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  本文關(guān)鍵詞： 腸出血性大腸桿菌O157:H7 ler基因 志賀毒素 突變 疫苗　出處：《吉林大學(xué)》2007年博士論文　論文類型：學(xué)位論文

【摘要】： 本研究首先建立腸出血性大腸桿菌O157:H7感染的動物模型,利用絲裂霉素對小鼠進(jìn)行腹腔注射預(yù)處理,并通過灌胃接種和腹腔注射兩種途徑,建立了O157:H7感染的動物模型。然后對EHEC O157:H7 ler/stx基因缺失突變菌株的原噬菌體及其原始整合位點(diǎn)進(jìn)行分析。通過對Stx A亞基毒性活性區(qū)和跨膜區(qū)的定點(diǎn)突變,消除或降低Stx的細(xì)胞毒性,并將Stx的無毒性或弱毒性突變體導(dǎo)入O157:H7 ler/stx基因缺失突變菌株,構(gòu)建O157重組弱毒疫苗菌株。利用小鼠模型和vero細(xì)胞對O157重組弱毒疫苗菌株的vero細(xì)胞毒性、對小鼠的致病性、重組菌株的穩(wěn)定性,以及被動免疫和主動免疫保護(hù)性進(jìn)行了研究。 結(jié)果表明,O157重組弱毒疫苗菌株具有良好的穩(wěn)定性,對vero細(xì)胞的毒性作用降低了約104倍,喪失了對小鼠模型的致病性,能有效縮短小鼠感染O157:H7強(qiáng)毒株后的排毒時間(免疫組小鼠第6天后糞便內(nèi)即檢測不到強(qiáng)毒株,而未免疫對照組第13天仍可檢測到)。用疫苗菌株對母鼠免疫,免疫母鼠所生的乳鼠通過吸吮母乳,能夠獲得良好的被動免疫保護(hù)作用(F25和F105的免疫保護(hù)率分別為75.2%和83.0%);通過腹腔注射的途徑對小鼠進(jìn)行免疫和攻毒,結(jié)果也表明O157重組弱毒疫苗菌株對小鼠具有良好的免疫保護(hù)作用(F25和F105的免疫保護(hù)作用分別為11/20和14/20)。
[Abstract]:In this study, the animal model of enterohemorrhagic Escherichia coli O157: H7 infection was established. The mice were pretreated by intraperitoneal injection of mitomycin, and were injected intraperitoneally and intraperitoneally. The animal model of O157: H7 infection was established. The phage and its original integration site of EHEC O157: H7 ler/stx gene deletion mutant strain were analyzed. The site-directed mutations of the toxic active region and transmembrane region of Stx A subunit were carried out. The cytotoxicity of Stx was eliminated or reduced, and the non-toxic or weakly toxic mutants of Stx were introduced into O157: H7 ler/stx gene deletion mutant. A recombinant attenuated vaccine strain O157 was constructed. The pathogenicity, stability, passive immunity and active immune protection of O157 recombinant attenuated vaccine strain were studied by using mouse model and vero cells to study the vero cytotoxicity of O157 recombinant attenuated vaccine strain. The results showed that the recombinant attenuated vaccine strain of YO157 had good stability, the toxicity to vero cells was reduced by about 104-fold, and the pathogenicity of the strain was lost to the mouse model. It can effectively shorten the detoxification time of mice infected with O157: H7 virulent strain (the mice in the immunized group could not detect the virulent strain in feces after 6 days, while the mice in the unimmunized control group could still be detected on the 13th day after inoculation with the vaccine strain. The immune protection rates of F25 and F105 were 75.2% and 83.0 by sucking breast milk, respectively, and the mice were immunized and poisoned by intraperitoneal injection. The results also showed that the recombinant attenuated strain O157 had a good protective effect on mice. The protective effects of F25 and F105 on mice were 11/20 and 14 / 20 respectively.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2007
【分類號】：R392

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本文編號：1498572