Fagerlund Annette, Lindbäck Toril, Storset Anne K, Granum Per Einar, Hardy Simon P
Department of Food Safety and Infection Biology, Norwegian School of Veterinary Science, PO Box 8146 Dep., N-0033 Oslo, Norway.
Microbiology (Reading). 2008 Mar;154(Pt 3):693-704. doi: 10.1099/mic.0.2007/014134-0.
The mechanism by which Bacillus cereus causes diarrhoea is unknown. Three putative enterotoxins have been proposed, haemolysin BL (Hbl), cytotoxin K and non-haemolytic enterotoxin (Nhe). Both Hbl and Nhe are three-component cytotoxins and maximal cytotoxicity of Nhe against epithelia is dependent on all three components. However, little is known of the mechanism of cytotoxicity. Markers of plasma membrane disruption, namely propidium iodide uptake, loss of cellular ATP and release of lactate dehydrogenase (LDH), were observed in epithelia exposed to Nhe from culture supernatants of B. cereus, but not in those exposed to supernatants from a mutant strain lacking NheB and NheC. Consistent with an exogenous cause of membrane damage, purified Nhe components combined to form large conductance pores in planar lipid bilayers. The inhibition of LDH release by osmotic protectants and the increase in cell size caused by Nhe indicate that epithelia lyse following osmotic swelling. Nhe and Hbl show sequence homology, and Hbl component B has remarkable structural similarities to cytolysin A (ClyA), with both structures possessing an alpha-helix bundle and a unique subdomain containing a hydrophobic beta-hairpin. Correspondingly, we show that Nhe has haemolytic activity against erythrocytes from a variety of species. We propose that the common structural and functional properties indicate that the Hbl/Nhe and ClyA families of toxins constitute a superfamily of pore-forming cytotoxins.
蜡样芽孢杆菌引起腹泻的机制尚不清楚。已提出三种假定的肠毒素，即溶血素BL（Hbl）、细胞毒素K和非溶血肠毒素（Nhe）。Hbl和Nhe都是三组分细胞毒素，Nhe对上皮细胞的最大细胞毒性取决于所有三个组分。然而，对细胞毒性机制知之甚少。在暴露于蜡样芽孢杆菌培养上清液中的Nhe的上皮细胞中观察到质膜破坏的标志物，即碘化丙啶摄取、细胞ATP丧失和乳酸脱氢酶（LDH）释放，但在暴露于缺乏NheB和NheC的突变株上清液的上皮细胞中未观察到。与膜损伤的外源性原因一致，纯化的Nhe组分在平面脂质双分子层中结合形成大电导孔。渗透保护剂对LDH释放的抑制以及Nhe引起的细胞大小增加表明上皮细胞在渗透性肿胀后裂解。Nhe和Hbl显示出序列同源性，Hbl组分B与溶细胞素A（ClyA）具有显著的结构相似性，两者的结构都具有α-螺旋束和一个包含疏水β-发夹的独特亚结构域。相应地，我们表明Nhe对多种物种的红细胞具有溶血活性。我们提出，共同的结构和功能特性表明Hbl/Nhe和ClyA毒素家族构成了一个成孔细胞毒素超家族。
