Analysis of human cytomegalovirus in the healthy human carrier

Taylor-Wiedeman, Jean (1992). Analysis of human cytomegalovirus in the healthy human carrier. PhD thesis The Open University.

Abstract

Much circumstantial evidence has pointed to peripheral blood leukocytes as one site of persistence of Human Cytomegalovirus (HCMV) in healthy carriers. However, the exact population of peripheral blood cells that carry HCMV and to what extent they express HCMV gene products in not known. I have examined the sites of HCMV persistence in the peripheral blood of healthy carriers. Analysis of pure cell populations by the use of the polymerase chain reaction (PCR), sensitive to between 1 and 10 copies of the HCMV genome, showed that the predominant site of persistence was the monocyte. In addition, analysis of healthy seronegative subjects revealed that a significant number (30%) also harbored HCMV. Finally, study of granulocytes demonstrated no evidence of persistent HCMV.

Expression of HCMV during persistence was also analyzed, by using reverse transcription PCR (RT-PCR) with a sensitivity of between 1 and 100 infected fibroblasts. RNA from monocytes showed no evidence of polyadenylated immediate early (IE) or late transcripts. In contrast, in vitro differentiated monocyte-derived macrophages (MDM) did show evidence of HCMV gene expression with the class of HCMV genes expressed dependent on the method of differentiation. MDM treated with hydrocortisone (HC) and phorbol 12-myristate 13-acetate, expressed only lEI, but not IE2, glycoprotein B (gB) or phosphoprotein 28 (pp28) transcripts. Whereas, MDM treated with granulocyte-macrophage colony stimulating factor and HC expressed lEI, IE2 and gB, but not pp28 transcripts. In both cases, cocultivation experiments did not show plaques. Therefore, in the healthy carrier, persistence of HCMV in monocytes is independent of HCMV lytic gene expression, but in vitro differentiation of monocytes to MDM induced endogenous HCMV transcription consistent with the known permissivity of in vivo differentiated macrophages to HCMV infection.

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