Genetic Characterization of a Panel of Diverse HIV-1 Isolates at Seven International Sites.
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HIV-1 subtypes and drug resistance are routinely tested by many international surveillance groups. However, results from different sites often vary. A systematic comparison of results from multiple sites is needed to determine whether a standardized protocol is required for consistent and accurate data analysis. A panel of well-characterized HIV-1 isolates (N = 50) from the External Quality Assurance Program Oversight Laboratory (EQAPOL) was assembled for evaluation at seven international sites. This virus panel included seven subtypes, six circulating recombinant forms (CRFs), nine unique recombinant forms (URFs) and three group O viruses. Seven viruses contained 10 major drug resistance mutations (DRMs). HIV-1 isolates were prepared at a concentration of 107 copies/ml and compiled into blinded panels. Subtypes and DRMs were determined with partial or full pol gene sequences by conventional Sanger sequencing and/or Next Generation Sequencing (NGS). Subtype and DRM results were reported and decoded for comparison with full-length genome sequences generated by EQAPOL. The partial pol gene was amplified by RT-PCR and sequenced for 89.4%-100% of group M viruses at six sites. Subtyping results of majority of the viruses (83%-97.9%) were correctly determined for the partial pol sequences. All 10 major DRMs in seven isolates were detected at these six sites. The complete pol gene sequence was also obtained by NGS at one site. However, this method missed six group M viruses and sequences contained host chromosome fragments. Three group O viruses were only characterized with additional group O-specific RT-PCR primers employed by one site. These results indicate that PCR protocols and subtyping tools should be standardized to efficiently amplify diverse viruses and more consistently assign virus genotypes, which is critical for accurate global subtype and drug resistance surveillance. Targeted NGS analysis of partial pol sequences can serve as an alternative approach, especially for detection of low-abundance DRMs.
Published Version (Please cite this version)10.1371/journal.pone.0157340
Publication InfoBusch, Michael P; Chen, Y; Chow, WZ; Denny, Thomas Norton; Gao, Feng; Hackett, J; ... Zeng, P (2016). Genetic Characterization of a Panel of Diverse HIV-1 Isolates at Seven International Sites. PLoS One, 11(6). pp. e0157340. 10.1371/journal.pone.0157340. Retrieved from http://hdl.handle.net/10161/13343.
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Professor in Medicine
Thomas N. Denny, MSc, M.Phil, is the Chief Operating Officer of the Duke Human Vaccine Institute (DHVI) and the Center for HIV/AIDS Vaccine Immunology (CHAVI), and a Professor of Medicine in the Department of Medicine at Duke University Medical Center. He is also an Affiliate Member of the Duke Global Health Institute. He has recently been appointed to the Duke University Fuqua School of Business Health Sector Advisory Council. Previously, he was an Associate Professor of Pathology, Laboratory M
Professor of Medicine
Dr. Feng Gao is Professor of Medicine at Duke University. The Gao laboratory has a long-standing interest in elucidating the origins and evolution of human and simian inmmunodeficiency viruses (HIV and SIV), and in studying HIV/SIV gene function and pathogenic mechanisms from the evolutionary perspective. These studies have led to new strategies to better understand HIV origins, biology, pathogenesis and drug resistance, and to design new AIDS vaccines.
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