Development of Hepatitis C Virus Genotyping by Real-Time PCR Based on the NS5B Region
dc.contributor.author | Nakatani, Sueli M. | |
dc.contributor.author | Santos, Carlos A. | |
dc.contributor.author | Riediger, Irina N. | |
dc.contributor.author | Krieger, Marco A. | |
dc.contributor.author | Duarte, Cesar A. B. | |
dc.contributor.author | Lacerda, Marco A. | |
dc.contributor.author | Biondo, Alexander W. | |
dc.contributor.author | Carilho, Flair J. | |
dc.contributor.author | Ono-Nita, Suzane K. | |
dc.contributor.department | Medicine, School of Medicine | en_US |
dc.date.accessioned | 2020-05-19T12:06:24Z | |
dc.date.available | 2020-05-19T12:06:24Z | |
dc.date.issued | 2010-04-13 | |
dc.description.abstract | Hepatitis C virus (HCV) genotyping is the most significant predictor of the response to antiviral therapy. The aim of this study was to develop and evaluate a novel real-time PCR method for HCV genotyping based on the NS5B region. Methodology/Principal Findings Two triplex reaction sets were designed, one to detect genotypes 1a, 1b and 3a; and another to detect genotypes 2a, 2b, and 2c. This approach had an overall sensitivity of 97.0%, detecting 295 of the 304 tested samples. All samples genotyped by real-time PCR had the same type that was assigned using LiPA version 1 (Line in Probe Assay). Although LiPA v. 1 was not able to subtype 68 of the 295 samples (23.0%) and rendered different subtype results from those assigned by real-time PCR for 12/295 samples (4.0%), NS5B sequencing and real-time PCR results agreed in all 146 tested cases. Analytical sensitivity of the real-time PCR assay was determined by end-point dilution of the 5000 IU/ml member of the OptiQuant HCV RNA panel. The lower limit of detection was estimated to be 125 IU/ml for genotype 3a, 250 IU/ml for genotypes 1b and 2b, and 500 IU/ml for genotype 1a. Conclusions/Significance The total time required for performing this assay was two hours, compared to four hours required for LiPA v. 1 after PCR-amplification. Furthermore, the estimated reaction cost was nine times lower than that of available commercial methods in Brazil. Thus, we have developed an efficient, feasible, and affordable method for HCV genotype identification. | en_US |
dc.eprint.version | Final published version | en_US |
dc.identifier.citation | Nakatani SM, Santos CA, Riediger IN, Krieger MA, Duarte CAB, Lacerda MA, et al. (2010) Development of Hepatitis C Virus Genotyping by Real-Time PCR Based on the NS5B Region. PLoS ONE 5(4): e10150. https://doi.org/10.1371/journal.pone.0010150 | en_US |
dc.identifier.uri | https://hdl.handle.net/1805/22801 | |
dc.language.iso | en_US | en_US |
dc.publisher | Public Library of Science | en_US |
dc.relation.isversionof | 10.1371/journal.pone.0010150 | en_US |
dc.relation.journal | PLoS ONE | en_US |
dc.rights | Attribution 4.0 International | * |
dc.rights.uri | http://creativecommons.org/licenses/by/4.0/ | * |
dc.source | Publisher | en_US |
dc.subject | Polymerase chain reaction | en_US |
dc.subject | Genotyping | en_US |
dc.subject | Hepatitis C virus | en_US |
dc.subject | Untranslated regions | en_US |
dc.subject | Brazil | en_US |
dc.subject | RNA extraction | en_US |
dc.subject | Reverse transcription | en_US |
dc.subject | Phylogenetic analysis | en_US |
dc.title | Development of Hepatitis C Virus Genotyping by Real-Time PCR Based on the NS5B Region | en_US |
dc.type | Article | en_US |
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