Sensitivity and specificity of a universal primer set for the rapid diagnosis of dengue virus infections by polymerase chain reaction and nucleic acid hybridization.
A set of sense and anti-sense oligomeric DNA primers, degenerate in the third "wobble" base position of codons so as to match all known dengue virus sequences, was evaluated as universal primers in a polymerase chain reaction (PCR) assay for the rapid diagnosis of dengue virus infections. Virus-specific complementary DNA (cDNA) was prepared by reverse transcription (RT) of total RNA extracted from serum. Amplified cDNA was identified by nucleic acid hybridization with four serotype-specific, oligomeric DNA probes. Using sera from patients admitted with dengue fever, RT/PCR followed by nucleic acid hybridization using radiolabeled probes was 68% sensitive (50/74; 95% confidence interval [CI] = 57-78%) and 100% specific. Chemiluminescent detection of hybridized products was 62% sensitive (26/42; 95% CI = 46-75%). Using specimens from which a virus isolate had been obtained, RT/PCR followed by nucleic acid hybridization with radiolabeled probes was 80% sensitive (40/50; 95% CI = 69-91%) and 100% specific. The results suggest that RT/PCR using degenerate primers is a sensitive and specific method for the detection of dengue viruses in clinical specimens.
Henchal
E A
EA
Department of Virus Diseases, Walter Reed Army Institute of Research, Washington, DC.
Polo
S L
SL
Vorndam
V
V
Yaemsiri
C
C
Innis
B L
BL
Hoke
C H
CH
D015342:DNA Probes / Q000737:chemistry; D003715:Dengue / Q000175:diagnosis*; D003716:Dengue Virus / Q000235:genetics* / Q000302:isolation & purification; D004587:Electrophoresis, Agar Gel; D006801:Humans; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction; D011237:Predictive Value of Tests; D012367:RNA, Viral / Q000097:blood*; D014158:Transcription, Genetic
D015342:DNA Probes; D012367:RNA, Viral