一株耐硒壶瓶碎米荠内生菌分离、鉴定及其体外硒代谢研究

【背景】壶瓶碎米荠对硒具有超积累能力,并主要以硒代胱氨酸的形式存在,与已有的硒超积累植物显著不同,其硒超积累机制不明。【目的】从硒超积累植物壶瓶碎米荠(Cardamine hupingshanensis)体内分离耐硒内生菌,并对其进行鉴定和体外硒代谢特征研究,为壶瓶碎米荠超积累硒的机制研究提供参考。【方法】从壶瓶碎米荠新鲜叶片中分离纯化耐硒内生菌株,对其进行生理生化特征及16S rRNA基因序列分析鉴定,并对其进行亚硒酸钠培养代谢。【结果】获得一株耐硒内生菌CSN-1,被鉴定为甲基营养型芽孢杆菌(Bacillus methylotrophicus),培养液中硒含量低(Se 1.5 mg/L)时其吸光度值较对照组高,硒含量高(Se 10 mg/L)时其吸光度值较对照组低;代谢后的上清液中硒主要以Se~(4+)存在,而菌体中硒主要是硒代胱氨酸(SeCys_2)。【结论】硒超积累植物壶瓶碎米荠叶片体内存在甲基营养型芽孢杆菌(Bacillus methylotrophicus)CSN-1,具有将亚硒酸钠转化为硒代胱氨酸的能力,低浓度的硒对该内生菌的生长具有一定的促进作用,而高浓度的硒则会抑制该内生菌的生长。

Isolation, identification and selenite metabolism of a selenium-tolerant endophyte from Cardamine hupingshanensis

[Background] Cardamine hupingshanensis is a novel selenium (Se)-hyperaccumulating plant in China with the predominate Se species as selenocystine (SeCys2), which is different with other Se-hyperaccumulating plants in USA, eg. Astragalus bisulcatus, Stanleya pinnata. [Objective] To explore the role of microorganism during Se-hyperaccumulation in Cardamine hupingshanensis, isolation, identification and Se metabolism in vitro of a Se-tolerant endophyte from C. hupingshanensis were conducted. [Methods] A Se-tolerant endophyte was isolated and purified from fresh leaves of C. hupingshanensis as CSN-1. The physiological and biochemical analysis and 16S rRNA gene sequence analysis were carried out. Moreover, the endophyte CSN-1 was cultured in selenite medium to study its Se metabolism. [Results] The Se-tolerant endophyte CSN-1 was identified as Bacillus methylotrophicus. The selenite culture revealed that the medium absorbance was higher than that in controls under 1.5 mg Se/L medium, but lower under 10 mg Se/L medium. The metabolized Se speciation was Se4+ in supernants, but SeCys2 in bacteria deposits. [Conclusion] The Se-tolerant endophyte Bacillus methylotrophicus CSN-1 presented in Se-hyperaccumulating plant, Cardamine hupingshanensis, and could transform selenite into SeCys2. The low concentration of Se could promote the growth of Bacillus methylotrophicu CSN-1, but negative effects could be observed at high Se levels.