Screening Of Swainsonine-producing Endophytic Fungi In Locoweed And The Test Of Bacteriostatic Activity In The Fermentation Broth

Endophytic fungi in its life cycle refers to a period of time or the whole life period in the plant tissue of the host, without causing obvious symptoms kind of the host plant. Some researches found that the secondary metabolites of endophytic fungi contain many bioactive substances.Locoweed is composed of Astragalas and Oxytropis, which can cause animals characterized nerve disorders after poisoning.Swainsonine(SW) is the main toxin for locoweed, a kind of indolizidine alkaloids. Named it swainsonine because it has been separated from Swainsonia salsula. Swainsonine, it has very broad medicinal prospects in many medicinal field, especially its huge potential antitumor activity which attracted wide attention of scholars at home and abroad. By the researches of domestic and foreign scholars to locoweed, endophytic fungi of isolation, identification and fermented product testing ,they found that swainsonine-producing endophytic fungi existing in locoweed, such as Embellisia bacteria (sp).This research aimed at locoweed serious disasters of qinghai province and Inner Mongolia, and acquisition of the locoweed samples: Oxytropis glabra, oxytropis ochrocephala bunge, Oxytropis kansuensis and Astragalus variabilis.Then endophytic fungi separation, purification, identification and spawn preservation were proceed. The antibacterial activities element in the Fermented product of endophytic fungi was analysed, in order to identify swaisonine-producing endophytic fungi, and then proceed biological fermentation method of the separated endophytic fungi strains to detect its fermentation liquor and mycelia for different polarity segment of antibacterial activities. lay the foundation for the development and utilization of the endophytic fungi of Chinese locoweed. This research has achieved following results:1.Screening swaisonine-producing endophytic fungiSeparate and purify the endophytic fungi according to different parts of locoweed. Then analysed the element in the fermented products of endophytic fungi by thin layer chromatography (TLC) and gas chromatography (GC), it showed that the secondary metabolites of two endophytic fungi strains contain swainsonine: the swainsonine content in XJ-H-1 and XJ-J-1 fermentation liquor are 16.7338 mg/L and 4.2092 mg/L respectively. 2. Identification of swaisonine-producing endophytic fungiCombining the morphological observation of hyphae and spore and the analysis of 5.8 SrDNA - ITS sequencing to Identify the two swaisonine-producing endophytic fungi strains. The two swaisonine-producing endophytic fungi strains have minor differences in morphology, but in 5.8 SrDNA - ITS sequence level, their gene homology is exactly the same as high as 100%. XJ-J-1 and XJ-H-1 are Fusarium proliferatum, their homology reached 99%. Through the analysis of Phylogenetic Tree, the two swaisonine-producing endophytic fungi strains have close kin to Fusarium proliferatum, self-check approval ratings as high as 90%. Combined with description of morphology in The Fungus Appraisal Manual, XJ-H-1 and XJ-J-1 are Fusarium proliferatum.3. Antibacterial activity researches in fermented product of the endophytic fungiUse the swaisonine-producing endophytic fungi strains(Fusarium proliferatum) for fermentation, extract the fermentation products with petroleum ether, chloroform, ethyl acetate, n-butyl alcohol in turn, then via disc agar diffusion, MIC and MBC method to test its antibacterial activity in fermented and mycelia of different polarity segment. The result shows: the ethyl acetate polarity segment of the fermenting has good antibacterial activity, and the bacteriostatic circle diameter of bacillus subtilis, staphylococcus aureus, salmonella and e. coli are 1.66 cm, 1.984 cm, 1.902 cm, 1.902 cm respectively. The MIC are 0.39 mg/mL, 3.125 mg/mL, 1.56 mg/mL, 0.78 mg/mL respectively. For its MBC are 3.125 mg/mL,12.5 mg/mL,6.25 mg/mL,6.25 mg/mL respectively.