柬埔寨青蒿素耐药性恶性疟原虫的新型表型检测方法：体外和体内药物反应研究。
Novel phenotypic assays for the detection of artemisinin-resistant Plasmodium falciparum malaria in Cambodia: in-vitro and ex-vivo drug-response studies.
机构信息
Malaria Molecular Epidemiology Unit, Institut Pasteur du Cambodge, Phnom Penh, Cambodia.
出版信息
Lancet Infect Dis. 2013 Dec;13(12):1043-9. doi: 10.1016/S1473-3099(13)70252-4. Epub 2013 Sep 11.
BACKGROUND
Artemisinin resistance in Plasmodium falciparum lengthens parasite clearance half-life during artemisinin monotherapy or artemisinin-based combination therapy. Absence of in-vitro and ex-vivo correlates of artemisinin resistance hinders study of this phenotype. We aimed to assess whether an in-vitro ring-stage survival assay (RSA) can identify culture-adapted P falciparum isolates from patients with slow-clearing or fast-clearing infections, to investigate the stage-dependent susceptibility of parasites to dihydroartemisinin in the in-vitro RSA, and to assess whether an ex-vivo RSA can identify artemisinin-resistant P falciparum infections.
METHODS
We culture-adapted parasites from patients with long and short parasite clearance half-lives from a study done in Pursat, Cambodia, in 2010 (registered with ClinicalTrials.gov, number NCT00341003) and used novel in-vitro survival assays to explore the stage-dependent susceptibility of slow-clearing and fast-clearing parasites to dihydroartemisinin. In 2012, we implemented the RSA in prospective parasite clearance studies in Pursat, Preah Vihear, and Ratanakiri, Cambodia (NCT01736319), to measure the ex-vivo responses of parasites from patients with malaria. Continuous variables were compared with the Mann-Whitney U test. Correlations were analysed with the Spearman correlation test.
FINDINGS
In-vitro survival rates of culture-adapted parasites from 13 slow-clearing and 13 fast-clearing infections differed significantly when assays were done on 0-3 h ring-stage parasites (10·88% vs 0·23%; p=0·007). Ex-vivo survival rates significantly correlated with in-vivo parasite clearance half-lives (n=30, r=0·74, 95% CI 0·50-0·87; p<0·0001).
INTERPRETATION
The in-vitro RSA of 0-3 h ring-stage parasites provides a platform for the molecular characterisation of artemisinin resistance. The ex-vivo RSA can be easily implemented where surveillance for artemisinin resistance is needed.
FUNDING
Institut Pasteur du Cambodge and the Intramural Research Program, NIAID, NIH.
背景
在青蒿素单一疗法或青蒿素联合疗法中，恶性疟原虫对青蒿素的耐药性会延长寄生虫清除半衰期。缺乏青蒿素耐药的体外和体内相关指标，阻碍了对这种表型的研究。我们旨在评估体外环早期生存检测（RSA）是否可以识别清除速度较慢或较快的感染患者中的培养适应恶性疟原虫分离株，研究体外 RSA 中对二氢青蒿素的寄生虫的阶段依赖性敏感性，并评估体外 RSA 是否可以识别青蒿素耐药恶性疟原虫感染。
方法
我们从 2010 年柬埔寨菩萨省的一项研究中培养了清除半衰期长和短的患者的寄生虫（在 ClinicalTrials.gov 注册，编号 NCT00341003），并使用新型体外生存检测来探索清除速度较慢和较快的寄生虫对二氢青蒿素的阶段依赖性敏感性。2012 年，我们在柬埔寨菩萨省、柏威夏省和腊塔纳基里省进行了前瞻性寄生虫清除研究中实施了 RSA（NCT01736319），以测量疟疾患者寄生虫的体外反应。连续变量采用曼-惠特尼 U 检验进行比较。采用 Spearman 相关检验分析相关性。
结果
当在 0-3 小时环早期寄生虫上进行检测时，来自 13 例清除速度较慢和 13 例清除速度较快的感染的培养适应寄生虫的体外存活率差异有统计学意义（10.88%比 0.23%；p=0.007）。体外存活率与体内寄生虫清除半衰期显著相关（n=30，r=0.74，95%CI 0.50-0.87；p<0.0001）。
结论
0-3 小时环早期寄生虫的体外 RSA 为青蒿素耐药性的分子特征提供了一个平台。在需要进行青蒿素耐药监测的地方，体外 RSA 可以很容易地实施。
资金
柬埔寨巴斯德研究所和美国国立卫生研究院 NIAID 内部研究计划。
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