贡嘎蝠蛾幼虫肠道真菌多样性分析

Analysis of fungal diversity in intestines of Hepialus gonggaensis larvae

摘要:

【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TaqⅠ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。

Abstract:

[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 in-testinal 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 MspⅠ, HaeⅢand TaqⅠenzymes, 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 Mor-tierellaone 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.