表皮葡萄球菌agrC特异结合多肽对聚氯乙烯材料表面细菌生物膜形成的作用研究

发布时间：2018-04-15 18:57

  本文选题：表皮葡萄球菌 + agrC　； 参考：《昆明医科大学》2017年硕士论文

【摘要】：[目的]研究表皮葡萄球菌agrC特异结合多肽对聚氯乙烯材料表面细菌生物膜形成的作用,为合成agrC为靶点治疗表皮葡萄球菌生物膜形成相关感染的特异性药物奠定基础。[方法]1.采用微量板半定量法确定agrC特异结合多肽的最佳抑菌浓度;2.采用微量板半定量法观察agrC特异结合多肽对表皮葡萄球菌不同生长时期生物膜形成能力的影响;3.利用扫描电镜观察PVC材料表面细菌生物膜的表面结构;4.利用激光共聚焦显微镜观察PVC材料表面细菌群落数量、细菌生物膜形成的厚度和细菌生物膜的三维结构。[结果]1.微量板半定量法结果显示表皮葡萄球菌agrC特异结合多肽的最佳抑菌浓度为 800μg/ml;2.微量板半定量法结果显示当agrC特异结合多肽浓度为800 μg/ml时,其在12h时对细菌生物膜的形成有明显抑制作用;3.扫描电镜观察显示表皮葡萄球菌ATCC35984在培养6h时,实验组与对照组均可见散在小的细菌团块;在培养12h时,实验组细菌团块明显少于对照组;在18h时两组均可见细菌团块相互连接成片状、塔状结构;在24h时两组均可见成熟生物膜结构,结构中可见大量无定形细胞外基质填充;在30h时,两组可见生物膜结构,但有部分呈分解状。表皮葡萄球菌ATCC12228在培养6、12、18、24和30h时,实验组和对照组细菌均未见生物膜形成。 ,4.激光共聚焦显微镜观察显示,实验组加入agrC特异结合多肽与对照组加入无关肽,培养12h时观察实验组PVC材料表面细菌明显少于对照组,且以死菌居多;而在6、18、24h两组细菌量未见明显差别,均以活菌居多。培养12h时观察实验组细菌生物膜厚度明显少于对照组,其余时间点生物膜厚度未见明显差别。[结论]1.表皮葡萄球菌与agrC特异结合多肽的抑菌强度之间存在剂量效应关系;2. agrC特异结合多肽在生物膜形成的聚集阶段对表皮葡萄球菌生物膜的形成有明显抑制作用:agrC特异结合多肽能抑制表皮葡萄球菌生物膜的形成能力;agrC特异结合多肽可以阻碍表皮葡萄球菌进一步聚集,使细菌团块疏松,抑制生物膜的形成;agrC特异结合多肽可以在细菌生物膜形成的聚集阶段抑制细菌增殖,加速细菌死亡,影响细菌生物膜形成的厚度。3. agrC特异结合多肽可以为治疗表皮葡萄球菌生物膜形成相关感染的特异性药物奠定基础,它作为预防用药可能更为有效,对成熟生物膜的抑制作用不明显。
[Abstract]:[objective] to study the effect of agrC binding polypeptide of Staphylococcus epidermidis on bacterial biofilm formation on the surface of polyvinyl chloride (PVC), so as to lay a foundation for the synthesis of agrC as a target for the treatment of staphylococcus epidermidis biofilm formation related infection.[methods] 1.The optimum inhibitory concentration of agrC specific binding peptide was determined by microplate semi-quantitative method.The effect of agrC specific binding peptide on biofilm formation of Staphylococcus epidermidis at different growth stages was observed by microplate semi-quantitative method.The surface structure of bacterial biofilm on the surface of PVC was observed by scanning electron microscope (SEM).The number of bacterial community, the thickness of bacterial biofilm formation and the three-dimensional structure of bacterial biofilm on the surface of PVC were observed by laser confocal microscope.[result] 1.The results of microplate semi-quantitative analysis showed that the best inhibitory concentration of agrC specific binding polypeptide was 800 渭 g / ml ~ (2) 路ml ~ (-1) 路L ~ (-1) 路L ~ (-1).The results of microplate semi-quantitative assay showed that when the concentration of specific binding peptide of agrC was 800 渭 g/ml, it could inhibit the formation of bacterial biofilm significantly at 12 h.Scanning electron microscope (SEM) observation showed that the ATCC35984 of Staphylococcus epidermidis was scattered in the small bacterial mass in both the experimental group and the control group at 6 h, and at 12 h, the bacterial mass in the experimental group was significantly lower than that in the control group.At 18 h, the bacterial masses were connected to each other into flakes and tower structures, mature biofilm structures were observed in both groups at 24 h, and a large number of amorphous extracellular matrices were found in the structures. At 30 h, biofilm structures were observed in both groups.But some of them are decomposed.No biofilm formation was observed in the experimental group and control group after culture of Staphylococcus epidermidis ATCC12228 at 612U 1824 and 30h.Laser confocal microscopy showed that the number of bacteria on the surface of PVC in the experimental group was significantly lower than that in the control group, and the number of dead bacteria was more than that in the control group after the addition of specific binding peptide of agrC to the control group and the addition of unrelated peptides to the control group.However, there was no significant difference in bacterial quantity between the two groups at 6: 18 ~ 24 h, and most of them were live bacteria.The bacterial biofilm thickness in the experimental group was significantly lower than that in the control group at 12 h, but there was no significant difference in the other time points.[conclusion] 1.There is a dose-effect relationship between the bacteriostatic strength of Staphylococcus epidermidis and agrC specific binding polypeptide 2.The inhibitory effect of agrC specific binding polypeptide on the formation of biofilm of Staphylococcus epidermidis during the aggregation stage of biofilm formation is significant.Hetero-binding polypeptides can inhibit the formation of staphylococcus epidermidis biofilm. The agrC specific binding peptide can inhibit the further accumulation of Staphylococcus epidermidis.The agglomeration and inhibition of the formation of biofilm can inhibit the proliferation of bacteria and accelerate the death of bacteria during the aggregation stage of biofilm formation.The thickness of bacterial biofilm formation. 3. The specific binding polypeptide of agrC can lay a foundation for the treatment of biofilm formation related infection of Staphylococcus epidermidis, and it may be more effective as a preventive drug.The inhibitory effect on mature biofilm was not obvious.
【学位授予单位】：昆明医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R515

