Rapid detection and quantification of Ebola Zaire virus by one-step real-time quantitative reverse transcription-polymerase chain reaction.
Given that Ebola virus causes severe hemorrhagic fever in humans with mortality rates as high as 90%, rapid and accurate detection of this virus is essential both for controlling infection and preventing further transmission. Here, a one-step qRT-PCR assay for rapid and quantitative detection of an Ebola Zaire strain using GP, VP24 or VP40 genes as a target is introduced. Routine assay conditions for hydrolysis probe detection were established from the manufacturer's protocol used in the assays. The analytical specificity and sensitivity of each assay was evaluated using in vitro synthesized viral RNA transcripts. The assays were highly specific for the RNA transcripts, no cross-reactivity being observed among them. The limits of detection of the assays ranged from 102to 103copies per reaction. The assays were also evaluated using viral RNAs extracted from cell culture-propagated viruses (Ebola Zaire, Sudan and Reston strains), confirming that they are gene- and strain-specific. The RT-PCR assays detected viral RNAs in blood samples from virus-infected animal, suggesting that they can be also a useful method for identifying Ebola virus in clinical samples.
Department of Biochemistry, Graduate School of Medicine, Konkuk University, Seoul 143-701, Korea.|Department of Virology and Immunology, Texas Biomedical Research Institute 7620 NW Loop 410, San Antonio, TX78227, USA.
Young-Tae
YT
Ro
Department of Virology and Immunology, Texas Biomedical Research Institute 7620 NW Loop 410, San Antonio, TX78227, USA.
Anysha
A
Ticer
Department of Virology and Immunology, Texas Biomedical Research Institute 7620 NW Loop 410, San Antonio, TX78227, USA.
Ricardo
R
Carrion
Department of Virology and Immunology, Texas Biomedical Research Institute 7620 NW Loop 410, San Antonio, TX78227, USA.
Jean L
JL
Patterson
D000818:Animals; D001769:Blood / Q000821:virology; D029043:Ebolavirus / Q000235:genetics / Q000302:isolation & purification*; D019142:Hemorrhagic Fever, Ebola / Q000175:diagnosis* / Q000821:virology*; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D014759:Viral Envelope Proteins / Q000235:genetics; D014763:Viral Matrix Proteins / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics
C446938:VP24 protein, Ebola virus; C080981:VP40 protein, virus; D014759:Viral Envelope Proteins; D014763:Viral Matrix Proteins; D014764:Viral Proteins; C113189:envelope glycoprotein, Ebola virus
Ebola Zaire virus; VP24; VP40 gene detection; one-step qRT-PCR