Abstract: Objective Objective To analyze the polymorphism of histidine rich protein 2（HRP II）gene in Plasmodium falciparum （Pfhrp2）from falciparum malaria patients in Yunnan Province，so as to lay the foundation for studying the defection of antigen genes of Plasmodium. Methods Methods The filter paper blood samples and related information of falciparum malaria cases reported were obtained in Yunnan Province from August 2012 to September 2015. Under the guidance of the specific primers，the exon2 regions in Pfhrp2 gene in P. falciparum from DNA samples were amplified by PCR，and the PCR products were sequenced. The sequences of exon2 region in Pfhrp2 gene were blasted by comparing with the reference sequences AY816237，AY816240，and AY816301. Next，the polymorphism of the sequence in exon2 region of Pfhrp2 gene was analyzed by MEGA 5.04 software. The conserved sites and genetic distances between sequences were calculated by using the software as well，and the clustering tree was drawn according to the genetic distances between the amino acid sequences. Results Results A total of 218 bloods samples from the falciparum malaria cases in 15 prefectures of Yunnan Province were collected，and the sources of infection included Yun? nan，Africa and Myanmar. The PCR results showed that the exon2 regions in Pfhrp2 genes of 155 samples were positive by am? plification and their products were sequenced successfully. The sequence analysis showed that the length range of the amino acid residues of exon2 region in Pfhrp2 gene was from 115 aa to 298 aa，the average length was 239.7 aa. There was no statistically significance among the means of the amino acid residues of the isolates from Africa（239.9 aa），Myanmar （239.5 aa）and Yun? nan（241.6 aa） （F = 0.025，P > 0.05） . All the 155 amino acid sequences ended with type 12 repeat，98.1%（152/155）of them started with type 1 repeat and 1.9%（3/155）of them started with type 2. Type 2 presented most frequently repeat in all the se? quences and the average repeat times were 12.9. The homologous locus of the DNA sequences in exon2 regions of the 155 Pfhrp2 genes was 894 bp，among which the conservative sites accounted for 20.6%（186/894），and the variable sites for 78.2%（699/ 894） . The genetic distances between the sequences of Africa isolates ranged from 0 to 0.741，and those of the Myanmar and Yun? nan isolates were 0-0.948 and 0-0.750，respectively. The cluster analysis showed that all the 155 sequences clustered into 3 cat? egories on genetic distances between amino acid sequences according to the size of the amino acid sequence length. At the same level，the sequences had approximate lengths and amino acid repeat types. Conclusion Conclusion The sequence of exon2 region in Pfhrp2 gene of P. falciparum from falciparum malaria cases in Yunnan Province is highly polymorphic，the P. falciparum iso? lates are clustered mainly according to the size of the amino acid sequence of exon2 region in Pfhrp2 gene.

Key words: Falciparum malaria；Plasmodium falciparum；Histidine ? rich protein Ⅱ（HRP II）；Pfhrp2 gene；Polymor? phism；Yunnan Province

摘要： 目的 目的 分析云南省恶性疟病例疟原虫虫株的富组氨酸蛋白Ⅱ （Histidine?rich protein?2，HRPⅡ） 基因 （Pfhrp2） 序 列的多态性， 为研究疟原虫抗原基因缺失奠定基础。方法 方法 搜集2012年8月-2015年9月云南省恶性疟现症病例的滤纸 血样和相关信息， 用PCR技术扩增血样中恶性疟原虫Pfhrp2基因exon2区并测序， 测序成功序列与AY816237、 AY816240、 AY816301参比序列比对； 用Mega 5.04分析Pfhrp2基因exon2区序列多态性， 计算序列间保守位点、 遗传距离 等； 根据氨基酸序列间遗传距离绘制聚类树状图。结果 结果 共收集云南省15个州 （市） 的恶性疟现症病例血样218份， 病例 感染来源地包括云南本地、 非洲、 缅甸等流行区。其中155份Pfhrp2基因exon2区扩增阳性和测序成功， 编码氨基酸数在 115～298之间， 平均为239.7 aa， 非洲 （239.9 aa）、 缅甸 （239.5 aa）、 云南 （241.6 aa） 感染虫株的氨基酸残基均数差异无统计 学意义 （F = 0.025， P > 0.05）。所有氨基酸残基序列均以12型重复作为结尾， 以1型、 2型重复为起始， 比例各占98.1% （152/155） 和1.9% （3/155）， 2型重复次数最多， 为12.9次。155条Pfhrp2 基因exon2区DNA序列的同源位点为894 bp， 保 守位点占20.8% （186/894）， 变异位点占78.2% （699/894）， 非洲虫株序列间遗传距离为0.000～0.741， 缅甸为0.000～ 0.948， 云南为0.000～0.750。所有155条序列按氨基酸序列大小聚类成3个大类， 同一个层级的序列具有近似的序列长 度和氨基酸重复类型。结论 结论 云南省恶性疟现症病例所感染疟原虫的Pfhrp2基因exon2区存在高度多态性， 恶性疟原虫 株主要按Pfhrp2基因exon2区的氨基酸序列长度进行聚类。

关键词: 恶性疟； 恶性疟原虫； 富组氨酸蛋白Ⅱ； Pfhrp2基因； 多态； 云南省