Introduction:HTLV-1 and HTLV-2 are human retroviruses that primarily infect immune system T cells, causing them to lose their ability to defend the organism.These viruses are transmitted through breastfeeding, unprotected sex, and blood.Diagnosis of the infection is based on the detection of specific antibodies through serological tests, in which screening tests (ELISA or chemiluminescence) are performed; once positive, a Western Blot test is done as a confirmatory test.However, many cases are indeterminate by serological tests, making it difficult to reach a definitive diagnosis.Therefore, it is necessary to develop molecular assays to assist in the confirmation and typing of these cases.Objectives: To evaluate the performance of two molecular assays under development, based on real-time PCR for the detection and typing of HTLV-1 and HTLV-2.Methodology: Two molecular assays based on real-time PCR were tested: assay 1 and assay 2 with the detection and typing of HTLV-1 and 2 based on the pol and tax regions for each target.Both moleculars assays detect HTLV-1 by FAM fluorescence, while HTLV-2 is detected by RED (Assay 1) and VIC (Assay 2) fluorescence.Ribonuclease P (RNase P) was included as an internal control in both assays, being detected by Cy5 dye.To evaluate the performance of both molecular assays, an analysis was performed using three serial dilutions of an HIV-based Virus-Like Particle (VLP) that serves as a positive control in the assays.The VLP was constructed from a plasmid into which fragments of the HTLV-1 and HTLV-2 pol and tax genes had been inserted.In addition, commercial negative control based on synthetic DNA was tested to assess RP Ct values for both tests.Ethics committee approval was not required because a synthetic virus was used. Results:The Assay 1 showed mean Ct values of 28.9, 32.5, and 34.3 for the first, second, and third dilutions of HTLV-1, respectively.For HTLV-2, the mean Ct values were 30.2, 33.6, and 35.8.In Assay 2, the mean Ct values for HTLV-1 were 29.7, 32.5, and 35.2 across the three dilutions, whereas for HTLV-2 they were 31.3,34.1, and 37.1, respectively.The negative control evaluated by RP had a mean Ct of 30.4 in Assay 1, and in Assay 2 the mean Ct was 31.1, with nonspecificity for HTLV-1 in some replicates. Conclusion:The two molecular assays showed similar sensitivity for HTLV-1 and HTLV-2, demonstrating that they are promising assays for molecular diagnosis, viral typing, and effective clinical and epidemiological management.
Brandão et al. (Thu,) studied this question.