Using NS5B Sequencing for Hepatitis C Virus Genotyping Reveals Discordances with Commercial Platforms

Chueca, Natalia; Rivadulla, I; Lovatti, Ruben; Reina, Gabriel; Blanco, Ana; Fernández-Caballero, Jose Ángel; Cardeñoso, Laura; Rodríguez-Granjer, Javier; Fernandez‐Alonso, M.; Aguilera, Antonio; Alvarez, Marta; Galán, Juan Carlos; García, Féderico

doi:10.1371/journal.pone.0153754

Cited by 35 publications

(24 citation statements)

References 28 publications

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“…we also found some related to the methods mainly employed herein for HCV genotyping; this limitation refers to the correct viral subtype determination of genotype 1, especially when comparing commercial to reference methods based on sequencing and phylogenetic analyses of the NS5B region of HCV. [34][35][36][37][38][39][40] In this regard, the inclusion of HCV genotyping by sequencing-based technologies of different viral genome regions may notably improve viral subtype knowledge, which is still important for choosing the most appropriate treatment regimen and for determining resistance-associated substitutions (RAS). A final limitation is that our study lacks immigration data.…”

Section: Discussionmentioning

confidence: 99%

Prevalence and distribution of hepatitis C virus genotypes in Spain during the 2000‐2015 period (the GEHEP 005 study)

Aguilera

Navarro

Rodríguez‐Frías

et al. 2017

Journal of Viral Hepatitis

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We report the largest study on the prevalence and distribution of HCV genotypes in Spain (2000-2015), and we relate them with clinical, epidemiological and virological factors. Patients from 29 hospitals in 10 autonomous communities (Andalusia, Aragon, Castilla-Leon, Catalonia, Galicia, Canary Islands, Madrid Community, Valencian Community, Murcia Region and Basque Country) have been studied. Annual distribution of HCV genotypes and subtypes, as well as gender, age, transmission route, HIV and/or HBV coinfection, and treatment details were recorded. We included 48595 chronically HCV-infected patients with the following characteristics: median age 51 years (IQR, 44-58), 67.9% male, 19.1% HIV-coinfected, 23.5% HBV-coinfected. Parenteral transmission route was the most frequent (58.7%). Genotype distribution was 66.9% GT1 (24.9% subtype 1a and 37.9% subtype 1b), 2.8% GT2, 17.3% GT3, 11.4% GT4 and 0.1% GT5 and 0.02% GT6. LiPA was the most widely HCV genotyping test used (52.4%). HCV subtype 1a and genotypes 3 and 4 were closely associated with male gender, parenteral route of infection and HIV and HBV coinfection; in contrast, subtype 1b and genotype 2 were associated with female gender, nonparenteral route and mono-infection. Age was related to genotype distribution, and different patterns of distribution and biodiversity index were observed between different geographical areas. Finally, we describe how treatment and changes in transmission routes may have affected HCV genotype prevalence and distribution patterns. We present the most recent data on molecular epidemiology of hepatitis C virus in Spain. This study confirms that genotype distributions vary with age, sex, HIV and HBV coinfection and within geographical areas and epidemiological groups.

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Section: Discussionmentioning

confidence: 99%

Prevalence and distribution of hepatitis C virus genotypes in Spain during the 2000‐2015 period (the GEHEP 005 study)

Aguilera

Navarro

Rodríguez‐Frías

et al. 2017

Journal of Viral Hepatitis

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“…Thus, the true prevalence of GT6 could potentially be slightly underestimated when the strains are misclassified as GT1 when using 5'-non coding region analysis. Using sequencing approaches based on more informative genomic regions should be promoted, particularly to document transmission in at-risk populations 20 . For all sequenced strains, concordant phylogenic analyses whatever the chosen region, NS3, NS5A or NS5B were obtained and most sequences clustered with GenBank available data.…”

Section: Discussionmentioning

confidence: 99%

Genetic diversity of genotype 6 HCV infections in France: epidemiology and consequences for treatment strategy

Pronier¹,

Helene²,

Celine³

et al. 2018

Journal of Hepatology

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“…However, more concerning are cases of genotype misclassification because these often remain unnoticed until the clinical consequences of misclassification are evident, for example, after treatment failure with an HCV genotype 1-specific DAA combination. Recent studies have shown a substantial number of HCV genotype/subtype misclassifications, occurring in 2% to more than 50% of patients, depending on the genotyping assay and population studies (9)(10)(11)(12). However, it should be stressed that the great majority of misclassifications account for erroneous assignment of the HCV genotype 1 subtype, occasionally with important clinical consequences (9)(10)(11)(12).…”

