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首页> 外文期刊>Infection and immunity >Characterization of Anthrolysin O, the Bacillus anthracis Cholesterol-Dependent Cytolysin
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Characterization of Anthrolysin O, the Bacillus anthracis Cholesterol-Dependent Cytolysin

机译：炭疽芽孢杆菌炭疽杆菌胆固醇依赖性胞溶素的表达

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We characterized the expression of a putative toxin of Bacillus anthracis, a member of the cholesterol-dependent cytolysin (CDC) family, which includes listeriolysin O, perfringolysin O, and streptolysin O. We named this cytotoxin anthrolysin O (ALO). Although B. anthracis expresses minimal hemolytic activity in clinical settings, we show that Sterne strain 7702 expresses hemolytic activity when grown in brain heart infusion broth or in other rich bacteriologic media, but it secretes barely detectable amounts of hemolysin when grown in Luria-Bertani (LB) broth. Glucose supplementation of LB broth increases the amount of secreted hemolytic activity. Expression of hemolytic activity is maximal during mid- to late-log phase and decreases in the stationary phase. These observations are supported, in part, by semiquantitative reverse transcriptase PCR of alo mRNA. Hemolytic activity in growth supernatants was increased in the presence of reducing agent and almost totally inhibited in a dose-dependent manner by cholesterol; both of these activities are characteristic of a CDC toxin. A mutant of Sterne strain 7702, strain UT231, in which the alo gene was deleted and replaced by a kanamycin cassette, secreted barely detectable hemolytic activity into the growth medium. When strain UT231 was complemented in trans with native alo on a low-copy-number plasmid [strain UT231(pUTE554)], it regained the ability to secrete hemolytic activity, indicating that ALO is the major hemolysin secreted by this strain of B. anthracis in rich media in vitro. To further support the alo gene product being a hemolysin, recombinant B. anthracis ALO (rALO) purified from Escherichia coli was extremely active against washed human erythrocytes, with complete hemolysis detected at ～30 molecules of rALO per erythrocyte. Considering the virulence roles of CDCs for other gram-positive bacteria, we speculate that ALO may have a role in anthrax virulence.
机译：我们表征了炭疽杆菌的推定毒素的表达，该毒素是胆固醇依赖性溶细胞素（CDC）家族的成员，其中包括李斯特菌溶血素O，穿孔素溶血素O和链球菌溶血素O。 （ALO）。虽然 B。炭疽在临床环境中仅表现出最小的溶血活性，我们显示，Sterne菌株7702在脑心浸液或其他丰富的细菌培养基中生长时具有溶血活性，但在Luria-Bertani中生长时几乎无法检测到溶血素的分泌（LB）汤。葡萄糖补充LB肉汤可增加分泌的溶血活性。在对数中期到后期，溶血活性的表达最大，而在稳定期则降低。这些观察结果部分得到 alo mRNA的半定量逆转录酶PCR的支持。在存在还原剂的情况下，生长上清液中的溶血活性增加，并且几乎完全被剂量依赖性的胆固醇抑制。这两种活性都是CDC毒素的特征。斯特恩（Sterne）菌株7702突变株UT231，其中的 alo 基因被删除并被卡那霉素盒替代，在生长培养基中几乎无法检测到溶血活性。在低拷贝数质粒[UT231（pUTE554）]上，当UT231菌株在 trans 中用天然 alo 进行补充时，它恢复了分泌溶血活性的能力，表明ALO是这种 B菌株分泌的主要溶血素。炭疽病为了进一步支持 alo 基因产物为溶血素，重组 B。从大肠杆菌中纯化出的炭疽ALO（rALO）对洗涤后的人红细胞具有极强的活性，每个红细胞中约有30个rALO分子可检测到完全溶血。考虑到CDC对其他革兰氏阳性细菌的毒力作用，我们推测ALO可能在炭疽毒力中起作用。

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