Optimisation of a quantitative polymerase chain reaction-based strategy for the detection and quantification of human herpesvirus 6 DNA in patients undergoing allogeneic haematopoietic stem cell transplantation

Author(s):Show less -

Miriam YH Ueda ^[1] ; Paulo G Alvarenga ^[2] ; Juliana M Real ^[3] ; Eloisa de Sá Moreira ^[4] ; Aripuanã Watanabe ^[5] ; Ana Maria Passos-Castilho ^[6] ; Matheus Vescovi ^[7] ; Yana Novis ^[8] ; Vanderson Rocha ^[9] ; Adriana Seber ^[10] ; Jose SR Oliveira ^[11] ; Celso A Rodrigues ; Celso FH Granato ^[13]

Affiliation:Show less -

^[1] Universidade Federal de São Paulo - Brasil

^[2] Universidade Federal de São Paulo - Brasil

^[3] Hospital Sírio Libanês. Instituto de Ensino e Pesquisa. Centro de Oncologia - Brasil

^[4] Dendrix Research - Brasil

^[5] Universidade Federal de São Paulo - Brasil

^[6] Universidade Federal de São Paulo - Brasil

^[7] Universidade Federal de São Paulo - Brasil

^[8] Hospital Sírio Libanês. Instituto de Ensino e Pesquisa. Centro de Oncologia - Brasil

^[9] Hospital Sírio Libanês. Instituto de Ensino e Pesquisa. Centro de Oncologia - Brasil

^[10] Instituto de Oncologia Pediátrica - Brasil

^[11] Universidade Federal de São Paulo - Brasil

^[13] Universidade Federal de São Paulo - Brasil

• Document type: Journal article
• Source: Memórias do Instituto Oswaldo Cruz; v. 110, n. 4, p. 461-467, 2015-06-02.

Abstract

Human herpesvirus 6 (HHV-6) may cause severe complications after haematopoietic stem cell transplantation (HSCT). Monitoring this virus and providing precise, rapid and early diagnosis of related clinical diseases, constitute essential measures to improve outcomes. A prospective survey on the incidence and clinical features of HHV-6 infections after HSCT has not yet been conducted in Brazilian patients and the impact of this infection on HSCT outcome remains unclear. A rapid test based on real-time quantitative polymerase chain reaction (qPCR) has been optimised to screen and quantify clinical samples for HHV-6. The detection step was based on reaction with TaqMan® hydrolysis probes. A set of previously described primers and probes have been tested to evaluate efficiency, sensitivity and reproducibility. The target efficiency range was 91.4% with linearity ranging from 10-106 copies/reaction and a limit of detection of five copies/reaction or 250 copies/mL of plasma. The qPCR assay developed in the present study was simple, rapid and sensitive, allowing the detection of a wide range of HHV-6 loads. In conclusion, this test may be useful as a practical tool to help elucidate the clinical relevance of HHV-6 infection and reactivation in different scenarios and to determine the need for surveillance. (AU)