利用大气发生器培养嗜二氧化碳菌 Genbag-CO2 来评估体外恶性疟原虫对标准抗疟药物的敏感性。
Use of the atmospheric generators for capnophilic bacteria Genbag-CO2 for the evaluation of in vitro Plasmodium falciparum susceptibility to standard anti-malarial drugs.
机构信息
Unité de Recherche en Biologie et Epidémiologie Parasitaires, Institut de Recherche Biomédicale des Armées--antenne de Marseille, Marseille, France.
出版信息
Malar J. 2011 Jan 14;10:8. doi: 10.1186/1475-2875-10-8.
BACKGROUND
The aim of this study was to evaluate the cultivation system in which the proper atmospheric conditions for growing Plasmodium falciparum parasites were maintained in a sealed bag. The Genbag® system associated with the atmospheric generators for capnophilic bacteria Genbag CO2® was used for in vitro susceptibility test of nine standard anti-malarial drugs and compared to standard incubator conditions.
METHODS
The susceptibility of 36 pre-identified parasite strains from a wide panel of countries was assessed for nine standard anti-malarial drugs (chloroquine, quinine, mefloquine, monodesethylamodiaquine, lumefantrine, dihydroartemisinin, atovaquone and pyrimethamine) by the standard 42-hour 3H-hypoxanthine uptake inhibition method using the Genbag CO2® system and compared to controlled incubator conditions (5% CO2 and 10% O2).
RESULTS
The counts per minute values in the control wells in incubator atmospheric conditions (5% CO2 and 10% O2) were significantly higher than those of Genbag® conditions (2738 cpm vs 2282 cpm, p < 0.0001). The geometric mean IC50 estimated under the incubator atmospheric conditions was significantly lower for atovaquone (1.2 vs 2.1 nM, p = 0.0011) and higher for the quinolines: chloroquine (127 vs 94 nM, p < 0.0001), quinine (580 vs 439 nM, p < 0.0001), monodesethylamodiaquine (41.4 vs 31.8 nM, p < 0.0001), mefloquine (57.5 vs 49.7 nM, p = 0.0011) and lumefantrine (23.8 vs 21.2 nM, p = 0.0044). There was no significant difference of IC50 between the 2 conditions for dihydroartemisinin, doxycycline and pyrimethamine.To reduce this difference in term of anti-malarial susceptibility, a specific cut-off was estimated for each drug under Genbag® conditions by regression. The cut-off was estimated at 77 nM for chloroquine (vs 100 nM in 10% O2), 611 nM for quinine (vs 800 nM), 30 nM for mefloquine (vs 30 nM), 61 nM for monodesethylamodiaquine (vs 80 nM) and 1729 nM for pyrimethamine (vs 2000 nM).
CONCLUSIONS
The atmospheric generators for capnophilic bacteria Genbag CO2® is an appropriate technology that can be transferred to the field for epidemiological surveys of drug-resistant malaria. The present data suggest the importance of the gas mixture on in vitro microtest results for anti-malarial drugs and the importance of determining the microtest conditions before comparing and analysing the data from different laboratories and concluding on malaria resistance.
背景
本研究旨在评估在密封袋中维持恶性疟原虫生长适宜大气条件的培养系统。Genbag®系统与嗜二氧化碳细菌 Genbag CO2®大气发生器联合用于 9 种标准抗疟药物的体外药敏试验，并与标准孵育箱条件进行比较。
方法
使用 Genbag CO2®系统对来自广泛国家的 36 株预先鉴定的寄生虫株进行 9 种标准抗疟药物（氯喹、奎宁、甲氟喹、去乙基阿莫地喹、青蒿琥酯、二氢青蒿素、阿托伐醌和乙胺嘧啶）的药敏试验，并用标准 42 小时 3H-次黄嘌呤摄取抑制法进行比较，并与控制孵育箱条件（5%CO2和 10%O2）进行比较。
结果
孵育箱大气条件（5%CO2和 10%O2）对照孔每分钟计数（cpm）值明显高于 Genbag®条件（2738 cpm 对 2282 cpm，p<0.0001）。在孵育箱大气条件下估计的几何平均 IC50 值对于阿托伐醌明显较低（1.2 对 2.1 nM，p=0.0011），而对于喹啉类药物：氯喹（127 对 94 nM，p<0.0001）、奎宁（580 对 439 nM，p<0.0001）、去乙基阿莫地喹（41.4 对 31.8 nM，p<0.0001）、甲氟喹（57.5 对 49.7 nM，p=0.0011）和青蒿琥酯（23.8 对 21.2 nM，p=0.0044）较高。二氢青蒿素、多西环素和乙胺嘧啶在两种条件下的 IC50 无显著差异。为了降低抗疟药物敏感性方面的这种差异，通过回归估计了 Genbag®条件下每种药物的特定截止值。氯喹的截止值估计为 77 nM（10%O2 时为 100 nM），奎宁为 611 nM（800 nM），甲氟喹为 30 nM（30 nM），去乙基阿莫地喹为 61 nM（80 nM），乙胺嘧啶为 1729 nM（2000 nM）。
结论
嗜二氧化碳细菌 Genbag CO2®大气发生器是一种合适的技术，可以转移到现场进行抗疟药物耐药性的流行病学调查。本数据表明，气体混合物对体外药敏试验结果的重要性，以及在比较和分析来自不同实验室的数据并得出疟疾耐药性结论之前，确定药敏试验条件的重要性。
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