Abstract
BackgroundCCR5-using (r5) HIV-1 predominates during asymptomatic disease followed by occasional emergence of CXCR4-using (x4) or dual tropic (r5x4) virus. We examined the contribution of the x4 and r5 components to replicative fitness of HIV-1 isolates.MethodsDual tropic r5x4 viruses were predicted from average HIV-1 env sequences of two primary subtype C HIV-1 isolates (C19 and C27) and from two patient plasma samples (B12 and B19). Chimeric Env viruses with an NL4-3 backbone were constructed from the B12 and B19 env sequences. To determine replicative fitness, these primary and chimeric dual tropic HIV-1 were then competed against HIV-1 reference isolates in U87.CD4 cells expressing CXCR4 or CCR5 or in PBMCs ± entry inhibitors. Contribution of the x4 and r5 clones within the quasispecies of these chimeric or primary HIV-1 isolates were then compared to the frequency of x4, r5, and dual tropic clones within the quasispecies as predicted by phenotypic assays, clonal sequencing, and 454 deep sequencing.ResultsIn the primary HIV-1 isolates (C19 and C27), subtype C dual tropic clones dominated over x4 clones while pure r5 clones were absent. In two subtype B chimeric viruses (B12 and B19), r5 clones were >100-fold more abundant than x4 or r5/x4 clones. The dual tropic C19 and C27 HIV-1 isolates outcompeted r5 primary HIV-1 isolates, B2 and C3 in PBMCs. When AMD3100 was added or when only U87.CD4.CCR5 cells were used, the B2 and C3 reference viruses now out-competed the r5 component of the dual tropic C19 and C27. In contrast, the same replicative fitness was observed with dualtropic B12 and B19 HIV-1 isolates relative to x4 HIV-1 A8 and E6 or the r5 B2 and C3 viruses, even when the r5 or x4 component was inhibited by maraviroc (or AMD3100) or in U87.CD4.CXCR4 (or CCR5) cells.ConclusionsIn the dual tropic HIV-1 isolates, the x4 replicative fitness is higher than r5 clones but the x4 or x4/r5 clones are typically at low frequency in the intrapatient virus population. Ex vivo HIV propagation promotes outgrowth of the x4 clones and provides an over-estimate of x4 dominance in replicative fitness within dual tropic viruses.Electronic supplementary materialThe online version of this article (doi:10.1186/s12981-015-0066-7) contains supplementary material, which is available to authorized users.
Highlights
CCR5-using (r5) human immunodeficiency virus (HIV)-1 predominates during asymptomatic disease followed by occasional emergence of CXCR4-using (x4) or dual tropic (r5x4) virus
We examined the impact of both the “CXCR4”-using and “CCR5”using clones to the replicative fitness of r5x4 HIV-1 isolates in CD4+/CCR5+ cells, CD4+/CXCR4+ cells, and in the mixed cell population found in peripheral blood mononuclear cells (PBMC)
It is important to note that the propagation of primary HIV-1 isolates on PHA-activated PBMCs commonly enriches for CXCR4-using virus even in HIV-1 populations with a low frequency x4 compared to r5 variants [57]
Summary
CCR5-using (r5) HIV-1 predominates during asymptomatic disease followed by occasional emergence of CXCR4-using (x4) or dual tropic (r5x4) virus. Human immunodeficiency virus entry into host cells involves binding to CD4 and one of two coreceptors, namely CCR5 and CXCR4, the two viral phenotypes, the r5- and x4-using viruses. Env gp120/gp form trimer spikes that mediate virus attachment to the receptor complex on the cell surface (reviewed in [14]). Upon interaction with CD4, the V3 loop of gp120 can engage the second extracellular loop of CCR5 (or CXCR4) while a gp120 conformational change exposes a second Env site (comprised in part by the C2 region) to bind with the N terminus of the co-receptor
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