Huang Fang, Yan He, Xue Jing-Bo, Cui Yan-Wen, Zhou Shui-Sen, Xia Zhi-Gui, Abeyasinghe Rabindra, Ringwald Pascal, Zhou Xiao-Nong
National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention, Chinese Center for Tropical Diseases Research, WHO Collaborating Center for Tropical Diseases, National Centre for International Research On Tropical Diseases, Ministry of Science and Technology, Key Laboratory of Parasite and Vector Biology, Ministry of Health, Shanghai, China.
World Health Organization Country Office in Philippines, Manila, Philippines.
Malar J. 2021 Feb 6;20(1):73. doi: 10.1186/s12936-021-03613-5.
The emergence and spread of multidrug resistance poses a significant risk to malaria control and eradication goals in the world. There has been no indigenous malaria cases reported in China since 2017, and China is approaching national malaria elimination. Therefore, anti-malarial drug resistance surveillance and tracking the emergence and spread of imported drug-resistant malaria cases will be necessary in a post-elimination phase in China.
Dried blood spots were obtained from Plasmodium falciparum-infected cases returned from Africa to China between 2012 and 2015, prior to anti-malarial drug treatment. Whole DNA were extracted and known polymorphisms relating to drug resistance of pfcrt, pfmdr1 gene, and the propeller domain of pfk13 were evaluated by nested PCR and sequencing. The haplotypes and prevalence of these three genes were evaluated separately. Chi-squared test and Fisher's exact test were used to evaluate differences among the different sub-regions of Africa. A P value < 0.05 was used to evaluate differences with statistical significance. The maps were created using ArcGIS.
A total of 731 P. falciparum isolates were sequenced at the pfcrt locus. The wild type CVMNK was the most prevalent haplotype with prevalence of 62.8% and 29.8% of the isolates showed the triple mutant haplotype CVIET. A total of 434 P. falciparum isolates were successfully sequenced and pfmdr1 allelic variants were observed in only codons 86, 184 and 1246. Twelve haplotypes were identified and the prevalence of the wild type pfmdr1 NYD was 44.1%. The single mutant pfmdr1 in codons 86 and 184 was predominant but the haplotype NYY with single mutation in codon 1246 was not observed. The double mutant haplotype YFD was common in Africa. About 1,357 isolates were successfully sequenced of pfk13-propeller domain, the wild type was found in 1,308 samples (96.4%) whereby 49 samples (3.6%) had mutation in pfk13. Of 49 samples with pfk13 mutations, 22 non-synonymous and 4 synonymous polymorphic sites were confirmed. The A578S was the most common mutation in pfk13-propeller domain and three mutations associated with artemisinin resistance (M476I, R539T, P553L) were identified in three isolates.
This study provides evidence that could give insight into potential issues with anti-malarial drug resistance to inform national drug policy in China in order to treat imported cases.
多重耐药性的出现和传播对全球疟疾控制和消除目标构成重大风险。自2017年以来，中国未报告本土疟疾病例，中国正接近全国消除疟疾目标。因此，在中国消除疟疾后的阶段，开展抗疟药物耐药性监测以及追踪输入性耐药疟疾病例的出现和传播将很有必要。
采集2012年至2015年间从非洲返回中国且在接受抗疟药物治疗前感染恶性疟原虫病例的干血斑。提取全基因组DNA，通过巢式PCR和测序评估与pfcrt、pfmdr1基因耐药性以及pfk13螺旋桨结构域相关的已知多态性。分别评估这三个基因的单倍型和流行率。采用卡方检验和Fisher精确检验评估非洲不同次区域之间的差异。P值<0.05用于评估具有统计学意义的差异。使用ArcGIS绘制地图。
共对731株恶性疟原虫分离株的pfcrt基因座进行了测序。野生型CVMNK是最常见的单倍型，流行率为62.8%，29.8%的分离株显示三重突变单倍型CVIET。共成功测序434株恶性疟原虫分离株，仅在第86、184和1246密码子观察到pfmdr1等位基因变体。鉴定出12种单倍型，野生型pfmdr1 NYD的流行率为44.1%。第86和184密码子的单突变pfmdr1占主导，但未观察到第1246密码子单突变的单倍型NYY。双突变单倍型YFD在非洲很常见。成功对约1357株pfk13螺旋桨结构域进行了测序，1308个样本（96.4%）中发现野生型，49个样本（3.6%）的pfk13发生突变。在49个具有pfk13突变的样本中，确认了22个非同义多态性位点和4个同义多态性位点。A578S是pfk13螺旋桨结构域最常见的突变，在三个分离株中鉴定出与青蒿素耐药性相关的三个突变（M476I、R539T、P553L）。
本研究提供的证据有助于深入了解抗疟药物耐药性的潜在问题，为中国治疗输入性病例的国家药物政策提供参考。
