Regional Medical Research Center, Indian Council of Medical Research, Bhubaneswar, Odisha, India.
Mem Inst Oswaldo Cruz. 2011 Feb;106(1):78-84. doi: 10.1590/s0074-02762011000100013.
The global emergence and spread of malaria parasites resistant to antimalarial drugs is the major problem in malaria control. The genetic basis of the parasite's resistance to the antimalarial drug chloroquine (CQ) is well-documented, allowing for the analysis of field isolates of malaria parasites to address evolutionary questions concerning the origin and spread of CQ-resistance. Here, we present DNA sequence analyses of both the second exon of the Plasmodium falciparum CQ-resistance transporter (pfcrt) gene and the 5' end of the P. falciparum multidrug-resistance 1 (pfmdr-1) gene in 40 P. falciparum field isolates collected from eight different localities of Odisha, India. First, we genotyped the samples for the pfcrt K76T and pfmdr-1 N86Y mutations in these two genes, which are the mutations primarily implicated in CQ-resistance. We further analyzed amino acid changes in codons 72-76 of the pfcrt haplotypes. Interestingly, both the K76T and N86Y mutations were found to co-exist in 32 out of the total 40 isolates, which were of either the CVIET or SVMNT haplotype, while the remaining eight isolates were of the CVMNK haplotype. In total, eight nonsynonymous single nucleotide polymorphisms (SNPs) were observed, six in the pfcrt gene and two in the pfmdr-1 gene. One poorly studied SNP in the pfcrt gene (A97T) was found at a high frequency in many P. falciparum samples. Using population genetics to analyze these two gene fragments, we revealed comparatively higher nucleotide diversity in the pfcrt gene than in the pfmdr-1 gene. Furthermore, linkage disequilibrium was found to be tight between closely spaced SNPs of the pfcrt gene. Finally, both the pfcrt and the pfmdr-1 genes were found to evolve under the standard neutral model of molecular evolution.
疟原虫对抗疟药物产生耐药性是全球疟疾控制面临的主要问题。疟原虫对青蒿素类药物氯喹（CQ）耐药的遗传基础已有充分的研究，这使得对疟原虫野外分离株的分析能够解决与 CQ 耐药性起源和传播有关的进化问题。在此，我们对来自印度奥里萨邦 8 个不同地区的 40 株恶性疟原虫分离株的疟原虫 CQ 耐药转运蛋白（pfcrt）基因第二外显子和疟原虫多药耐药 1（pfmdr-1）基因 5'端的 DNA 序列进行了分析。首先，我们对这两个基因中的 pfcrt K76T 和 pfmdr-1 N86Y 突变进行了基因分型，这两个突变是与 CQ 耐药性主要相关的突变。我们进一步分析了 pfcrt 基因 72-76 密码子中的氨基酸变化。有趣的是，在总共 40 个分离株中，有 32 个分离株同时存在 K76T 和 N86Y 突变，这些分离株要么是 CVIET 或 SVMNT 单倍型，而其余 8 个分离株是 CVMNK 单倍型。总共观察到 8 个非同义单核苷酸多态性（SNP），其中 6 个在 pfcrt 基因中，2 个在 pfmdr-1 基因中。在 pfcrt 基因中发现了一个研究较少的 SNP（A97T），在许多恶性疟原虫样本中高频出现。使用群体遗传学方法对这两个基因片段进行分析，我们发现 pfcrt 基因的核苷酸多样性明显高于 pfmdr-1 基因。此外，pfcrt 基因紧密间隔的 SNP 之间存在紧密的连锁不平衡。最后，pfcrt 和 pfmdr-1 基因均被发现遵循分子进化的标准中性模型。