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confidence: 99%

“…When a probe-based HCV genotyping assay fails to determine the HCV genotype (or HCV subtype) and if capacities allow, laboratories often turn to population Sanger sequencing of a carefully selected, usually relatively short part of the HCV genome using commercial sequencing-based assays or in-house sequencing protocols (10). Although population sequencing is considered more accurate than probe-based assays, in certain circumstances, population sequencing can even be inferior (10).…”

mentioning

confidence: 99%

“…Although population sequencing is considered more accurate than probe-based assays, in certain circumstances, population sequencing can even be inferior (10). The main disadvantage of using population sequencing for HCV genotyping is that it can determine only the most prevalent genomic variant(s) and thus underperforms in the case of simultaneous infection with different HCV genotypes/subtypes, colloquially known as mixed infection.…”

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Commentary: Next-Generation Sequencing: a Diagnostic One-Stop Shop for Hepatitis C?

Poljak

2016

J Clin Microbiol

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Before starting chronic hepatitis C treatment, the viral genotype/subtype has to be accurately determined and potentially coupled with drug resistance testing. Due to the high genetic variability of the hepatitis C virus, this can be a demanding task that can potentially be streamlined by viral whole-genome sequencing using Hepatitis C virus (HCV) is well-known for its high genetic variability, with seven distinct major HCV genotypes and nearly 70 officially assigned subtypes (1). For this reason, even though revolutionary progress has recently been made with direct-acting antivirals (DAAs) against HCV with high efficacy rates and an improved safety profile, there is still no universal antiviral treatment available that works equally effectively against all HCV genotypes (and especially subtypes) and across all patient populations (2, 3). Recent approval by the U.S. Food and Drug Administration (on 28 June 2016) of the first all-oral, pan-genotypic, single-tablet regimen for treating HCV-infected adults with and without cirrhosis based on a combination of sofosbuvir (NS5B inhibitor) and velpatasvir (pan-genotypic NS5A inhibitor) is an important step forward, but only time will tell whether the excellent results obtained in the clinical trials across all six major HCV genotypes and their subtypes (4, 5) will be reproduced in real-life settings.Accurate determination of the HCV genotype and selected HCV subtypes before treatment initiation has been mandatory for more than 2 decades in order to select the most appropriate treatment regimen and to decide on its duration and the use of ribavirin, and it is here to stay for at least the next few years (6). A few sequencing-based and several non-sequencing-based HCV genotyping assays are commercially available and are used worldwide for routine determination of the HCV genotype and selected subtypes. The non-sequencing-based HCV genotyping assays are mainly founded on reverse hybridization or real-time PCR. Because both assay groups belong to DNA probe-based technologies, they occasionally fail to provide unambiguous results, particularly when the genetic diversity of the target is high, as in the case of HCV. Even with the last versions of commercial non-sequencing-based HCV genotyping assays, the HCV genotype/subtype fails to be assigned in 5 to 10% of patients (7,8). The failure to obtain an unambiguous genotyping result by probe-based assays is frustrating, but it can be overcome by the use of a supplementary test (9). However, more concerning are cases of genotype misclassification because these often remain unnoticed until the clinical consequences of misclassification are evident, for example, after treatment failure with an HCV genotype 1-specific DAA combination. Recent studies have shown a substantial number of HCV genotype/subtype misclassifications, occurring in 2% to more than 50% of patients, depending on the genotyping assay and population studies (9-12). However, it should be stressed that the great majority of misclassifications account for erroneous ...

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Using NS5B Sequencing for Hepatitis C Virus Genotyping Reveals Discordances with Commercial Platforms

Cited by 35 publications

References 28 publications

Prevalence and distribution of hepatitis C virus genotypes in Spain during the 2000‐2015 period (the GEHEP 005 study)

Prevalence and distribution of hepatitis C virus genotypes in Spain during the 2000‐2015 period (the GEHEP 005 study)

Genetic diversity of genotype 6 HCV infections in France: epidemiology and consequences for treatment strategy

Commentary: Next-Generation Sequencing: a Diagnostic One-Stop Shop for Hepatitis C?

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