Laboratory of Emerging Pathogens, Division of Emerging and Transfusion Transmitted Diseases, Center for Biologics Evaluation and Research, Food and Drug Administration, Silver Spring, MD, 20993, USA.
Department of Biological Environmental and Occupational Health Sciences, School of Public Health, College of Health Sciences, University of Ghana, Legon, Ghana.
Malar J. 2019 Apr 2;18(1):116. doi: 10.1186/s12936-019-2743-9.
Malaria remains a global public health problem responsible for 445,000 deaths in 2016. While microscopy remains the mainstay of malaria diagnosis, highly sensitive molecular methods for detection of low-grade sub-microscopic infections are needed for surveillance studies and identifying asymptomatic reservoirs of malaria transmission.
The Plasmodium falciparum genome sequence was analysed to identify high copy number genes that improve P. falciparum parasite detection in blood by RT-PCR. Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1)-specific primers were evaluated for P. falciparum detection in hospital-based microscopically positive dried blood spots and field-acquired whole blood from asymptomatic individuals from Ghana.
PfEMP1 outperformed the Pf18S sequence for amplification-based P. falciparum detection. PfEMP1 primers exhibited sevenfold higher sensitivity compared to Pf18S primers for parasite genomic DNA. Probit analysis established a 95% detection threshold of 9.3 parasites/mL for PfEMP1 compared to 98.2 parasites/mL for Pf18S primers. The PfEMP1 primers also demonstrated superior clinical sensitivity, identifying 100% (20/20) of dried blood spot samples and 70% (69/98) of asymptomatic individuals as positive versus 55% (11/20) and 54% (53/98), respectively, for Pf18S amplification.
These results establish PfEMP1 as a novel amplification target for highly sensitive detection of both acute infections from filter paper samples and submicroscopic asymptomatic low-grade infections.
疟疾仍是一个全球性的公共卫生问题，2016 年导致 44.5 万人死亡。虽然显微镜检查仍然是疟疾诊断的主要方法，但需要高度敏感的分子方法来检测低水平的亚微观感染，以便进行监测研究和确定疟疾传播的无症状储主。
分析恶性疟原虫基因组序列，以确定高拷贝数基因，通过 RT-PCR 提高血液中恶性疟原虫寄生虫的检测。评估疟原虫红细胞膜蛋白 1（PfEMP1）特异性引物在基于医院的显微镜阳性干血斑和加纳无症状个体现场采集的全血中对恶性疟原虫的检测。
PfEMP1 在基于扩增的恶性疟原虫检测中优于 Pf18S 序列。与 Pf18S 引物相比，PfEMP1 引物对寄生虫基因组 DNA 的灵敏度高 7 倍。概率分析建立了 PfEMP1 的 95%检测阈值为 9.3 个寄生虫/mL，而 Pf18S 引物的检测阈值为 98.2 个寄生虫/mL。PfEMP1 引物还表现出优越的临床灵敏度，在干血斑样本中，100%（20/20）和无症状个体中 70%（69/98）为阳性，而 Pf18S 扩增分别为 55%（11/20）和 54%（53/98）。
这些结果确立了 PfEMP1 作为一种新的扩增靶标，可高度敏感地检测滤纸样本中的急性感染和亚微观无症状低度感染。
