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https://www.arca.fiocruz.br/handle/icict/37055
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2020-11-14
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- INI - Artigos de Periódicos [3460]
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MULTIPLEX REAL-TIME PCR FOR THE DETECTION AND QUANTITATION OF HTLV-1 AND HTLV-2 PROVIRAL LOAD: ADDRESSING THE ISSUE OF INDETERMINATE HTLV RESULTS
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Affilliation
University College Dublin. National Virus Reference Laboratory. Dublin, Ireland
University College Dublin. Centre for Research in Infectious Diseases. Dublin, Ireland.
University College Dublin. National Virus Reference Laboratory. Dublin, Ireland
University College Dublin. Centre for Research in Infectious Diseases. Dublin, Ireland.
Fundação Oswaldo Cruz. Instituto de Pesquisa Clínica Evandro Chagas. Laboratório de Pesquisa em Patogenia Viral. Rio de Janeiro, RJ, Brasil.
Fundação Oswaldo Cruz. Instituto de Pesquisa Clínica Evandro Chagas. Laboratório de Pesquisa Clínica em Neuroinfecções. Rio de Janeiro, RJ, Brasil.
University College Dublin. Centre for Research in Infectious Diseases. Dublin, Ireland / Fundação Oswaldo Cruz. Instituto de Pesquisa Clínica Evandro Chagas. Laboratório de Pesquisa Clínica em Neuroinfecções. Rio de Janeiro, RJ, Brasil.
University College Dublin. National Virus Reference Laboratory. Dublin, Ireland / University College Dublin. Centre for Research in Infectious Diseases. Dublin, Ireland.
University College Dublin. Centre for Research in Infectious Diseases. Dublin, Ireland.
University College Dublin. National Virus Reference Laboratory. Dublin, Ireland
University College Dublin. Centre for Research in Infectious Diseases. Dublin, Ireland.
Fundação Oswaldo Cruz. Instituto de Pesquisa Clínica Evandro Chagas. Laboratório de Pesquisa em Patogenia Viral. Rio de Janeiro, RJ, Brasil.
Fundação Oswaldo Cruz. Instituto de Pesquisa Clínica Evandro Chagas. Laboratório de Pesquisa Clínica em Neuroinfecções. Rio de Janeiro, RJ, Brasil.
University College Dublin. Centre for Research in Infectious Diseases. Dublin, Ireland / Fundação Oswaldo Cruz. Instituto de Pesquisa Clínica Evandro Chagas. Laboratório de Pesquisa Clínica em Neuroinfecções. Rio de Janeiro, RJ, Brasil.
University College Dublin. National Virus Reference Laboratory. Dublin, Ireland / University College Dublin. Centre for Research in Infectious Diseases. Dublin, Ireland.
Abstract
Background: Routine diagnosis of Human T Lymphotropic virus (HTLV) infection is primarily serologically based; however the proportion of unresolved and indeterminate Western blot results range from 0.02% to 50% in endemic areas. Objectives: To validate a sensitive in-house quantitative multiplex real-time assay (mqRT-PCR), capable of detecting and quantifying HTLV-1 and HTLV-2, and use it to differentiate unresolved serological profiles, and monitor infection in HTLV-1 infected patients. Study Design: The mqRT-PCR was designed as a single-tube assay. Quantitative results were reported as copy number of HTLV provirus per 106 cells and the numbers of cells were calculated based on the quantitative result for albumin, of which there are 2 copies/cell. Assay standards were amplified from HTLV-1 infected MT-2 cells and HTLV-2 transfected CEM cells. Blood samples were obtained from HTLV seropositive former blood donors. Results: The mqRT-PCR assay was efficient (98.8–101.2%), reproducible (coefficient of variance < 5%) and sensitive to 1 copy for HTLV-1, HTLV-2 and Albumin. The assay resolved the infection profile in 16/17 patients, with undetermined subtype, all of which were reassigned as HTLV-1 infections. In addition, the average PVL detected in patients suffering from HTLV-1 associated HAM/TSP (n = 23, 13,450 copies/106 cells) was significantly higher than those detected in asymptomatic carriers (n = 21, 6665 copies/106 cells). Conclusions: We propose a new testing algorithm for the laboratory diagnosis of HTLV infection, which includes HTLV specific mqRT-PCR for resolving HTLV serological results. Furthermore, quantitation of PVL load by real-time PCR may be useful in assessing the link between infection and disease, and in monitoring patients undergoing therapy.
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