Isolation,Identification And Metabolites Analysis Of Actinomycetes Associated With Tropical Marine Sponge

Marine sponge-associated actinomycetes are an important part of marine microorganisms community.The secondary metabolites produced by marine sponge-associated actinomycetes have a high rate of structural innovation,which is an important resource for the discovery of new drug lead compounds.Hainan island is surrounded by the sea,and there are many kinds of marine sponges,which provide rich materials for the study of marine sponge-associated actinomycetes.In this paper,marine sponge samples were collected from the sea area of Wenchang,Hainan,the symbiotic actinomycetes were isolated,diversity of their strains was analyzed,the antibacterial strains were found,and the target secondary metabolites of the active strain were discussed.The main results are as follows:A total of 50 strains of marine actinomycetes were isolated from four kinds of marine sponge samples.According to the morphological differences of colonies,22 representative characteristic strains were selected and classified into 5 suborders,6 families and 7 genera by 16 S r RNA sequence analysis,including Streptomyces,Micromonospora,Rhodococcus,Saccharopolyspora,Isoptericola,Mycobacteria and Krasilnikoviella.Krasilnikoviella was isolated from marine sponge for the first time.Antibacterial activity test showed that 48%of the isolated strains showed antibacterial activity,and 24% of the isolated strains showed anti-plant filamentous fungal activity.Five new species of marine actinomycetes such as HNM0563,HNM0569,HNM0574,HNM0575 and HNM0581 were identified by polyphasic approach.HNM0563 and HNM0569 were identified as novel species of the genus Rhodococcus,for which the names Rhodococcu spongiae sp.nov.and Rhodococcu wenchangensis sp.nov.are proposed respectively;HNM0574 and HNM0575 were identified as novel species of of the genus Streptomyces,for which the names Streptomyces aquaticus sp.nov.and Streptomyces wenchangensis sp.nov.are proposed respectively;HNM0581 was identified as a novel species of Micromonospora,for which the name Micromonospora marinus sp.nov.are proposed.The new species strain HNM0575 was selected for mass fermentation,and eight media were selected to screen the best fermentation conditions,and the fermentation products were analyzed.Eleven compounds were isolated from the fermentation products by silica gel column,sephadex LH-20 gel column and HPLC chromatography.The structures of three compounds were determined by mass spectrometry and nuclear magnetic resonance analysis,among which compound 5 was formononetin,compound 6was daidzein and compound 11 was Oxopropaline G(?-carboline alkaloid).The test of antibacterial activity showed that compound 4 had antibacterial activity against Methicillin-resistant Staphylococcus aureus and Escherichia coli.Compound 6 showed antibacterial activity against Escherichia coli.