本文關(guān)鍵詞： 鮑曼不動(dòng)桿菌 噬菌體 脈沖場(chǎng)凝膠電泳 多位點(diǎn)序列分型 生物學(xué)特性 全基因組 小鼠肺炎 噬菌體治療　出處：《中南大學(xué)》2014年碩士論文　論文類型：學(xué)位論文

【摘要】：目的：耐藥性鮑曼不動(dòng)桿菌已成為醫(yī)院主要的病原體,且很容易通過(guò)交叉感染在醫(yī)院內(nèi)爆發(fā)流行,威脅患者的生命。尋找有效的預(yù)防及治療的方法是臨床亟待解決的問(wèn)題。噬菌體被認(rèn)為是針對(duì)耐藥菌防治的一種有效手段。本論文對(duì)噬菌體消毒和治療的有效性進(jìn)行了初步研究,為防治多耐藥鮑曼不動(dòng)桿菌感染提供新的思路。 方法：本研究根據(jù)鮑曼不動(dòng)桿菌耐藥嚴(yán)重的現(xiàn)狀,對(duì)收集到的40株耐藥性鮑曼不動(dòng)桿菌進(jìn)行脈沖場(chǎng)凝膠電泳和多位點(diǎn)序列分型；并以這些菌為指示菌分離噬菌體；選取其中一株噬菌體IME-AB2進(jìn)行生物學(xué)特性和全基因組分析；并對(duì)該噬菌體在菌液及物體表面的消毒效果進(jìn)行了探究,采用噬菌體吸入治療小鼠多耐藥鮑曼不動(dòng)桿菌肺部感染,研究其體內(nèi)治療效果。 結(jié)果：40株耐藥性鮑曼不動(dòng)桿菌顯示對(duì)大多數(shù)抗生素耐藥,包括含酶抑制劑和碳?xì)涿瓜╊�。通過(guò)脈沖場(chǎng)凝膠電泳可將細(xì)菌分為A、B、C共3個(gè)種克隆型,B型26(65%)株,為主要的克隆型。多位點(diǎn)序列分型中發(fā)現(xiàn)細(xì)菌共分13個(gè)ST型,以ST208為最原始ST型,ST218、ST195、ST369、ST451、ST238、ST368、ST469均由ST208分化而來(lái),共24株(占60%)。本研究成功分離出16株耐藥性鮑曼不動(dòng)桿菌噬菌體,對(duì)40株菌的裂解覆蓋率達(dá)78%。選取一株噬菌體IME-AB2進(jìn)行研究,該株噬菌體滴度高達(dá)1011pfu/ml,可以快速吸附到宿主菌表面,潛伏期和爆發(fā)時(shí)間短,在LB培養(yǎng)基中可以4℃長(zhǎng)期保存,反映出該株噬菌體裂解的高效能和穩(wěn)定性。電鏡觀察顯示這株噬菌頭部呈二十面體結(jié)構(gòu),含有尾部結(jié)構(gòu),屬于肌尾噬菌體。噬菌體IME-AB2基因組是一條環(huán)狀的雙鏈DNA分子,全長(zhǎng)43665bp,全基因組平均GC含量為37.5%,編碼82個(gè)蛋白質(zhì)基因。全基因組序列已在NCBI GenBank注冊(cè)(GenBank Accession number:JX976549)。該株噬菌體能在4小時(shí)內(nèi)使混濁菌液變澄清,并且能使器物表面的細(xì)菌數(shù)量明顯減少。在通過(guò)吸入治療小鼠肺炎的研究中,小鼠肺部接種細(xì)菌后30min,4h,24h給予噬菌體治療,分別得到100%,50%,20%的生存率；以不同MOI(感染復(fù)數(shù))進(jìn)行治療,MOI=1時(shí)生存率100%,MOI=0.1時(shí)生存率80%, MOI=0.01時(shí)生存率60%,MOI=0.001時(shí)生存率50%,PBS組生存率只有10%。治療的小鼠肺泡灌洗液中的細(xì)菌明顯少于非治療組,噬菌體數(shù)量沒(méi)有明顯差異,小鼠肺臟病理結(jié)果顯示噬菌體療效顯著。 結(jié)論：噬菌體對(duì)耐藥性鮑曼不動(dòng)桿菌具有良好的體內(nèi)、外抗菌作用,為臨床開(kāi)發(fā)防治耐藥性鮑曼不動(dòng)桿菌感染的制劑提供實(shí)驗(yàn)基礎(chǔ)。圖16幅,表13個(gè),參考文獻(xiàn)41篇。
[Abstract]:Objective: the drug-resistant Acinetobacter baumannii has become a major pathogen in hospitals and is prone to outbreaks in hospitals through cross-infection. It is an urgent clinical problem to find effective prevention and treatment methods. Bacteriophage is considered to be an effective means for the prevention and treatment of drug-resistant bacteria. This paper is effective in bacteriophage disinfection and treatment. A preliminary study was carried out, To provide a new idea for the prevention and treatment of multidrug resistant Acinetobacter baumannii infection. Methods: according to the current situation of severe drug resistance of Acinetobacter baumannii, 40 strains of Acinetobacter baumannii were detected by pulsed field gel electrophoresis (PGE) and multilocus sequence typing, and the bacteriophages were isolated from 40 strains of Acinetobacter baumannii. One of the bacteriophages IME-AB2 was selected for biological characteristics and genome analysis, and the disinfection effect of the bacteriophage on the surface of the bacteriophage and the object was studied. The bacteriophage inhalation was used to treat the pulmonary infection of Acinetobacter baumannii in mice. To study the therapeutic effect in vivo. Results 40 strains of Acinetobacter baumannii showed resistance to most antibiotics, including enzyme inhibitors and hydrocarbamene. In this study, 16 strains of drug-resistant Acinetobacter baumannii were successfully isolated from ST208. The cleavage coverage of 40 strains was 78. A phage IME-AB2 was selected to study the bacteriophage titer of 1011pfur / ml. The phage titer could be rapidly adsorbed to the surface of host bacteria, the incubation period and outbreak time were short, and it could be stored in LB medium at 4 鈩，

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