Development of a TaqMan® RT-PCR assay without RNA extraction step for the detection and quantification of African Chikungunya viruses

被引:225
作者
Pastorino, B
Bessaud, M
Grandadam, M
Murri, S
Tolou, HJ
Peyrefitte, CN
机构
[1] Inst Trop Med, Serv Sante Armees, Unite Virol Trop, Lab Associe,Ctr Natl Reference Arbovirus, F-13998 Marseille, France
[2] Univ Mediterranee, EA 3292, IFR 48, Marseille, France
[3] Inst Pasteur, Ctr Natl Reference Arbovirus, F-69365 Lyon 07, France
关键词
alphavirus; Chikungunya virus; heat-release; direct one-step TaqMan((R)) RT-PCR;
D O I
10.1016/j.jviromet.2004.11.002
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Chikungunya virus (CHIKV), a member of the alphavirus genus, is of considerable public health concern in Southeast Asian and African countries. However, despite serological evidence, the diagnosis of this arthropod-borne human disease is confirmed infrequently and needs to be improved. In fact, illness caused by CHIKV can be confused with diseases such as dengue or yellow fever, based on the similarity of the symptoms, and laboratory confirmation of suspected cases is required to launch control measures during an epidemic. Moreover, no quantitative molecular tool is described to study CHIKV replication or detection in clinical samples and cell culture supernatants. In this study, a specific and sensitive CHIKV one-step TaqMan(R) RT-PCR assay was developed as a tool for the diagnosis of African CHIKV as well as a rapid indicator of active infection by quantifying viral load. This study also showed that a simple heat viral RNA release during the reverse transcription step constituted an alternative to the conventional RNA extraction method. (C) 2004 Elsevier B.V. All rights reserved.
引用
收藏
页码:65 / 71
页数:7
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