Please use this identifier to cite or link to this item: http://docs.prosentient.com.au/prosentientjspui/handle/1/10606
Title: Multiplex Droplet Digital PCR Assay for Quantification of Human T-Cell Leukemia Virus Type 1 Subtype c DNA Proviral Load and T Cells from Blood and Respiratory Exudates Sampled in a Remote Setting.
Authors: Yurick, David
Khoury, Georges
Clemens, Bridie
Loh, Liyen
Pham, Hai
Kedzierska, Katherine
Einsiedel, Lloyd
Purcell, Damian
Affiliation: Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity at The University of Melbourne, Parkville, VIC, Australia..
Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity at The University of Melbourne, Parkville, VIC, Australia..
Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity at The University of Melbourne, Parkville, VIC, Australia..
Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity at The University of Melbourne, Parkville, VIC, Australia..
Baker Heart and Diabetes Institute, Alice Springs, NT, Australia..
Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity at The University of Melbourne, Parkville, VIC, Australia..
Baker Heart and Diabetes Institute, Alice Springs, NT, Australia.. Department of Medicine, Alice Springs Hospital, Alice Springs, NT, Australia..
Department of Microbiology and Immunology, The Peter Doherty Institute for Infection and Immunity at The University of Melbourne, Parkville, VIC, Australia dfjp@unimelb.edu.au..
Issue Date: Feb-2019
Citation: Journal of clinical microbiology 2019-02; 57(2)
Abstract: During human T-cell leukemia virus type 1 (HTLV-1) infection, the frequency of cells harboring an integrated copy of viral cDNA, the proviral load (PVL), is the main risk factor for progression of HTLV-1-associated diseases. Accurate quantification of provirus by droplet digital PCR (ddPCR) is a powerful diagnostic tool with emerging uses for monitoring viral expression. Current ddPCR techniques quantify HTLV-1 PVL in terms of whole genomic cellular material, while the main targets of HTLV-1 infection are CD4+ and CD8+ T cells. Our understanding of HTLV-1 proliferation and the amount of viral burden present in different compartments is limited. Recently a sensitive ddPCR assay was applied to quantifying T cells by measuring loss of germ line T-cell receptor genes as method of distinguishing non-T-cell from recombined T-cell DNA. In this study, we demonstrated and validated novel applications of the duplex ddPCR assay to quantify T cells from various sources of human genomic DNA (gDNA) extracted from frozen material (peripheral blood mononuclear cells [PBMCs], bronchoalveolar lavage fluid, and induced sputum) from a cohort of remote Indigenous Australians and then compared the T-cell measurements by ddPCR to the prevailing standard method of flow cytometry. The HTLV-1 subtype c (HTLV-1c) PVL was then calculated in terms of extracted T-cell gDNA from various compartments. Because HTLV-1c preferentially infects CD4+ T cells, and the amount of viral burden correlates with HTLV-1c disease pathogenesis, application of this ddPCR assay to accurately measure HTLV-1c-infected T cells can be of greater importance for clinical diagnostics and prognostics as well as monitoring therapeutic applications.
URI: http://docs.prosentient.com.au/prosentientjspui/handle/1/10606
DOI: 10.1128/JCM.01063-18
Type: Journal Article
Subjects: HTLV-1
T cells
ddPCR
induced sputum
peripheral blood
proviral load
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