title: Assessment of a Novel Automatic Real-Time PCR Assay on the Cobas 4800 Analyzer as a Screening Platform for Hepatitis C Virus Genotyping in Clinical Practice: Comparison with Massive Sequencing.
abstract: The unequivocal identification of hepatitis C virus (HCV) subtypes 1a/1b and genotypes 2 to 6 is required for optimizing the effectiveness of interferon-free, direct-acting antiviral therapies. We compared the performance of a new real-time HCV genotyping assay used on the Cobas 4800 system (C4800) with that of high-resolution HCV subtyping (HRCS). In total, 502 samples were used, including 184 samples from chronic HCV patients (from routine laboratory activity during April 2016), 5 stored samples with double HCV genotype infections for testing the limitations of the method, and 313 samples from a screening protocol implemented in our hospital (from May to August 2016) based on the new method to further determine its genotyping accuracy. A total of 282 samples, including 171 from April 2016 (the 13 remaining had too low of a viral load for HRCS), 5 selected with double infections, and 106 from screening, were analyzed by both methods, and 220 were analyzed only by the C4800. The C4800 correctly subtyped 125 of 126 1a/1b samples, and the 1 remaining sample was reported as genotype 1. The C4800 correctly genotyped 38 of 45 non-1a/1b samples (classified by HRCS), and it reported the remaining 7 samples as indeterminate. One hundred two of 106 non-1a/1b genotype samples that were identified using the C4800 for screening were confirmed by HRCS. In the 4 remaining samples, 3 were correctly reported as genotype 1 (without defining the subtype) and 1 was reported as indeterminate. None of the samples were misgenotyped. Four of 7 samples with double HCV infections were correctly genotyped by the C4800. Excluding the 5 selected double-infected samples, the C4800 showed 95.7% concordant results for genotyping HCVs 2 to 6 and 1a/1b subtyping, and 99.2% concordance for subtyping 1a/1b single infections in clinical samples. To improve laboratory workflow, we propose using the C4800 as a first-line test for HCV genotyping and 1a/1b classification, followed by transferring non-1a/1b samples to a center where HRCS is available, if further characterization is needed.
authors.affiliation: Liver Pathology Unit, Departments of Biochemistry and Microbiology, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.
authors.forename: Leonardo
authors.initials: L
authors.lastname: Nieto-Aponte
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain josep.quer@vhir.org frarodri@vhebron.net.|Viral Hepatitis Laboratory, Liver Unit-Internal Medicine, Vall d'Hebron Research Institute (VHIR), Hospital Universitari Vall d'Hebron (HUVH), Barcelona, Spain.
authors.forename: Josep
authors.initials: J
authors.lastname: Quer
authors.affiliation: Liver Pathology Unit, Departments of Biochemistry and Microbiology, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain.
authors.forename: Alicia
authors.initials: A
authors.lastname: Ruiz-Ripa
authors.affiliation: Liver Pathology Unit, Departments of Biochemistry and Microbiology, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain.
authors.forename: David
authors.initials: D
authors.lastname: Tabernero
authors.affiliation: Liver Pathology Unit, Departments of Biochemistry and Microbiology, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.
authors.forename: Carolina
authors.initials: C
authors.lastname: Gonzalez
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain.|Viral Hepatitis Laboratory, Liver Unit-Internal Medicine, Vall d'Hebron Research Institute (VHIR), Hospital Universitari Vall d'Hebron (HUVH), Barcelona, Spain.|Roche Diagnostics SL, Sant Cugat del Vallès, Spain.
authors.forename: Josep
authors.initials: J
authors.lastname: Gregori
authors.affiliation: Liver Pathology Unit, Departments of Biochemistry and Microbiology, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Vila
authors.affiliation: Liver Pathology Unit, Departments of Biochemistry and Microbiology, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.
authors.forename: Miriam
authors.initials: M
authors.lastname: Asensio
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain.|Viral Hepatitis Laboratory, Liver Unit-Internal Medicine, Vall d'Hebron Research Institute (VHIR), Hospital Universitari Vall d'Hebron (HUVH), Barcelona, Spain.
authors.forename: Damir
authors.initials: D
authors.lastname: Garcia-Cehic
authors.affiliation: Liver Pathology Unit, Departments of Biochemistry and Microbiology, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.
authors.forename: Gerardo
authors.initials: G
authors.lastname: Ruiz
authors.affiliation: Viral Hepatitis Laboratory, Liver Unit-Internal Medicine, Vall d'Hebron Research Institute (VHIR), Hospital Universitari Vall d'Hebron (HUVH), Barcelona, Spain.
authors.forename: Qian
authors.initials: Q
authors.lastname: Chen
authors.affiliation: Viral Hepatitis Laboratory, Liver Unit-Internal Medicine, Vall d'Hebron Research Institute (VHIR), Hospital Universitari Vall d'Hebron (HUVH), Barcelona, Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Ordeig
authors.affiliation: Viral Hepatitis Laboratory, Liver Unit-Internal Medicine, Vall d'Hebron Research Institute (VHIR), Hospital Universitari Vall d'Hebron (HUVH), Barcelona, Spain.
authors.forename: Meritxell
authors.initials: M
authors.lastname: Llorens
authors.affiliation: Liver Pathology Unit, Departments of Biochemistry and Microbiology, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.
authors.forename: Montserrat
authors.initials: M
authors.lastname: Saez
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain.|Liver Unit, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.
authors.forename: Juan I
authors.initials: JI
authors.lastname: Esteban
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain.|Liver Unit, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.
authors.forename: Rafael
authors.initials: R
authors.lastname: Esteban
authors.affiliation: Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain.|Liver Unit, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.
authors.forename: Maria
authors.initials: M
authors.lastname: Buti
authors.affiliation: Microbiology Department, Hospital Universitari Vall d'Hebron (HUVH), Vall d'Hebron Research Institute (VHIR), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain.
authors.forename: Tomas
authors.initials: T
authors.lastname: Pumarola
authors.affiliation: Liver Pathology Unit, Departments of Biochemistry and Microbiology, Hospital Universitari Vall d'Hebron (HUVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain josep.quer@vhir.org frarodri@vhebron.net.|Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Instituto de Salud Carlos III, Madrid, Spain.|Viral Hepatitis Laboratory, Liver Unit-Internal Medicine, Vall d'Hebron Research Institute (VHIR), Hospital Universitari Vall d'Hebron (HUVH), Barcelona, Spain.
authors.forename: Francisco
authors.initials: F
authors.lastname: Rodriguez-Frias
mesh_terms: D057205:Automation, Laboratory / Q000379:methods*; D060005:Genotyping Techniques / Q000379:methods*; D016174:Hepacivirus / Q000145:classification* / Q000235:genetics*; D019698:Hepatitis C, Chronic / Q000175:diagnosis / Q000821:virology; D006801:Humans; D008403:Mass Screening / Q000379:methods*; D011446:Prospective Studies; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA / Q000379:methods*
keywords: diagnostics; genotype identification; hepatitis C virus; molecular subtyping