Department of Parasitology, Faculty of Medicine, Molecular Biology of Malaria and Opportunistic Parasites Research Unit, Chulalongkorn University, Bangkok, Thailand.
Clin Microbiol Infect. 2011 Oct;17(10):1484-91. doi: 10.1111/j.1469-0691.2011.03507.x. Epub 2011 May 20.
Saliva and urine from malaria-infected individuals contain trace amounts of Plasmodium DNA, and therefore, could be used as alternative specimens for diagnosis. A nested PCR targeting the mitochondrial cytochrome b gene (Cytb-PCR) of four human malaria species and Plasmodium knowlesi was developed and tested with 693 blood samples from febrile patients living in diverse malaria-endemic areas of Thailand, and compared with microscopy and nested PCR targeting small-subunit rRNA (18S-PCR). Cytb-PCR was 16% and 39.8% more sensitive than 18S-PCR and microscopy, respectively, in detecting all of these malarial species in blood samples. Importantly, 34% and 17% of Plasmodium falciparum and Plasmodium vivax mono-infections, respectively, detected by microscopy were, in fact, mixed P. falciparum and P. non-falciparum infections. Analysis of matched blood, saliva and urine from 157 individuals showed that microscopy and Cytb-PCR of saliva yielded no significant difference in detecting P. falciparum and P. vivax. However, Cytb-PCR of saliva was more sensitive than microscopy for diagnosis of mixed-species infections. A combination of Cytb-PCR of saliva and of urine significantly outperformed microscopy (p 0.0098 for P. falciparum, p 0.006 for P. vivax, and p 0.0002 for mixed infections). Furthermore, Plasmodium malariae and P. knowlesi could also be identified in saliva and urine with this method. Therefore, the Cytb-PCR developed herein offers a high potential for the use of both saliva and urine for malaria diagnosis, with a sensitivity comparable with or superior to that of microscopy.
疟疾病人唾液和尿液中含有痕量的疟原虫 DNA，因此可作为替代标本用于诊断。本研究建立并检测了针对人疟原虫四种疟原虫和疟原虫 knowlesi 的线粒体细胞色素 b 基因（Cytb-PCR）的巢式 PCR，并与来自泰国不同疟疾流行地区的发热患者的 693 份血样进行了比较，与针对小亚单位 rRNA（18S-PCR）的显微镜检查和巢式 PCR 进行了比较。Cytb-PCR 在检测所有这些血液样本中的疟原虫时，比 18S-PCR 和显微镜检查分别分别提高了 16%和 39.8%的灵敏度。重要的是，显微镜检查检测到的 34%和 17%的恶性疟原虫和间日疟原虫单感染实际上是混合性疟原虫和非恶性疟原虫感染。对 157 名个体的匹配血液、唾液和尿液的分析表明，唾液的显微镜检查和 Cytb-PCR 在检测恶性疟原虫和间日疟原虫方面没有显著差异。然而，唾液 Cytb-PCR 对混合感染的诊断比显微镜检查更敏感。唾液 Cytb-PCR 和尿液 Cytb-PCR 的联合使用显著优于显微镜检查（恶性疟原虫：p 0.0098，间日疟原虫：p 0.006，混合感染：p 0.0002）。此外，该方法还可用于在唾液和尿液中鉴定出疟原虫和疟原虫 knowlesi。因此，本文建立的 Cytb-PCR 为利用唾液和尿液进行疟疾诊断提供了很高的潜力，其灵敏度与显微镜检查相当或优于显微镜检查。
