从Giemsa染色的血膜中扩增间日疟原虫DNA方法探讨

DNA Amplification of Plasmodium vivax Parasites from Giemsa-stained Blood Smears

目的建立从Giemsa染色的血膜中提取疟原虫DNA的方法。方法分别采用Na2HPO4法和Chelex-100离子交换法,经过反复优化,提取血膜中的DNA,进行套式PCR扩增鉴定PvMSP-1等位基因型。结果用Na2HPO4法和Chelex-100离子交换法分别提取40张不同类型的间日疟原虫阳性血膜DNA,未染色的厚血膜全部扩增出目的基因条带,而染色的薄血膜未能扩增出目的基因条带。两种方法均可检测红细胞间日疟原虫感染率≥0.01%的血涂片。结论从多年保存的标准血膜中提取疟原虫DNA进行基因型鉴别是可行的。

Objective To develop methods of extracting DNA from malaria parasites on Giemsa-stained blood smears. Methods Improved Na2HPO4 method and Chelex-100 ion-exchange technique were used to extract DNA from Giemsa-stained or unstained blood smears. Nested PCR was employed for amplification and identification of allelotypes in the Plasmodium vivax merozoite surface protein-1 （PvMSP-1）. Results Target DNA bands appeared in all samples of unstained thick blood smears, while no DNA bands were visible in the fixed and stained thin smears. Both methods identified PvMSP-1 alleles from smears with parasitemia of ≥0.01%, Conclusion It is feasible to identify PvMSP-1 alleles from Giemsa-stained blood smear.