Performance of the Applied Biosystems ViroSeq Human Immunodeficiency Virus Type 1 (HIV-1) Genotyping System for Sequence-Based Analysis of HIV-1 in Pediatric Plasma Samples

Cunningham, Shawn; Ank, Bonnie J.; Lewis, Dan; Lü, Wei; Wantman, Michael; J, Dileanis; Jackson, J. Brooks; Palumbo, Paul; Krogstad, Paul; Eshleman, Susan H.

doi:10.1128/jcm.39.4.1254-1257.2001

Cited by 49 publications

(30 citation statements)

References 12 publications

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“…Sample cross-contamination is minimized in the ViroSeq system by using a single nonnested PCR for amplification, and by using a dUTP/UNG contamination control system. There was no evidence of sample cross-contamination in this study or in previous reports (2,4).…”

Section: Discussioncontrasting

confidence: 43%

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Sensitivity and Specificity of the ViroSeq Human Immunodeficiency Virus Type 1 (HIV-1) Genotyping System for Detection of HIV-1 Drug Resistance Mutations by Use of an ABI PRISM 3100 Genetic Analyzer

Eshleman

Crutcher²,

Petrauskene³

et al. 2005

J Clin Microbiol

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The ViroSeq human immunodeficiency virus type 1 (HIV-1) genotyping system is an integrated system for identification of drug resistance mutations in HIV-1 protease and reverse transcriptase (RT). Reagents are included for sample preparation, reverse transcription, PCR amplification, and sequencing. Software is provided to assemble and edit sequence data and to generate a drug resistance report. We determined the sensitivity and specificity of the ViroSeq system for mutation detection using an ABI PRISM 3100 genetic analyzer with a set of clinical samples and recombinant viruses. Twenty clinical plasma samples (viral loads, 1,800 to 10,500 copies/ml) were characterized by cloning and sequencing individual viral variants. Twelve recombinant-virus samples (viral loads, approximately 2,000 to 5,000 copies/ml) were also prepared. Eleven recombinant-virus samples contained drug resistance mutations as 40% mixtures. One recombinant-virus sample contained an insertion at codon 69 in RT (100% mutant). Plasma and recombinant-virus samples were analyzed using the ViroSeq system. Each sample was analyzed on three consecutive days at each of three testing laboratories. The sensitivity of mutation detection was 99.65% for the clinical plasma samples and 99.7% for the recombinant-virus preparations. The specificity of mutation detection was 99.95% for the clinical samples and 100% for the recombinant-virus mixtures. The base calling accuracy of the 3100 instrument was 99.91%. Mutations in clinical plasma samples and recombinant-virus samples were detected with high sensitivity and specificity, including mutations present as mixtures. This report supports the use of the ViroSeq system for identification of drug resistance mutations in HIV-1 protease and RT genes.

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Section: Discussioncontrasting

confidence: 43%

“…The system reports the evidence of resistance to FDA-approved drugs based on the detection of antiretroviral drug resistance mutations. Previous studies report a high level of performance of the system for analysis of both subtype B and non-subtype B samples (2,4,7).…”

mentioning

confidence: 99%

Sensitivity and Specificity of the ViroSeq Human Immunodeficiency Virus Type 1 (HIV-1) Genotyping System for Detection of HIV-1 Drug Resistance Mutations by Use of an ABI PRISM 3100 Genetic Analyzer

Eshleman

Crutcher²,

Petrauskene³

et al. 2005

J Clin Microbiol

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“…These subgenomic regions included (1) a partial pol sequence spanning the region encoding HIV-1 protease and the first 335 amino acids of reverse transcriptase, which corresponds to the sequence produced by ViroSeq, [38][39][40][41] nt positions 2,253-3,554; (2) partial env sequences spanning the region encoding the gp120 V1C5 region, [42][43][44] nt positions 6,570-7,757; (3) ''product 2'' spanning the 3¢-end of gag and almost the entire pol, 45 nt positions 1,486-5,058; and (4) ''product 4'' spanning vpu, env, nef, and TATA-box in the U3 region of 3¢-LTR, 45 nt positions 5,967-9,517. In addition, combinations of the targeted subregions included gag + pol, gag + env, pol + env, gag + pol + env, and product 2 + product 4.…”

