Background
Characterization of HIV-1 persistence in both CD4+ T cell and myeloid reservoirs within systemic tissues and the brain are goals of the cure field. Clade D HIV-1 infects both CD4+ T cells and macrophages. Thus, we have employed SHIV encoding a clade D transmitted/founder HIV-1 Env to study reservoirs in ART-suppressed rhesus macaques (RM).
Methods
Ten RM were infected with barcoded SHIV.D.191859, started on ART 10 weeks post-infection, and continued through necropsy at 9 (n=5) or 20 months (n=5) of ART. Blood and tissue samples were tested for viral load, barcode clonotype distribution, total and intact proviral HIV DNA, and RNA/DNAscope.
Results
Plasma viremia peaked at week 2 post-infection (median 2.5 x 106 copies/mL) with barcode sequencing indicating a multiplicity of infection between 45-400 unique clonotypes. At week 4, viral loads ranged from 105-107 copies/mL in plasma and 102-103 copies/mL in CSF. ART was initiated during setpoint viremia (median 2 x 104 copies/mL) at week 10 and durably suppressed in plasma and CSF.
Reservoir quantification at necropsy showed persistence of SHIV.D within CD4+ T cells and myeloid cells systemically (blood, 5 lymph nodes, spleen). In animals necropsied after 9 months on ART, total and intact proviral DNA in was ~10X higher in CD4+ T cells (median 3.5 x 103 total copies, 1.3 x 103 intact copies/106 cells) than in myeloid cells (median 1.2 x 102 total copies, 45 intact copies/106 cells) across compartments. Animals necropsied at 20 months showed similar ratios, with ~10X lower absolute measures. Within the brain, all necropsied animals had evidence of SHIV RNA and DNA by immunohistochemistry; 6/10 RM had detectable SHIV proviral DNA within purified brain myeloid cells.
Sequencing of barcode clonotypes from plasma and CSF during viremia, and within the brain and systemic tissues at necropsy, showed substantial overlap between viruses infecting CD4+ T cells and myeloid cells systemically, and between the brain and systemic tissues.
Conclusions
The dual CD4+ T cell- and macrophage-tropic SHIV.D persists on ART in myeloid and CD4+ T cells within the brain and systemic tissues. With 10 weeks of virus replication prior to ART, there was no compartmentalization of viruses between the brain and systemic tissues, and none evolved on suppressive ART. Between 9 and 20 months of ART, brain and systemic reservoirs decrease in size but remain detectable. Together, these results inform the use of SHIV.D infected RM for persistence and cure studies.
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