Ambiguous Nucleotide Calls From Population-based Sequencing of HIV-1 are a Marker for Viral Diversity and the Age of Infection

Kouyos, Roger D.; Wyl, Viktor von; Yerly, Sabine; Böni, Jürg; Rieder, Peter; Joos, Béda; Taffé, Patrick; Shah, Cyril; Bürgisser, Philippe; Klimkait, Thomas; Weber, Rainer; Hirschel, Bernard; Cavassini, Matthias; Rauch, Andri; Battegay, Manuel; Vernazza, Pietro; Bernasconi, Enos; Ledergerber, Bruno; Bonhoeffer, Sebastian; Günthard, Huldrych F.

doi:10.1093/cid/ciq164

Cited by 129 publications

(180 citation statements)

References 18 publications

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“…Because of the retrospective nature of our analysis, two different PCR systems, sequencing systems, and software packages to call ambiguous bases were used over time, which is an inherent technical limitation of the SAM. However, the methodological diversity in our study was still lower than in related studies (14)(15)(16). Since Ragonnet-Cronin et al (15) demonstrated that differences in ambiguous base calling were at least negligible in the commonly used threshold range of 15 to 25%, we consider the sequencing software to have only a minor influence on the overall frequency of ambiguous base calling.…”

Section: Discussionmentioning

confidence: 60%

“…In the previous studies, sequence ambiguity thresholds of 0.45% (15), 0.47% (16), and 0.5% (14) were described to best identify infections acquired within Յ155 days (15) or Յ365 days (14, 16) with accuracies of 79.9% (15), 79.7% (16), and 70.9% (14). Our analysis is in accordance with these reports, yielding a sequence ambiguity threshold of 0.5%, a mean duration of recency of 208 days, and an accuracy of 76.7%.…”

Section: Discussionmentioning

confidence: 99%

“…HIV-1 diversity is considered to be a candidate nonserological biomarker, as it increases during the course of infection in antiretroviral treatment (ART)-naive individuals. Viral diversity can be determined by single-genome sequencing (11), next-generation sequencing (10), and high-resolution melting assays (12), but direct Sanger population sequencing can also reflect HIV-1 polymorphisms via ambiguous base calls (14)(15)(16).…”

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confidence: 99%

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Assessment of Ambiguous Base Calls in HIV-1 pol Population Sequences as a Biomarker for Identification of Recent Infections in HIV-1 Incidence Studies

et al. 2014

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An increase in the proportion of ambiguous base calls in HIV-1 pol population sequences during the course of infection has been demonstrated in different study populations, and sequence ambiguity thresholds to classify infections as recent or nonrecent have been suggested. The aim of our study was to evaluate sequence ambiguities as a candidate biomarker for use in an HIV-1 incidence assay using samples from antiretroviral treatment-naive seroconverters with known durations of infection (German HIV-1 Seroconverter Study). We used 2,203 HIV-1 pol population sequences derived from 1,334 seroconverters to assess the sequence ambiguity method (SAM). We then compared the serological incidence BED capture enzyme immunoassay (BED-CEIA) with the SAM for a subset of 723 samples from 495 seroconverters and evaluated a multianalyte algorithm that includes BED-CEIA results, SAM results, viral loads, and CD4 cell counts for 453 samples from 325 seroconverters. We observed a significant increase in the proportion of sequence ambiguities with the duration of infection. A sequence ambiguity threshold of 0.5% best identified recent infections with 76.7% accuracy. The mean duration of recency was determined to be 208 (95% confidence interval, 196 to 221) days. In the subset analysis, BED-CEIA achieved a significantly higher accuracy than the SAM (84.6 versus 75.5%, P < 0.001) and results were concordant for 64.2% (464/723) of the samples. Also, the multianalyte algorithm did not show better accuracy than the BED-CEIA (83.4 versus 84.3%, P ‫؍‬ 0.786). In conclusion, the SAM and the multianalyte algorithm including SAM were inferior to the BED-CEIA, and the proportion of sequence ambiguities is therefore not a preferable biomarker for HIV-1 incidence testing.

