Isolation And Screening Of Endophytes From Tripterygium Wilfordii With Insecticidal Activity

More and more research focus on the resources of medicinal plants endophytes. It isfound that endophytes have the characteristics the breadth and diversity and their metaboliteshave novel structure and diverse biological activity. Therefore endophytes have beenconcerned by a number of areas such as medicine, plant pathology and microbiology.Tripterygium wilfordii is a traditional Chinese medicinal plant, a variety of compounds thathave medical and agricultural activity are isolated from the root bark. In this study,Tripterygium endophytes were systematic separated and the insecticidal activity of strains wasscreened so as to obtain one or two strains which can produce high insecticidal activitycompounds. The main process and the results are as follows:1.183endophytes were separated from two kinds of tripterygium wilfordii plantmaterials in all, including111fungus,23actinomycetes and49bacteria. The proportion offungus was the lagest, accounting for60.65%of the total isolate strains number. Bacteriaaccounted for26.78%and actinomycetes accounted for12.57%. The numbers of endophytesin different tissues of Tripterygium wilfordii were obviously different. The most endophytes(89) were isolated from Tripterygium wilfordii root accounting for48.63%, secondly wasleaf,accounting for39.35%. The least endophytes were obtained from stems(22), accountingfor12.02%, in point of view different tissues of plants.2. The insecticidal activity against Artemia and3th instar larvae of Culex pipiens of183endophytes was determinated. The results showed that endophytes the mortality more than90%after24h against Artemia were34strains, they were11-3-8,11-3-37,11-6-58,11-6-76,11-6-73and so on, accounting for18.58%of the tested. There were19,11and3in roots,stems and leaves part, respectively, accounted for55.88%,32.35%and8.82%of the numberof active strains. Among the34strains, there were33fungus and an actinomycete. Theinsecticidal activity of183endophytes against larvae of Culex pipiens was determinated. Themortality rate more than70%after24h had18strains, they were11-3-37,11-6-58,11-6-76,11-6-71and so on, accounting for9.83%of the total number. Strains isolated from stem ofTripterygium wilfordii had no insecticidal activity against larvae of Culex pipiens.3. The insecticidal activity against Myzus persicae of the18strains was determinated using the dip worm immersion method. Three strains demonstrated high activity againstMyzus persicae.11-6-76had higher activity both intracellular and extracellular, the mortalityrates were78.64%and70.75%. The mortality rate of intracellular crude extractings of11-6-59and12-2-47were82.74%and76.97%respectively. The LC50of intracellularmetabolites crude extracts of11-3-37,11-6-76and12-2-47against Myzus persicae were34.8mg/mL,31.2mg/mL and46.4mg/mL respectively.4. Using four different chromogenic reagents, the alkaloids of18Tripterygium wilfordiiendophytes fermentation broth and mycelium extracts was determined.The results showedthat the crude extracts of intracellular metabolites of11-3-37and11-6-76was positive. It wasdeterminated preliminary that the two strains contain alkaloids. Further, the amount ofalkaloids in the mycelium of the two strains were determined by HPLC, the amount were:8μg/g and40μg/g.5. Basing on the morphological characteristics and physiological and biochemicalcharacteristics,11-6-76was preliminarily identified as Streptomyces Albosporus sp..