Generic RT-nested-PCR for detection of flaviviruses using degenerated primers and internal control followed by sequencing for specific identification.
Flaviviruses are a widespread and numerous group of arboviruses that can cause serious illness in humans. The continuous and slow spread of certain flaviviruses, such as Dengue viruses, and the recent entry and spread of West Nile virus to the American continent, point to the need to control these infections. This control requires the use of suitable techniques for diagnostic and surveillance programmes. A generic RT-nested-PCR that is, theoretically, able to detect each member of the group has been designed. The identification of the detected virus is carried out by sequencing. The introduction of an internal control would reduce the number of false negative results and could be used to quantify the viral load in clinical samples where the method works well.
Sánchez-Seco
M P
MP
Laboratory of Arboviruses and Imported Viral Diseases, Diagnostic Microbiology Service, National Center for Microbiology, Instituto de Salud Carlos III, Majadahonda, Madrid, Spain. paz.sanchez@isciii.es
Rosario
D
D
Domingo
C
C
Hernández
L
L
Valdés
K
K
Guzmán
M G
MG
Tenorio
A
A
D005188:False Negative Reactions; D005416:Flavivirus / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D018177:Flavivirus Infections / Q000175:diagnosis*; D006801:Humans; D010802:Phylogeny; D012367:RNA, Viral / Q000032:analysis* / Q000235:genetics; D012015:Reference Standards; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D017422:Sequence Analysis, DNA
D012367:RNA, Viral