Artemisinin resistance in Plasmodium falciparum is associated with an altered temporal pattern of transcription (CGH)

Artemisinin resistance in Plasmodium falciparum malaria has emerged in western Cambodia. This is a major threat to global plans to control and eliminate malaria as the artemisinins are a key component of antimalarial treatment throughout the world. Using DNA microarrays we identify key features of a transcriptional profile that are associated with the delayed parasite clearance phenotype. These include reduced expression of several basic metabolic and cellular pathways in the early stages, and increased expression of essentially all functionalities associated with protein metabolism in the later stages of P. falciparum intraerythrocytic development. This is consistent with the reduced ring stage susceptibility that characterizes artemisinin resistant P. falciparum. This modulation of the P. falciparum intraerythrocytic transcriptome may result from differential expression of several regulatory proteins such as transcription factors of chromatin remodeling associated factors. In addition, the artemisinin resistant phenotype is strongly associated with a specific pattern of copy number variations, some of which are linked with differential expression of several regulatory proteins such as histone 4 and zinc permease. This study reports the first global transcriptional survey of artemisinin resistant parasites and provides a set of candidate genes for further investigation. 6 P. falciparum parasites (field isolates) which are either Artemsinin resistant or sensitive from 3 study sites (Pailin in Cambodia, Xepon in Laos, Mae Sot in Thailand) were sampled and harvested for genomic DNA. gDNA from a total of 6 samples were extracted by phenol chloroform. Synthesis of labelled target DNA was carried out as previously described: Mackinnon, M.J. et al. Comparative transcriptional and genomic analysis of Plasmodium falciparum field isolates. PLoS Pathog 5, e1000644 (2009), and used in comparative genomic microarray hybridizations (CGH).

恶性疟原虫疟疾中的青蒿素（artemisinin）抗性已在柬埔寨西部出现。这对全球疟疾防控与消除计划构成重大威胁，因为青蒿素类药物是全球抗疟治疗的核心组成部分。本研究借助DNA微阵列（DNA microarray）技术，鉴定出与恶性疟原虫延迟清除表型相关的转录组特征谱关键特征。这些特征包括：在恶性疟原虫红细胞内发育早期，多条基础代谢与细胞通路的表达水平下调；而在发育后期，几乎所有与蛋白质代谢相关的功能通路表达均上调。该结果与青蒿素抗性恶性疟原虫的典型特征——环状体阶段易感性降低——相符。恶性疟原虫红细胞内转录组的这种调控模式，可能源于多种调控蛋白的差异表达，例如染色质重塑相关因子的转录因子。此外，青蒿素抗性表型与特定的拷贝数变异模式显著相关，其中部分变异与组蛋白H4（histone 4）、锌通透酶（zinc permease）等调控蛋白的差异表达存在关联。本研究首次完成了青蒿素抗性疟原虫的全球转录组分析，并为后续研究提供了一组候选基因。本研究从柬埔寨拜林、老挝谢奔、泰国湄索3个研究地点采集了6株恶性疟原虫野外分离株，其中部分为青蒿素抗性株，部分为敏感株，用于基因组DNA（gDNA）的提取。总计6份样本的基因组DNA均通过酚-氯仿法提取。标记靶DNA的合成按照此前发表的方法进行：Mackinnon MJ等人于2009年发表在《PLOS Pathogens》第5卷e1000644的《恶性疟原虫野外分离株的转录组与基因组比较分析》一文，合成的标记靶DNA将用于比较基因组微阵列杂交（CGH）实验。