缓症链球菌外毒素的致病性研究

A study on the pathogenesis of Streptococcus mitis exotoxin

目的 从中毒性休克综合征爆发流行现场获取的病原菌 ,进行增菌培养 ,从病原菌分泌物中分离、纯化外毒素蛋白 ,以对其致病性进行实验研究。方法 将现场分离的缓症链球菌增菌培养后 ,所得上清液分别经 2 0 %、40 %、6 0 %、80 %硫酸铵盐析 ,进一步经快速蛋白液相色谱系统 (FPLC)最终纯化 ,分别皮下注射新西兰兔 ,观察有否发热。引起新西兰兔对大肠杆菌内毒素致死性休克的敏感性实验 :先给实验组新西兰兔皮下注射纯化的链球菌外毒素 10 μg ,4h后静脉注射 10 μg大肠杆菌内毒素 ,对照组仅注射 10 μg大肠杆菌内毒素。 结果 经FPLC最终纯化的 2 0 %硫酸铵盐析蛋白 (分子量 340 0 0 )有致热性 ,可使新西兰兔平均体温升高 1℃左右 ,病理见脾脏有丝分裂增加。纯化的链球菌外毒素可增加新西兰兔对大肠杆菌内毒素致死性休克的敏感性 :实验组动物出现休克的临床表现 ,分别于 16～ 2 9h内死亡 ;而仅注射 10 μg大肠杆菌内毒素的对照组动物无上述表现。

Objective To study the isolation, purification and pathogenesis of Streptococcus mitis pyrogenic exotoxin causing toxic shock syndrome. Methods Streptococcus mitis isolated from patients′ throat were shaking cultivated. After being centrifuged, the supernatant fluid of the culture was precipitated with 20%, 40%, 60%, 80%(NH 4) 2SO 4 respectively and the fast protein liquid chromatography(FPLC) was used for the final step of purification. Rabbits receiving subcutaneous injection with respective purified proteins were monitored daily for fever. The ability of the purified proteins to enhance the susceptibility of the rabbits to lethal Escherichia coli endotoxin shock is recorded, when the endotoxin was injected intravenously 4 hours after administration of 10 μg Streptococcus mitis exotoxin. Results Only the protein precipitated by 20% (NH 4) 2SO 4 (molecular weight is 34 000) from culture supernatant fluid was pyrogenic for rabbits(average temperature increase near 1℃), and it can also cause the proliferation of rabbit splenocytes(mitogenicity). All the animals receiving subcutaneous injection of exotoxin containing purified proteins precipitated with higher concentrations of (NH 4) 2SO 4 died within 16～29 hours after intravenous injection of the Escherichia coli endotoxin,demonstrating the enhanced susceptibility of the animals to lethal endotoxin shock. The control rabbits displayed none of these effects. Conclusion Streptococcus mitis exotoxin is a novel streptococcal pyrogenic exotoxin.