Using High-Throughput Sequencing to Characterize the Development of the Antibody Repertoire During Infections: A Case Study of HIV-1

2017 ◽  
pp. 245-263 ◽  
Author(s):  
Felix Breden ◽  
Corey T. Watson
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Antibody Repertoire ◽  
Hiv 1

scholarly journals Modelling the Evolutionary Dynamics of Viruses within Their Hosts: A Case Study Using High-Throughput Sequencing

2012 ◽  
Vol 8 (4) ◽  
pp. e1002654 ◽  
Author(s):  
Frédéric Fabre ◽  
Josselin Montarry ◽  
Jérôme Coville ◽  
Rachid Senoussi ◽  
Vincent Simon ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Evolutionary Dynamics

scholarly journals MiDRMpol: A High-Throughput Multiplexed Amplicon Sequencing Workflow to Quantify HIV-1 Drug Resistance Mutations against Protease, Reverse Transcriptase, and Integrase Inhibitors

Viruses ◽  
2019 ◽  
Vol 11 (9) ◽  
pp. 806
Author(s):  
Shambhu G. Aralaguppe ◽  
Anoop T. Ambikan ◽  
Manickam Ashokkumar ◽  
Milner M. Kumar ◽  
Luke Elizabeth Hanna ◽  
...  
Keyword(s):  
Drug Resistance ◽  
High Throughput ◽  
Large Scale ◽  
High Throughput Sequencing ◽  
Polymorphism Analysis ◽  
Resistance Mutations ◽  
Subtype C ◽  
Bioinformatics Pipeline ◽  
Drug Resistance Mutations ◽  
Hiv 1

The detection of drug resistance mutations (DRMs) in minor viral populations is of potential clinical importance. However, sophisticated computational infrastructure and competence for analysis of high-throughput sequencing (HTS) data lack at most diagnostic laboratories. Thus, we have proposed a new pipeline, MiDRMpol, to quantify DRM from the HIV-1 pol region. The gag-vpu region of 87 plasma samples from HIV-infected individuals from three cohorts was amplified and sequenced by Illumina HiSeq2500. The sequence reads were adapter-trimmed, followed by analysis using in-house scripts. Samples from Swedish and Ethiopian cohorts were also sequenced by Sanger sequencing. The pipeline was validated against the online tool PASeq (Polymorphism Analysis by Sequencing). Based on an error rate of <1%, a value of >1% was set as reliable to consider a minor variant. Both pipelines detected the mutations in the dominant viral populations, while discrepancies were observed in minor viral populations. In five HIV-1 subtype C samples, minor mutations were detected at the <5% level by MiDRMpol but not by PASeq. MiDRMpol is a computationally as well as labor efficient bioinformatics pipeline for the detection of DRM from HTS data. It identifies minor viral populations (<20%) of DRMs. Our method can be incorporated into large-scale surveillance of HIV-1 DRM.

scholarly journals High-Throughput Sequencing of the Zebrafish Antibody Repertoire

Science ◽  
2009 ◽  
Vol 324 (5928) ◽  
pp. 807-810 ◽  
Author(s):  
J. A. Weinstein ◽  
N. Jiang ◽  
R. A. White ◽  
D. S. Fisher ◽  
S. R. Quake
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Antibody Repertoire

scholarly journals Streamlined Subpopulation, Subtype, and Recombination Analysis of HIV-1 Half-Genome Sequences Generated by High-Throughput Sequencing

mSphere ◽  
2020 ◽  
Vol 5 (5) ◽  
Author(s):  
Bhavna Hora ◽  
Naila Gulzar ◽  
Yue Chen ◽  
Konstantinos Karagiannis ◽  
Fangping Cai ◽  
...  
Keyword(s):  
Genome Sequencing ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Vaccine Development ◽  
Consensus Sequence ◽  
Cost Effective ◽  
Recombination Analysis ◽  
Genome Sequences ◽  
Single Genome ◽  
Hiv 1

ABSTRACT High-throughput sequencing (HTS) has been widely used to characterize HIV-1 genome sequences. There are no algorithms currently that can directly determine genotype and quasispecies population using short HTS reads generated from long genome sequences without additional software. To establish a robust subpopulation, subtype, and recombination analysis workflow, we amplified the HIV-1 3′-half genome from plasma samples of 65 HIV-1-infected individuals and sequenced the entire amplicon (∼4,500 bp) by HTS. With direct analysis of raw reads using HIVE-hexahedron, we showed that 48% of samples harbored 2 to 13 subpopulations. We identified various subtypes (17 A1s, 4 Bs, 27 Cs, 6 CRF02_AGs, and 11 unique recombinant forms) and defined recombinant breakpoints of 10 recombinants. These results were validated with viral genome sequences generated by single genome sequencing (SGS) or the analysis of consensus sequence of the HTS reads. The HIVE-hexahedron workflow is more sensitive and accurate than just evaluating the consensus sequence and also more cost-effective than SGS. IMPORTANCE The highly recombinogenic nature of human immunodeficiency virus type 1 (HIV-1) leads to recombination and emergence of quasispecies. It is important to reliably identify subpopulations to understand the complexity of a viral population for drug resistance surveillance and vaccine development. High-throughput sequencing (HTS) provides improved resolution over Sanger sequencing for the analysis of heterogeneous viral subpopulations. However, current methods of analysis of HTS reads are unable to fully address accurate population reconstruction. Hence, there is a dire need for a more sensitive, accurate, user-friendly, and cost-effective method to analyze viral quasispecies. For this purpose, we have improved the HIVE-hexahedron algorithm that we previously developed with in silico short sequences to analyze raw HTS short reads. The significance of this study is that our standalone algorithm enables a streamlined analysis of quasispecies, subtype, and recombination patterns from long HIV-1 genome regions without the need of additional sequence analysis tools. Distinct viral populations and recombination patterns identified by HIVE-hexahedron are further validated by comparison with sequences obtained by single genome sequencing (SGS).

