Plasmodium falciparum-like Parasites Infecting Wild African Apes Do Not Represent a Recurrent Source of Human Malaria

Wild-living chimpanzees and gorillas harbor a multitude of Plasmodium spp., including six of the subgenus Laverania, one of which is the progenitor of P. falciparum. Despite the magnitude of this natural reservoir, it is unknown whether apes represent a recurrent source of human infections. Here, we used Plasmodium species-specific PCR, single genome amplification (SGA) and 454 sequencing to screen humans from remote areas of Cameroon for ape Laverania infections. Among 1,403 blood samples, we found 1,000 to be positive for Plasmodium mitochondrial (mt)DNA, all of which contained human parasites as determined by sequencing and/or restriction enzyme digestion. To exclude low abundance infections, we subjected 514 samples to 454 sequencing, targeting a region of the mtDNA genome that distinguishes ape from human Laverania species. Developing algorithms capable of differentiating rare Plasmodium variants from 454 sequencing error, we identified mono- and mixed-species infections with P. falciparum, P. malariae and/or P. ovale. However, none of the human samples contained ape Laverania parasites, including the gorilla precursor of P. falciparum. To characterize further the diversity of P. falciparum in Cameroon, we used SGA to amplify 3.4 kb mtDNA fragments from 229 infected humans. Phylogenetic analysis identified 62 new variants, all of which clustered with extant P. falciparum strains, providing further evidence that P. falciparum emerged following a single gorilla-to-human transmission. Thus, unlike P. knowlesi infected macaques in southeast Asia, African apes harboring Laverania parasites do not serve as a recurrent source of human malaria, a finding of import to ongoing control and eradication measures.

野生黑猩猩与大猩猩自然携带着多种疟原虫属（Plasmodium）物种，其中包含6种拉韦朗疟原虫亚属（Laverania）成员，其中一种是恶性疟原虫（P. falciparum）的祖先株。尽管这一天然储存宿主的规模庞大，但类人猿是否会反复成为人类疟疾感染的传染源仍未明确。本研究采用疟原虫种特异性聚合酶链反应（PCR）、单基因组扩增（SGA）与454测序技术，对喀麦隆偏远地区的人类开展类人猿来源的拉韦朗疟原虫感染筛查。在1403份血液样本中，共1000份检出疟原虫线粒体DNA（mtDNA）阳性；经测序及/或限制性酶切验证，所有阳性样本均携带人类疟原虫。为排除低丰度感染，我们对514份样本进行454测序，靶向区分类人猿与人类拉韦朗疟原虫物种的线粒体基因组区域。通过开发可区分罕见疟原虫变异株与454测序误差的算法，我们鉴定出恶性疟原虫、三日疟原虫（P. malariae）及/或卵形疟原虫（P. ovale）的单一感染与混合感染病例。然而，所有人类样本均未检出类人猿来源的拉韦朗疟原虫，包括恶性疟原虫的大猩猩祖先株。为进一步解析喀麦隆地区恶性疟原虫的遗传多样性，我们通过单基因组扩增技术，从229份人类感染样本中扩增得到3.4千碱基对（kb）的线粒体DNA片段。系统发育分析共鉴定出62种新的变异株，所有变异株均与现存恶性疟原虫菌株聚类，进一步证实恶性疟原虫起源于单次大猩猩向人类的跨种传播事件。因此，与东南亚地区受诺氏疟原虫（P. knowlesi）感染的猕猴不同，携带拉韦朗疟原虫的非洲类人猿并不会反复成为人类疟疾的传染源。这一发现对当前正在推进的疟疾防控与根除工作具有重要意义。