分支杆菌噬菌体Leo生物学特性及抗结核作用

发布时间：2018-05-31 17:06

本文选题：分支杆菌噬菌体Leo + 生物学特性　；参考：《中国人兽共患病学报》2015年03期

【摘要】：目的明确分支杆菌噬菌体Leo的生物学特性及其抗结核作用,为"鸡尾酒疗法"筛选候选噬菌体。方法采用双层平板法制备噬菌体Leo,观察噬菌斑形态;电镜观察噬菌体颗粒形态;提取噬菌体DNA,限制性内切酶酶切分析确定其核酸类型;以不同MOI扩增噬菌体Leo,确定最佳MOI及最低MOI;通过一步生长曲线,确定潜伏期及裂解量;采用终点滴定反浊法测定耻垢分支杆菌对噬菌体Leo的耐受突变率;检测噬菌体Leo对酒精、温度的耐受性及在不同pH下的裂解能力;通过体外杀结核分枝杆菌实验,明确噬菌体Leo对结核分枝杆菌的杀灭作用。结果 Leo噬菌斑圆形透明,边缘清晰,含对称的多面体头部,直径(70±3.0)nm,可弯曲的尾部,尾长(211±31.7)nm;其基因组能被限制性内切酶HindⅢ、BglⅠ切开;最佳MOI为0.0 0001,最低MOI为0.0 001,潜伏期为150min,裂解量74,耐受突变率为10-7。Leo对温度、酒精敏感,在pH 7.4和pH 5.0时均能裂解宿主菌。Leo作用于结核分支杆菌72h后,Leo组菌量明显低于空白组(P0.05)。结论分支杆菌噬菌体Leo属长尾噬菌体科,双链DNA噬菌体,能够杀灭结核分支杆菌,可作为"鸡尾酒疗法"的候选噬菌体。
[Abstract]:Objective to identify the biological characteristics and anti tuberculosis effect of Mycobacterium phage Leo, to screen candidate phage for "cocktail therapy". Method using double plate method to prepare phage Leo, observe phage plaque morphology, observe the morphology of phage particles, extract phage DNA, restriction endonuclease digestion analysis to determine its nucleic acid type; The phage Leo was amplified with MOI to determine the best MOI and the lowest MOI; the incubation period and the amount of lysis were determined by one step growth curve. The resistance mutation rate of Mycobacterium phage to phage Leo was determined by the endpoint titration anti turbid method; the resistance of phage Leo to alcohol, temperature and the ability to crack under different pH were detected. Bacteriophage test, clear bacteriophage Leo killing effect of phage on Mycobacterium tuberculosis. Results Leo phage plaque is transparent, edge clear, symmetrical polyhedral head, diameter (70 + 3) nm, curved tail, tail length (211 + 31.7) nm; its genome can be restricted by Hind III, Bgl I incision, the optimum MOI is 0.00001, the lowest MOI is 0.0001, latent MOI is 0.0001, latent MOI is 0.0001, latent MOI is potential 0.0001. The lowest MOI is 0.0001. The lowest MOI is 0.0001. The lowest MOI is 0.0001. The lowest MOI is 0.0001. The lowest MOI is 0.0001, latent MOI is latent and latent MOI is potential 0.0001. Latent MOI is 0.0001, latent MOI potential 0.0001. Latent MOI is 0.0001. Latent MOI potential 0.0001. Latent MOI potential 0.0001. Latent MOI potential 0.0001. Latent MOI potential 0.0001. Latent MOI potential 0.0001. Latent MOI potential 0.0001 The volt period was 150min, the amount of lysis was 74, the tolerance mutation rate was 10-7.Leo to temperature, alcohol sensitive, pH 7.4 and pH 5 could cleave the host bacterium.Leo to act on the Mycobacterium tuberculosis 72h, and the amount of bacteria in Leo group was obviously lower than that of the blank group (P0.05). Conclusion the Mycobacterium phage is a long tail phage, double stranded DNA phage can kill Mycobacterium tuberculosis. It can be used as a candidate phage for "cocktail therapy".
【作者单位】：重庆医科大学附属第一医院呼吸与危重症医学科;成都市第三人民医院呼吸内科;重庆市长寿区人民医院呼吸内科;
【基金】：国家“十二五”重大专项课题(2012ZX10003009) 国家临床重点专科专项经费资助项目(No.2012-649)资助~~
【分类号】：R378.911

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