摘要
目的： 建立在同一次扩增中即可鉴别患者感染的疟原虫虫种的复式ＰＣＲ检测方法。方法： 根据恶性疟原虫（Ｐ．ｆ．）中度重复基因序列ｐＢＲＫ１－１４ 和间日疟原虫（Ｐ．ｖ．）线粒体细胞色素Ｃ氧化酶基因ＣＯＩＩＩ合成引物，采用经优化的ＰＣＲ反应体系， 对疟原虫ＤＮＡ模板进行扩增。结果： Ｐ．ｆ．与Ｐ．ｖ．分别被扩增出２０６ 和３７０ ｂｐ 大小的ＤＮＡ片段，与人白细胞ＤＮＡ 无交叉；用该反应体系至少可检测出原虫血症为５×１０－ ７的Ｐ．ｆ．感染和１．０２×１０－ ６ Ｐ．ｖ．感染； 自云南疟疾流行区采集的７８３份滤纸干血滴样本中， 复式ＰＣＲ法阳性检出率为８５．８％ ， 误诊率为０， 漏诊率为０．１％ ， 而镜检法依次分别为８４．９％ 、３．１％ 和１．０％ ， 两者符合率为９５．８％ 。结论： 本复式ＰＣＲ检测疟原虫较镜检敏感、特异， 适用于我国恶性疟与间日疟混合流行区的疟疾诊断、流行病学调查、药物的疗效考核和献血员的筛选等。
AIM: To establish a sensitive and specific PCR\|based method to detect Plasmodium falciparum and P.vivax in blood samples in a single amplification reaction. METHODS: Malaria parasite DNA in blood was amplified by the multiplex polymerase chain reaction using two sets of primers derived from the P.f. moderately\|repetitive DNA sequence and COIII gene of P.v. RESULTS: A 206\| bp product for P.f. and a 370\| bp product for P.v. were amplified by multiplex PCR, being able to detect parasitemia level as low as 5×10\+\{-7\} for P.f. and 1\^02×10\+\{-6\} for P.v. and having no cross\|reaction with human leucocyte DNA. A total of 783 blood samples on the filter paper collected from patients attending to malaria clinics in malaria endemic areas were detected. The positive rate of multiplex PCR was 85\^8%, the misdiagnosis rate was 0, and the under\|diagnosis rate was 0\^1%, while these three rates of microscopic examination were 84\^9%, 3\^1% and 1\^0%, respectively. The concordance between the two methods was 95\^8%. CONCLUSION: The multiplex PCR method made the malaria detection more sensitive and specific than the microscopic examination and should be suitable for the diagnosis of malaria in mixed endemic areas, large\|scale epidemiological studies, follow\|up of drug treatment and donor blood screenig.\;
出处
《中国寄生虫学与寄生虫病杂志》
CAS
CSCD
北大核心
1999年第6期380-383,共4页
Chinese Journal of Parasitology and Parasitic Diseases
关键词
恶性疟原虫
间日疟原虫
聚合酶链反应
Multiplex PCR, detection, Plasmodium falciparum, Plasmodium vivax
