HPV6), HPV11, HPV16, HPV18, HPV31, HPV45, HPV52, and HPV58 and E6 and E7 sequences of HPV33, HPV35, HPV39, HPV51, HPV56, and HPV59 in type- and gene-specific real-time multiplex PCR assays. Specimens were also tested for the presence of L1 sequences using two versions of the Roche Linear Array genotyping assay. Measures of concordance of a modified version of the Linear Array and the standard Linear Array PCR assay were evaluated. With specimen DNA extraction using the Qiagen Spin blood kit held as the constant, multiplex PCR assays detect more HPV-positive specimens for the 14 HPV types common to both than either version of the Linear Array HPV genotyping assay. Type-specific agreements between the assays were good, at least 0.838, but were often driven by negative agreement in HPV types with low prevalence, as evidenced by reduced proportions of positive agreement. Overall HPV status agreements ranged from 0.615 for multiplex PCR and standard Linear Array to 0.881 for multiplex PCR and modified Linear Array. An alternate DNA extraction technique, that used by the Qiagen MinElute kit, impacted subsequent HPV detection in both the multiplex PCR and Linear Array assays.
authors.affiliation: Vaccine Basic Research Department, Merck & Co, Inc, West Point, Pennsylvania 19454-1099, USA. christine_roberts@merck.com
authors.forename: Christine C
authors.initials: CC
authors.lastname: Roberts
authors.forename: Ryan
authors.initials: R
authors.lastname: Swoyer
authors.forename: Janine T
authors.initials: JT
authors.lastname: Bryan
authors.forename: Frank J
authors.initials: FJ
authors.lastname: Taddeo
mesh_terms: D036022:Capsid Proteins / Q000235:genetics; D004279:DNA, Viral / Q000302:isolation & purification*; D005260:Female; D005836:Genitalia, Female / Q000821:virology; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods; D009856:Oncogene Proteins, Viral / Q000235:genetics; D027383:Papillomaviridae / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D030361:Papillomavirus Infections / Q000175:diagnosis* / Q000821:virology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013048:Specimen Handling / Q000379:methods*; D014773:Virology / Q000379:methods
chemical_list: D036022:Capsid Proteins; D004279:DNA, Viral; C114580:HPV L1 protein, Human papillomavirus; D009856:Oncogene Proteins, Viral