Rapid detection of human pathogenic orthobunyaviruses.
Modern detection and identification tools can help to provide answers to urgent questions about the incidence, prevalence, and epidemiology of currently emerging diseases. We developed highly sensitive one-step TaqMan reverse transcription-PCR assays with sensitivities ranging from 10(4) to 10(1) molecules for 11 human pathogens of the orthobunyaviruses. We compared the performances of these assays on three currently available cyclers (ABI-PRISM 7700, LightCycler, and SmartCycler). The assay for Oropouche virus (OROV) was tested using sera collected from days 1 to 5 after onset of OROV disease and was found to be greatly superior to an established nested PCR system. A mean copy number of 1.31 x 10(7) OROV RNA/ml of serum was detected. Diagnostic RNA detection can be used as early as day 1 after onset of OROV disease. The use of a mobile SmartCycler and a hands-on time of less than 3 h could help to intensify outbreak surveillance and control, especially in field studies.
Weidmann
Manfred
M
Department of Virology, Institute of Medical Microbiology and Hygiene, University of Freiburg, 79104 Freiburg, Germany. weidmann@ukl.uni-freiburg.de
Rudaz
Veronique
V
Nunes
Marcio R T
MR
Vasconcelos
Pedro F C
PF
Hufert
Frank T
FT
D000818:Animals; D002043:Bunyaviridae / Q000145:classification / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D002044:Bunyaviridae Infections / Q000175:diagnosis* / Q000821:virology; D002460:Cell Line; D002522:Chlorocebus aethiops; D017931:DNA Primers; D006801:Humans; D012367:RNA, Viral / Q000097:blood; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D012840:Simbu virus / Q000145:classification / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification; D019914:Taq Polymerase; D013997:Time Factors; D014709:Vero Cells
D017931:DNA Primers; D012367:RNA, Viral; D019914:Taq Polymerase