Vibrio sp.形成的微生物被膜对厚壳贻贝稚贝附着的影响

发布时间：2018-06-23 14:25

本文选题：弧菌 + 厚壳贻贝　；参考：《上海海洋大学》2017年硕士论文

【摘要】：厚壳贻贝(Mytilus coruscus)是我国主要的经济养殖贝类,同时也是我国海洋污损生物的主要类群之一,在我国沿海有广大的分布区域。厚壳贻贝主要通过附着在附着基(网绳、潮间带岩石、堤岸设施表等)的表面生活,附着过程对于厚壳贻贝来说是极其重要生命历程,当环境因素发生不利变化时,厚壳贻贝会通过自身行为,切断足丝并重新选择附着基再次附着,因此研究附着基对厚壳贻贝附着的影响对于增进厚壳贻贝的养殖产业和防御厚壳贻贝在污损生物方面的作用均具有重要的意义。以往的研究发现自然微生物被膜是厚壳贻贝附着的高效天然诱导物,且作为自然微生物被膜的重要成员之一的海洋细菌,是影响厚壳贻贝稚贝附着的重要因素。同时以往的研究结果也显示,海洋细菌中具有极大比例的海洋弧菌(Vibrio),海洋弧菌是最早定植在自然微生物被膜表面的细菌类群之一。因此,本文以海洋弧菌和厚壳贻贝为研究对象,探究了海洋弧菌微生物被膜对厚壳贻贝稚贝的附着的影响,并分析了海洋弧菌微生物被膜的代谢产物中蛋白及糖的差异。主要包括下列内容:1.不同来源海洋弧菌形成的微生物被膜对厚壳贻贝稚贝附着的影响为探讨广泛存在于近海环境中的海洋弧菌和贝类附着的相互作用关系,本研究通过从自然微生物被膜和厚壳贻贝成体肠道内分离了海洋弧菌,测定其种属及亲缘性,调查了这些不同来源弧菌形成的微生物被膜与厚壳贻贝稚贝附着的调控作用。结果表明,测试弧菌形成的微生物被膜密度随着初始细菌密度的增加呈现不同程度的增加。源于自然微生物被膜和贻贝肠道内的10株弧菌均能显著促进厚壳贻贝稚贝的附着,不同菌株形成微生物被膜的诱导活性存在显著差异,稚贝在Vibrio sp.17微生物被膜上的附着率最高,为67%±2%,附着率变化范围为17%-67%。微生物被膜对稚贝附着诱导活性与被膜密度呈显著相关性,其中弧菌Vibrio crassostreae ECSMB14106的相关性系数最高,为0.8992。此外,微生物被膜的诱导活性与弧菌的来源无关。本实验初步探明了海洋弧菌对厚壳贻贝稚贝附着的影响,有助于进一步理解微生物被膜调控厚壳贻贝的附着分子机理。2.不同稚贝诱导活性的弧菌微生物被膜生物学特性分析实验选取10~8 cells/ml的密度下形成的微生物被膜,结果发现诱导活性最高的菌株为Vibrio sp.17,其次为V.crassostreae ECSMB14106、第三为Vibrio sp.13、最低为Vibrio sp.19。本研究应用了共聚焦显微技术、电子显微镜扫描技术、SDS-PAGE蛋白分析技术以及胞外多糖染色技术分析了四株弧菌形成微生物被膜的生物学特性。共聚焦结果发现四株细菌形成的被膜膜厚存在显著差异,膜厚与稚贝附着相关。扫描电镜结果发现菌株Vibrio sp.17和V.crassostreae ECSMB14106的细菌形态呈短圆杆状,菌株Vibrio sp.19呈长杆状,而菌株Vibrio sp.13生长聚集成花瓣状。SDS-PAGE蛋白技术分析菌膜总蛋白发现,四株细菌的菌膜总蛋白种类不同,其中Vibrio sp.17的蛋白条带最多,有31个条带。而胞外多糖染色结果发现,Vibrio sp.17的多糖含量最高。本研究从多个角度分析了诱导活性不同的同种细菌形成的微生物被膜的生物学特性,发现了其膜厚、蛋白差异、胞外多糖含量之间的规律,为探究影响厚壳贻贝稚贝附着的主要诱导因子奠定了一定基础。
[Abstract]:Mytilus coruscus is the main economic culturing shellfish in China, and it is also one of the main groups of marine fouling organisms in China. There are broad distribution areas in the coastal areas of China. The thick shell mussels are mainly surface living by attaching to the attachment base (net, intertidal rock, dike setting and so on), and the attachment process for thick shell mussels It is an extremely important life course. When the environmental factors are disadvantageous, the thick shell mussels will cut off the foot silk and re select the attachments to attach again through their own behavior. Therefore, the effect of the attachment group on the adhesion of the thick shell mussels has the effect on promoting the culture industry of the thick shell mussel and preventing the fouling of the thick shell mussels. It is of great significance. Previous studies have found that natural microorganism membrane is an efficient natural inducer attached to mussels, and marine bacteria, one of the important members of the natural microorganism membrane, is an important factor affecting the attachment of juvenile shells of mussels. Vibrio (Vibrio), Vibrio marine vibrio is one of the earliest bacteria colonized on the surface of natural microbes. Therefore, the effects of Vibrio marine Vibrio and thick shell mussel on the attachment of juvenile marine Vibrio mussels were investigated, and the proteins in the metabolites of the membrane of Vibrio marine Vibrio were analyzed. The main contents of the difference are as follows: 1. the influence of the microorganism formed by different sources of Vibrio marine Vibrio on the attachment of the juvenile shells of mussels is to explore the interaction relationship between marine Vibrio and