SYBR Green–based quantitation of human T-lymphotropic virus type 1 proviral load in Peruvian patients with neurological disease and asymptomatic carriers: Influence of clinical status, sex, and familial relatedness

Abstract

To evaluate the human T-lymphotropic virus type 1 (HTLV-1) proviral DNA load in patients with HTLV-1–associated myelopathy/tropical spastic paraparesis (HAM/TSP) and asymptomatic HTLV-1 carriers, a SYBR Green–based real-time quantitative polymerase chain reaction (qPCR) assay was developed. HTLV–1 proviral DNA in peripheral blood mononuclear cells (PBMCs) was quantified using primers targeting the pX region and the HTLV-1 copy number normalized to the amount of ERV-3 (Endogenous Retrovirus 3) cellular DNA. Thirty-three asymptomatic HTLV-1 carriers (ACs) and 39 patients with HAM/TSP were enrolled. Some participants were relatives of HAM/TSP cases (16 ACs and 7 patients with HAM/TSP). On multiple linear regression analysis, the authors found a significant association between clinical status and HTLV-1 proviral load (P < .01), but only among women. ACs showed a median proviral load of 561 copies per 10⁴ PBMCs (interquartile range: 251–1623). In HAM/TSP patients, the median proviral load was 1783 (1385–2914). ACs related to HAM/TSP patients presented a relatively high proviral load (median 1152); however, the association between relatedness to a HAM/TSP patient and proviral load was not significant (P = .1). In HAM/TSP patients, no association was found between proviral load and disease duration, progression or severity. The fact that the effect of HAM/TSP upon the HTLV-1 proviral load differed between sexes and the finding of a high proviral load among asymptomatic relatives of HAM/TSP patients suggest that not yet identified genetic or environmental factors influence the pathogenesis of HTLV-1 infection.

Keywords:

• human T-lymphotropic virus 1
• paraparesis, PBMC
• Peru
• proviral load
• tropical spastic

This work was supported by the Directorate-General for Development Cooperation (DGDC) of the Belgian Government (framework agreement 02, project 95501) and by a specific “Own Initiative on HTLV-1 in Peru,” sponsored by the Flemish Interuniversity Council (VLIR).

The authors thank Afilio Tello, their health worker, for his care for all patients with HTLV-1 infection; Giovanni López for his technical assistance; and Dr. Daniel Clark for critical reading of the manuscript. The authors also thank Dr. Thérèse Astier-Gin (CNRS UMR5097, Université Victor Ségalen Bordeaux2, France), who kindly provided the HTLV-1 plasmid p4.39 for the proviral load assays.