本文選題：鼠疫耶爾森氏菌　切入點：質(zhì)粒相互作用　出處：《安徽醫(yī)科大學(xué)》2014年碩士論文　論文類型：學(xué)位論文

【摘要】：鼠疫是由鼠疫耶爾森氏菌(以下簡稱鼠疫菌)引起的一種自然疫源性烈性傳染病。在人類歷史上,曾經(jīng)發(fā)生過3次世界鼠疫大流行,給人類帶來過深重災(zāi)難。新中國成立后，鼠疫疫情雖得到有效控制，但近年來，世界鼠疫疫情呈上升趨勢，2000年世界衛(wèi)生組織將鼠疫列為重新抬頭的傳染病。我國鼠疫自然疫源地分布廣泛，動物鼠疫流行面積不斷擴(kuò)大，人間鼠疫病例不斷增加。此外，鼠疫菌又是一種典型的生物戰(zhàn)劑和生物恐怖劑。因此，對鼠疫菌致病機(jī)制的研究對國民健康、社會經(jīng)濟(jì)發(fā)展和國家安全具有重要意義。 鼠疫菌是由假結(jié)核耶爾森氏菌（以下簡稱假結(jié)核菌）O:1b血清型進(jìn)化而來。典型的鼠疫菌含有3個毒力質(zhì)粒（pMT1、pPCP1和pCD1），其中pCD1是從假結(jié)核菌遺傳下來的固有質(zhì)粒，而pMT1和pPCP1則是通過水平轉(zhuǎn)移從外界獲得的質(zhì)粒。鼠疫菌引起嚴(yán)重的致死性疾病，而假結(jié)核菌只引起溫和的腸道疾病，這說明新獲得質(zhì)粒與鼠疫菌的強(qiáng)致病性密切相關(guān)。那么，新獲得質(zhì)粒與固有質(zhì)粒之間是否存在相互作用呢？若存在相互作用，其對致病性的影響又是如何呢？為了回答上述兩個問題，本文基于田鼠型鼠疫菌201質(zhì)粒缺失突變株，通過毒力分析、基因敲除和蛋白表達(dá)分析等策略，研究了質(zhì)粒間相互作用及其與致病性的關(guān)系。 鼠疫菌201株，除含有pPCP1、pMT1、pCD1外，還含有一個隱性質(zhì)粒pCRY。本研究基于鼠疫菌201株及其15個不同質(zhì)粒缺失株，通過生存分析評價了它們的毒力。結(jié)果表明，pCD1質(zhì)粒是鼠疫菌致病必不可少的，pMT1質(zhì)粒對小鼠有一定的毒力，pPCP1和pCRY單獨或共存時毒力很低。臟器存活分析結(jié)果顯示，所有含pCD1質(zhì)粒的鼠疫菌株pCD1+pMT1+pCRY+、pPCP1+pCD1+pMT1+、pPCP1+pCD1+pCRY+、pPCP1+pCD1+、pCD1+pCRY+、pCD1+在肝、脾臟器中能分離到活菌，但pCD1+pMT1+例外，這個結(jié)果表明pMT1可能對pCD1質(zhì)粒具有抑制作用，而pPCP1或pCRY卻能抵消pMT1的這種抑制作用。此后，我們結(jié)合SDS-PAGE電泳和質(zhì)譜分析的方法，對pCD1質(zhì)粒編碼的三型分泌系統(tǒng)外膜蛋白（Yops）的分泌情況進(jìn)行了測定，發(fā)現(xiàn)pCD1+pMT1+與其它含pCD1質(zhì)粒的菌株相比，YopN和LcrV的表達(dá)明顯受到抑制，，再次證明pMT1對pCD1質(zhì)粒有抑制作用，而這種抑制作用會因其它質(zhì)粒的存在而消失。 為了研究質(zhì)粒間互作與致病性的關(guān)系，我們采用基因敲除的策略構(gòu)建了pPCP1+pCD1+pMT1+pla、pPCP1+pMT1+pla、pCD1+pMT1+ymt和pMT1+ymt等基因缺失突變株。通過比較這些突變株與未突變株的毒力，證明在pCD1質(zhì)粒缺失的情況下，鼠毒素起主要的毒力作用，同時pla具有降解鼠毒素的作用。pPCP1+pCD1+pMT1+pla與pPCP1+pCD1+pMT1+相比，在小鼠臟器中存活能力明顯降低，說明了pPCP1促進(jìn)鼠疫菌在宿主體內(nèi)的感染能力主要由pla負(fù)擔(dān)。pCRY除可能抵消pMT1的抑制作用外，其他功能尚不清楚，但pPCP1、pMT1和pCD1卻是鼠疫菌保持完整毒力所必需的。
[Abstract]:Yersinia pestis is a natural infectious disease caused by Yersinia pestis. In human history, there have been three world plague pandemics, which have brought serious disasters to mankind. After the founding of New China, Although the plague epidemic situation has been effectively controlled, in recent years, the world plague epidemic situation is on the rise. In 2000, the World Health Organization listed the plague as a newly rising infectious disease. In China, the plague natural foci are widely distributed, and the epidemic area of animal plague is constantly expanding. In addition, Yersinia pestis is a typical biological warfare agent and bioterrorist agent. Therefore, the study of the pathogenic mechanism of Yersinia pestis is of great significance to national health, social and economic development and national security. Yersinia pestis evolved from the serotype O: 1b of pseudotuberculosis. Typical Yersinia pestis contains three virulent plasmids, pMT1, pPCP1 and pCD1, in which pCD1 is an inherent plasmid inherited from pseudotuberculosis. PMT1 and pPCP1 are plasmids obtained from outside through horizontal transfer. Yersinia pestis causes serious fatal diseases, while pseudotuberculosis causes mild intestinal diseases, which indicates that the newly obtained plasmids are