Development of a panel of well-characterized human immunodeficiency virus type 1 isolates from newly diagnosed...

The aim of this study was the development of a panel constituted by well-defined HIV-1 strains of different genetic forms, with a particular focus on isolates from acute and recent infections. Fourteen HIV-1 isolates, including four from acute and five from recent infections, were expanded in peripheral blood mononuclear cells. SI phenotype, coreceptors use, and TCID(50)/ml were determined. V3 net charge was calculated. Near full-length genomes were amplified by RT-nested PCR in four overlapping segments. Phylogenetic analyses were performed with neighbor-joining trees and bootscanning. Analysis of cysteine residues, lengths of variable regions, and potential N-linked glycosylation sites in gp120 and gp41 was performed. Viral stocks were produced. Thirteen strains were NSI/R5 and one SI/R5,X4. TCID(50)/ml ranged between 10(4.6) and 10(6). V3 net charge was <+5 in 12 sequences and +5 in two sequences. Near full-length HIV-1 genomes analysis identified viruses of the following genetic forms: eight subtype B, three subtype C, two CRF02_AG, and one subtype G. Cysteine residues that form the V1,V2,V3, and V4 loops were highly conserved. The number of potential N-linked glycosylation sites in gp120 and gp41 ranged between 24-29 and 4-6, respectively. Seven potential N-linked glycosylation sites in gp120 and three in gp41 were conserved. V1, V2, V4, and V5 variable regions exhibited substantial length variation. In addition, an analysis of transmitted and natural resistance to current antiretroviral drugs in these strains was performed. It is worth mentioning that the 13S mutation in the V3 sequence, associated with resistance to maraviroc, was observed in a subtype B strain that harbored resistance mutations to nucleoside reverse transcriptase inhibitors and to T20. The availability of a panel including strains from acute and recent infections should be a valuable resource for optimizing and standardizing vaccine candidate assessment. Near full-length genome characterization may be necessary for evaluating clade-specific reactivities.