ゲノムバイオロジーを基盤とする黄色ブドウ球菌の分子疫学と危害評価

近年来，许多新的葡萄球菌肠毒素（SEs）被报道。已知某些SE基因与可移动的遗传元件如致病性岛、噬菌体和质粒有关。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。有一种可能性是，这些可移动的遗传元素在金黄色葡萄球菌作为病原体的进化中发挥了重要作用。为了调查这些新发现的SEs的病理生物学和流行病学，我们进行了如下所示的研究。利用4套多重PCR技术建立了18种葡萄球菌超抗原毒素基因的综合检测系统。对69株食物中毒分离株和97株健康人鼻签分离株进行超抗原毒素基因分型，发现32种超抗原毒素基因型，表明许多金黄色葡萄球菌分离株含有多重毒素基因。毒素基因型与毒素基因编码移动遗传元件谱之间关系的进一步分析表明，毒素基因编码移动遗传元件的组合可能在确定金黄色葡萄球菌超抗原毒素基因型中起作用（微生物学通报，46:191-198,2005）。我们利用转座体策略确定了sed、selj和selj编码质粒p196的完整核苷酸序列。该质粒编码青霉素抗性操纵子和cd抗性操纵子，以及三种SE （SE1）基因。我们描述了一种新的葡萄球菌肠毒素（SE）样推定毒素SE1R。SE1R蛋白表现出显著的T细胞刺激活性。MHC II类分子是SER刺激T细胞所必需的。SER刺激T细胞携带受体Vβ3、11、12、13.2和14。这些结果表明SER作为一种超抗原（感染与免疫72:3664,2004）。我们研究了一种新型葡萄球菌肠毒素（SE）样毒素P型（SE1P）的生物学特性。当给药浓度为0.4 pM或更高时，SE1P诱导了大量的增殖反应，并从人T细胞中产生细胞因子IL-2、IFN-γ、TNF-α和IL-4。辅助细胞上MHC II类分子的表达是SE1P刺激T细胞所必需的。SE1P选择性地刺激大量携带受体Vβ 5.1、6、8、16、18和21.3的人T细胞。这些结果表明SE1P作为一种超抗原。SE1P在麝香鼩催吐试验中被证明是催吐的，尽管剂量相对较高（《感染与免疫》，73:in press,2005）。我们构建并表达了无毒突变体SEC (mSEC)，并研究了缺乏超抗原活性的mSEC是否能免疫金黄色葡萄球菌感染。小鼠用mSEC免疫，并用活的金黄色葡萄球菌攻毒。mscs免疫小鼠脏器内细菌计数明显低于对照组，存活率明显高于对照组。mSEC免疫强烈诱导t -辅助性2型抗体、免疫球蛋白G1和免疫球蛋白G2b的产生。无超抗原特性的mSEC免疫对金黄色葡萄球菌感染具有保护作用，其保护作用可能通过IL-10和IL-4诱导和IFN-γ的下调以及SEC中和抗体介导（感染与免疫，73:174-180,2005）。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.We have developed a comprehensive detection system for 18 kinds of classical and newly described staphylococcal superantigenic toxin genes using four sets of multiplex PCR. Superantigenic toxin genotyping of Staphylococcus aureus for 69 food poisoning isolates and 97 healthy human nasal swab isolates revealed 32 superantigenic toxin genotypes and showed that many S.aureus isolates harbored multiple toxin genes. Ana … More lysis of the relationship between toxin genotypes and toxin genes encoding profiles of mobile genetic elements suggests its possible role in determining superantigenic toxin genotypes in S.aureus as combinations of toxin gene-encoding mobile genetic elements (FEMS Microbiology Letters,246 : 191-198,2005).2.We have determined complete nucleotides sequence of sed, selj and selj encoding plasmid, p196, using transposomics strategy. This plasmid is encoding penicillin resistance operon and Cd-resistance operon, in addition to three kinds of SE (SE1) genes.3.We characterized a novel staphylococcal entertoxin (SE)-like putative toxin SE1R. The SE1R protein showed significant T cell stimulation activity. MHC class II molecules were required for T cell stimulation by SER. SER stimulated T cells bearing receptors Vβ3,11,12,13.2, and 14. These results suggested that SER acts as a superantigen (Infection and Immunity 72 : 3664,2004).4.We investigated the biological properties of a novel staphylococcal enterotoxin (SE)-like toxin type P (SE1P). SE1P induced a substantial proliferative response and the production of cytokines IL-2, IFN-γ, TNF-α, and IL-4 from human T cells when administered at a concentration of 0.4 pM or more. The expression of MHC class II molecules on accessory cells was required for T cell stimulation by SE1P. SE1P selectively stimulated a vast number of human T cells bearing receptors Vβ 5.1,6,8,16,18, and 21.3. These results indicated that SE1P acts as a superantigen. SE1P proved to be emetic in the house musk shrew emetic assay, although at a relatively high dose (Infection and Immunity, 73 : in press,2005).5.We constructed and expressed a non-toxic mutant SEC (mSEC) and investigated whether immunization with mSEC, devoid of superantigenic activity, can protect against S.aureus infection. Mice were immunized with mSEC and challenged with viable S.aureus. Bacteria counts in the organs in mSEC-immunized mice were significantly lower and the survival rate was higher than those of the control group. Immunization with mSEC induced strongly the production of T-helper 2 type antibodies, immunoglobulin G1 and immunoglobulin G2b. Immunization with mSEC devoid of superantigenic properties provides protection against S.aureus infection and the protection might be mediated by the IL-10 and IL-4 induction and down-regulation of IFN-γ, as well as SEC neutralizing antibodies (Infection and Immunity, 73 : 174-180,2005). Less