本文選題：金黃色葡萄球菌 + 生物膜�。� 參考：《第三軍醫(yī)大學(xué)》2006年碩士論文

【摘要】： 金黃色葡萄球菌(Staphylococcus aureus)是醫(yī)院感染的重要病原菌之一,也是生物膜感染常見(jiàn)的病原菌,在ICU導(dǎo)管相關(guān)感染敗血癥的致病原中,金黃色葡萄球菌占15.4%,居第二位。形成生物膜的金黃色葡萄球菌具有高度的耐藥性并能逃避免疫系統(tǒng)的攻擊,其感染易慢性化并難于控制,因此對(duì)金黃色葡萄球菌等臨床重要病原菌的生物膜形成機(jī)制的研究已成為抗感染研究領(lǐng)域的熱點(diǎn)。 目前對(duì)金黃色葡萄球菌生物膜的形成機(jī)制仍不清楚。初步研究顯示生物膜的形成首先是由細(xì)胞壁相關(guān)因子調(diào)節(jié)的細(xì)胞表面的附著,隨后細(xì)菌主要通過(guò)ica操縱子表達(dá)產(chǎn)物共同催化合成的多糖胞間粘附素(polysaccharide intercellular adhesin, PIA)介導(dǎo),細(xì)胞增殖并相互聚集成細(xì)胞團(tuán)塊,繼而形成生物膜。該過(guò)程中有多個(gè)基因參與調(diào)控生物膜的形成。其中革蘭氏陽(yáng)性菌中σ因子編碼基因sigB的表達(dá)可能是生物膜形成的關(guān)鍵調(diào)控因素之一。它除可通過(guò)直接調(diào)節(jié)控制生物膜形成的ica操縱子的表達(dá)外,尚可影響ica的調(diào)控基因sarA和/或agr的表達(dá)。但新近采用對(duì)特定的金黃色葡萄球菌實(shí)驗(yàn)室菌株敲除sigB基因的研究發(fā)現(xiàn),sigB缺失的菌株在某些條件下仍能形成生物膜。鑒于這些研究多局限于幾株特定的實(shí)驗(yàn)室菌株,為明確sigB基因在金黃色葡萄球菌形成生物膜中的意義,本研究以第三軍醫(yī)大學(xué)第一臨床醫(yī)學(xué)院2004年7月-2004年12月分離的野生菌株為研究對(duì)象,在明確其耐藥及生物膜形成情況的基礎(chǔ)上,PCR擴(kuò)增分析金黃色葡萄球菌中生物膜形成相關(guān)調(diào)節(jié)基因的存在情況;篩選可形成生物膜的菌株,采用融合PCR的方法構(gòu)建金黃色葡萄球菌sigB同源重組質(zhì)粒,為建立sigB缺失的野生金黃色葡萄球菌模型奠定基礎(chǔ)。 研究分三部分進(jìn)行: 1.臨床分離金黃色葡萄球藥物菌敏感性和形成生物膜的檢測(cè) 2.金黃色葡萄球菌生物膜形成相關(guān)基因的PCR分析 3.金黃色葡萄球菌sigB同源重組質(zhì)粒的構(gòu)建 1.方法 1.1金黃色葡萄球菌敏感性(最小抑菌濃度)用瓊脂稀釋法測(cè)定;生物膜形成的檢測(cè)用96孔板番紅染色法。 1.2 icaAD、icaBC、sar、agr和sigB基因采用PCR法擴(kuò)增,并對(duì)產(chǎn)物進(jìn)行測(cè)序和
[Abstract]:Staphylococcus aureus (Staphylococcus aureus) is one of the important pathogens of nosocomial infection and a common pathogen of biofilm infection. In the pathogeny of ICU catheter related septicemia, Staphylococcus aureus occupies 15.4%, ranking second. Staphylococcus aureus forming biofilm has high resistance and can escape immunity. The infection of the system is easy to be chronicity and difficult to control. Therefore, the study of the mechanism of the biofilm formation of clinical important pathogens such as Staphylococcus aureus has become a hot spot in the field of anti infection research.
The formation mechanism of Staphylococcus aureus biofilm is still unclear. Preliminary study showed that the formation of biofilm was first attached to cell surface regulated by cell wall related factors, and then bacteria mainly catalyzed by ica operon to co catalyze the polysaccharide intercellular adhesin, PIA In this process, there are several genes involved in regulating the formation of biofilms. In gram positive bacteria, the expression of the sigma factor encoding gene sigB may be one of the key regulatory factors of biofilm formation. It can be used to control the ICA of biofilm by direct regulation. The expression of the operon can affect the expression of the regulatory gene sarA and / or agr of the ICA. However, recent studies on the knockout of the sigB gene of a specific Staphylococcus aureus laboratory strain have found that the sigB missing strain can still form a biofilm under certain conditions. The significance of the sigB gene in the formation of Staphylococcus aureus in the biofilm was studied. This study was based on the study of the wild strains isolated from the first clinical medicine hospital of Third Military Medical University in December July 2004. On the basis of its drug resistance and biofilm formation, PCR amplification was used to analyze the regulation of biofilm formation in Staphylococcus aureus. The existence of the gene, screening the strains that can form biofilm, and using the fusion method of PCR to construct the homologous recombinant plasmid of Staphylococcus aureus sigB, lay the foundation for the establishment of the sigB deletion of the Wild Staphylococcus aureus model.
The study is divided into three parts:
1. clinical isolation of Staphylococcus aureus drug sensitivity and biofilm formation detection
PCR analysis of 2. genes related to biofilm formation in Staphylococcus aureus
Construction of 3. homologous recombination plasmid of Staphylococcus aureus sigB
1. method
1.1 the sensitivity (minimum inhibitory concentration) of Staphylococcus aureus was determined by agar dilution method, and the detection of biofilm formation was made by 96 hole plate red staining.
1.2 the icaAD, icaBC, SAR, agr and sigB genes were amplified by PCR and sequenced.
【學(xué)位授予單位】：第三軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2006
【分類號(hào)】：R346

【引證文獻(xiàn)】

相關(guān)碩士學(xué)位論文 前1條

1 李蕾;金黃色葡萄球菌isdA缺失株的構(gòu)建及其免疫活性的研究[D];吉林大學(xué);2012年

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本文編號(hào)：2073115