Taxonomic Analysis Of Two Novel Bacteria And Screening Of Antagonistic Bacteria Against Pathogenic Fungi Of Orchid

For a long time,the study of novel bacteria is a hot topic.The novel bacteria can not only enrich microbial resources,also can discover new functional genes,which has great prospects and potential application.Phylogenetic analysis of 16 S rRNA gene sequence revealed that two endophytic bacteria isolated from Kumtag plants were potential new species.Polyphasic taxonomy was used to clarify their classification status.(1)Polyphasic taxonomic approach were used to characterize strain N5SSJ16.The cells were Gram-negative,aerobic,non-motile,non-spore-forming,rods.On TSA agar,colonies were circular with regular margins,convex,yellow and opaque.The strains grew in the presence of 0-4%(w/v)NaCl(optimum growth in the absence of NaCl),at pH 7-9(optimum,pH 7)and at 12-50?.The major fatty acids were iso-C15 : 0(53.20%)?summed feature 3(C16 : 1?7c and/or C16 : 1?6c)(20.94%)?summed feature 9(C16 : 010-methyl and/or iso-C17 : 1 ?9c)(9.98%)and iso-C17 : 03-OH(4.15%)and the content of iso-C17 : 03-OH was different from Parapedobacter species.The major respiratory quinone was MK-7.The major polar lipids were Phosphatidylethanolamine(PE),unidentified aminophospholipid(UPAL),two unknown lipids(UL).The existence of UPAL and UL makes strain N5SSJ16 different from Parapedobacter species.The DNA G+C content was 45.8 mol%.16 S rRNA gene sequences analysis revealed strain N5SSJ16 has close relationship with Parapedobacter species,and the closest neighbours being P.composti 4M40T(96.5% similarity)and P.luteus DSM 4M29T(95.9% similarity).On the basis of chemotaxonomic,phylogenetic and phenotypic evidence results,strain N5SSJ16 T represents a novel member of the genus Parapedobacter,for which the name Parapedobacter deserti sp.nov.is proposed.The type strain is N5SSJ16 T.(2)A moderately halophilic bacterium,designated R4HLG17 T was characterized using a polyphasic taxonomic approach.The cells were Gram-negative,aerobic,non-motile,non-spore-forming rods,non-pigmentation.On ISP5 agar,colonies were circular,yellow,edge neat.The strains grew in the presence of 0-16%(w/v)NaCl(optimum,5-6),at pH 5-9(optimum,pH 7)and at 16-45?.The major fatty acids were Summed Feature 8(C18 : 1?7c and/or C18 : 1?6c)(34.04%)?Summed Feature 3(C16 : 1?7c and/or C16 : 1?6c)(24.57%)?C16 : 0(16.85%)?C12 : 03-OH(8.32%)?cyclo-C19 : 0?8c(6.36%)and the content of cyclo-C19:0 ?8c is different from genus species of Halomonadaceae.The major respiratory quinone was Q-9.Phylogenetic analysis based on 16 S RNA gene sequences showed that the strain was a member of the family Halomonadaceae.The highest similiarity between R4HLG17 and Halomonas lutea KCTC 12847 T was 95.2%.On the basis of the chemotaxonomic,phylogenetic and phenotypic evidence,strain R4HLG17 T represents a novel genus and species preliminarily.Phytophthora cactorum Colletotrichum gloeosporioides,Fusarium oxysporum and Fusarium solani are four pathogenic fungi which can cause orchid diseases.In order to screen bacterial strains with high antagonism against the four fungal pathogens,120 strains kept in our laboratory were tested using the method of dual culture on plate.Preliminarily screening results showed that 8 strains had antagonistic activity against Phytophthora cactorum,8 strains against C.gloeosporioides,7 strains against F.oxysporum and 10 strains against F.solani.After secondary screening,5 strains could distinctly inhibit C.gloeosporioides,F.oxysporum and F.solani and strain GT312 was the best one.Phylogenetic analysis of 16 S rRNA gene sequence revealed that GT312 had 99.93% similarity with Bacillus amyloliquefacien type strain FZB42 T.Based on the morphologic and physiological characteristics,strian GT312 was identified as Bacillus amyloliquefaciens.