本文選題：宋內(nèi)氏志賀菌　切入點(diǎn)：生物學(xué)特性　出處：《西南大學(xué)》2011年碩士論文　論文類型：學(xué)位論文

【摘要】：志賀氏菌屬(Shigella spp.)是一類能夠通過侵襲腸上皮細(xì)胞引起患者典型性痢疾癥狀(發(fā)熱、腹痛、腹瀉)的食源性細(xì)菌。宋內(nèi)氏志賀菌(S. sonnei)是痢疾流行的主要病原菌之一。由于各個地區(qū)環(huán)境不同,宋內(nèi)氏志賀菌呈現(xiàn)很多差別。本課題通過四株來源于不同的志賀菌(S09071、S09072、S09076、S09077)的比較研究,對志賀菌致病臨床防疫以及流行病學(xué)的研究提供一定的指導(dǎo)意義。通過毒力志賀氏菌大質(zhì)粒缺失突變體庫構(gòu)建可以更好的研究毒力大質(zhì)粒與染色體基因組之間的相互作用,也同時為志賀氏菌致病機(jī)理的研究打下基礎(chǔ)。 論文首先對四株宋內(nèi)氏志賀菌(S. sonnei) S09071、S09072、S09076、S09077從形態(tài)、血清凝集反應(yīng)、毒力評價、生長特征、生理生化特性、耐藥性六個方面做了詳細(xì)比較研究；其次我們做了毒力基因檢測、雙向電泳、全菌蛋白表達(dá)譜的比較,從分子的角度進(jìn)一步比較研究了四株菌；最后對蛋白表達(dá)圖譜的差異蛋白做了MALDI-TOF/TOF質(zhì)譜鑒定。另一方面本論文還建立了志賀菌毒力大質(zhì)粒大片段敲除的方法。通過這些研究,取得了以下主要結(jié)果： (1)四株宋內(nèi)氏志賀菌形態(tài)呈直桿狀,無芽胞、莢膜、鞭毛及菌毛,菌體表面形態(tài)凹凸不平,大小為0.5～0.7×1～2μm。菌落在LB培養(yǎng)基上呈大圓形、扁平、光滑濕潤、無色、半透明、邊緣較整齊,大小約為2.5 mm。四株菌菌體和菌落形態(tài)幾乎無差別。 (2)生長曲線比較發(fā)現(xiàn)S09071生長最快,S09076、S09077其次,S09072菌株生長最慢。 (3)生化特性略有差別,S09076、S09077可以利用鼠李糖、L-巖糖、熊果甙、七葉靈發(fā)酵而S09071、S09072不能利用。 (4)耐藥性比較發(fā)現(xiàn)S09071、S09072比S09076、S09077兩株菌有更強(qiáng)的耐藥性,其中S09072耐藥性最強(qiáng)。 (5)毒力評價實(shí)驗(yàn)發(fā)現(xiàn)只有S09072具有上皮細(xì)胞侵襲能力,而S09071、S09076、S09077則沒有侵襲力。 (6)毒力基因檢測發(fā)現(xiàn)S09071、S09076、S09077部分毒力相關(guān)基因發(fā)生丟失；只有S09072毒力相關(guān)基因完整。 (7)通過比較蛋白質(zhì)組學(xué)研究發(fā)現(xiàn)S09071、S09072全菌蛋白表達(dá)圖譜比較一致,而S09076、S09077全菌蛋白表達(dá)圖譜也趨于一致。S09071、S09072與S09076、S09077之間有48個明顯的蛋白差異點(diǎn),質(zhì)譜鑒定得到47個分值有效的蛋白點(diǎn),對應(yīng)了43個蛋白,如GadA、GadB、OmpA、TatB等,這些蛋白在細(xì)菌代謝、毒力發(fā)揮以及耐受不同的環(huán)境等過程中扮演重要作用。 (8)成功建立了志賀氏菌毒力大質(zhì)粒大片段敲除方法,為志賀菌毒力大質(zhì)粒的功能以及志賀氏菌的致病機(jī)理研究奠定了基礎(chǔ)。
[Abstract]:Shigella spp.is a group of typical dysentery symptoms (fever, abdominal pain) that can be caused by the invasion of intestinal epithelial cells. Shigella sonneiensis is one of the leading pathogens of dysentery. There are many differences in Shigella sonnei. A comparative study was conducted on four strains of Shigella s09071, S09072, S09076 and S09077, respectively. It can be used to study the interaction between virulence plasmids and chromosomal genomes by constructing large plasmid deletion mutants of Shigella virulent. It also lays a foundation for the study of the pathogenic mechanism of Shigella. In this paper, four strains of Shigella sonneiensis S09071C S09072C S09076N S09076S09077 were studied in detail in terms of morphology, serum agglutination reaction, virulence evaluation, growth characteristics, physiological and biochemical characteristics, and drug resistance. Two dimensional electrophoresis, the comparison of the whole bacterial protein expression profile, from the molecular point of view, further comparative study of four strains of bacteria; Finally, the differential protein of protein expression map was identified by MALDI-TOF/TOF mass spectrometry. On the other hand, the method of large plasmid fragment knockout of Shigella virulence was established. (1) the four strains of Shigella sonnei showed straight rod shape, no spores, capsule, flagella and fimbriae, and the surface morphology of the bacteria was uneven, and the size of the bacteria was 0. 5 0. 7 脳 1. 2 渭 m. The colony on LB medium was large, flat, smooth, moist, colorless and translucent. The edges were neat and the size was about 2.5 mm. there was almost no difference in the morphology of the four strains of bacteria and colony. The growth curve of S09071 was compared with that of S09071. The growth rate of S09071 was the fastest, and that of S09076-S09077 was the slowest. (3) the biochemical characteristics of S09076 and S09077 could be used to ferment rhamnose L-, arbutin and aescectin, but S09071 and S09072 could not be used. The drug resistance of S09071 / S09072 was stronger than that of S09076 / S09077, and the resistance of S09072 was the strongest. 5) virulence evaluation showed that only S09072 had the ability of invasion of epithelial cells, while S09071 and S09076 of S09076-S09077 had no invasiveness. The virulence genes of S09071and S09076S09076-S09077 were found to be lost, but only S09072 was intact. (7) through comparative proteomics studies, we found that the protein expression profiles of S09071 and S09072 were consistent, while that of S09076-S09077 was similar to that of S09071-S09072 and S09076-S09077.There were 48 distinct protein differences between S09071-S09072 and S09076-S09077, and 47 effective protein spots were identified by mass spectrometry. They correspond to 43 proteins, such as GadAl, GadBX, OmpAMP TatB, which play an important role in bacterial metabolism, virulence and tolerance to different environments. The large fragment knockout method of Shigella virulent plasmids was successfully established, which laid a foundation for the function of virulent plasmids of Shigella and the pathogenic mechanism of Shigella.
【學(xué)位授予單位】：西南大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R378

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相關(guān)期刊論文 前10條

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本文編號：1614733