Malaria Branch, Division of Parasitic Diseases and Malaria, Center for Global Health, Centers for Disease Control and Prevention, Atlanta, Georgia, USA
School of Biology, Georgia Institute of Technology, Atlanta, Georgia, USA.
Antimicrob Agents Chemother. 2018 Mar 27;62(4). doi: 10.1128/AAC.02474-17. Print 2018 Apr.
The recent advances in next-generation sequencing technologies provide a new and effective way of tracking malaria drug-resistant parasites. To take advantage of this technology, an end-to-end Illumina targeted amplicon deep sequencing (TADS) and bioinformatics pipeline for molecular surveillance of drug resistance in , called laria esistance urveillance (MaRS), was developed. TADS relies on PCR enriching genomic regions, specifically target genes of interest, prior to deep sequencing. MaRS enables researchers to simultaneously collect data on allele frequencies of multiple full-length drug resistance genes (, , , , , and the cytochrome gene), as well as the mitochondrial genome. Information is captured at the individual patient level for both known and potential new single nucleotide polymorphisms associated with drug resistance. The MaRS pipeline was validated using 245 imported malaria cases that were reported to the Centers for Disease Control and Prevention (CDC). The chloroquine resistance CV genotype (mutations underlined) was observed in 42% of samples, the highly pyrimethamine-resistant triple mutant in 92% of samples, and the sulfadoxine resistance mutation SAA in 26% of samples. The NSND genotype was found in 40% of samples. With the exception of two cases imported from Cambodia, no artemisinin resistance alleles were identified, and 99% of patients carried parasites susceptible to atovaquone-proguanil. Our goal is to implement MaRS at the CDC for routine surveillance of imported malaria cases in the United States and to aid in the adoption of this system at participating state public health laboratories, as well as by global partners.
新一代测序技术的最新进展为追踪抗疟药物耐药寄生虫提供了一种新的有效方法。为了利用这项技术，我们开发了一种端到端的 Illumina 靶向扩增子深度测序（TADS）和用于药物抗性分子监测的生物信息学管道，称为拉氏抗性监测（MaRS）。TADS 依赖于 PCR 对基因组区域进行富集，特别是对感兴趣的靶基因进行富集，然后进行深度测序。MaRS 使研究人员能够同时收集多个全长药物抗性基因（ 、 、 、 、 和细胞色素基因）以及线粒体基因组的等位基因频率数据。在个体患者层面上，该系统可以捕获与药物抗性相关的已知和潜在新单核苷酸多态性的信息。该 MaRS 管道使用向疾病预防控制中心（CDC）报告的 245 例输入性疟疾病例进行了验证。观察到 42%的样本中存在氯喹抗性 CV 基因型（下划线突变），92%的样本中存在高度耐嘧啶的 三重突变体，26%的样本中存在磺胺多辛抗性 突变 SAA。40%的样本中发现了 NSND 基因型。除了从柬埔寨输入的两例病例外，未发现青蒿素耐药 等位基因，99%的患者携带对阿托伐醌-丙磺舒敏感的寄生虫。我们的目标是在美国疾病预防控制中心实施 MaRS 以对输入性疟疾病例进行常规监测，并帮助参与的州公共卫生实验室以及全球合作伙伴采用该系统。
