Performance of the ViroSeq HIV-1 genotyping system v2.0 on HIV-1 strains circulating in Senegal

Thiam, Moussa; Diop-Ndiaye, Halimatou; Kébé, Khadim; Vidal, Nicole; Diakhate-Lô, Rokhaya; Diouara, Abou Abdallah Malick; Leye, Nafissatou; Ndiaye, O.; Sow, A.I.; Ngom-Gueye, Ndeye Fatou; Mboup, Souleymane; Touré-Kâne, Coumba

doi:10.1016/j.jviromet.2012.11.044

Cited by 14 publications

(11 citation statements)

References 15 publications

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“…This nucleic acid or signal amplification assays rely on the use of sequence specific primers and/or probes. Therefore, HIV-1 increased heterogeneity may affect assay performance as the presence of natural polymorphisms in the target regions may reduce or inhibit hybridization thereby compromising the reliability of viral load quantification and genotyping test [51], [52]. Our “in-house” genotyping method was first designed using primers validated for subtype B.…”

Section: Discussionmentioning

confidence: 99%

“…In fact, the five recombinants variants (BG, BC and BF) were successfully sequenced with our genotyping system. ViroSeq HIV-1 Genotyping System v2.0 has shown a decrease in performance on HIV-1 non-B strains in certain countries however, in Panama this diagnostic test has proven to perform well [52]. Recent studies evaluating viral load diagnostics test on current HIV genetic complexity found a lower correlations with subtypes C samples [53].…”

Section: Discussionmentioning

confidence: 99%

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Molecular Epidemiology of HIV-1 in Panama: Origin of Non-B Subtypes in Samples Collected from 2007 to 2013

et al. 2014

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Phylogenetic studies have suggested that the HIV-1 epidemic in the Americas is mainly dominated by HIV subtype B. However, countries of South America and the Caribbean have recently reported changes in their circulating HIV-1 genetic profiles. The aim of this study was to characterize the molecular profile of the HIV-1 epidemic in Panama by the analysis of 655 polymerase gene (pol) sequences that were obtained from HIV-infected Panamanians diagnosed between 1987 and 2013. Blood samples were collected from recently infected, antiretroviral drug-naïve and treatment-experienced subjects since mid-2007 to 2013. Viral RNA from plasma was extracted and sequences of HIV protease and reverse transcriptase genes were obtained. Bootscanning and phylogenetic methods were used for HIV subtyping and to trace the putative origin of non-B subtype strains. Our results showed that HIV-1 infections in Panama are dominated by subtype B (98.9%). The remaining 1.1% is represented by a diverse collection of recombinant variants including: three URFs_BC, one CRF20_BG, and one CRF28/29_BF, in addition to one subtype F1 and one subtype C, none of which were previously reported in Panama. The non-B subtype variants detected in Panama were probably introduced from Brazil (subtype F1 and CRF28/29_BF), Cuba (CRF20_BG), Dominican Republic (URFs_BC) and India (subtype C). Panama is the geographical vertex that connects the North with South America and the Caribbean through trade and cultural relations, which may explain the observed introductions of non-B subtype HIV-1 variants from both the Caribbean and South America into this Central American country.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Molecular Epidemiology of HIV-1 in Panama: Origin of Non-B Subtypes in Samples Collected from 2007 to 2013

et al. 2014

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“…Unlike conventional qPCR, an a priori understanding of all probe-binding site variations does not need to be known; PANDAA can adapt de novo secondary polymorphisms within the probebinding site. By virtue of the adaptation process, PANDAA is a subtype agnostic assay, an issue that has contributed to the high failure rate of commercial Sanger sequencing assays for non-B subtypes (33,34). As our algorithm can incorporate either global or regional subtype prevalence data, an assay that incorporates local HIV-1 sequence diversity in a specific geographical region can be readily designed, which may facilitate local research efforts in LMICs.…”

Section: Discussionmentioning

confidence: 99%

PANDAA-monium: Intentional violations of conventional qPCR design enables rapid, HIV-1 subtype-independent drug resistance SNP detection

