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A genomic epidemiological research on a drug resistant malaria spreading in Asian countries.

关于在亚洲国家传播的耐药疟疾的基因组流行病学研究。

基本信息

批准号：
16406012
负责人：
KANO Shigeyuki
金额：
$ 8.51万
依托单位：
International Medical Centre of Japan
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2006
项目状态：
已结题

项目摘要

A mutation from Lys to Thr in the codon 76 of Plasmodium falciparum chloroquine resistant transporter protein (PfCRT) is associated with CQ resistance, and the genotype is used as a molecular marker to monitor the distribution and frequency of the CQ resistant malaria. In the present study, selection of the resistant strain in mixed cultures of a drug resistant strain (K1 which shows K76T) and a susceptible strain (FCR-3 which shows 76K) under a low chloroquine pressure (<80nM) was nicely observed in vivo using real-time PCR analysis.Then, we analyzed the frequency of the mutation in codon 72-76 of the pfcrt gene and 3 microsatellite (MS) DNA markers flanking the pfcrt, using P. falciparum isolates from the Philippines, Thailand, and southern part of Vietnam. Thirty-nine isolates from Thailand showed identical K76T but 6 out of 13 isolates from the Philippines showed mixed type. Twenty-seven of 39 isolates from Vietnam showed CQ susceptible genotype coding CVMNK, while other 10 did CQ resistance type (CVIET, CVIDT, CVMDT). The other 2 isolates were proved to be mixed with CQ susceptible/resistant.In the further analysis of the Vietnam isolates, the MS DNA markers of the CQ susceptible isolates were revealed to be highly polymorphic, while those of CQ resistant isolates were less polymorphic. One of the CQ resistant isolates (CVIET) had a different MS DNA pattern from K1 (CQ resistant strain; CVIET) from Thailand. The other CQ resistant isolate types (CVIDT, CVMDT) shared the same MS DNA patterns and these patterns were different from those of the CQ susceptible isolates nor K1. These results showed that the origin of the CQ resistant isolates in Vietnam might not be due to an introduction of the isolates from Thailand. Instead, it is suggested that the CQ resistant mutations possibly occurred within Vietnam.

恶性疟原虫氯喹抗性转运蛋白（PfCRT）第76位密码子Lys突变为Thr与CQ抗性相关，该基因型可作为监测CQ抗性疟疾分布和频率的分子标记。在本研究中，使用实时PCR分析在体内很好地观察了在低氯喹压力（<80nM）下耐药菌株（K1显示K76T）和敏感菌株（FCR-3显示76K）的混合培养物中耐药菌株的选择。然后，我们分析了pfcrt基因的密码子72-76和3个微卫星（MS）的突变频率 pfcrt 两侧的 DNA 标记，使用来自菲律宾、泰国和越南南部的恶性疟原虫分离株。来自泰国的 39 个分离株显示出相同的 K76T，但来自菲律宾的 13 个分离株中有 6 个显示出混合型。来自越南的 39 株分离株中有 27 株表现出编码 CVMNK 的 CQ 易感基因型，而另外 10 株则表现出 CQ 耐药型（CVIET、CVIDT、CVMDT）。另外2株被证明与CQ敏感/耐药混合。在对越南分离株的进一步分析中，发现CQ敏感分离株的MS DNA标记具有高度多态性，而CQ抗性分离株的MS DNA标记多态性较低。其中一种 CQ 抗性分离株 (CVIET) 与来自泰国的 K1（CQ 抗性菌株；CVIET）具有不同的 MS DNA 模式。其他 CQ 抗性分离株类型（CVIDT、CVMDT）具有相同的 MS DNA 模式，这些模式与 CQ 易感分离株和 K1 不同。这些结果表明，越南 CQ 抗性分离株的起源可能不是由于从泰国引入分离株所致。相反，有人认为 CQ 抗性突变可能发生在越南境内。