Analytical and clinical evaluation of the Abbott RealTime hepatitis B sequencing assay.
Long-term nucleoside analogue (NA) treatment leads to selection for drug-resistant mutations in patients undergoing hepatitis B virus (HBV) therapy. The Abbott RealTime HBV Sequencing assay (Abbott assay; Abbott Molecular Inc., Des Plaines, IL, USA) targets the reverse transcriptase region of the polymerase gene and as such has the ability to detect NA resistance-associated mutations in HBV.



We evaluated the analytical performance of the Abbott assay and compared its diagnostic performance to that of a laboratory-developed nested-PCR and sequencing method.



The analytical sensitivity of the Abbott assay was determined using a serially-diluted WHO International Standard. To validate the clinical performances of the Abbott assay and the laboratory-developed assay, 89 clinical plasma samples with various levels of HBV DNA were tested using both assays.



The limit of detection of the Abbott assay, was 210IU/ml and it successfully detected mutations when the mutant types were present at levels ≥20%. Among 89 clinical specimens, 43 and 42 were amplification positive in the Abbott and laboratory-developed assays, respectively, with 87.6% overall agreement (78/89; 95% confidence interval [CI], 78.6-93.4). The Abbott assay failed to detect the minor mutant populations in two specimens, and therefore overall concordance was 85.3% (76/89), and the kappa value was 0.79 (95% CI, 0.67-0.90).



The Abbott assay showed comparable diagnostic performance to laboratory-developed nested PCR followed by direct sequencing, and may be useful as a routine method for detecting HBV NA resistance-associated mutations in clinical laboratory settings.
Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea.
Hee Jae
HJ
Huh
Center for Clinical Medicine, Samsung Biomedical Research Institute, Samsung Medical Center, Seoul, Republic of Korea.
Ji-Youn
JY
Kim
Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea.
Myoung-Keun
MK
Lee
Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea.
Nam Yong
NY
Lee
Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea.
Jong-Won
JW
Kim
Department of Laboratory Medicine and Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea. Electronic address: changski@skku.edu.
Chang-Seok
CS
Ki
D024882:Drug Resistance, Viral; D006515:Hepatitis B virus / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*; D019694:Hepatitis B, Chronic / Q000821:virology; D006801:Humans; D008826:Microbial Sensitivity Tests / Q000379:methods*; D020125:Mutation, Missense; D016133:Polymerase Chain Reaction; D012194:RNA-Directed DNA Polymerase / Q000235:genetics*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods*
D012194:RNA-Directed DNA Polymerase
Direct sequencing; HBV; Mutations; Nested PCR; Resistance