Studies On The Diversity Of Co-existing Microorganisms And Their Secondary Metabolites

The unique ecological environment of the ocean brings up various biological species,and generates novel,abundant,and bioactive metabolites,which provides enormous biological sources for developing new drugs.At present,there are plenty of reports on the bioactive substances from sponge,coral,sea urchin,actiniae and sea hare associated microorganisms.But there are quite few reports on marine flatworms.Therefore,it is of great significance to study the important but long-time ignored creature associated microorganisms.The microorganisms from the marine flatworm Planocera sp.were isolated,purified and identified in the study.Totally 141 strains of bacteria including 45 strains of actinomyces were obtained and three novel bacterial strains which were named as Pseudovibrio hongkongensis UST20140214-015 BT,Pseudovibrio stylochi UST20140214-052 T and Deinococcus planocerae XY-FW106 T.These microorganisms were shown to separately belong to the following five phylogenetic groups,Proteobacteria,Bacteroidetes,Firmicutes,Deinococcus-Thermus and Actinobacteria.The results demonstrated the diversity and novelty of Planocera sp.associated microorganisms.Analysis of 16 S rRNA gene sequence of XY-FW106 T suggested it belonged to Deinococcus genus and it was similar to Deinococcus aetherius ST0316T(94.7%).The optimal growth condition of the strain XY-FW106 T are 25–30 °C and pH 7.2–7.4 without NaCl.Strain XY-FW106 T contained unidentified glycolipids,phosphoglycolipids,phospholipids and lipids,which differed from those of its closest related species.Menaquinone MK-8 was identified as themajor respiratory quinone and the predominant cellular fatty acids were found to be Summed Feature 3(C16:1 ?7c/C16:1 ?6c),C16:0,iso-C15:0,and Summed Feature 8(C18:1 ?7c/C18:1 ?6c).The DNA G+C content was determined to be 70.2 mol%.The biochemical and chemotaxonomic results together suggested that the strain showed the common characteristics with its gene,such as non-spore,non-motility,aerobic and UV resistant.However,it reveals its peculiarity which can grow in 5% of NaCl compared to the highest similarity strain Deinococcus aetherius ST0316 T which only grow without NaCl.Hence,strain XY-FW106 T identification represents a new species for which the name Deinococcus planocerae sp.nov.is proposed(Genbank No.KT886059).Based on bioactivity screening of Methicillin-resistant Staphylococcus aureus(MRSA)resistance and HeLa cell cytotoxicity,and liquid chromatography-mass spectrometry(UPLC-MS)analysis,the target strain Streptomyces samsunensis XY-FW47 with better bioactivity and varied chemical diversity was selected to proceed its secondary metabolites isolation,identification and bioactivity study.Scale fermentation of strain XY-FW47 was performed,and four compounds of geldanamycin and its derivatives were isolated and purified which was identified to be 4,5-dihydro-17-O-demethylgeldanamycin(1),17-O-demethylgeldanamycin(2),geldanamycin(3)and 4,5-dihydrogeldanamycin(4),and compound 1 was novel geldanamycin derivatives.The bioactivities of these compounds were evaluated by various screening models including anti-MRSA,antitumor,and anti-fungal assays.Compound 1 revealed non-ideal activity,while compound 3 exhibited significant anti-HeLa cell activity with EC5011.2 ?g/mL concentration and anti-fungal activity with MIC 2.4 ?g/mL as well.From the preliminary analysis of structure-activity relationship(SAR),it is speculated that the introduction of C4-C5 double bond and C17-O-demethyl)had a positive effect on improving HeLa cytotoxicity and anti-fungal activities of geldanamycins.Taken together,this study enriches the microorganisms source with the antibiotics of geldanamycins,which will provide technical support and basic information for the study on microorganisms associated with marineflatworms in China.