Background:  Infection initiated by a single HIV-1 variant gives rise to a heterogeneous population driven by immune selection, adaptation to distinct cellular niches, and isolation in distinct compartments within a host. For example, multiple env variants can represent a mixture of CXCR4- and CCR5-tropic virus, and because CXCR4 is differentially expressed on naïve and memory T cells and macrophages, viral populations with different co-receptor phenotypes may be compartmentalized in these distinct cell types. To assess the degree of compartmentalization of CXCR4- and CCR5-tropic virus, we employed 2 strategies, one that exploits the differential decay characteristics of virus emerging from cell types with potentially different half-lives in patients initiating HAART, and the analysis of genetic linkage between determinants of tropism in V3 sequences and distal genetic markers.

Methods:  Viral tropism was predicted based on V3 sequences using a position-specific scoring matrix. HTA was used to track the differential rates of loss of V3 and V4/V5 variants in longitudinal plasma samples drawn frequently upon initiation of HAART. Linkage analysis in R5/X4 populations was done on sequences generated using Single Genome Amplification and sequencing of V1 through the 3' R. Linkage disequilibrium analysis was done using the DnaSP software and Bayesian phylogenetic analysis.

Results:  Variant decay analysis carried out on one subject with a R5/X4 mixed tropic population did not show differential decay of either V3 or V4/V5 env variants, suggesting that CXCR4- and CCR5-tropic V3 variants were not compartmentalized in cell types with different half-lives. Linkage analysis of 10 additional subjects with R5/X4 tropic populations revealed varying degrees of linkage between specific V3 sequences and markers in gp120, gp41, nef and the long terminal repeat (LTR). Markers in gp120 were in high linkage disequilibrium (R^2) with specific V3 sequences; however, a precipitous decline in linkage disequilibrium was observed between the V3 loop and markers in gp41 and beyond.

Conclusions:  The results indicate that CCR5- and CXCR4-tropic populations, as predicted by their V3 sequences, are not compartmentalized in cell types with distinct half-lives. The lack of genetic linkage outside of the regions proximal to V3 also suggests that R5 and X4 variants can recombine, again suggesting mixing. These results suggest that to the extent X4 viruses populate naïve T cells, some mixing continues to occur in memory T cells.