Development of a Versatile, Near Full Genome Amplification and Sequencing Approach for a Broad Variety of HIV-1 Group M Variants.

Banin, Andrew N.; Tuen, Michael; Bimela, Jude S.; Tongo, Marcel; Zappile, Paul; Khodadadi-Jamayran, Alireza; Nanfack, Aubin J.; Meli, Josephine; Wang, Xiaohong; Mbanya, Dora; Ngogang, Jeanne; Heguy, Adriana; Nyambi, Phillipe N.; Fokunang, Charles; Duerr, Ralf

Near full genome sequencing (NFGS) of HIV-1 is required to assess the genetic composition of HIV-1 strains comprehensively. Population-wide, it enables a determination of the heterogeneity of HIV-1 and the emergence of novel/recombinant strains, while for each individual it constitutes a diagnostic instrument to assist targeted therapeutic measures against viral components. There is still a lack of robust and adaptable techniques for efficient NFGS from miscellaneous HIV-1 subtypes. Using rational primer design, a broad primer set was developed for the amplification and sequencing of diverse HIV-1 group M variants from plasma. Using pure subtypes as well as diverse, unique recombinant forms (URF), variable amplicon approaches were developed for NFGS comprising all functional genes. Twenty-three different genomes composed of subtypes A (A1), B, F (F2), G, CRF01_AE, CRF02_AG, and CRF22_01A1 were successfully determined. The NFGS approach was robust irrespective of viral loads (≥306 copies/mL) and amplification method. Third-generation sequencing (TGS), single genome amplification (SGA), cloning, and bulk sequencing yielded similar outcomes concerning subtype composition and recombinant breakpoint patterns. The introduction of a simple and versatile near full genome amplification, sequencing, and cloning method enables broad application in phylogenetic studies of diverse HIV-1 subtypes and can contribute to personalized HIV therapy and diagnosis.

GENOMES; HETEROGENEITY; GENES; BLOOD plasma; PHYLOGENY

Viruses (1999-4915), 2019, Vol 11, Issue 4, p317

1999-4915

Academic Journal

10.3390/v11040317