耐药性鲍曼不动杆菌噬菌体特性和基因组及应用效果评价

发布时间：2018-02-17 05:05

  本文关键词： 鲍曼不动杆菌 噬菌体 脉冲场凝胶电泳 多位点序列分型 生物学特性 全基因组 小鼠肺炎 噬菌体治疗　出处：《中南大学》2014年硕士论文　论文类型：学位论文

【摘要】：目的：耐药性鲍曼不动杆菌已成为医院主要的病原体,且很容易通过交叉感染在医院内爆发流行,威胁患者的生命。寻找有效的预防及治疗的方法是临床亟待解决的问题。噬菌体被认为是针对耐药菌防治的一种有效手段。本论文对噬菌体消毒和治疗的有效性进行了初步研究,为防治多耐药鲍曼不动杆菌感染提供新的思路。 方法：本研究根据鲍曼不动杆菌耐药严重的现状,对收集到的40株耐药性鲍曼不动杆菌进行脉冲场凝胶电泳和多位点序列分型；并以这些菌为指示菌分离噬菌体；选取其中一株噬菌体IME-AB2进行生物学特性和全基因组分析；并对该噬菌体在菌液及物体表面的消毒效果进行了探究,采用噬菌体吸入治疗小鼠多耐药鲍曼不动杆菌肺部感染,研究其体内治疗效果。 结果：40株耐药性鲍曼不动杆菌显示对大多数抗生素耐药,包括含酶抑制剂和碳氢霉烯类。通过脉冲场凝胶电泳可将细菌分为A、B、C共3个种克隆型,B型26(65%)株,为主要的克隆型。多位点序列分型中发现细菌共分13个ST型,以ST208为最原始ST型,ST218、ST195、ST369、ST451、ST238、ST368、ST469均由ST208分化而来,共24株(占60%)。本研究成功分离出16株耐药性鲍曼不动杆菌噬菌体,对40株菌的裂解覆盖率达78%。选取一株噬菌体IME-AB2进行研究,该株噬菌体滴度高达1011pfu/ml,可以快速吸附到宿主菌表面,潜伏期和爆发时间短,在LB培养基中可以4℃长期保存,反映出该株噬菌体裂解的高效能和稳定性。电镜观察显示这株噬菌头部呈二十面体结构,含有尾部结构,属于肌尾噬菌体。噬菌体IME-AB2基因组是一条环状的双链DNA分子,全长43665bp,全基因组平均GC含量为37.5%,编码82个蛋白质基因。全基因组序列已在NCBI GenBank注册(GenBank Accession number:JX976549)。该株噬菌体能在4小时内使混浊菌液变澄清,并且能使器物表面的细菌数量明显减少。在通过吸入治疗小鼠肺炎的研究中,小鼠肺部接种细菌后30min,4h,24h给予噬菌体治疗,分别得到100%,50%,20%的生存率；以不同MOI(感染复数)进行治疗,MOI=1时生存率100%,MOI=0.1时生存率80%, MOI=0.01时生存率60%,MOI=0.001时生存率50%,PBS组生存率只有10%。治疗的小鼠肺泡灌洗液中的细菌明显少于非治疗组,噬菌体数量没有明显差异,小鼠肺脏病理结果显示噬菌体疗效显著。 结论：噬菌体对耐药性鲍曼不动杆菌具有良好的体内、外抗菌作用,为临床开发防治耐药性鲍曼不动杆菌感染的制剂提供实验基础。图16幅,表13个,参考文献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，

本文编号：1517283