白念珠菌菌丝相与酵母相ERG11基因差异性分析

Difference of ERG11 gene between hyphal and yeast forms of Candida aIbicans

目的探讨白念珠菌菌丝相与酵母相ERG11基因间的差异。方法将16株同一亲本来源、对氟康唑敏感性不同的白念珠菌（CA-1-CA-9、CA-11-CA-17）进行菌丝相与酵母相培养后，分别抽提基因组DNA。设计引物扩增ERG11基因，扩增产物分别用AccⅠ、MunⅠ、AnⅡ消化，电泳后比较两相细胞RFLP图谱，同时用下游引物P2对ERG11基因扩增产物进行反向测序。结果AccⅠ、MunⅠ、AftⅡRFLP图谱及基因测序结果均显示菌株CA-7、CA-8、CA-14、CA-16、CA-17的菌丝相与酵母相间存在差异。两相细胞ERG11基因在1547、1587、1617三位点存在差异，位点编号按照基因库中ERG11基因序列编号（登录号X13296）。结论白念珠菌菌丝相与酵母相ERG11基因间存在差异，在进行白念珠菌体外研究时，选用其菌丝相更能反映体内真实情况。

Objective To compare the differences of ERG11 gene between the hyphal and yeast forms of Candida albicans. Methods Sixteen C albicans strains （ CA-1, CA-2, … CA-9, CA-11, CA-12, … CA-17 ）, which were all isolated from a HIV-infected patient and had different sensitivity to fluconazole, were cultured in both hyphal and yeast forms. Genomic DNA was extracted from both forms of these strains. Primers were designed to amplify their ERG11 gene. The PCR products were digested by restriction endonucleases Acc I , Mun ] and Afl Ⅱ, respectively, and after agarose gel electrophoresis, RFLP fingerprints were compared between the two forms. Meanwhile, the PCR products were sequenced with the downstream primer （ P2 ）. Restdts Differences were found in the ERG11 gene between the two forms in 5 strains by both RFLP analysis and gene sequencing. The digestion of PCR products with Acc Ⅰ resulted in two fragments in the yeast form of strains CA-7 and CA-8 and in the hyphal form of strains CA-14, CA-16 and CA-17, but one fragment in the hyphal form of strains CA-7 and CA-8 and in the yeast form of strains CA-14, CA-16 and CA-17. The RFLP pattern resulted from Mun Ⅰ was coincident with that from Aft Ⅱ, which presented three fragments in the yeast form of strains CA-7 and CA-8 and in the hyphal form of strains CA-14, CA-16 and CA-17, but one fragment in the hyphal form of strains CA-7 and CA-8 and in the yeast form of strains CA-14, CA-16 and CA-17. Gene sequencing revealed a C→T or T→C mutation at 1547 site, C/T→T or T→C/T mutation at 1 587 site and an A→A/G or A/G→A mutation at 1617 site, of the ERG11 gene （GeneBank accession number： X13296） between the two forms of the five strains. Conclusion There are differences in ERG11 gene between the hyphal and yeast forms of C albicans.