Cholesterol-dependent enrichment of understudied erythrocytic stages of human Plasmodium parasites
Cholesterol-dependent enrichment of understudied erythrocytic stages of human Plasmodium parasites
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DOI：
10.1038/s41598-020-61392-6
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发表时间：
2020-03-12
影响因子：
4.6
通讯作者：
Guler, Jennifer L.
中科院分区：
文献类型：
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作者：
Brown, Audrey C.;Moore, Christopher C.;Guler, Jennifer L.
对于胞内病原体，宿主细胞提供所需的保护和营养。细胞内寄生虫研究的一个主要挑战是收集从宿主污染中分离出来的大量寄生虫。这种情况的例子是疟疾寄生虫，它花了相当大的一部分生命周期内的红细胞作为环，滋养体，和寄生虫，然后外出和再入侵。红细胞疟原虫寄生虫形式难富仍然研究不足，由于高主机污染相对于低寄生虫数量。在这里，我们提出了一种方法，分离所有阶段的疟原虫感染的红细胞，通过裂解和去除未感染的红细胞。链球菌溶血素O-Percoll（SLOPE）方法对以前无法获得的形式有效，包括疟疾感染患者的循环环和青蒿素诱导的静止寄生虫。SLOPE可在多种培养基配方下用于多种寄生虫种属，对寄生虫生存能力缺乏可测量的影响。我们证明红细胞膜胆固醇水平调节SLO对未感染宿主细胞的优先裂解，从而调节SLOPE的有效性。SLOPE富集环阶段样品的靶向代谢组学证实，与非富集样品相比，寄生虫衍生代谢物增加，污染宿主物质减少。由于其他细胞内细菌和原生动物消耗胆固醇，SLOPE具有改善疟原虫以外生物研究的潜力。
For intracellular pathogens, the host cell provides needed protection and nutrients. A major challenge of intracellular parasite research is collection of high parasite numbers separated from host contamination. This situation is exemplified by the malaria parasite, which spends a substantial part of its life cycle inside erythrocytes as rings, trophozoites, and schizonts, before egress and reinvasion. Erythrocytic Plasmodium parasite forms refractory to enrichment remain understudied due to high host contamination relative to low parasite numbers. Here, we present a method for separating all stages of Plasmodium-infected erythrocytes through lysis and removal of uninfected erythrocytes. The Streptolysin O-Percoll (SLOPE) method is effective on previously inaccessible forms, including circulating rings from malaria-infected patients and artemisinin-induced quiescent parasites. SLOPE can be used on multiple parasite species, under multiple media formulations, and lacks measurable impacts on parasite viability. We demonstrate erythrocyte membrane cholesterol levels modulate the preferential lysis of uninfected host cells by SLO, and therefore modulate the effectiveness of SLOPE. Targeted metabolomics of SLOPE-enriched ring stage samples confirms parasite-derived metabolites are increased and contaminating host material is reduced compared to non-enriched samples. Due to consumption of cholesterol by other intracellular bacteria and protozoa, SLOPE holds potential for improving research on organisms beyond Plasmodium.