Department of International Affairs and Tropical Medicine, Tokyo Women's Medical University School of Medicine, 8-1 Kawada-cho, Shinjuku-ku, Tokyo 162-8666, Japan.
Malar J. 2012 Mar 28;11:92. doi: 10.1186/1475-2875-11-92.
In Plasmodium falciparum, resistance to chloroquine (CQ) is conferred by a K to T mutation at amino acid position 76 (K76T) in the P. falciparum CQ transporter (PfCRT). To date, at least 15 pfcrt genotypes, which are represented by combinations of five amino acids at positions 72-76, have been described in field isolates from various endemic regions. To identify novel mutant pfcrt genotypes and to reveal the genetic relatedness of pfcrt genotypes, a large-scale survey over a wide geographic area was performed.
Sequences for exon 2 in pfcrt, including known polymorphic sites at amino acid positions 72, 74, 75 and 76, were obtained from 256 P. falciparum isolates collected from eight endemic countries in Asia (Bangladesh, Cambodia, Lao P.D.R., the Philippines and Thailand), Melanesia (Papua New Guinea and Vanuatu) and Africa (Ghana). A haplotype network was constructed based on six microsatellite markers located -29 kb to 24 kb from pfcrt in order to examine the genetic relatedness among mutant pfcrt genotypes.
In addition to wild type (CVMNK at positions 72-76), four mutant pfcrt were identified; CVIET, CVIDT, SVMNT and CVMNT (mutated amino acids underlined). Haplotype network revealed that there were only three mutant pfcrt lineages, originating in Indochina, Philippines and Melanesia. Importantly, the Indochina lineage contained two mutant pfcrt genotypes, CVIET (n = 95) and CVIDT (n = 14), indicating that CVIDT shares a common origin with CVIET. Similarly, one major haplotype in the Melanesian lineage contained two pfcrt genotypes; SVMNT (n = 71) and CVMNT (n = 3). In Africa, all mutant pfcrt genotypes were the CVIET of the Indochina lineage, probably resulting from the intercontinental migration of CQ resistance from Southeast Asia.
The number of CQ-mutant lineages observed in this study was identical to that found in previous studies. This supports the hypothesis that the emergence of novel CQ resistance is rare. However, in the mutant pfcrt genotypes, amino acid changes at positions 72, 74 and 75 appear to have recently been generated at least several times, producing distinct pfcrt mutant genotypes. The occurrence of new mutations flanking K76T may yield stronger resistance to CQ and/or a higher fitness than the original pfcrt mutant.
在恶性疟原虫中，对氯喹（CQ）的耐药性是由疟原虫 CQ 转运蛋白（PfCRT）中氨基酸位置 76 的 K 突变为 T（K76T）引起的。迄今为止，已在来自不同流行地区的现场分离株中描述了至少 15 种 pfcrt 基因型，这些基因型由位置 72-76 的五个氨基酸的组合表示。为了鉴定新的突变 pfcrt 基因型并揭示 pfcrt 基因型的遗传相关性，在广泛的地理区域内进行了大规模调查。
从来自亚洲（孟加拉国、柬埔寨、老挝人民民主共和国、菲律宾和泰国）、美拉尼西亚（巴布亚新几内亚和瓦努阿图）和非洲（加纳）的 8 个流行地区采集的 256 株恶性疟原虫分离株中获得 pfcrt 外显子 2 的序列，包括氨基酸位置 72、74、75 和 76 的已知多态性位点。构建了一个基于位于 pfcrt 上游-29 kb 至 24 kb 处的六个微卫星标记的单倍型网络，以检查突变 pfcrt 基因型之间的遗传相关性。
除了野生型（72-76 位的 CVMNK）外，还鉴定出了四种突变 pfcrt；CVIET、CVIDT、SVMNT 和 CVMNT（带下划线的突变氨基酸）。单倍型网络显示，只有三种突变 pfcrt 谱系起源于印度支那、菲律宾和美拉尼西亚。重要的是，印度支那谱系包含两种突变 pfcrt 基因型，CVIET（n = 95）和 CVIDT（n = 14），表明 CVIDT 与 CVIET 具有共同的起源。同样，美拉尼西亚谱系中的一个主要单倍型包含两种 pfcrt 基因型；SVMNT（n = 71）和 CVMNT（n = 3）。在非洲，所有的突变 pfcrt 基因型都是印度支那谱系的 CVIET，这可能是由于东南亚 CQ 耐药性的洲际迁移。
本研究中观察到的 CQ 突变谱系数量与之前的研究相同。这支持了新的 CQ 耐药性出现很少的假设。然而，在突变 pfcrt 基因型中，位置 72、74 和 75 的氨基酸变化似乎最近至少发生了几次，产生了不同的 pfcrt 突变基因型。K76T 侧翼新突变的发生可能比原始 pfcrt 突变赋予更强的 CQ 耐药性和/或更高的适应性。