Section: Analyzed Subgenomic Regions Of the Hiv-1c Genomementioning

confidence: 99%

Importance of Viral Sequence Length and Number of Variable and Informative Sites in Analysis of HIV Clustering

Novitsky

Moyo

Lei

et al. 2015

AIDS Research and Human Retroviruses

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To improve the methodology of HIV cluster analysis, we addressed how analysis of HIV clustering is associated with parameters that can affect the outcome of viral clustering. The extent of HIV clustering and tree certainty was compared between 401 HIV-1C near full-length genome sequences and subgenomic regions retrieved from the LANL HIV Database. Sliding window analysis was based on 99 windows of 1,000 bp and 45 windows of 2,000 bp. Potential associations between the extent of HIV clustering and sequence length and the number of variable and informative sites were evaluated. The near full-length genome HIV sequences showed the highest extent of HIV clustering and the highest tree certainty. At the bootstrap threshold of 0.80 in maximum likelihood (ML) analysis, 58.9% of near full-length HIV-1C sequences but only 15.5% of partial pol sequences (ViroSeq) were found in clusters. Among HIV-1 structural genes, pol showed the highest extent of clustering (38.9% at a bootstrap threshold of 0.80), although it was significantly lower than in the near full-length genome sequences. The extent of HIV clustering was significantly higher for sliding windows of 2,000 bp than 1,000 bp. We found a strong association between the sequence length and proportion of HIV sequences in clusters, and a moderate association between the number of variable and informative sites and the proportion of HIV sequences in clusters. In HIV cluster analysis, the extent of detectable HIV clustering is directly associated with the length of viral sequences used, as well as the number of variable and informative sites. Near full-length genome sequences could provide the most informative HIV cluster analysis. Selected subgenomic regions with a high extent of HIV clustering and high tree certainty could also be considered as a second choice.

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“…The genotypic assays used to monitor HIV drug resistance include bulk sequencing (6,13,23,29,34), oligonucleotide ligation assay (11,14), line probe assay (37), microarray hybridization (38), heteroduplex formation (1), allele-specific PCR (S. Palmer, 11th Conference on Retroviruses and Opportunistic Infections), and single-genome sequencing (30). Other than allele-specific PCR, these assays are limited in their sensitivities for the detection of drug-resistant variants present at low frequencies.…”

Section: Discussionmentioning

confidence: 99%

Sensitive Phenotypic Detection of Minor Drug-Resistant Human Immunodeficiency Virus Type 1 Reverse Transcriptase Variants

et al. 2005

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Detection of drug-resistant variants is important for the clinical management of human immunodeficiency virus type 1 (HIV-1) infection and for studies on the evolution of drug resistance. Here we show that hybrid elements composed of the Saccharomyces cerevisiae retrotransposon Ty1 and the reverse transcriptase (RT) of HIV-1 are useful tools for detecting, monitoring, and isolating drug-resistant reverse transcriptases. This sensitive phenotypic assay is able to detect nonnucleoside reverse transcriptase inhibitor-resistant RT domains derived from mixtures of infectious molecular clones of HIV-1 in plasma and from clinical samples when the variants comprise as little as 0.3 to 1% of the virus population. Our assay can characterize the activities and drug susceptibilities of both known and novel reverse transcriptase variants and should prove useful in studies of the evolution and clinical significance of minor drug-resistant viral variants.

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Performance of the Applied Biosystems ViroSeq Human Immunodeficiency Virus Type 1 (HIV-1) Genotyping System for Sequence-Based Analysis of HIV-1 in Pediatric Plasma Samples

Cited by 49 publications

References 12 publications

Sensitivity and Specificity of the ViroSeq Human Immunodeficiency Virus Type 1 (HIV-1) Genotyping System for Detection of HIV-1 Drug Resistance Mutations by Use of an ABI PRISM 3100 Genetic Analyzer

Sensitivity and Specificity of the ViroSeq Human Immunodeficiency Virus Type 1 (HIV-1) Genotyping System for Detection of HIV-1 Drug Resistance Mutations by Use of an ABI PRISM 3100 Genetic Analyzer

Importance of Viral Sequence Length and Number of Variable and Informative Sites in Analysis of HIV Clustering

Sensitive Phenotypic Detection of Minor Drug-Resistant Human Immunodeficiency Virus Type 1 Reverse Transcriptase Variants

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