Division of Cell and Molecular Biology, Imperial College London, London, UK.
Malar J. 2010 Jan 29;9:35. doi: 10.1186/1475-2875-9-35.
Malaria transmission is now recognized as a key target for intervention. Evaluation of the Plasmodium oocyst burden in the midguts of Anopheles spp. is important for many of assays investigating transmission. However, current assays are very time-consuming, manually demanding and patently subject to observer-observer variation.
This report presents the development of a method to rapidly, accurately and consistently determine oocyst burdens on mosquito midguts using GFP-expressing Plasmodium berghei and a custom-written macro for ImageJ. The counting macro was optimized and found to be fit-for-purpose by performing gametocyte membrane feeds with parasite infected blood. Dissected midguts were counted both manually and using the automated macro, then compared. The optimized settings for the macro were then validated by using it to determine the transmission blocking efficacies of two anti-malarial compounds - dehydroepiandrosterone sulphate and lumefantrine, in comparison to manually determined analysis of the same experiment.
Concurrence of manual and macro counts was very high (R2 = 0.973) and reproducible. Estimated transmission blocking efficacies between manual and automated analysis were highly concordant, indicating that dehydroepiandrosterone sulphate has little or no transmission blocking potential, whilst lumefantrine strongly inhibits sporogony.
Recognizing a potential five-fold increase in throughput, the resulting reduction in personnel costs, and the absence of inter-operator/laboratory variation possible with this approach, this counting macro may be a benefit to the malaria community.
疟疾传播现已被认为是干预的关键目标。评估按蚊中疟原虫卵囊负担对于许多研究传播的检测非常重要。然而，目前的检测方法非常耗时，对人力要求高，明显存在观察者间的差异。
本报告介绍了一种使用表达 GFP 的恶性疟原虫孢子虫和 ImageJ 编写的自定义宏来快速、准确和一致地确定蚊子中肠卵囊负担的方法。通过用感染寄生虫的血液进行配子体膜喂养来优化计数宏，并发现其适用于目的。手动和使用自动化宏对解剖的中肠进行计数，然后进行比较。然后，使用该宏来确定两种抗疟化合物 - 硫酸脱氢表雄酮和盐酸左旋咪唑的阻断传播效率，并与手动分析相同实验进行比较，从而验证宏的优化设置。
手动计数和宏计数的一致性非常高（R2 = 0.973）且可重现。手动和自动分析估计的传播阻断效率高度一致，表明硫酸脱氢表雄酮几乎没有或没有阻断传播的潜力，而盐酸左旋咪唑强烈抑制孢子生殖。
考虑到通量可能增加五倍，人员成本降低，并且这种方法不存在操作员/实验室之间的差异，这种计数宏可能对疟疾社区有益。
Zotero 插件
