Aly Ahmed S I, Deveci Gozde, Yilmaz Ilknur, Abraham Amanah, Golshan Aneesa, Hart Robert J
Beykoz Institute of Life Sciences and Biotechnology, Bezmialem Vakif University, Istanbul, Turkey; School of Public Health and Tropical Medicine, Department of Tropical Medicine, Tulane University;
Beykoz Institute of Life Sciences and Biotechnology, Bezmialem Vakif University, Istanbul, Turkey.
J Vis Exp. 2019 May 30(147). doi: 10.3791/55688.
Recent advances in genetics and systems biology technologies have promoted our understanding of the biology of malaria parasites on the molecular level. However, effective malaria parasite targets for vaccine and chemotherapy development are still limited. This is largely due to the unavailability of relevant and practical in vivo infection models for human Plasmodium species, most notably for P. falciparum and P. vivax. Therefore, rodent malaria species have been extensively used as practical alternative in vivo models for malaria vaccine, drug targeting, immune response, and functional characterization studies of conserved Plasmodiumspp. genes. Indeed, rodent malaria models have proven to be invaluable, especially for exploring mosquito transmission and liver stage biology, and were indispensable for immunological studies. However, there are discrepancies in the methods used to evaluate the phenotypes of transgenic and wild-type asexual and sexual blood-stage parasites. Examples of these discrepancies are the choice of an intravenous vs. intraperitoneal infection of rodents with blood-stage parasites and the evaluation of male gamete exflagellation. Herein, we detail standardized experimental methods to evaluate the phenotypes of asexual and sexual blood stages in transgenic parasites expressing reporter-gene or wild-type rodent malaria parasite species. We also detail the methods to evaluate the phenotypes of malaria parasite mosquito stages (gametes, ookinetes, oocysts, and sporozoites) inside Anopheles mosquito vectors. These methods are detailed and simplified here for the lethal and non-lethal strains of P. berghei and P. yoelii but can also be applied with some adjustments to P. chabaudi and P. vinckei rodent malaria species.
遗传学和系统生物学技术的最新进展促进了我们在分子水平上对疟原虫生物学的理解。然而，用于疫苗和化疗开发的有效疟原虫靶点仍然有限。这主要是由于缺乏针对人类疟原虫物种，尤其是恶性疟原虫和间日疟原虫的相关且实用的体内感染模型。因此，啮齿动物疟原虫物种已被广泛用作疟原虫疫苗、药物靶向、免疫反应以及保守疟原虫属基因功能表征研究的实用替代体内模型。事实上，啮齿动物疟原虫模型已被证明具有极高价值，特别是在探索蚊虫传播和肝脏期生物学方面，并且对于免疫学研究不可或缺。然而，用于评估转基因和野生型无性及有性血液期疟原虫表型的方法存在差异。这些差异的例子包括用血液期疟原虫对啮齿动物进行静脉注射与腹腔注射感染的选择以及雄配子体出丝现象的评估。在此，我们详细介绍了用于评估表达报告基因的转基因寄生虫或野生型啮齿动物疟原虫物种无性和有性血液期表型的标准化实验方法。我们还详细介绍了评估按蚊媒介体内疟原虫蚊虫期（配子、动合子、卵囊和子孢子）表型的方法。这里针对伯氏疟原虫和约氏疟原虫的致死和非致死菌株详细且简化了这些方法，但经过一些调整后也可应用于查巴迪疟原虫和温氏疟原虫啮齿动物疟原虫物种。
