Background: Maximizing the response of specific HIV-1 cellular reservoirs to anti-retroviral therapy is critical for achieving the goal of minimizing or eliminating viral evolution.

Methods: Peripheral blood mononuclear cells from 29 individuals off anti-retroviral therapy were collected and stained with combinations of T-cell (CD4, CD45RO, HLA-DR, CD62L, CCR7) and monocyte (CD14, CD16) immunophenotypic markers followed by ultrasensitive fluorescence in situ hybridization (UFISH) for HIV-1 gag-pol mRNA. Plasma viral load was determined using the Roche Amplicor system (limit 50 copies). Three (3) individuals were placed on anti-retroviral therapy for 48 wks consisting of 2 nucleoside inhibitors and a protease inhibitor to study the response of HIV-1 to therapy in specific cellular reservoirs.

Results: We found that HIV-1 infected T-cells are larger than uninfected T-cells (13.4 uM vs 10.6 uM, p < 0.001, Mann-Whitney) therefore they do not appear in normal lymphocyte gates by light scatter. In addition, monocytes (MM) with macrophage differentiation (CD14⁺, CD16⁺) exhibit greater HIV-1 transcriptional activity than CD14⁺ monocytes lacking CD16. Last, marked variability exists in HIV-1 transcriptional activity in T-cell subsets. The percentage of HIV-1 infected CD4⁺, HLA-DR⁺, CCR7⁺ T-cells was greater than the percentage CD4⁺, CD45RO⁺, HLA-DR⁺ T-cells or CD4⁺, CD45RO⁺, HLA-DR- (resting) T-cells. Transcriptional activity in the naive (CD4⁺, CD45RO^-) T-cell was at or below the limits of detection. Viral decay in the CD14⁺, CD16⁺ MM (Pearson Correlation 0.84, p = 0.00004) and in the CD4⁺, HLA-DR⁺, CCR7⁺ T-cell (Pearson Correlation 0.6, p = 0.017) reservoirs correlated with viral decay of plasma viral load in the 2 individuals that became undetectable (< 50 copies) and the 1 individual with persistently elevated plasma viral load (48,012).

Conclusions: Here, we report HIV-1 transcriptional activity in previously uncharacterized subpopulations of monocytes and T-cells. Because changes in transcriptional activity in these reservoirs correlates with changes in plasma viral load, these reservoirs most likely represent the predominant cellular reservoirs of HIV-1 in infected individuals. Tailoring antiretroviral therapy to minimize viral replication in these and other more persistent reservoirs should aid in controlling viral evolution.