输入性恶性疟原虫Pfcrt基因76位点多态性分析

Analysis of K76T Pfcrt genetic polymorphism in imported Plasmodium falciparum

目的了解输入性恶性疟原虫氯喹抗性转运蛋白基因(Pfcrt基因)K76T点突变发生情况。方法采集2008-2012年从非洲、东南亚等疟疾流行区回国人员恶性疟现症患者血样,根据恶性疟原虫Pfcrt基因序列设计巢式PCR引物,以血样中的恶性疟原虫DNA为模板,进行巢式PCR RFLP分析。结果92份输入性恶性疟现症患者血样中,突变型Pfcrt基因50份,占54.3%;野生型Pfcrt基因42份,占45.7%。采自非洲国家(尼日利亚、赤道几内亚、利比里亚等9国)回国人员的70份血样中,野生型Pfcrt基因37份,突变型Pfcrt基因33份,分别占52.9%和47.1%;采自东南亚国家(缅甸和印度尼西亚)回国人员的22份血样中,野生型Pfcrt基因5份,突变型Pfcrt基因17份,分别占22.7%和77.3%。不同地区回国人员Pfcrt基因突变发生率差异有统计学意义(χ2=6.12,P<0.05)。结论来自不同流行区的恶性疟原虫分离株Pfcrt基因突变发生率也不同,Pfcrt基因K76T位点突变检测在输入性恶性疟原虫氯喹抗性监测中具有应用价值。

Objective To identify the incidence of the K76T mutation in Pfcrt genes of imported Plasmodium falciparum. Methods The blood samples were collected from returnees infected with P. falciparum in endemic areas of Southeast Asia and Af- rica from 2008 to 2012. According to the Pfcrt gene sequence of P. falciparum, nested PCR primers were designed. Nested ＇PCR- RFLP was applied with falciparum DNA in the blood samples as templates. Results Among 92 blood samples of P. falciparum, the mutant Pfcrt alleles were found in 50 samples （54.3%）, and the wild type Pfcrt alleles were found in 42 samples （45.7%）. There were 33 samples （47.1%） with mutant Pfcrt alleles and 37 samples （52.9%） with wild type, respectively, from Africa. There were 17 samples （77.3%） with mutant Pfcrt alleles and 5 samples （22.7%） with wild type, respectively, from Southeast Asia. There was a significant difference between the Africa group and the Southeast Asia group （ x2 = 6.12, P 〈 0.05）. Conclu- sion The incidence of Pfcrt gene mutation is different among P. falciparum isolates from different regions. Therefore, Pfcrt K76T has an application value in the surveillance of the imported falciparum chloroquine- resistance.