closely related to the strong pathogenicity of Yersinia pestis. Is there any interaction between the newly obtained plasmids and the inherent plasmids? If there is interaction, how does it affect pathogenicity? In order to answer the above two questions, the relationship between plasmids interaction and pathogenicity was studied by virulence analysis, gene knockout and protein expression analysis based on plasmids deletion mutants of Yersinia pestis 201. 201 strains of Yersinia pestis, in addition to pPCP1, pMT1, pCD1, also contained a recessive plasmid pCRY. based on 201 strains of Yersinia pestis and 15 different plasmid deletion strains, The virulence of pCD1 plasmid was evaluated by survival analysis. The results showed that pCD1 plasmid was essential to Yersinia pestis. The virulence of pPCP1 plasmid to mice was very low when pPCP1 and pCRY co-existed, and the organ survival analysis showed that the virulence of pCD1 plasmid was very low. All Yersinia pestis strains containing pCD1 plasmids pCD1 pMT1 pCRY pPCP1 pCD1 pMT1 pPCP1 pCD1 pCRY pPCP1 pCD1 pCD1 pCRY pCD1 can be isolated from the organs of liver and spleen, except pCD1 pMT1. The results indicate that pMT1 may have inhibitory effect on pCD1 plasmid. However, pPCP1 or pCRY could counteract the inhibition of pMT1. After that, we determined the secretion of the outer membrane protein of the three type secretory system encoded by pCD1 plasmids by SDS-PAGE electrophoresis and mass spectrometry. It was found that the expression of YOPN and LcrV in pCD1 pMT1 was significantly inhibited compared with other strains containing pCD1 plasmids. It was proved that pMT1 had inhibitory effect on pCD1 plasmids, and this inhibition would disappear due to the existence of other plasmids. In order to study the relationship between plasmids interaction and pathogenicity, pPCP1 pCD1 pMT1 plapPCP1 pMT1 plapCD1 pMT1 ymt and pMT1 ymt were constructed by gene knockout strategy. The results showed that the murine toxin had the main virulence in the absence of pCD1 plasmid, and that pla could degrade the mouse toxin. Compared with pPCP1 pCD1 pMT1, the survival ability of pCD1 pMT1 pla was significantly lower than that of pPCP1 pCD1 pMT1. The results showed that the ability of pPCP1 to promote the infection of Yersinia pestis in the host was mainly affected by pla. The other functions were not clear except that pla might counteract the inhibitory effect of pMT1, but pPCP1, pMT1 and pCD1 were necessary for Yersinia pestis to maintain their complete virulence.
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R516.8

【參考文獻(xiàn)】

相關(guān)期刊論文 前3條

1 梁少生,林新勤,楊光華,秦石英,楊勤保,江超穗;一起大型水電站庫區(qū)鼠疫暴發(fā)流行的調(diào)查分析[J];中國地方病防治雜志;2001年03期

2 周冬生;楊瑞馥;;鼠疫研究進(jìn)展與展望[J];解放軍醫(yī)學(xué)雜志;2010年10期

3 韋蝶心;宋志忠;;鼠疫耶爾森菌遺傳學(xué)研究進(jìn)展[J];中國媒介生物學(xué)及控制雜志;2010年01期

相關(guān)博士學(xué)位論文 前1條

1 趙向娜;鼠疫耶爾森氏菌噬菌體Yep-phi基因組研究及其受體鑒定[D];中國人民解放軍軍事醫(yī)學(xué)科學(xué)院;2013年

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