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Section: Discussionmentioning

confidence: 60%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Assessment of Ambiguous Base Calls in HIV-1 pol Population Sequences as a Biomarker for Identification of Recent Infections in HIV-1 Incidence Studies

et al. 2014

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“…In this protocol, patients are offered early ART. After 1 y of suppressed viremia (,40 HIV-1 RNA copies/ ml plasma), patients can chose to interrupt ART (14)(15)(16)(17)(18). Study details are listed under http://www.clinicaltrials.gov; ID NCT00537966.…”

Section: Study Populationmentioning

confidence: 99%

CD8+ T Cells Are Activated in an Antigen-Independent Manner in HIV-Infected Individuals

Bastidas

Graw

Smith

et al. 2014

The Journal of Immunology

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Hyperactivation of T cells, particularly of CD8+ T cells, is a hallmark of chronic HIV 1 (HIV-1) infection. Little is known about the antigenic specificities and the mechanisms by which HIV-1 causes activation of CD8+ T cells during chronic infection. We report that CD8+ T cells were activated during in vivo HIV-1 replication irrespective of their Ag specificity. Cytokines present during untreated HIV-1 infection, most prominently IL-15, triggered proliferation and expression of activation markers in CD8+ T cells, but not CD4+ T cells, in the absence of TCR stimulation. Moreover, LPS or HIV-1–activated dendritic cells (DCs) stimulated CD8+ T cells in an IL-15–dependent but Ag-independent manner, and IL-15 expression was highly increased in DCs isolated from viremic HIV-1 patients, suggesting that CD8+ T cells are activated by inflammatory cytokines in untreated HIV-1 patients independent of Ag specificity. This finding contrasts with CD4+ T cells whose in vivo activation seems biased toward specificities for persistent Ags. These observations explain the higher abundance of activated CD8+ T cells compared with CD4+ T cells in untreated HIV-1 infection.

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“…A variety of methods for assessing HIV diversity are being explored as tools for estimating HIV incidence, including the quantification of ambiguous nucleotide calls and other computational approaches based on analyses of data from traditional or next-generation sequencing (13,14). We recently developed an assay that quantifies the level of genetic diversity in HIV populations without the use of sequencing (12,15).…”

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confidence: 99%

HIV Diversity as a Biomarker for HIV Incidence Estimation: Including a High-Resolution Melting Diversity Assay in a Multiassay Algorithm

et al. 2014

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pMultiassay algorithms (MAAs) can be used to estimate cross-sectional HIV incidence. We previously identified a robust MAA that includes the BED capture enzyme immunoassay (BED-CEIA), the Bio-Rad Avidity assay, viral load, and CD4 cell count. In this report, we evaluated MAAs that include a high-resolution melting (HRM) diversity assay that does not require sequencing. HRM scores were determined for eight regions of the HIV genome (2 in gag, 1 in pol, and 5 in env). The MAAs that were evaluated included the BED-CEIA, the Bio-Rad Avidity assay, viral load, and the HRM diversity assay, using HRM scores from different regions and a range of region-specific HRM diversity assay cutoffs. The performance characteristics based on the proportion of samples that were classified as MAA positive by duration of infection were determined for each MAA, including the mean window period. The cross-sectional incidence estimates obtained using optimized MAAs were compared to longitudinal incidence estimates for three cohorts in the United States. The performance of the HRM-based MAA was nearly identical to that of the MAA that included CD4 cell count. The HRM-based MAA had a mean window period of 154 days and provided cross-sectional incidence estimates that were similar to those based on cohort follow-up. HIV diversity is a useful biomarker for estimating HIV incidence. MAAs that include the HRM diversity assay can provide accurate HIV incidence estimates using stored blood plasma or serum samples without a requirement for CD4 cell count data.

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Ambiguous Nucleotide Calls From Population-based Sequencing of HIV-1 are a Marker for Viral Diversity and the Age of Infection

Cited by 129 publications

References 18 publications

Assessment of Ambiguous Base Calls in HIV-1 pol Population Sequences as a Biomarker for Identification of Recent Infections in HIV-1 Incidence Studies

Assessment of Ambiguous Base Calls in HIV-1 pol Population Sequences as a Biomarker for Identification of Recent Infections in HIV-1 Incidence Studies

CD8+ T Cells Are Activated in an Antigen-Independent Manner in HIV-Infected Individuals

HIV Diversity as a Biomarker for HIV Incidence Estimation: Including a High-Resolution Melting Diversity Assay in a Multiassay Algorithm

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