Abstract Body

Single cell.methods have enhanced the resolution at which cells in blood and tissues can be studied in people with HIV (PWH). Yet, capturing the native multi-omics state of cells obtained from multiple tissue compartments and from rare and vulnerable cells in cerebrospinal fluid (CSF) while avoiding artifacts that may arise from cryopreservation remains a challenge for single cell studies, more so in resource limited settings.

To demonstrate single cell studies of fresh cells from PWH are logistically feasible in a resource limited setting, we built upon the opportunities of the RV254/SEARCH010 and RV304/SEARCH013 studies enrolling people with acute and chronic HIV (AHI & CHI) in Bangkok, Thailand where uptake of optional procedures including leukapheresis, lumbar puncture (LP), gut biopsy, and lymph node (LN) biopsy is high. Fresh cells were isolated from gut and LN, CSF, and blood over 2 days from an ART naïve PWH with CHI (Participant 1, Figure 1A), and from gut and blood from a PWH on suppressive ART initiated during AHI (Participant 2). The 10X genomics platform was used locally to generate single cell transcriptome and T-cell/B-cell receptor data from fresh specimens within hours of sampling. Seurat suite was used for analysis.

Multi-omics single cell data was obtained for 28,400 freshly isolated lymph node cells, 5,968 gut cells, and 5,614 CSF cells from Participant 1 (Figure 1B). We also leveraged flow cytometry cell sorting capabilities of fresh T follicular helper cells from LN for multi-omics single cell profiling of 5,712 cells. In addition, multi-omics profiling was performed on 8,876 blood and 6,488 gut cells obtained on the same day from Participant 2. To enhance detection of HIV viral transcripts in the CHI ART naïve participant, we generated an individualized near full-length patched viral sequence to align sequencing reads and detected HIV transcript containing cells in all compartments with heterogenous single cells either producing high or low HIV transcripts. Notably, all HIV transcript containing cells in the CSF were identified in inferred CD4 memory T cells (Figure 1C).

We demonstrate the logistical feasibility of generating single cell multi-omics data from fresh cells from blood, CSF, LN, and gut in PWH in Bangkok. A personalized HIV mapping approach can be used to pinpoint infected single cells in tissue compartments including the CNS. This will enable cross-compartmental multi-omics studies to further interrogate HIV reservoirs.