MacLeod

Rowley

Essex

2019

Preprint

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Global efforts to ensure that 90% of all HIV-infected people receiving antiretroviral therapy (ART) will be virally suppressed by 2020 could be crippled by increases in acquired and transmitted HIV drug resistance (HIVDR), which challenge ART efficacy. The long-term sustainability of ART treatment programs is contingent on effective HIVDR monitoring yet current Sanger sequencing genotypic resistance tests are inadequate for large-scale implementation in low-and middle-income countries (LMICs). A simple, rapid, affordable HIVDR diagnostic would radically improve the treatment paradigm in LMICs by facilitating informed clinical decision-making upon ART failure. Although point mutation assays can be broadly deployed in this context, the primary challenge arises from extensive sequence variation surrounding targeted drug resistance mutations (DRMs). Here, we systematically and intentionally violate the canonical principles of qPCR design to develop a novel assay, Pan-Degenerate Amplification and Adaptation (PANDAA), that mitigates the impact of DRM-proximal secondary polymorphisms on probe-based qPCR performance to enable subtype-independent, focused resistance genotyping. Using extremely degenerate primers with 3' termini overlapping the probe-binding site, the HIV-1 genome is adapted through site-directed mutagenesis to replace secondary polymorphisms flanking the target DRM during the initial qPCR cycles. We show that PANDAA can quantify key HIV DRMs present at ≥5% and has diagnostic sensitivity and specificity of 96.9% and 97.5%, respectively, to detect DRMs associated with ART failure. PANDAA is an innovative solution for HIVDR genotyping and is an advancement in qPCR technology that could be applicable to any scenario where target-proximal genetic variability has been a roadblock in diagnostic development.

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“…The use of this commercial test as a reference standard has been frequently described in previous studies [ 12 , 22 – 32 ] and is recommended in the WHO guidelines [ 9 ]. Although ViroSeq has been approved for HIV-1 subtype B strains only, and difficulties with non-B subtypes have been published earlier [ 33 – 35 ], other studies report that ViroSeq is also applicable to various HIV-1 subtypes and recombinant strains [ 29 , 36 – 39 ].…”

Section: Discussionmentioning

confidence: 99%

Performance of an In-House Human Immunodeficiency Virus Type 1 Genotyping System for Assessment of Drug Resistance in Cuba

et al. 2015

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As commercial human immunodeficiency virus type 1 drug resistance assays are expensive, they are not commonly used in resource-limited settings. Hence, a more affordable in-house procedure was set up taking into account the specific epidemiological and economic circumstances of Cuba. The performance characteristics of the in-house assay were evaluated using clinical samples with various subtypes and resistance patterns. The lower limit of amplification was determined on dilutions series of 20 clinical isolates and ranged from 84 to 529 RNA copies/mL. For the assessment of trueness, 14 clinical samples were analyzed and the ViroSeq HIV-1 Genotyping System v2.0 was used as the reference standard. The mean nucleotide sequence identity between the two assays was 98.7% ± 1.0. Additionally, 99.0% of the amino acids at drug resistance positions were identical. The sensitivity and specificity in detecting drug resistance mutations was respectively 94.1% and 99.5%. Only few discordances in drug resistance interpretation patterns were observed. The repeatability and reproducibility were evaluated using 10 clinical samples with 3 replicates per sample. The in-house test was very precise as nucleotide sequence identity among paired nucleotide sequences ranged from 98.7% to 99.9%. The acceptance criteria were met by the in-house test for all performance characteristics, demonstrating a high degree of accuracy. Subsequently, the applicability in routine clinical practice was evaluated on 380 plasma samples. The amplification success rate was 91% and good quality consensus sequences encoding the entire protease and the first 335 codons in reverse transcriptase could be obtained for 99% of the successful amplicons. The reagent cost per sample using the in-house procedure was around € 80 per genotyping attempt. Overall, the in-house assay provided good results, was feasible with equipment and reagents available in Cuba and was half as expensive as commercial assays.

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Performance of the ViroSeq HIV-1 genotyping system v2.0 on HIV-1 strains circulating in Senegal

Cited by 14 publications

References 15 publications

Molecular Epidemiology of HIV-1 in Panama: Origin of Non-B Subtypes in Samples Collected from 2007 to 2013

Molecular Epidemiology of HIV-1 in Panama: Origin of Non-B Subtypes in Samples Collected from 2007 to 2013

PANDAA-monium: Intentional violations of conventional qPCR design enables rapid, HIV-1 subtype-independent drug resistance SNP detection

Performance of an In-House Human Immunodeficiency Virus Type 1 Genotyping System for Assessment of Drug Resistance in Cuba

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