Validation of publicly-available software used in analyzing NGS data for HIV-1 drug resistance mutations and transmission networks in a Washington, DC, Cohort

Jair, Kamwing; McCann, Chase D.; Reed, Harrison; Castel, Amanda D.; Pérez‐Losada, Marcos; Wilbourn, Brittany; Greenberg, Alan E.; Jordan, Jeanne A.

doi:10.1371/journal.pone.0214820

Cited by 12 publications

(14 citation statements)

References 31 publications

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“…We detected high levels of clustering using haplotypes (32.8-64.2%, not including the V1V2 region), as also seen in other HIV-1 cohorts (24-65%) 29,35,[49][50][51][52][53][54][55][56][57][58][59][60][61] . Other studies, including one completed with Sanger sequencing in DC 28,31,36,52,62 , however, have found lower levels of clustering (7-17%), in agreement with those reported here for the consensus sequence phylogenetic clusters (pol(c) and env(c)) and the V1V2 region for haplotypes (9.0%). Therefore, a more comprehensive understanding of HIV-1 transmission events in DC has been achieved when evaluating multiple genes together, rather than primarily focusing on polymerase genes that are typically screened for DRMs in clinical settings or used in investigations at the local health department level.…”

Section: Pr/rt Int Envsupporting

confidence: 90%

“…A Drug Resistant Mutation (DRM) prevalence of 48.6-54.0% was detected in this subset of the DC Cohort, depending on use of consensus sequence or haplotypes. Lower DRM prevalence rates were previously reported for the DC area 28,29,34,36 (17.3-37.9% between 1994 and 2016). A much higher rate (66%) was reported in a smaller study of ART treatment-naïve and experienced pediatric patients in Rhode Island 37 .…”

Section: Pr/rt Int Envmentioning

confidence: 52%

“…# 18091050, Invitrogen, Carlsbad, CA) were used respectively and according to manufacturers' instructions. Multiple sets of HIV-1 specific primer pairs were used to target and amplify using polymerase-chain-reaction (PCR) the protease (PR), reverse transcriptase (RT), integrase (int), and envelope (env) HIV-1 genes (~43% of genome) 36 Sequence analyses. The raw sequence data for each patient were processed through HAPHPIPE (https:// github.com/gwcbi/haphpipe), a HAplotype reconstruction and PHylodynamics PIPEline for genome-wide assembly of viral consensus sequences and haplotypes 69 .…”

Section: Ethics Institutional Review Board (Irb071029) Approval Wasmentioning

confidence: 99%

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A cross-sectional study to characterize local HIV-1 dynamics in Washington, DC using next-generation sequencing

Gibson

Jair

Castel

et al. 2020

Sci Rep

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the DC Cohort Executive Committee † Washington, DC continues to experience a generalized HIV-1 epidemic. We characterized the local phylodynamics of HIV-1 in DC using next-generation sequencing (NGS) data. Viral samples from 68 participants from 2016 through 2017 were sequenced and paired with epidemiological data. Phylogenetic and network inferences, drug resistant mutations (DRMs), subtypes and HIV-1 diversity estimations were completed. Haplotypes were reconstructed to infer transmission clusters. Phylodynamic inferences based on the HIV-1 polymerase (pol) and envelope genes (env) were compared. Higher HIV-1 diversity (n.s.) was seen in men who have sex with men, heterosexual, and male participants in DC. 54.0% of the participants contained at least one DRM. The 40-49 year-olds showed the highest prevalence of DRMs (22.9%). Phylogenetic analysis of pol and env sequences grouped 31.9-33.8% of the participants into clusters. HIV-TRACE grouped 2.9-12.8% of participants when using consensus sequences and 9.0-64.2% when using haplotypes. NGS allowed us to characterize the local phylodynamics of HIV-1 in DC more broadly and accurately, given a better representation of its diversity and dynamics. Reconstructed haplotypes provided novel and deeper phylodynamic insights, which led to networks linking a higher number of participants. Our understanding of the HIV-1 epidemic was expanded with the powerful coupling of HIV-1 NGS data with epidemiological data.Despite recent reductions in HIV-1 prevalence in Washington, DC from 2.5% in 2013 1 to 1.8% in 2018, the United States (US) capital is still experiencing a generalized HIV-1 epidemic -as defined by the World Health Organization 2-4 . There were 340 newly diagnosed cases in DC in 2018, and the DC rate is five times higher than the national rate 3 . Blacks, men, men who have sex with men (MSM), and heterosexuals (HRH) account for the majority of people living with HIV-1 (PLWH) in DC 2,3 . However, ~20% of the newly diagnosed persons had an unknown risk for transmission in both 2016 and 2017 2,3 . Furthermore, the leading group (33.3%) of newly diagnosed cases was between the ages of 20-29 years old 3 . This same age group had the highest percentage (27.5%) of drug resistance mutations (DRM) at diagnosis, suggesting broader spread of HIV-1 drug resistant variants and potential concern for future therapeutic options, especially if these mutations are against first line antiretroviral (ART) drugs for newly infected individuals. With blacks and young adults being the most impacted groups of individuals for HIV-1 in DC, understanding the current HIV-1 phylodynamics can provide informative data to guide programs that prevent and reduce the incidence of HIV-1. Moreover, identifying potential transmission clusters amongst individuals in DC and their associated epidemiological features may help infer otherwise 'unknown' transmission modes and provide insight for more targeted prevention and intervention strategies.In 2011, the DC Cohort, a longitudinal observational NIH-funded co...

