Human pegivirus (HPgV; previously called GB virus C/hepatitis G virus) has limited pathogenicity, despite causing persistent infection, and is associated with prolonged survival in human immunodeficiency virus-infected individuals. Although HPgV RNA is found in and produced by T- and B-lymphocytes, the primary permissive cell type(s) are unknown. We quantified HPgV RNA in highly purified CD4+ and CD8+ T-cells, including naïve, central memory and effector memory populations, and in B-cells (CD19+), NK cells (CD56+) and monocytes (CD14+) using real-time reverse transcription-PCR. Single-genome sequencing was performed on viruses within individual cell types to estimate genetic diversity among cell populations. HPgV RNA was present in CD4+ and CD8+ T-lymphocytes (nine of nine subjects), B-lymphocytes (seven of ten subjects), NK cells and monocytes (both four of five). HPgV RNA levels were higher in naïve (CD45RA+) CD4+ cells than in central memory and effector memory cells (P<0.01). HPgV sequences were highly conserved among subjects (0.117±0.02 substitutions per site; range 0.58–0.14) and within subjects (0.006±0.003 substitutions per site; range 0.006–0.010). The non-synonymous/synonymous substitution ratio was 0.07, suggesting a low selective pressure. Carboxyfluorescein succinimidyl ester (CFSE)-labelled HPgV RNA-containing particles precipitated by a commercial exosome isolation reagent delivered CSFE to uninfected monocytes, NK cells and T- and B-lymphocytes, and HPgV RNA was transferred to PBMCs with evidence of subsequent virus replication. Thus, HPgV RNA-containing serum particles including microvesicles may contribute to delivery of HPgV to PBMCs in vivo, explaining the apparent broad tropism of this persistent human RNA virus.
In vitro differentiation of embryonic stem cells into lymphocyte precursors able to generate T and B lymphocytes in vivo.
