一株产冠菌素新菌种的分离与鉴定

Isolation and identification of a coronatine producing strain

【目的】从不同样本中分离筛选性能稳定的产冠菌素菌株。【方法】根据冠菌素引起植物叶片产生弥散性黄萎病、块茎膨大的特性,采集各种植物病叶、病枝及感病植物的土壤,采用穿刺法与系列稀释法分离筛选菌株;液相色谱测定菌株产生的冠菌素;在电子和光学显微镜下观察菌体形态;根据生理生化试验、(G+C)mol%含量、16S rDNA序列分析等对菌株进行鉴定;对分离提纯的发酵产物进行紫外、质谱和红外分析。【结果】菌株BBC933为革兰氏阴性菌,端生鞭毛,短杆状,无芽孢。在41℃下不生长,细胞内有聚β-羟基丁酸盐颗粒积累,没有精氨酸双水解酶和氧化酶,不能水解淀粉、明胶,不进行硝酸还原及反硝化作用,过氧化氢酶反应呈阳性。菌株的(G+C)mol%含量为67.2%,根据该菌株16S rDNA序列的同源性分析,构建系统发育树。【结论】菌株BCC933鉴定为洋葱伯克霍尔德氏菌(Burkholder cepacia),具有产冠菌素性能。国内外未曾见报道洋葱伯克霍尔德氏菌产冠菌素。

[ Objective] We screened and isolated coronatine-producing stains from various samples. [ Methods] The strains were isolated and selected from samples by the methods of streak plate and serial dilution. The samples were sick leaves/branches and soil in which plants got sick according to the symptoms of leaf blight disease and tuber enlargement. Coronatine production was determined by high performance liquid chromatography （HPLC）. The strain was characterized by the physiological and biochemical analysis,the determination of （G ＋ C） mol% contents and 16S rDNA sequencing. The molecule structure of the fermentation product was identified based on the data of ultraviolet spectrum, infrared spectrum and mass spectrum. [ Results] Strain BCC933 was gram-negative,polar flagella,short rod and non-spore-forming bacterium and accumulated poly-13-hydroxybutyrate （PHB）. It producted catalase, but not arginine dihydrolase nor oxidase,couldn＇t grow at temperature 41℃. It hadn＇t the abilities to hydrolyze starch and gelatine. No nitrate reduction and denitrification activity was detected. The （ G ＋ C） mo1% content was 67.2%. We analyzed 16S rDNA nucleotide sequence, and ascertained the phylogenetic position of the strain. [ Conclusion ] Strain BCC933 with coronatine biosynthesis ability was identified as Burkholderia cepacia,which hasn＇t been reported up to date.