贡嘎蝠蛾幼虫肠道真菌多样性分析

目的]分析贡嘎蝠蛾肠道(Hepialus gonggaensis)幼虫肠道真菌多样性.方法]采用常规分离与分子鉴定的方法和基于ITS(internal transcribed spacer)基因的RFLP方法,建立贡嘎蝠蛾幼虫肠道真菌的ITS克隆文库,分别用MspⅠ、HaeⅢ和Taq Ⅰ对205个阳性克隆的质粒酶切指纹图谱分析,结果显示有23个不同的RFLP操作分类单元(OTU),对这23个操作分类单元的阳性克隆子进行测序并绘制系统进化树.结果]结果显示贡嘎蝠蛾幼虫肠道内存在8个属的真菌类群.其中被孢霉属(Mortierella)和丝孢酵母属(Trichosporon)的丰度最高,分别占克隆文库的46.34%和40.00%,鉴定为肠道内的优势真菌类群.用常规分离与分子鉴定方法只获得隐球酵母(Cryptococcus magnus)、Geomyces sp和丝孢酵母(Trichosporon porosum)3个类群的真菌.结合常规分离法与RFLP法能够更有效的分析肠道微生物的多样性,获得更多更全面的微生物多样性信息.

Analysis of fungal diversity in intestines of Hepialus gonggaensis larvae

OBJECTIVE: Studied fungi diversity in the guts of larval Hepialus gonggaensis using culture-independent and traditional culturing methods. METHODS: For the culture-independent method, the total DNA of fungus was extracted from the intestinal contents and internal transcribed spacer (ITS) regions were amplified with fungal universal primers. A near-full length ITS gene library was constructed. Subsequently, the fingerprints of the microorganisms were analyzed by isolated plasmid and digestion with Msp I, HaeIII and Taq I enzymes, respectively. Restriction fragment length polymorphism (RFLP) analysis based on the fungal ITS sequences indicated that the library established includes 23 operational taxonomic units (OTUS) and a phylogenetic tree depicted the linkage of the isolated fungi. RESULTS: Abundant fungi were in the intestines of H. gonggaensis larvae, but their abundance was very different. The dominant fungi belonged to Mortierellaone and Trichosporon and accounted for 46.34 % and 39.02 % of the total ITS clones, respectively. Only three genera of fungi were identified from eight isolated fungal populations by traditional culturing methods. CONCLUSION: We could get more information by combined traditional culturing and molecular biology methods.