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Section: Pr/rt Int Envsupporting

confidence: 90%

Section: Pr/rt Int Envmentioning

confidence: 52%

Section: Ethics Institutional Review Board (Irb071029) Approval Wasmentioning

confidence: 99%

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A cross-sectional study to characterize local HIV-1 dynamics in Washington, DC using next-generation sequencing

Gibson

Jair

Castel

et al. 2020

Sci Rep

Self Cite

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“…Eight studies containing 855 samples from 821 persons met the inclusion criteria [18–25] (Figure 1; Table 1). These samples included 693 PR, 700 RT, and 449 IN NGS sequence sets.…”

Section: Resultsmentioning

confidence: 99%

“…Two studies had the lowest numbers of both usual and unusual mutations, and of the proportions of unusual mutations [21, 22]. Four studies had intermediate values for these three metrics [18, 20, 24, 25], and two had higher values [19, 23].…”

Section: Resultsmentioning

confidence: 99%

Analysis of unusual and signature APOBEC-mutations in HIV-1polnext-generation sequences

Tzou

Pond

Ávila‐Ríos

et al. 2019

Preprint

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IntroductionAt low mutation-detection thresholds, next generation sequencing (NGS) for HIV-1 genotypic resistance testing is susceptible to artifactual detection of mutations arising from PCR error and APOBEC-mediated G-to-A hypermutation.MethodsWe analyzed published HIV-1 pol Illumina NGS data to characterize the distribution of mutations at eight NGS thresholds: 20%, 10%, 5%, 2%, 1%, 0.5%, 0.2%, and 0.1%. At each threshold, we determined the proportion of amino acid mutations that were unusual (defined as having a prevalence <0.01% in HIV-1 group M sequences) or were signature APOBEC mutations.ResultsEight studies, containing 855 samples, in the NCBI Sequence Read Archive were analyzed. As detection thresholds were lowered, there was a progressive increase in the proportion of positions with both usual and unusual mutations and in the proportion of all mutations that were unusual. The median proportion of positions with an unusual mutation increased gradually from 0% at the 20% threshold to 0.3% at the 1% threshold and then exponentially to 1.3% (0.5% threshold), 6.9% (0.2% threshold), and 23.2% (0.1% threshold). In two of three studies with available plasma HIV-1 RNA levels, the proportion of positions with unusual mutations was negatively associated with virus levels. Although the complete set of signature APOBEC mutations was much smaller than that of unusual mutations, the former outnumbered the latter in one-sixth of the samples at the 0.5%, 1%, and 2% thresholds.ConclusionsThe marked increase in the proportion of positions with unusual mutations at thresholds below 1% and in samples with lower virus loads suggests that, at low thresholds, many unusual mutations are artifactual, reflecting PCR error or G-to-A hypermutation. Profiling the numbers of unusual and signature APOBEC pol mutations at different NGS thresholds may be useful to avoid selecting a threshold that is too low and poses an unacceptable risk of identifying artifactual mutations.

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High-throughput sequencing (HTS) for the analysis of viral populations

Pérez‐Losada

Arenas

Galán

et al. 2020

Infection, Genetics and Evolution

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Validation of publicly-available software used in analyzing NGS data for HIV-1 drug resistance mutations and transmission networks in a Washington, DC, Cohort

Cited by 12 publications

References 31 publications

A cross-sectional study to characterize local HIV-1 dynamics in Washington, DC using next-generation sequencing

A cross-sectional study to characterize local HIV-1 dynamics in Washington, DC using next-generation sequencing

Analysis of unusual and signature APOBEC-mutations in HIV-1polnext-generation sequences

High-throughput sequencing (HTS) for the analysis of viral populations

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