Browsing by Author "Ferreira, Isabella Anna Theresa Markham."

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Identifying the cellular HIV-1 reservoir in lymph nodes of antiretroviral therapy suppressed individuals.
(2019) Ferreira, Isabella Anna Theresa Markham.; Sigal, Alexander.
HIV-1 infection is suppressed but not cured in the face of antiretroviral therapy (ART). Pinpointing the cellular HIV-1 reservoir, which allows HIV-1 to persist, is key to the eradication of the virus. Lymph nodes are known to be a reservoir site for HIV-1 persistence, and we have assembled lymph nodes from a cross-sectional cohort of participants on suppressive ART to better understand the cellular HIV-1 reservoir. We developed a novel single-cell RNA-Seq methodology to identify the cellular HIV-1 reservoir in the lymph node compartment in ART suppressed individuals. HIV-1 positive cells from these lymph nodes were stained with anti-HIV- 1 antibodies and selected using flow cytometric sorting. Seq-Well, a high throughput single-cell RNA-Seq approach, was then performed to detect gag and env HIV-1 transcripts in individual cells, as well as the infected subtype using the cellular transcriptome. In parallel, the consensus near full length viral clone from the lymph node was sequenced and used for alignment. Using our methods for identifying HIV-1 infected cells from lymph nodes from chronically infected individuals, we have identified both known and novel putative host markers that are associated with persistent infection. These included co-expression of APOBEC3G, NFAT5, and NFKB2 in cells that contained HIV-1 mRNA. Our results show that cells with transcriptomes consistent with a T cell origin are the main infected population, and we are in the process of deeply characterizing the cell subtypes involved that also express markers of HIV-1 infection.