shellfish, which is widely existed in the near sea environment. This study separates the ocean from the natural microorganism membrane and the thick shell mussels into the intestinal tract. Vibrio, determination of its species and affinity, investigated the regulation of the adhesion of microbes formed by different sources of Vibrio and juvenile shellfish of mussels. The results showed that the membrane density of microbes formed by Vibrio was increased in varying degrees with the increase of initial bacterial density. The 10 strains of Vibrio can significantly promote the attachment of the juvenile shellfish of the mussel. There are significant differences in the induction activity of the biofilm of different strains. The attachment rate of the juvenile shells on the Vibrio sp.17 membrane is the highest, which is 67% + 2%, and the adhesion rate of the adhesion rate is significantly related to the adhesion induced activity of the 17%-67%. microbe to the juvenile shellfish and the density of the membrane. Among them, the correlation coefficient of Vibrio Vibrio crassostreae ECSMB14106 is the highest, which is 0.8992.. The induction activity of the microorganism membrane is not related to the source of Vibrio. This experiment preliminarily explored the effect of Vibrio marine Vibrio on the attachment of the juvenile shells of mussels mussels, which is helpful to further understand the molecular mechanism of the adhesion of the thick shell mussels by the microorganism membrane to understand the molecular mechanism of.2.. The biological characteristics of Vibrio microbes with different juveniles were analyzed by the membrane biological characteristics of 10~8 cells/ml. The results showed that the highest inducible activity was Vibrio sp.17, followed by V.crassostreae ECSMB14106, third was Vibrio sp.13, and the lowest was Vibrio sp.19.. The confocal microscopy was used in this study. Techniques, electron microscope scanning, SDS-PAGE protein analysis and extracellular polysaccharide staining were used to analyze the biological characteristics of the biofilm of four Vibrio strains. The confocal results showed that there were significant differences in the membrane thickness of the four bacteria, and the thickness of the membrane was closely related to the attachment of the larvae. The scanning electron microscope showed that the strain Vibrio sp.17 and V were found. The bacterial morphology of.Crassostreae ECSMB14106 was short and rod-shaped, and the strain Vibrio sp.19 was long rod like, and the strain Vibrio sp.13 grew to integrate the integrated petal.SDS-PAGE protein to analyze the total protein of the mycelia. The total protein species of the four strains of bacteria were different, of which the egg white bands of Vibrio sp.17 were the most and 31 bands were stained, and the extracellular polysaccharide was stained. The results showed that the polysaccharide content of Vibrio sp.17 was the highest. In this study, the biological characteristics of the membrane were analyzed from several angles, and the film thickness, protein difference and the content of extracellular polysaccharide were found. Basics.
【学位授予单位】：上海海洋大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S917.4

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