Diversity And Antimicrobial Activity Of Actinobacteria Isolated From Mangrove Rhizosphere Soil In Maowei Sea Of Guangxi

Objective To study the diversity and antimicrobial activity of actinobacteria isolated from mangrove rhizosphere soils in Maowei Sea Mangrove Reserve,Guangxi Zhuang Autonomous Region and lay the foundation for finding new antibacterial drugs.Methods Eight rhizosphere soil samples were collected from Maowei Sea Mangrove Reserve.Actinobacteria were isolated by using standard serial dilution plating method with eight different media.To analyze the diversity of actinobacteria,sequences of 16S r RNA gene obtained by PCR amplification were submitted to gene database for BLAST search and then the phylogenetic trees were constructed by adjacency method to analyze homologous species.The fermentation broths and mycelia of the actinobacterial strains were extracted by ethyl acetate and acetone,respectively.Two different samples were obtained,including the organic layer and acetone extraction.Samples were selected to evaluate antibacterial activities and mechanisms by paper disc diffusion method and a unique double fluorescent protein reporter system(p Dualrep2),respectively.TLC technology was used for preliminary analysis of active substances.Morphological and physiological and biochemical tests were performed on some strains.Results 261 actinobacterial strains were isolated from 8 mangrove-originated soil samples by culture-dependent methods.They were distributed in 19 genera affiliated to 10families within 6 orders.Microbacteriaceae includes five genera:Micrococcus,Arthrobacter,Sinomonas,Paenarthrobacter and Pseudarthrobacter;Nocardiaceae includes three genera:Nocardia,Rhodococcus and Gordonia;Streptomycetaceae includes two genera:Streptomyces and Kitasatospora;Mycobacteriaceae includes two genera:Mycobacterium and Mycolicibacterium;Thermomonosporaceae includes two genera:Actinomadura and Actinocorallia;Microbacteriaceae includes one genera:Agromyces;Intrasporangiaceae includes one genera:Intrasporangium;Micromonosporaceae includes one genera:Micromonospora;Streptosporangiaceae includes one genera:Microbispora;Nakamurellaceae includes one genera:Nakamurella and the dominant genera were Micromonospora and Streptomyces.The highest similarities of 16S r RNA gene sequence between the three strains of actinomyces and the validly described species Agromyces binzhouensis OAct353~Twere 98.29%,98.36%and 98.37%.The low similarities(<98.65%)indicated the three strains were potential new taxa.Antimicrobial assay showed that 32 out of 83 strains exhibited positive results in antibacterial assay,and the total positive rate reached to 39.00%.The double fluorescent protein reporter system screened that 1 strain(strain B441)in genus Streptomyces,and 1 strain(strain B704)in genus Micromonospora,induced Katushka2S expression as erythromycin did.Meanwhile,four strains(strains B475,B486,B98 and B353)in genus Streptomyces,induced DNA damage SOS response as levofloxacin did.TLC analysis showed that streptolyces B353,B98,B704 and B486 produced active substances against Staphylococcus aureus.Conclusion The study demonstrated that the actinobacteria from Maowei Sea Mangrove Reserve were high diversity,and thus deserved to be further explored to discover new actinobacterial taxa and antibiotics.