Species Diversity Analysis And Bioactive Components Of Actinobacteria From Hehua Hot Spring In Baoshan,Yunnan Province

Objective:To explore the species diversity,enzymatic characteristic and antibacterial activity of actinomycetes in Hehua hot spring and obtain some bioactive strains for further studies,which provides microbial basis for further study of potentially bioactive secondary metabolites from microorganisms in extreme environments.Methods:The actinomycetes were isolated and purified by plate dilution coating method and streak separation method,and identified by morphological,physiological,biochemical and molecular biological methods,based on 16 S r DNA gene sequence.The enzymatic characteristic of actinobacteria was tested according to the enzyme identification method as described in "Rapid Identification and Systematic Classification of Actinomycetes".The antibacterial activity of the ethyl acetate crude extract of microbial fermentation broth was screened by 48-well plate method.High performance liquid chromatography technology was used for screening of chemical diversity / novelty of secondary metabolites of these strains.Results:A total of 60 actinobacterial strains were isolated from Hehua hot spring,including of 10 species of Streptomyce,3 species of Micromonospora and 1 species of Nomura.Among these strains,a total of 15 strains can produce hydrogen sulfide and have amylase activity,14 strains can reduce nitrate,12 strains have chymosin-producing properties,10 of the strains have cellulase activity,and 3 strains have protease activity.Among these enzymatic strains,strainsS.ossamyceticus H13 and H28 showed extremely significant enzyme-producing characteristics.Moreover,a total of 6 strains affiliated to the genus Streptomyces exhibited antibacterial activity.Specifically,strains S.ossamyceticus H1,S.toxytricinis H20 and S.filipinensis H48 showed antibacterial activity against Staphylococcus aureus,and strains S.torulosus H14,S.ossamyceticus H43 and S.filipinensis H47 displayed antibacterial activity against Micrococcus luteus.More importantly,strains H47 and H14 have good antibacterial activity against S.aureus with MIC values ranging from 6.25 to 12.5 ?g/m L,and strain H47 showed excellent bacteriostatic effect on M.luteus with MIC value ranging from 3.125 to6.25 ?g/m L.In addition,a total of 6 strains of Streptomyces show good chemical diversity /novelty.Among them,strains S.ossamyceticus H1 and H43 may produce new secondary metabolites with antibacterial activity.Strains S.torulosus H14 may produce a class of unsaturated lipid varicose B analogues with strong antibacterial activity against S.aureus and M.luteus.A series of polyacetal compounds were found in the fermentation broth of strain S.toxytricini H20,which showed antibacterial activity against S.aureus.Last but not the least,although strains S.speibonae H8 and S.toxytricini H41 do not show antibacterial activity,both of them can produce some new secondary metabolites with large quantities,which can be considered as engineered bacteria.Conclusions:Hehua hot spring is rich in actinobacteria,and many strains have good biological activity,which deserves for further study.This research can provide data reference for the further exploitation of microorganisms from hot spring in Baoshan area,and is also significant for rational utilization and protection of microbes derived from hot springs in Yunnan province.