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Automated nucleic acid isolation methods for HDV viral load quantification can lead to viral load underestimation.

Abstract：

BACKGROUND:HDV infection is a cause of severe liver disease. Diagnosis and monitoring of HDV RNA are important to patient management. Since 2012, a WHO standard for HDV RNA quantification has been available; however, the impact of RNA extraction methods on HDV viral load quantification has never been evaluated. METHODS:The aim of this study was to compare four commonly used automated nucleic acid (NA) extraction methods (AmpliPrep, MagNA Pure, QIAcube QBK and QIAcube VRK) with a manual RNA extraction method (Instant Virus RNA/DNA kit) and evaluate the possible effect of each method on HDV RNA yield with subsequent amplification with the Robogene HDV assay. Serum samples from HDV-positive patients taken before treatment with pegylated interferon-α2a and at treatment weeks 12 and 48 were studied. RESULTS:The automated extraction methods MagNA Pure, Ampliprep and QIAcube VRK extraction led to about 10-fold lower HDV RNA values compared with the manual method of NA extraction, while the difference was smaller with QIAcube QBK (about 6-fold lower). The median viral load was 10,665 IU/ml for the manual method, 445 IU/ml for AmpliPrep, 3,209 IU/ml for MagNA Pure, 2,060 IU/ml for QIAcube QBK and 3,568 IU/ml for QIAcube VRK. Use of MagNA Pure led to misclassification of two on-treatment samples with low viral load as being false negative. CONCLUSIONS:The NA extraction method had a significant impact on the measured HDV viral loads determined by the commonly used Robogene assay, with the manual RNA method yielding consistently higher values of viral load. ClinicalTrials.gov Identifier: NCT00932971.

journal_name

Antivir Ther

journal_title

Antiviral therapy

authors

Bremer B,Anastasiou OE,Ciesek S,Wedemeyer H

doi

10.3851/IMP3281

subject

Has Abstract

pub_date

2019-01-01 00:00:00

pages

117-123

issue

2

eissn

1359-6535

issn

2040-2058

journal_volume

24

pub_type

杂志文章

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