Study On Separation, Diversity And Activity Of Endophytic Actinomycetes Isolated From Acanthopanax Senticosus Harms

Objectives To isolate the endophytic actinomycetes from medicinal plants, and preliminarily establish the resource pool of endophytic actinomycetes strains separated from Acanthopanax senticosus. In addition, to reveal the diversity of endophytic actinomycetes strains, their distribution in different tissues of the plant, and explore the antimicrobial activity of strains and the synthetic potential of the secondary metabolites.Methods Acanthopanax senticosus samples were collected from 5 sites(Benxi in Liaoning Province; Meihekou in Jilin Province; Liuhe in Jilin Province; Muling in Heilong-jiang Province; Xinglong in Hebei Province.). Rinsed by running water, surfacesterilized, split and broken according to the root, rhizome, stem, leaf and other parts. the plants were put in 8 different culture media. Endophytic actinomycetes were separated and transferred to the ISP2 plate. the strain was purified by plate streak and frozen at-80 ℃.Dereplicated repetitive strains by BOX-PCR. Quite different strains and strains with special morphology were selected to carry on 16 S r RNA gene sequencing and phylogenetic analysis. Furthermore, the genus of endophytic actinomycetes and the distribution of strains in different tissues of the Acanthopanax senticosus were obtained.Secondary metabolite biosynthesis genes were detected by PCR. Paper disc diffusion method was used to test the antimicrobial activity of strains.Results 1 300 endophytic actinomycetes were isolated, and 265 strains were obtained after removing the same clones by BOX-PCR. 43 representative strains belong to 9 genera,9 families, 7 suborder. Streptomyces and Micromonospora genus are the dominant genera.Separated from the root, rhizome, stem and leaf of actinomyces strains were 110, 93, 45 and 17 strains, The total isolated actinomycetes number obtained from the root and the rhizome of samples were far greater than other parts, 77 percents, while the leaf was the least, 6 percents. The isolated actinomycetes number obtained from different media were difference. Out of the 8 different culture media,LNMS culture medium displayed the best separation effect, 60 strains, while trehalose-mineral salt medium displayed the worst separation effect, 17 strains. 2 a variety of biological activities were showed, the antibacterial activities were wide and strong.78 strains showed antimicrobial activities. 3PKS I, PKS II, NRPS, Halo and CYP gene sequences were detected by PCR in 48, 123, 52,18 and 29 of tested strains, respectively. Overall, those secondary metabolite biosynthesis genes were found in 172 strains of the total cultured strains.Conclusions 1 Wild Acanthopanax senticosus has a wealth of endophytic actinomycetes and abundant species diversity, The total isolated from the root and the rhizome of plants were far more than other parts. 2 these strains can be the high quality resources for screening new antibiotic because of the good antibacterial activity. 3 Diverse genes and potentiality of secondary metabolite existed inthese strains,which can have the potential to produce a variety of types of active compounds, the new active and novel structure of the drug lead compounds can be discoveried based on the genes, so these strains were worth to research and application.