Browsing by Author "Ahmed, Manzoor"

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    Fast Dissemination of New HIV-1 CRF02/A1 Recombinants in Pakistan.
    (PLoS One, 2016) Chen, Yue; Hora, Bhavna; DeMarco, Todd; Shah, Sharaf Ali; Ahmed, Manzoor; Sanchez, Ana M; Su, Chang; Carter, Meredith; Stone, Mars; Hasan, Rumina; Hasan, Zahra; Busch, Michael P; Denny, Thomas N; Gao, Feng
    A number of HIV-1 subtypes are identified in Pakistan by characterization of partial viral gene sequences. Little is known whether new recombinants are generated and how they disseminate since whole genome sequences for these viruses have not been characterized. Near full-length genome (NFLG) sequences were obtained by amplifying two overlapping half genomes or next generation sequencing from 34 HIV-1-infected individuals in Pakistan. Phylogenetic tree analysis showed that the newly characterized sequences were 16 subtype As, one subtype C, and 17 A/G recombinants. Further analysis showed that all 16 subtype A1 sequences (47%), together with the vast majority of sequences from Pakistan from other studies, formed a tight subcluster (A1a) within the subtype A1 clade, suggesting that they were derived from a single introduction. More in-depth analysis of 17 A/G NFLG sequences showed that five shared similar recombination breakpoints as in CRF02 (15%) but were phylogenetically distinct from the prototype CRF02 by forming a tight subcluster (CRF02a) while 12 (38%) were new recombinants between CRF02a and A1a or a divergent A1b viruses. Unique recombination patterns among the majority of the newly characterized recombinants indicated ongoing recombination. Interestingly, recombination breakpoints in these CRF02/A1 recombinants were similar to those in prototype CRF02 viruses, indicating that recombination at these sites more likely generate variable recombinant viruses. The dominance and fast dissemination of new CRF02a/A1 recombinants over prototype CRF02 suggest that these recombinant have more adapted and may become major epidemic strains in Pakistan.
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    Streamlined Subpopulation, Subtype, and Recombination Analysis of HIV-1 Half-Genome Sequences Generated by High-Throughput Sequencing.
    (mSphere, 2020-10-14) Hora, Bhavna; Gulzar, Naila; Chen, Yue; Karagiannis, Konstantinos; Cai, Fangping; Su, Chang; Smith, Krista; Simonyan, Vahan; Shah, Sharaf Ali; Ahmed, Manzoor; Sanchez, Ana M; Stone, Mars; Cohen, Myron S; Denny, Thomas N; Mazumder, Raja; Gao, Feng
    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).