Quantitative Viral Outgrowth Assay to Measure the Functional SIV Reservoir in Myeloid Cells

The role of CD4+ T cells in HIV infection and the latent reservoir, that is, latently infected cells that harbor replication competent virus, has been rigorously assessed. We have previously reported a quantitative viral outgrowth assay (QVOA) for SIV that demonstrated the frequency of latently infected CD4+ T cells is approximately 1 in a million cells, similar to that of HIV infected individuals on ART. However, the frequency of productively infected monocytes in blood and macrophages in tissues has not been similarly studied. Myeloid cells are infected during acute HIV and SIV infection; however, unlike lymphocytes, they are resistant to the cytopathic effects of the virus. Moreover, tissue-resident macrophages have the ability to self-renew and persist in the body for months to years. Thus, tissue macrophages, once infected, have the characteristics of a stable viral reservoir. A better understanding of the number of productively infected macrophages is critical to understanding the role of infected myeloid cells as a viral reservoir. In order to assess the functional latent reservoir. we have developed specific QVOAs for monocytes in blood, and macrophages in spleen, BAL and brain, which are described in detail in this chapter.