溶血菌毒

Methods The clpE-deficient strain was constructed by LFH-PCR and identified by PCR and sequencing. The impact of clpE mutant on the virulence of S. pneumoniae was evaluated in a mouse model. In addition, we also studied the effect of clpE mutant on adherence and invasion of host cells. Real time RT-PCR was used to measure the mRNA expression levels of autolysin A, pneumococcal surface adhesion A, pneumolysin, pneumococcal surface protein A and neuraminidase The clpE gene was replaced completely by erm cassette.

用长臂同源多聚酶链式反应方法失活clpE基因，用PCR、测序鉴定缺失菌株，通过动物实验观察clpE基因缺失株毒力改变情况，同时用细胞实验比较clpE基因缺失株和野生菌对宿主细胞的粘附和侵袭能力，最后用实时荧光定量PCR分析自溶素(major autolysin A，lytA)、表面黏附素A(pneumococcal surface adhesion A，psaA)、溶血素(pneumolysin，ply)、肺炎球菌表面蛋白A（pneumococcal surface protein A， pspA和神经氨酸酶（neuraminidase， nanA）的表达。

Morphological conversion from spiralto coccoid forms occurred when HP was cultured with selenium and most of coccoid forms had fragmentary flagella and pilus; the effection of selenium to activity of urease, catalase, alkaline phosphatase is not exist or not evidence; selenium enable pET32a-vacA-E.coli BL21 which can express VacA produce yellow lipochrome, and the color become deeper with increasing of concentration of selenium; selenium can decrease expression of recombined VacA by restrain the growth of pET32a-vacA-E.coli,but can not influence activity and gene sequence of VacA; selenium can enhance the suppression effect of recombined VacA for gastric cancer cell, depress the index of cell multiplication and the forming rate of colony, decrease the number of cell in S period, increase total of cell in G period; After purified, the recombined VacA can be identified by VacA antibody.

体外实验结果表明：一定浓度的硒作用HP后，HP菌落数量随着硒浓度的增加而减少，菌落颜色由灰白色变为砖红色，溶血现象随硒浓度的增加和作用时间的延长而逐渐消失，HP的形态由弯曲形变为球形或椭圆形，鞭毛及菌毛出现脱落现象；硒对尿素酶、过氧化氢酶、碱性磷酸酶活性无影响或影响不明显；一定浓度的硒能使表达VacA的pET32a-vacA-E.coli BL21工程菌产生脂溶性砖红色色素，颜色随硒浓度增加而加深；一定浓度的硒通过抑制pET32a-vacA-E.coli BL21工程菌的生长，减少重组VacA表达量，但不影响其空泡毒活性及编码基因序列；纯化后的VacA重组蛋白可被VacA抗体识别，具有良好的抗原性。

We also discovered that selenium can decrease hematolysis and effects function of vacuole toxin of HP. morphological conversion from spiralto coccoid forms occurredwhen HP was caltured with selenium and most of coccoid forms had fragmentary flagella and pilus; the effection of selenium to activity of urease, catalase, alkaline phosphatase is not exist or not evidence; selenium enable pET32a-vacA-E. coli BL21 which can express VacA produce yellow lipochrome, and the color become deeper with increasing of concentration of selenium; selenium can decrease expression of recombined VacA by restrain the growth of pET32a-vacA-E. coli, but can not influence activity and gene equence of VacA; selenium can enhance the suppression effect of recombined VacA for gastric cancer cell, depress the index of cell multiplication and the forming rate of colony, decrease the number of cell in S period, increase total of cell in G period; After purified, the recombined VacA can be identified by VacA antibody.

结果：体外实验结果表明：一定浓度的硒作用HP后，HP菌落数量随着硒浓度的增加而减少，菌落颜色由灰白色变为砖红色，溶血现象随硒浓度的增加和作用时间的延长而逐渐消失，HP的形态由弯曲形变为球形或椭圆形，鞭毛及菌毛出现脱落现象；硒对尿素酶、过氧化氢酶、碱性磷酸酶活性无影响或影响不明显；一定浓度的硒能使表达VacA的pET32a-vacA-E.coli BL21工程菌产生脂溶性砖红色色素，颜色随硒浓度增加而加深；一定浓度的硒通过抑制pET32a-vacA-E.coli BL21工程菌的生长，减少重组VacA表达量，但不影响其空泡毒活性及编码基因序列；硒能增强重组VacA对胃癌细胞的抑制作用，使细胞分裂指数和集落形成率降低、S期细胞数量减少，G1期细胞总数增加；纯化后的VacA重组蛋白可被VacA抗体识别，具有良好的抗原性。

ABSTRACT: Objective To investigate the relation between enterococcus hemolysin and its virulence.

目的 探讨肠球菌溶血素与毒力的关系。

These results implied that hemolysin may contribute to the virulence of enterococci.

肠球菌的溶血素可能与其毒力有关。

The results showed that when the proportion of the resistant subpopulation within the h-VRSA was low, te level of the toxic factor and its pathogenicity to animals had no difference from that of the standard strain. If proportion of the resistant subpopulation increased the level of coagulase and lecithinase changing from positive from negative and the ring of hemolysis disappeared. The speed of proliferation of the organisms in the animal slowed down and the LD50 increased.

结果 当试验菌株中耐药亚群的比例较少时，其毒力因子及动物致病性检测结果与金葡菌标准菌株无显著差异；随着试验菌株中耐药亚群的比例增加，表现为血浆凝固酶、卵磷脂酶由阳性转为阴性，血琼脂平板上溶血环消失，细菌在动物体内增殖速度减慢，对动物的半数致死量(LD50)增加。

Methods Four pairs of primers were designed from toxin sub-unit A gene, O139 special gene, Haemolytic subunit A gene and Toxin coregulated pilus subunit A gene. Multiplex PCR was established to simultaneously detect those four genes.

分别针对霍乱弧菌肠毒素A亚单位基因、O139群特异基因、霍乱弧菌溶血素A亚单位基因和毒力协同调节菌毛A亚单位基因设计引物，建立多基因PCR方法。

I didn't watch TV last night, because it .

昨晚我没有看电视，因为电视机坏了。

Since this year, in a lot of villages of Beijing, TV of elevator liquid crystal was removed.

今年以来，在北京的很多小区里，电梯液晶电视被撤了下来。