口腔黏膜真菌鉴定方法的比较

比较常见用于黏膜真菌菌种鉴别的多种方法,探寻最佳的鉴别方法。采集230例普通人群口腔黏膜样本,分别用玉米吐温-80培养观察厚膜孢子法、糖发酵生化反应法、CHROMagar假丝酵母菌显色培养基法、ITS基因的PCR-RFLP(聚合酶链反应-限制性片段长度多态性)法、ITS测序菌种鉴定法,鉴别真菌各菌株。结果显示:有56例菌株至少通过1种方法检出真菌;玉米吐温-80分离培养假丝酵母菌37株;50例菌株ITS基因测序共鉴定出8个菌种,白假丝酵母菌(C.albicans)29株,近平滑假丝酵母菌(C.parapsilosis)10株,热带假丝酵母菌(C.tropicalis)5株,Candida metapsilosis 1株,Lodderomyces elongisporus 1株,克柔假丝酵母菌(Candida krusei)1株,乙醇假丝酵母菌(C.ethanolica)1株,季也蒙毕赤酵母菌(Pichia guilliermondii)2株;CHROMagar假丝酵母菌显色培养基法鉴定出3种菌株,分别是白假丝酵母菌、热带假丝酵母菌、近平滑假丝酵母菌;PCR-RFLP法检出5种菌株,分别是白假丝酵母菌、热带假丝酵母菌、近平滑假丝酵母菌、季也蒙毕赤酵母菌、克柔假丝酵母菌,与基因的测序鉴定一致率为91%;糖发酵生化反应法阳性标本占被检出真菌例数的46.4%(26/56)。结果表明:ITS基因的测序法可以准确鉴定真菌各个菌种;PCR-RFLP法能鉴定常见的菌种,但操作繁琐;CHROMagar假丝酵母菌显色培养基法能快速准确鉴别3种常见假丝酵母菌菌种;玉米吐温-80可以准确培养鉴别白假丝酵母菌;糖发酵生化反应法,缺乏足够的敏感度和特异性,难以准确鉴别各个菌种。

Comparison of Methods for Fungal Identification from Oral Mucosa

A total of 230 common crowd’s isolates of fungal strains from oral mucosa samples were collected, and identified with common mucosa fungal species identification methods such as corn TWEEN80 chlamydospore observation, sugar fermentation biochemical reaction, CHROMagar Candida chromogenic medium, PCR-RLFP of ITS gene, ITS sequencing, respectively for comparison to seek optimal identification method. The results showed that at least 56 cases of strains were identified with one method. 37 Candida strains were separated with corn TWEEN80; 50 cases of strains were identified as 8 strains based on ITS gene sequencing： 29 strains were C. albicans, 10 strains C. parapsilosis, 5 strains C.tropicalis, 1 strain C. metapsilosis, 1 strain Lodderomyces elongisporus, 1 strain C. krusei, 1 strain C. ethanolica, 2 strains Pichia guilliermond; 3 species were identified by CHROMagar Candida medium, including C. albicans, C. tropicalis,C. parapsilosis; 5 species were identified by PCR-RFLP, including C. albicans, C.tropicalis, C. parapsilosis, Pichia guilliermondii and C. krusei. The concordance detection ratio of PCR-RFLP and gene sequencing were 91%; the detected cases by biochemical reaction of sugar-positive specimens accounted for 46.4%（26/56） of total cases. Therefore, ITS gene sequencing could accurately identify various species of fungi; PCR-RFLP method could identify common species, but cumbersome to operate; CHROMagar Candida chromogenic medium was a fast and accurate method to identify three kinds of common Candida species; Corn TWEEN80 cultivation could accurately identify C. albicans; sugar fermentation biochemical reaction was lack of sufficient sensitivity and specificity, and was difficult to accurately identify various species.