紫外消毒致耐药基因水平转移规律和影响因素的研究

发布时间：2018-03-14 12:19

本文选题：抗性基因　切入点：抗性菌　出处：《天津大学》2016年硕士论文　论文类型：学位论文

【摘要】：大量研究显示现在的消毒技术不仅不能除去ARGs和ARB,反而会使水中ARGs和ARB比例增加。但ARGs发生水平转移和传播的原因,却未见研究报道。所以,本课题的目的是研究UV消毒对耐药菌和耐药基因(质粒)的灭活规律,以及消毒过程中损伤菌的形成规律;探讨UV消毒损伤菌的性质;明确游离质粒转化消毒损伤菌的形成条件和影响因素,为进一步提出一种有效控制ARGs水平转移和传播的消毒新工艺提供科学依据和技术支撑。研究内容如下:1.以E.coli 25922(RP4)为研究对象,观察UV消毒对其灭活及损伤菌形成的规律,对消毒释放耐药质粒转化活性,耐药基因转座子及耐药基因的影响,还对5种耐药菌的UV消毒抗性进行了比较。UV消毒对E.coli 25922(RP4)作用的先后顺序为:耐药菌损伤耐药菌灭活耐药质粒灭活耐药基因的转座子损伤耐药基因的去除。5种耐药菌(RP4)消毒抗性由强到弱为:S.aureus 6538约E.faecalis 33186Salmonell 50312约E.coli 25922P.aeruginosa 27853。2.采用透射电镜、拉曼光谱分析及生物化学的方法,观测了UV消毒致损伤菌的形态、物质结构、膜通透性和相关功能的改变。发现UV消毒可使细菌内容物减少,且β-半乳糖苷酶活性提高及拉曼光谱分析结果均提示消毒后细菌的膜通透性发生了改变.此外,菌体中相关抗氧化酶活性也有明显升高。3.以E.coli K12为待测目标菌,采用大滤膜法(MF)、多管发酵法(MTF)、酶底物法(EST)分别对臭氧损伤菌进行检测,计算其对臭氧损伤E.coli的检出率。结果发现:MTF在检测野生损伤菌方面表现出较高的灵敏性,其检出率最高可达100%,EST最多可检测4.76%损伤的E.coli,而MF检出率为0。4.通过实验室模拟水处理中的消毒处理,研究了游离ARGs转化UV消毒损伤菌的条件及其影响因素。结果发现:损伤菌转化频率较对照组高1~2个数量级。环境温度、pH值,作用时间、氨氮、CODMn、金属离子、受体菌浓度等因素虽对转化有影响,但实验组转化频率均比相应对照组高。以上结果说明:UV消毒促进了ARGs的释放,同时形成了大量具感受态性质的消毒损伤菌,这两者的存在促进了自然转化的发生。
[Abstract]:A large number of studies have shown that the current disinfection techniques not only can not remove ARGs and ARB, but also increase the proportion of ARGs and ARB in water. However, the causes of horizontal transfer and transmission of ARGs have not been reported. The purpose of this paper is to study the inactivation of drug-resistant bacteria and resistant genes (plasmids) in UV disinfection, and the formation of damaged bacteria in the process of disinfection, and to explore the properties of UV disinfection of drug-resistant bacteria and drug-resistant genes (plasmids). The forming conditions and influencing factors of free plasmids transformed into disinfectant damaged bacteria were determined. In order to provide scientific basis and technical support for a new disinfection process to effectively control the horizontal transfer and spread of ARGs, the research contents are as follows: 1. Taking E.coli 25922RP4) as the research object, the regularity of the inactivation and the formation of damaged bacteria by UV disinfection was observed. The effects on the transformation activity, transposon and drug resistance gene of drug resistance gene by disinfection and release of drug resistance plasmids. The sequence of UV disinfection resistance to E. coli 25922 (RP4) by UV disinfection was also compared. The sequence was as follows: drug resistant bacteria damage resistant bacteria, inactivated plasmid resistant plasmids, transposon damage resistance genes, removal of 5 drug resistance genes. The sterilizing resistance of RP4 was from strong to weak to that of: S.aureus 6538, E. faecalis 33186 Salmonell 50312, E. coli 25922P.aeruginosa 27853.2. transmission electron microscopy was used. The changes of morphology, material structure, membrane permeability and related functions of the damaged bacteria caused by UV disinfection were observed by Raman spectroscopy and biochemical methods. It was found that UV disinfection could reduce bacterial contents. Moreover, the increase of 尾 -galactosidase activity and Raman spectrum analysis indicated that the membrane permeability of bacteria changed after disinfection. In addition, the activity of antioxidant enzymes in bacteria also increased significantly. 3. E.coli K12 was used as the target bacteria. The ozone damaging bacteria were detected by large filter membrane method, multi-tube fermentation method and enzyme substrate method, respectively, and the detection rate of ozone damaged E. coli was calculated. The results showed that: MTF showed a high sensitivity in the detection of wild injured bacteria. The highest detectable rate of EST can be as high as 4.76% damage E.coli, while MF detection rate is 0.4. Disinfection treatment in simulated water treatment in laboratory, The conditions and influencing factors of free ARGs transformation for UV disinfection of damaged bacteria were studied. The results showed that the transformation frequency of the injured bacteria was 1 ~ 2 orders of magnitude higher than that of the control group. The environmental temperature, pH value, time of action, ammonia nitrogen, CODMn, metal ions, etc. Although the concentration of receptor bacteria had an effect on the transformation, the transformation frequency of the experimental group was higher than that of the corresponding control group. The existence of both promotes the occurrence of natural transformation.
【学位授予单位】：天津大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R123.6

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