春季油樟可培养内生真菌多样性及产黄酮功能初筛

为开发油樟内生菌资源并发现潜在活性成分,本研究以春季油樟健康植株的根、茎、叶为材料,采用组织培养法分离纯化可培养内生真菌,结合ITS序列和系统发育树分析菌株多样性,筛选具有产黄酮功能菌株,设计单因素和正交试验优化产黄酮效果最佳菌株的发酵条件。结果表明：从油樟中共分离到62株可培养内生真菌,划分为3门、7纲、14目、22科、28属、54种。其中优势门为子囊菌门（Ascomycota,RF为88.71%）,优势纲为座囊菌纲（Dothideomycetes,RF为45.16%）,优势目为格孢腔菌目（Pleosporales,RF为37.10%）,优势科为格孢菌科（Pleosporineae,RF为14.52%）、亚隔孢壳科（Didymellaceae,RF为12.90%）,优势属为链格孢属（Alternaria,RF为12.90%）、附球菌属（Epicoccum,RF为11.29%）。以纲为类群单位,根组织分离的16属20种21株真菌的系统发育可分为7个类群,茎分离的13属24种30株真菌则分为2个类群,叶的8属10种11株真菌则分为3个类群。多样性指数和均匀度系数结果表明,油樟根部内生真菌种群多样性相对较丰富,群落分布相对均匀。10株内生真菌具有产黄酮功能,5株来自根部,5株来自于茎部,隶属2门3纲5目8科9属,其中YZ-29菌株产黄酮能力相对较强。YZ-29菌株最优发酵条件为初始pH 7、时间7 d、温度30℃、转速175 r/min、装液量80 mL/150 mL。综上结果显示,内生真菌在油樟不同组织数量、种群及组成上存在差异,分离得到的功能性菌株值得深入挖掘和开发利用,为开展内生真菌在油樟抗氧化成分的生物合成机制方面的研究奠定基础。

Diversity and Flavonoid-Producing Function of Culturable Endophytic Fungi in Natural Spice Cinnamomum longepaniculatum (Gamble) N. Chao in Spring

In order to develop the resources of Cinnamomum longepaniculatum endophytes and discover its potential active components, this study utilized the roots, stems and leaves of healthy C. longepaniculatum plants as the materials to isolate and purify culturable endophytic fungi by the tissue culture method. Combined with the ITS sequence and system, the developmental tree was used to analyze the diversity of strains, screen strains with flavonoid-producing functions, and design single-factor and orthogonal experiments to optimize the fermentation conditions of strains with the best flavonoid-producing effect. Sixty-two culturable endophytic fungi isolated from C. longepaniculatum, which were divided into three phyla, seven classes, fourteen orders, twenty-two families, twenty-eight genera, and fifty-four species. Among them, the dominant phyla was Ascomycota (88.71%), the dominant class was Dothideomycetes (45.16%), the dominant order was Pleosporales (37.10%), and the dominant family was Pleosporineae (14.52%) and Didymellaceae (12.90%), the dominant genera were Alternaria (12.90%) and Epicoccum (11.29%). Using the class as a group unit, the phylogeny of the fungi isolated from the root tissues included sixteen genera, twenty species and twelve isolates that could be divided into seven groups. The fungi isolated from the stems contained thirteen genera, twenty-four species, and thirty isolates that could be divided into two groups. The fungi isolated from the leaves contained eight genera, ten species, and eleven isolates and could be divided into three groups. The results of the diversity index and evenness coefficient showed that the endophytic fungi population diversity in the roots of C. longepaniculatum was relatively rich, and the communal distribution was relatively even. Ten strains of endophytic fungi had the function of producing flavonoids: five from roots, five from stems, belonging to two phyla, three classes, five orders, eight families and nine genera. Among them, the YZ-29 strain had relatively strong flavonoid production capabilities. The optimal fermentation conditions for the YZ-29 strain to produce flavonoid were to be cultured at initial pH seven for 7 days at 30℃, with rotation speed of 175 r/min and liquid volume 80 mL/150 mL. The results above showed that there were differences in the number, population and composition of endophytic fungi in different tissues of C. longepaniculatum. The functional strains that were screened are worthy of further excavation, development and utilization. The results would lay a foundation for the biosynthesis of antioxidant components from endophytic fungi in C. longepaniculatum.