凋落物真菌Berkleasmium sp.及其螺二萘类化合物抗菌活性的研究

Studies on the Antimicrobial Activity of Litter Fungus Berkleasmium sp. and Its Spirobisnaphthalenes

为了分离鉴定森林凋落物中可培养的真菌,并发现具有抗菌活性的菌株,研究它们的主要抗菌代谢物,为进一步开发利用凋落物真菌资源提供备选菌株。本研究通过颗粒法分离森林凋落物中的可培养真菌,用内部转录间隔区(ITS)序列分析鉴定分离到的真菌,采用打孔药剂扩散法检测抗菌活性,利用柱层析法分离抗菌单体化合物,通过波谱学分析确定其化学结构。共分离得到63株真菌,它们隶属于36个属、48个分类单元,它们中有14株菌ITS相似性小于98.5%。通过抗菌活性筛选发现,11株真菌的乙酸乙酯提取物具有抗菌活性,其中Berkleasmium sp. SGSF622抗菌活性较强。通过PDA培养基对该株真菌进行发酵,获得乙酸乙酯提取物3 g。通过抗菌活性追踪分离,得到其含量最高的抗菌化合物622A。通过核磁共振及高分辨质谱分析,进一步确认该化合物为螺二萘类化合物Sch53825,它的分子式为CHClO.,其抗青枯劳尔氏菌的最低抑菌浓度为125~250μg/mL。研究首次在Berkleasmium属真菌中分离到化合物Sch53825,并首次发现该化合物具有抗植物病原菌活性,这为进一步开发利用森林凋落物真菌资源提供了参考。

The purpose of this study is to isolate and identify culturable fungi from forest litter,to find the strains with antimicrobial activity and to study their main antimicrobial metabolites,so as to provide alternative strains for further development and utilization of fungal resources in forest litter.This study used the particle separation method to isolate the culturable fungi in forest litter,internal transcribed spacer (ITS) sequence analyses to identify isolated fungi,well diffusion method to detect antimicrobial activity,column chromatography method to separate antimicrobial monomeric compounds,and spectrum analyses to determine their chemical structure of the monomeric compounds.A total of 63 fungal strains were isolated,belonging to36 genera and 48 taxa,among which,14 strains had less than 98.5%of ITS similarity.Ethyl acetate crude extracts of 11 strains had antimicrobial activity,and Berkleasmium sp.SGSF622 had strong antimicrobial activity.The fungus was fermented in PDA medium,and 3 g of ethyl acetate extract was obtained.By bioguided separation of antimicrobial activity,antimicrobial compound 622A with the highest content was obtained.By NMR and HR-MS analyses,this compound was further identified as Sch53825 and the molecular formula was CHClO.Its minimal inhibition concentration of antibacterial activity against Ralstonia solanacearum was 125-250μg/m L.It is the first time to separate Sch53825 from fungi of genus Berkleasmium.The study can lay a foundation for further development and utilization of fungal resources from forest litter.