Advanced yellow fever virus genome detection in point-of-care facilities and reference laboratories.
Reported methods for the detection of the yellow fever viral genome are beset by limitations in sensitivity, specificity, strain detection spectra, and suitability to laboratories with simple infrastructure in areas of endemicity. We describe the development of two different approaches affording sensitive and specific detection of the yellow fever genome: a real-time reverse transcription-quantitative PCR (RT-qPCR) and an isothermal protocol employing the same primer-probe set but based on helicase-dependent amplification technology (RT-tHDA). Both assays were evaluated using yellow fever cell culture supernatants as well as spiked and clinical samples. We demonstrate reliable detection by both assays of different strains of yellow fever virus with improved sensitivity and specificity. The RT-qPCR assay is a powerful tool for reference or diagnostic laboratories with real-time PCR capability, while the isothermal RT-tHDA assay represents a useful alternative to earlier amplification techniques for the molecular diagnosis of yellow fever by field or point-of-care laboratories.
Robert Koch Institute, Berlin, Germany. Domingo-CarrascoC@rki.de
Cristina
C
Domingo
Pranav
P
Patel
Jasmin
J
Yillah
Manfred
M
Weidmann
Jairo A
JA
Méndez
Emmanuel Rivalyn
ER
Nakouné
Matthias
M
Niedrig
D019411:Clinical Laboratory Techniques / Q000379:methods*; D016679:Genome, Viral; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D019095:Point-of-Care Systems; D012367:RNA, Viral / Q000235:genetics*; D012680:Sensitivity and Specificity; D014773:Virology / Q000379:methods; D015004:Yellow Fever / Q000175:diagnosis* / Q000821:virology; D015005:Yellow fever virus / Q000235:genetics / Q000302:isolation & purification*
D012367:RNA, Viral