Rapid and Sensitive Qualitative Duoplex Real-Time PCR Method for Discriminatory and Confirmatory Diagnosis of HTLV-1 and HTLV-2 Infections: Brazilian Multicentric Study

Frontiers in Medicine

article

Benedito Antonio Lopes da Fonseca¹  Augusto César Penalva de Oliveira²  Jerusa Smid²  Oswaldo Massaiti Takayanagui¹  Jorge Casseb²  Dimas Tadeu Covas³  Simone Kashima³  Mauricio Cristiano Rocha-Junior³  Evandra Strazza Rodrigues³  Svetoslav Nanev Slavov³  Tatiane Assone²  Maíra Pedreschi²  Debora Glenda Lima de La Roque³  Maisa Sousa⁶  Viviana Olavarria⁷  Bernardo Galvão-Castro⁷ 

[1] Department of Neurology, Ribeirão Preto Medical School, University of São Paulo;Laboratory of Medical Investigation LIM 56, Division of Dermatology, Medical School, University of São Paulo;Regional Blood Center of Ribeirão Preto, Ribeirão Preto Medical School, University of São Paulo;Center for Cell-Based Therapy CTC, Regional Blood Center of Ribeirão Preto, University of São Paulo;Graduate Program in Biosciences and Biotechnology, School of Pharmaceutical Sciences of Ribeirão Preto, University of São Paulo;Tropical Medicine Center, Federal University of Para;Centro de HTLV

关键词: HTLV-1;    HTLV-2;    molecular diagnosis;    real time PCR;    multiplex real time PCR;   

DOI  :  10.3389/fmed.2022.881630

学科分类：社会科学、人文和艺术（综合）

来源: Frontiers

【 摘 要 】

Human T cell lymphotropic virus (HTLV) is the caustive agent of two main conditions i. e., the HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and the adult T-cell leukemia/lymphoma (ATLL). HTLV diagnosis is based on serological and molecular approaches; however, an accurate and validated method is still needed. The objective of this study was to establish a rapid and sensitive molecular test to confirm and discriminate HTLV 1/2 types. The test validation was performed as a multicentric study involving HTLV confirmation centers throughout Brazil. Proviral DNA was extracted from whole blood and the amplification was performed using in-house designed primer and probe sets targeting the pol genomic region. An internal control to validate the extraction and amplification was also included. The limit of detection (LoD) of the assay was four copies/reaction for HTLV-1 and 10.9 copies/reaction for HTLV-2. The diagnostic sensitivity of the platform was 94.6% for HTLV-1, 78.6% for HTLV-2, and the specificity was 100% for both viruses. Cross-reactions of the test with human viruses including HAV, HBV, HCV, HIV-1/2, and parvovirus B19 were not observed. During the multicentric validation, the test was used to screen a total of 692 blood samples obtained from previously confirmed HTLV-positive individuals. From these, 91.1% tested positive being concordant with the previously obtained results. In conclusion, our duoplex-RT-PCR-HTLV1 /2 presented adequate efficiency for HTLV-1/2 differentiation showing high sensitivity and specificity. Therefore, it can be a suitable tool for confirmation of suspected and inconclusive HTLV cases, prenatal and pre-transplant diagnosis, in Brazil and in other countries HTLV-endemic countries.

【 授权许可】

CC BY