Studies on hemolysin of Anginosus group streptococci isolated from the human oral cavity

人口腔分离的心绞痛群链球菌溶血素的研究

• 批准号: 11671826
• 负责人: NAKAMURA Takeshi
• 金额: $ 1.22万
• 依托单位: Matsumoto Dental University, School of Dentistry
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11671826/
• 关键词: Oral streptococci; Anginosus group; S.anginosus; Hemolysin; Cytokine; 精製

Total 15 β hemolytic streptococcal strains were isolated from subgingival plaque samples (5 from patients with adult periodontal disease and 10 from healthy adults). Of these strains, 13 strains exhibited streptogram code of 0110434, indicating that they have close properties of reference strain of Streptococcus anginosus NCTC 10713. Furthermore, 12 strains out of the 13 ones reacted with anti G group streptococcal serum in slide latex agglutination test of Strept LA.These observations suggest that the isolated β hemolytic microorganisms are identical to Streptococcus anginosus which are considered to distribute widely in the oral cavity. The hemolysin of this strain was detected in the extracellular culture fluids and the much more production was observed in general anaerobic medium (GAM) than in brain heart infusion or Todd Hewitt broth.Purification of the hemolysin in the culture supernatant was carried out by concentration by ammonium sulfate (60% saturation), ion-exchange chromatography, gel filtration, and hydroxyapatite chromatography by 625 fold with the recovery of 33.6%. The purified hemolysin formed diffuse bands between 50 to 60 kDa in SDS-PAGE.Isoelectric point was 4.2 and it was inactivated by heating at 70℃ for 10 min. The optimum pH for hemolysis was 7.5. The activity was enhanced 2-3 fold by 2 mercaptoethanol or dithiothreitol. The hemolytic spectrum of erythrocytes was found to be wide ; human, bovine, horse, sheep, guinea pig, and rabbit erythrocytes were lyzed at almost the same degree by this hemolysin. The other properties were quite similar to those of S.anginosus NCTC 10713.Both purified hemolysin samples from the isolated strain and reference strain (NCTC 10713) induced the production of TNF-α and IL-6 by peritoneal macrophages of BALB/c mice.

总共从尺寸斑块样品中分离出15个β溶血链球菌菌株（成人牙周疾病患者5例，健康成年患者10例）。在这些菌株中，有13个菌株暴露于0110434的链球菌代码，表明它们具有链球菌链球菌的参考菌株NCTC NCTC 10713的近距离菌株。此外，在13个菌株中，有12种与抗G组链球菌血清反应的13个菌株在Slide slide slide乳胶凝集测试链球菌的链球菌中反应。这些观察结果表明，孤立的β溶解微生物与链球菌的链球菌相同，这些链球菌被认为在口腔中分布广泛。在细胞外培养液中检测到该菌株的血素蛋白，并且在一般的厌氧培养基（GAM）中观察到的产生要多得多，而不是在大脑心脏输注或Todd Hewitt Groth中观察到。通过硫酸铵（60％满意度），离子 - 交换色谱，凝胶过滤和羟基磷灰石色谱法通过625倍以33.6％的恢复，通过硫酸铵，离子 - 交换色谱，凝胶过滤和羟基磷灰石色谱法对培养上清液中的纯化纯化。在SDS-PAGE的50至60 kDa之间形成纯化的血素蛋白形成的弥漫带。异电点为4.2，并通过在70℃中加热10分钟而被灭活。溶血的最佳pH值为7.5。 2-3倍通过2倍乙醇或二硫代硫醇增强了2-3倍。发现红细胞的溶血光谱宽。人类，牛，马，绵羊，几内亚猪和兔红细胞几乎通过这种hemolysin lyz。其他特性与NCTC链球菌10713的特性非常相似。从隔离菌株和参考应变（NCTC 10713）中纯化的纯种血素蛋白样品（NCTC 10713）诱导了BALB/C小鼠的腹膜巨噬细胞的TNF-α和IL-6的产生。