肺炎克雷伯菌裂解性噬菌体KP-ZS5的分离及鉴定  

Isolation of Klebsiella pneumoniae lytic phage KP-ZS5 and its identification

作　　者：彭秀玲 邵瑞瑞 田艳 黄孝彬 陈绍海 文胜 陈峥宏[1,4] 崔古贞 齐晓岚 洪伟[1] PENG Xiuling;SHAO Ruirui;TIAN Yan;HUANG Xiaobing;CHEN Shaohai;WEN Sheng;CHEN Zhenghong;CUI Guzhen;QI Xiaolan;HONG Wei(Key Laboratory of Endemic and Ethnic Diseases,Ministry of Education&Key Laboratory of Medical Molecular Biology of Guizhou Province&Key Laboratory of Pathogenic Biology,Guizhou Medical Universities,Guiyang 550004,Guizhou,China;School of Biology and Engineering,Guizhou Medical University,Guiyang 550025,Guizhou,China;Department of Public Art,Guizhou Polytechnic Of Construction,Qingzhen 551400,Guizhou,China;Department of Microbiology,School of Basic Medicine,Guizhou Medical University,Guiyang 550025,Guizhou,China)

机构地区：[1]贵州医科大学地方病与少数民族疾病教育部重点实验室&贵州省分子生物学重点实验室&贵州省普通高等学校病原生物学特色重点实验室,贵州贵阳550004 [2]贵州医科大学生物与工程学院,贵州贵阳550025 [3]贵州建设职业技术学院公共艺术教研室,贵州清镇市551400 [4]贵州医科大学基础医学院微生物学教研室,贵州贵阳550025

出　　处：《贵州医科大学学报》2023年第3期259-265,共7页Journal of Guizhou Medical University

基　　金：2021年国家级大学生创新创业训练项目(202110660013);贵州省自然科学基金重点项目(〔2020〕1Z067)。

摘　　要：目的肺炎克雷伯菌裂解性噬菌体KP-ZS5的分离及鉴定。方法以18株临床分离肺炎克雷伯菌为宿主菌,采用双层平板法从水样中分离肺炎克雷伯菌噬菌体并进行纯化,透射电镜观察噬菌体形态,提取噬菌体DNA及蛋白并测定其生理生化特性。结果成功分离到1株长尾肺炎克雷伯菌噬菌体,并命名为KP-ZS5,pH稳定性实验表明KP-ZS5不耐强酸(pH≤3);限制性片段酶切多样性(RFLP)分析表明,KP-ZS5基因组中存在3个BglⅡ酶切位点,预计总基因组大小为20000~30000 bp;裂解谱实验表明,KP-ZS5可以感染33%(6/18)的临床肺炎克雷伯菌株;SDS-PAGE结果显示,在KP-ZS5外壳蛋白由3种蛋白组成,外壳蛋白分子量为35~48 kDa。结论成功分离出1株长尾肺炎克雷伯菌噬菌体KP-ZS5,该噬菌体具有较广的宿主范围和pH值耐受性,外壳蛋白组成简单,仅含3种分子量为35~48 KDa的组分,具有在临床治疗肺炎克雷伯菌感染的潜力。Objective To isolate and identify the Klebsiella pneumoniae lytic phage KP-ZS5A.Methods Using 18 clinical isolates of K.pneumoniae as the host bacteria,the K.pneumoniae phage was isolated and purified from water samples by double-layer plate method.The morphological features of the phage were examined by electron microscopy,its DNA and protein were extracted and its physiological and biochemical characteristics determined.Results A long-tailed K.pneumoniae phage,KP-ZS5,was successfully isolated.The pH stability test showed that KP-ZS5 was not resistant to strong acid(pH≤3).The Restriction Fragment Length Polymorphism(RFLP)experiment showed that three BglⅡdigestion sites were in the KP-ZS5 genome,and the total genome size was expected between 20000-30000 bp.The host spectrum test showed that KP-ZS5 can infect 6/18(33%)clinical isolated K.pneumoniae.The SDS-PAGE results showed that,KP-ZS5 consisted of three coat proteins and its molecular weights ranged from 35-48 kDa.Conclusion A long-tailed K.pneumoniae phage,KP-ZS5,was successfully isolated from the surrounding rivers of the Affiliated Hospital of Guizhou Medical University.The phage has a broad host range and pH tolerance,a simple shell protein composition containing only three components with molecular weights of 35-48 kDa,and has potential for the clinical treatment of K.pneumoniae infections.

关 键 词：肺炎克雷伯菌 噬菌体 生理生化特征 双层平板法 噬菌体疗法 

分 类 号：R378.996[医药卫生—病原生物学]