SENSITIVITY AND SPECIFICITY OF A UNIVERSAL PRIMER SET FOR THE RAPID DIAGNOSIS OF DENGUE VIRUS-INFECTIONS BY POLYMERASE CHAIN-REACTION AND NUCLEIC-ACID HYBRIDIZATION
SENSITIVITY AND SPECIFICITY OF A UNIVERSAL PRIMER SET FOR THE RAPID DIAGNOSIS OF DENGUE VIRUS-INFECTIONS BY POLYMERASE CHAIN-REACTION AND NUCLEIC-ACID HYBRIDIZATION
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DOI：
10.4269/ajtmh.1991.45.418
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发表时间：
1991-10-01
影响因子：
3.3
通讯作者：
HOKE, CH
中科院分区：
文献类型：
--
作者：
HENCHAL, EA;POLO, SL;HOKE, CH
一组正义和反义寡聚DNA引物，简并在第三个“摆动”碱基位置的密码子，以匹配所有已知的登革病毒序列，被评估为通用引物的聚合酶链反应（PCR）检测登革病毒感染的快速诊断。通过从血清中提取的总RNA的逆转录（RT）制备病毒特异性互补DNA（cDNA）。扩增的cDNA通过与四种不同类型特异性寡聚DNA探针的核酸杂交来鉴定。使用登革热患者的血清，RT/PCR，然后使用放射性标记的探针进行核酸杂交，灵敏度为68%（50/74; 95%置信区间[CI] = 57-78%），特异性为100%。杂交产物的化学发光检测的灵敏度为62%（26/42; 95%CI = 46-75%）。使用已获得病毒分离株的标本，RT/PCR后用放射性标记探针进行核酸杂交的灵敏度为80%（40/50; 95% CI = 69-91%），特异性为100%。结果表明，简并引物RT/PCR是一种敏感、特异的登革病毒检测方法。
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.