多重PCR快速检测恶性疟和间日疟的研究

目的 建立一种简便快速、能同时检测恶性疟和间日疟的核酸检测方法。方法 针对两种疟原虫18S rRNA基因设计2对(3条引物),优化引物浓度与退火温度,建立可扩增出两种疟原虫基因片段的多重PCR。并进行最低检测限确定和临床标本检测,以镜检法为金标准分析灵敏度和特异度等指标。结果 该方法可扩增出431 bp(恶性疟原虫)和341 bp(间日疟原虫)基因片段,最低检测限为10²copies/反应,检测临床标本的结果与镜检法无差别(P＞0.05),敏感度为93.55%,特异度为70.83%,阳性预测值为89.23%,阴性预测值为80.95%。结论 所建立的多重PCR方法可快速检测疟疾感染并鉴别分型,灵敏度高,值得推广。

Multiplex PCR for detection of Plasmodium falciparum and Plasmodium vivax

A multiplex PCR to detect Plasmodium falciparum （P. falciparum） and 191asmodiurn vivax （P. vivax） was established simply and rapidly in our study. Two pairs of primers （3 strips） were designed according to the sequences of the 18S rRNA of the two kinds of Plasmodium. The reacting system was optimized by analysis on the different primer concen- tration and annealing temperature. The patients＇ blood samples were tested with the optimized multiplex PCR, and the limit of detection of this assay was estimated. The sensitivity and specificity were calculated using microscopy as the gold standard. The sizes of amplification products of multiplex PCR amplifying genomic DNA of P. falciparum and P. vivax were 431 bp and 341 bp, respectively. The limit of detection of this assay was 102 copies/reaction. The clinical sensitivity was 93.55 % and the specificity was 70.83%. The positive predictive value （PPV） was 89.23% and the negative predictive value （NPV） was 80.95%. So we concluded that the multiplex PCR with high sensitivity could detect and identify malaria rapidly, which is suit- able for primary Centers for Disease Prevention and Control.