Häussler S, Nimtz M, Domke T, Wray V, Steinmetz I
Institute of Medical Microbiology, Hannover Medical School, Germany.
Infect Immun. 1998 Apr;66(4):1588-93. doi: 10.1128/IAI.66.4.1588-1593.1998.
Burkholderia pseudomallei is the causative agent of melioidosis, an infectious disease, which is increasingly recognized as an important public health problem in various tropical regions. This study describes the identification and characterization of a heat-stable extracellular toxin of B. pseudomallei. After cultivation of B. pseudomallei in liquid media, the heated cell-free supernatant was concentrated by ultrafiltration. The concentrate exhibited a cytotoxic and hemolytic activity which showed remarkable resistance against alkaline and acidic treatments. For further purification, reversed-phase chromatography using a fast-performance liquid chromatography system was performed. After elution with an acetonitrile gradient, a single cytotoxic and hemolytic peak was detected. Structural characterization of the toxin was performed by a combination of mass spectrometric and nuclear magnetic resonance spectroscopic techniques. A highly purified glycolipid, 2-O-alpha-L-rhamnopyranosyl-alpha-L-rhamnopyranosyl-beta-hydroxytetradec anoyl-beta-hydroxytetradecanoate (Rha-Rha-C14-C14), with a molecular mass of 762 Da was identified. The purified exolipid showed a time- and dose-dependent cytotoxic effect on phagocytic (HL60) and nonphagocytic (HeLa) cell lines. In addition, a time- and dose-dependent hemolysis of erythrocytes from various species was observed. The toxin structure makes a detergentlike action most probable. Interestingly, the cytotoxic and hemolytic activities of the glycolipid could be neutralized by albumin. Future studies will concentrate on the role of this exolipid as a virulence factor in the pathogenesis of melioidosis.
类鼻疽伯克霍尔德菌是类鼻疽的病原体，类鼻疽是一种传染病，在各个热带地区日益被视为一个重要的公共卫生问题。本研究描述了类鼻疽伯克霍尔德菌一种热稳定细胞外毒素的鉴定和特性。将类鼻疽伯克霍尔德菌在液体培养基中培养后，经加热的无细胞上清液通过超滤浓缩。浓缩物表现出细胞毒性和溶血活性，对碱性和酸性处理具有显著抗性。为进一步纯化，使用快速高效液相色谱系统进行反相色谱分析。用乙腈梯度洗脱后，检测到一个单一的细胞毒性和溶血峰。通过质谱和核磁共振光谱技术相结合对毒素进行结构表征。鉴定出一种高度纯化的糖脂，2-O-α-L-鼠李糖基-α-L-鼠李糖基-β-羟基十四烷酰-β-羟基十四烷酸酯（Rha-Rha-C14-C14），分子量为762 Da。纯化的细胞外脂质对吞噬细胞系（HL60）和非吞噬细胞系（HeLa）表现出时间和剂量依赖性的细胞毒性作用。此外，观察到对来自不同物种红细胞的时间和剂量依赖性溶血。毒素结构最有可能产生类似去污剂的作用。有趣的是，该糖脂的细胞毒性和溶血活性可被白蛋白中和。未来的研究将集中于这种细胞外脂质作为类鼻疽发病机制中毒力因子的作用。
