恶性疟原虫现场样品HRP-Ⅱ基因部分序列分析

ANALYSIS ON THE POLYMORPHISM OF PLASMODIUM FALCIPARUM HRP-Ⅱ GENE

目的 分析和比较不同地理株恶性疟原虫现场样品及我国恶性疟原虫现场样品与实验室培养虫株在HRP Ⅱ基因上的多态性。 方法 分别以在中国感染的恶性疟病人和在非洲感染的恶性疟病人全血为材料 ,PCR扩增HRP Ⅱ基因片段 ,扩增的产物分别克隆于 pUCm T载体并进行测序 ,用GENEDOC软件比较分析核苷酸序列及其推导的氨基酸序列的同源性。 结果 从中国海南省和云南省恶性疟病人血样中扩增出序列和长度完全相同的片段 ,大小为44 7bp ;从在非洲感染的恶性疟病人血样中扩增出 813bp的片段 ;中国恶性疟原虫实验室培养株相应核苷酸序列长度为870bp。从中国恶性疟病人血样扩增出的HRP Ⅱ基因片段序列与从非洲恶性疟病人血样扩增出的HRP Ⅱ基因片段序列及我国恶性疟原虫培养株相应的核苷酸序列相比 ,不但有多个长短不等序列的缺失和插入 ,还有多个碱基发生了突变 ;比较推导的氨基酸序列 ,除了有多个长短不等氨基酸序列的缺失和插入外 ,其他氨基酸残基没有发生改变。 结论 恶性疟原虫HRP Ⅱ基因在不同地理株间存在差异 ,在同一地理株的实验室培养株与现场样品间也存在差异 。

Objective To analyze the polymorphism of Plasmodium falciparum HRP-Ⅱ gene in field samples with different geographic origin or between field strains and cultured isolate. Methods HRP- Ⅱ gene fragments of P. falciparum were amplified by PCR from blood samples of patients infected with P. falciparum in China and in Africa, respectively, then the amplified fragments were cloned into vector pUCm-T and their nucleotide sequences were determined. The sequences were compared and analyzed using GENEDOC software. Results A fragment of 447 bp was amplified from all blood samples from patients infected with P. falciparum in China and their sequences were also identical. However, a 813 bp fragment was amplified from blood samples of patients infected with P. falciparum in Africa. Alignment of their sequences revealed that not only different length of sequences were deleted and a eighteen-oligonucleotide was inserted in the HRP-Ⅱ gene fragment amplified from blood of patients infected in China, but also ten nucleotides were different between two amplified nucleotide sequences. There was no difference in the two deduced amino acid sequences except a few deletions and/or insertions of different length. Alignment of sequence from Chinese falciparum malaria patients with published sequence from cultured Chinese strain of P. falciparum also revealed deletion/insertion of several different length of sequences and several nucleotide mutation. Conclusion There was considerable difference in the HRP-II gene, mainly in length of sequence, not only in field strains with different geographic origin but also between field samples and cultured strain of P. falciparum.