Comparative genomics of staphylococcal enterotoxin gene family

近年来，许多新的葡萄球菌肠毒素（SE）被报道。已知某些SE基因与移动的遗传元件如致病岛、原噬菌体和质粒相关。这些事实表明，超抗原毒素基因在葡萄球菌菌株之间水平转移。这些移动的遗传元件可能在S.金黄色葡萄球菌作为病原体。为了研究这些新发现的Ses的病理生物学和流行病学，我们进行了以下研究。为了检测和表征致病岛的未知SE基因，我们发展了一种新的方法，称为致病岛扫描。根据几个S.金黄色葡萄球菌中，我们设计的PCR引物扩增整个区域的致病岛。对扩增子进行限制性片段长度多态性（RFLP）分析。利用毒力岛扫描/RFLP技术， ...更多信息 SEB相关的致病岛至少可分为5类。同时，我们还用鸟枪测序法测定了selj/selr编码质粒pF 5的全序列。该质粒除编码selj和selr外，还编码两个新的肠毒素基因ses和set。此外，我们还建立了几种诊断方法，包括SEs的免疫-PCR敏感检测法、S.金黄色葡萄球菌，建立了SE/SE 1基因的多重PCR检测方法.我们确定了两个新的葡萄球菌肠毒素SES和SET。在MHC II^+抗原呈递细胞（APC）存在的情况下，重组（r）SES以T细胞受体（TCR）Vβ9特异性方式特异性刺激人T细胞。rSET在MHC II类^+ APC存在的情况下也能刺激T细胞，尽管在反应性T细胞中未发现其Vβ偏斜。随后，我们研究了SES和SET的催吐活性。我们还研究了SEIR以确定灵长类动物的呕吐活性。这种毒素在以前的研究中被确定，但没有检查灵长类动物是否具有催吐活性。rSES以100 μg/kg剂量灌胃给药5小时内，4只猴中有2只出现呕吐反应。在一只猴中，rSET在100 μg/kg剂量下诱导了延迟反应（给药后24小时），在另一只猴中，反应发生在给药后5天。此外，我们研究了SEA引起的呕吐的机制，使用一个小的呕吐动物模型，家麝鼩（臭鼩）。SEA可诱导肠道5-羟色胺（5-HT）的释放，迷走神经传入神经元上的5-HT_3受体是SEA致吐的关键。少

In recent years, many new staphylococcal enterotoxins (SEs) have been reported. It has been known that certain SE genes are associated with mobile genetic elements such as pathogenicity islands, prophages, and plasmids. These facts imply that superantigenic toxin genes are transferred horizontally between staphylococcal strains. There is a possibility that these mobile genetic elements have played an important role in the evolution of S. aureus as a pathogen. To investigate pathobiology and epidemiology of these newly identified Ses, we had undertaken studies shown in below.1. To detect and characterize unknown SE gene encoding pathogenicity islands, we have developed a new method, named pathogenicity islands scanning. Based on full genome sequences of several S. aureus, we designed PCR primers to amplify entire region of pathogenicity islands. Amplicons were subjected to restriction fragments length polymorphism (RFLP) analysis. Using pathogenicity islands scanning/RFLP, We have found … More that SEB-related pathogenicity islands are classified into at least 5 groups. Also, we have determined complete nucleotide sequence of selj/selr encoding plasmid pF5 using shotgun sequencing strategy. This plasmid is encoding two novel enterotoxin gene, ses and set, in addition to selj and selr. In addition, we developed several diagnostic methods including immuno-PCR sensitive detection of SEs, cultivation/PCR of sensitive detection of S. aureus, and multiplex PCR for comprehensive detection of SE/SE1 genes.2. We characterized two novel staphylococcal entertoxins SES and SET. Recombinant (r)SES specifically stimulated human T cells in T cell receptor (TCR) Vβ9-specific manner in the presence of MHC class II^+ antigen presenting cells (APCs). rSET also stimulated T cells in the presence of MHC class II^+ APC, although its Vβ skewing was not found in reactive T cells. Subsequently, we examined the emetic activity of SES and SET. We also studied SEIR to determine emetic activity in primates. This toxin was identified in previous studies but not examined in terms of possessing emetic activity for primates. rSES induced emetic reactions in 2 of 4 monkeys at a dose of 100 μg/kg within 5 hours of intragastric administration. In one monkey, rSET induced a delayed reaction (24 hours post-administration) at a dose of 100 μg/kg and in the other one, reaction occurred 5 days post-administration. Also, we investigated a mechanism of SEA-induced emesis using a small emetic animal model, House musk shrew (Suncus murinus). SEA induces serotonin (5-hydroxytryptamine, 5-HT) release in intestine, and that the 5-HT3 receptor on vagal afferent neurons are essential for SEA induced emesis. Less