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目的分离纯化水仙根茎中的内生真菌,并进行菌群组成及其多样性分析。方法采用表面消毒、划线法从水仙鳞茎和根中分离内生真菌,基于形态学特征和核糖体转录间隔区(ITS-rDNA)序列分析,对分离株进行分类鉴定;以内生真菌的多样性指数(H)和均匀度指数(E)为指标,分析水仙内生真菌的菌群组成及其多样性。结果共分离到18株内生真菌,其中16株来自根部,2株来自鳞茎,分别占总分离菌株的88.89%和11.11%。PCR扩增其中14株真菌ITS-rDNA,得到大小约500 bp片段,用BLAST软件对其测序结果进行相似性比对,发现4株为青霉属Penicillium,相似性95%~99%;3株为曲霉属Aspergillus,相似性99%~100%;3株为喙枝孢属Rhinocladiella,相似性为99%;其他4株分别为木霉属Trichoderma、链格孢属Alternaria、赤霉属Gibberella和镰孢霉属Fusarium,相似性都为99%。青霉菌为水仙内生真菌优势菌属。通过构建系统发育树发现这14株内生菌明显分为2大类群,Shannon多样性指数为1.778,均匀度为0.913 7。结论水仙内生真菌菌群组成具有一定程度的多样性和差异性。
Objective To isolate and purify the endophytic fungi in the rhizome of narcissus, and to analyze the composition and diversity of the flora. Methods Endophytic fungi were isolated from bulb and root of Narcissus by surface disinfection and scribing method. The isolates were classified and identified based on their morphological characteristics and ITS-rDNA sequence analysis. The endophytic fungi Index (H) and evenness index (E) were used as indexes to analyze the flora composition and diversity of endophytic fungi in Narcissus. Results A total of 18 strains of endophytic fungi were isolated, of which 16 were from the roots and 2 from the bulbs, accounting for 88.89% and 11.11% of the total isolates, respectively. Fourteen strains of ITS-rDNA were amplified by PCR, and a fragment of about 500 bp was obtained. By BLAST software, the similarity of ITS-rDNA sequences was found. Four strains were Penicillium with similarities of 95% -99%. Three strains Aspergillus, the similarity is 99% ~ 100%; 3 strains are Rhinocladiella, the similarity is 99%; the other 4 strains are Trichoderma, Alternaria, Gibberella and Sickle Fusarium, similarity is 99%. Penicillium dactyloides endophytic dominant bacteria genus. Through the construction of phylogenetic tree, the 14 endophytic bacteria were found to be divided into two groups, Shannon’s diversity index was 1.778, and the evenness was 0.9137. Conclusion The composition of endophytic fungi flora of Narcissus has a certain degree of diversity and diversity.