Characterization of Samples Identified as Hepatitis C Virus Genotype 1 without Subtype by Abbott RealTi m e HCV Genotype II Assay Using the New Abbott HCV Genotype Plus RUO Test

Abstract

Hepatitis C virus (HCV) genotyping continues to be relevant for therapeutic strategies. Some samples are reported as genotype 1 (gt 1) without subtype by the Abbott RealTime HCV Genotype II (GT II) test. To characterize such samples further, the Abbott HCV Genotype Plus RUO (Plus) assay, which targets the core region for gt 1a, gt 1b, and gt 6 detection, was evaluated as a reflex test in reference to NS5B or 5′-untranslated region (UTR)/core region sequencing. Of 3,626 routine samples, results of gt 1 without subtype were received for 171 samples (4.7%), accounting for 11.5% of gt 1 specimens. The Plus assay and sequencing were applied to 98 of those samples. NS5B or 5′-UTR/core region sequencing was successful for 91/98 specimens (92.9%). Plus assay and sequencing results were concordant for 87.9% of specimens (80/91 samples). Sequencing confirmed Plus assay results for 82.6%, 85.7%, 100%, and 89.3% of gt 1a, gt 1b, gt 6, and non-gt 1a/1b/6 results, respectively. Notably, 12 gt 6 samples that had been identified previously as gt 1 without subtype were assigned correctly here; for 25/28 samples reported as “not detected” by the Plus assay, sequencing identified the samples as gt 1 with subtypes other than 1a/1b. The genetic variability of HCV continues to present challenges for the current genotyping platforms regardless of the applied methodology. Samples identified by the GT II assay as gt 1 without subtype can be further resolved and reliably characterized by the new Plus assay.

Reliable resolution of ambiguous hepatitis C virus genotype 1 results with the Abbott HCV Genotype Plus RUO assay

“…While commercial genotyping assays use sub-genomic regions such as core or NS5B in addition to the more conserved 5′ untranslated (5′NC) region, the high genetic variability and small differences between genotypes and subtypes still remain a challenge for both real-time PCR and line probe-based HCV genotyping assays. This also applies to HCV-1 subtyping 11–21 . In case of ambiguous results, the use of a second genotyping method may help guiding treatment selection.…”

“…However, even this method is not able to resolve every individual sample, e.g. due to failure of amplification 11,13,17,23 . Furthermore, the procedure is considered impractical for most clinical laboratories because it is time-consuming, less sensitive 21 , may be technically challenging, and does not readily allow for detecting mixed-type infections.…”

Indeterminate genotypes of hepatitis C virus by the Abbott RealTime HCV Genotype II assay in Morocco. About eight cases resolved by a sequencing method

“…A similar study reported among 3626 samples, a rate of 4.7% (171) of genotypes 1 without subtype. Sequencing was applied to 98 of these samples and was successful for 92.9% of the specimen (91/98) …”

“…Hence, they developed the Abbott RealTi m e HCV Genotype Plus RUO assay. Recent studies assessed this new add‐on to the HCV genotyping kit, which resolves several indeterminate genotypes 1a, 1b, and 6 …”

Ultra-Deep Sequencing Characterization of HCV Samples with Equivocal Typing Results Determined with a Commercial Assay

“…The additional HCV genome targets used for subtyping are in the core region for the Versant HCV Genotype 2.0 and in the NS5B region for the Abbott RealTime HCV Genotype v 2.0. Furthermore, Abbott improved the performance of RealTime HCV Genotype II (targeting the 5′ UTR and NS5B regions), with the ABBOTT Genotype PLUS RUO test, where the core region is used to further characterize genotype 1 unsubtyped samples [7]. Despite the technical improvements in HCV genotyping tools, indeterminate, mixed and unspecified subtype results are obtained in a small but not irrelevant proportion of samples in the daily clinical practice [8].…”