【参考文献】

相关期刊论文 前10条

1 成鹏;叶联华;;agrC对表皮葡萄球菌生物膜形成作用机制的研究进展[J];国际生物医学工程杂志;2016年04期

2 王曦;黄云超;周永春;刘德权;吕志勇;叶联华;陈颖;杨加鹏;成鹏;王正玺;;雌二醇对乳腺癌假体植入术后表皮葡萄球菌生物膜形成的影响研究[J];中国修复重建外科杂志;2016年07期

3 邱良婷;叶联华;黄云超;;生物材料植入感染与表皮葡萄球菌生物膜形成的相关基因调控[J];国际生物医学工程杂志;2016年01期

4 杨永长;肖代雯;陈亮;刘华;喻华;黄文芳;;临床分离表皮葡萄球菌附属基因调节因子agr基因多态性分析[J];中国实验诊断学;2015年04期

5 叶联华;黄云超;杨达宽;赵光强;刘馨;郭凤丽;周友全;;表皮葡萄球菌ica操纵子与聚氯乙烯材料表面细菌生物膜形成的关系[J];中华医院感染学杂志;2010年24期

6 叶联华;许赓;黄云超;郭凤丽;赵光强;雷玉洁;;溴代呋喃酮对胸心外科聚氯乙烯材料表面表皮葡萄球菌生物膜形成的影响[J];中国胸心血管外科临床杂志;2010年04期

7 叶联华;黄云超;李高峰;杨达宽;张勇;巫正伟;;表皮葡萄球菌生物被膜结构观察[J];中华微生物学和免疫学杂志;2009年07期

8 唐俊妮;周锐;王红宁;曾志光;;附属基因调节子在葡萄球菌生物被膜形成与感染中的作用[J];中南大学学报(医学版);2008年11期

9 欧元祝;朱于莉;陈洁敏;秦智强;江娟;杨晓梅;瞿涤;;表皮葡萄球菌AtlE蛋白介导生物膜起始黏附的相关机制[J];复旦学报(医学版);2006年05期

10 朱于莉;杨晓梅;李娜;江娟;陈洁敏;秦智强;李敏;吕元;魏武;瞿涤;;trap基因表达在表皮葡萄球菌生物膜形成及附属基因调节(agr)系统活化中的作用[J];复旦学报(医学版);2006年05期

相关博士学位论文 前3条

1 李学军;金黄色葡萄球菌agrC结合小肽的筛选及其功能研究[D];第三军医大学;2009年

2 王重振;ClpP蛋白酶在表皮葡萄球菌生物膜形成中的作用[D];复旦大学;2008年

3 李敏;附属基因调节系统（agr）-Ⅰ型对表皮葡萄球菌生物膜形成调节关系的研究[D];复旦大学;2005年

相关硕士学位论文 前1条

1 成鹏;表皮葡萄球菌agrC蛋白原核表达载体的构建及蛋白表达[D];昆明医科大学;2016年

，

本文编号：1755366