High-throughput sequencing identifies HIV-1-replication- and latency-related miRNAs in CD4+ T cell lines

2017 ◽  
Vol 162 (7) ◽  
pp. 1933-1942 ◽  
Author(s):  
Xiangyun Lu ◽  
Jin Yang ◽  
Haibo Wu ◽  
Zongxing Yang ◽  
Changzhong Jin ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Lines ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Cd4 T Cell ◽  
T Cell Lines ◽  
Hiv 1

scholarly journals High-throughput sequencing of integrated HIV-1 reveals novel proviral structures

2019 ◽  
Vol 5 ◽  
pp. 25-26
Author(s):  
K. Joseph ◽  
E. Halvas ◽  
L. Brandt ◽  
S. Patro ◽  
J. Rausch ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Hiv 1

scholarly journals Hiding in plain sight: structure and sequence analysis reveals the importance of the antibody DE loop for antibody-antigen binding

2020 ◽  
Author(s):  
Simon P. Kelow ◽  
Jared Adolf-Bryfogle ◽  
Roland L. Dunbrack
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Data Bank ◽  
Antigen Binding ◽  
Sequence Variability ◽  
Antigen Binding Site ◽  
Sequencing Studies ◽  
Variable Domains ◽  
Complementarity Determining Regions ◽  
Hiv 1

AbstractAntibody variable domains contain “complementarity determining regions” (CDRs), the loops that form the antigen binding site. CDRs1-3 are recognized as the canonical CDRs. However, a fourth loop sits adjacent to CDR1 and CDR2 and joins the D and E strands on the antibody v-type fold. This “DE loop” is usually treated as a framework region, even though mutations in the loop affect the conformation of the CDRs and residues in the DE loop occasionally contact antigen. We analyzed the length, structure, and sequence features of all DE loops in the Protein Data Bank, as well as millions of sequences from HIV-1 infected and naïve patients. We refer to the DE loop as H4 and L4 in the heavy and light chain respectively. Clustering the backbone conformations of the most common length of L4 (6 residues) reveals four conformations: two κ-only clusters, one λ-only cluster, and one mixed κ/λ cluster. The vast majority of H4 loops are length-8 and exist primarily in one conformation; a secondary conformation represents a small fraction of H4-8 structures. H4 sequence variability exceeds that of the antibody framework in naïve human high-throughput sequences, and both L4 and H4 sequence variability from λ and heavy germline sequences exceed that of germline framework regions. Finally, we identified dozens of structures in the PDB with insertions in the DE loop, all related to broadly neutralizing HIV-1 antibodies, as well as antibody sequences from high-throughput sequencing studies of HIV-infected individuals, illuminating a possible role in humoral immunity to HIV-1.

scholarly journals Increased acquired protease inhibitor drug resistance mutations in minor HIV-1 quasispecies from infected patients suspected of failing on national second-line therapy in South Africa

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Adetayo Emmanuel Obasa ◽  
Anoop T. Ambikan ◽  
Soham Gupta ◽  
Ujjwal Neogi ◽  
Graeme Brendon Jacobs
Keyword(s):  
South Africa ◽  
Drug Resistance ◽  
Reverse Transcriptase ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Second Line ◽  
Viral Quasispecies ◽  
Resistance Mutations ◽  
Drug Resistance Mutations ◽  
Hiv 1

Abstract Background HIV-1C has been shown to have a greater risk of virological failure and reduced susceptibility towards boosted protease inhibitors (bPIs), a component of second-line combination antiretroviral therapy (cART) in South Africa. This study entailed an evaluation of HIV-1 drug resistance-associated mutations (RAMs) among minor viral populations through high-throughput sequencing genotypic resistance testing (HTS-GRT) in patients on the South African national second-line cART regimen receiving bPIs. Methods During 2017 and 2018, 67 patient samples were sequenced using high-throughput sequencing (HTS), of which 56 samples were included in the final analysis because the patient’s treatment regimen was available at the time of sampling. All patients were receiving bPIs as part of their cART. Viral RNA was extracted, and complete pol genes were amplified and sequenced using Illumina HiSeq2500, followed by bioinformatics analysis to quantify the RAMs according to the Stanford HIV Drug Resistance Database. Results Statistically significantly higher PI RAMs were observed in minor viral quasispecies (25%; 14/56) compared to non-nucleoside reverse transcriptase inhibitors (9%; 5/56; p = 0.042) and integrase inhibitor RAM (4%; 2/56; p = 0.002). The majority of the drug resistance mutations in the minor viral quasispecies were observed in the V82A mutation (n = 13) in protease and K65R (n = 5), K103N (n = 7) and M184V (n = 5) in reverse transcriptase. Conclusions HTS-GRT improved the identification of PI and reverse transcriptase inhibitor (RTI) RAMs in second-line cART patients from South Africa compared to the conventional GRT with ≥20% used in Sanger-based sequencing. Several RTI RAMs, such as K65R, M184V or K103N and PI RAM V82A, were identified in < 20% of the population. Deep sequencing could be of greater value in detecting acquired resistance mutations early.

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