广州市环境及空调冷却水中釜山军团菌的分离与鉴定  

Identification and characterization of LegioneUa busanensis strains isolated from environmental and air conditioning cooling water in Guangzhou

作　　者：范晓姗 胡朝晖[1] 顾全[1] 赵利伟[1] 严慧[1] 朱庆义[1] 

机构地区：[1]广州金域医学检验中心有限公司,510330

出　　处：《中华微生物学和免疫学杂志》2014年第2期96-99,共4页Chinese Journal of Microbiology and Immunology

基　　金：国家自然科学基金资助项目（31270049;31300004）

摘　　要：目的对广州市环境水源及中央空调冷却水中分离的两株疑似嗜肺军团菌进行分子生物学鉴定。方法通过培养特征、生化反应特性、军团菌属特异性引物PCR鉴定、PCR-酶切分型方法，16SrRNA基因、mip基因和rpoB基因序列分析，确定两株疑似嗜肺军团菌的分类地位。结果两株疑似嗜肺军团菌均为革兰氏阴性杆菌，在BCYEa平板上生长，36℃培养48h可见白色菌落，氧化酶、明胶液化与马尿酸水解实验均为阳性，尿素酶、硝酸盐还原实验阴性，按表型特征极易鉴定为嗜肺军团菌。军团菌属特异性引物PCR扩增阳性，PCR-酶切分型结果为单-226bp片段。16SrRNA基因、mip基因和rpoB基因序列比对与系统进化分析显示两株菌与釜山军团菌最相似。结论这是我国首次分离得到釜山军团菌，且同时存在于自然环境中和空调冷却塔水中；由于其生化特征，易错误鉴定为嗜肺军团菌，PCR-酶切分型与多基因鉴定的方法可有效鉴定釜山军团菌。Objective To identify two suspected Legionella pneumophila （L. pneumophila ） strains isolated from environmental water and air conditioning eooling water systems in Guangzhou city. Methods The two strains were identified by their eultural eharaeteristies, biochemical test, Legionella-spe- eifie primer PCR identification, PCR-enzymatie digestion analysis, 16S rRNA, mip and rpoB gene sequen- cing analysis. Results The two suspected L. pneumophila isolates were identified as gram-negative bacillus appeared as white colonies on BCYEct-agar after incubation for 48 hours at 36~C. Both isolates were positive for oxidase, gelatinase and hydrolysis of hippurate, and negative for urease activity and nitrate reduction. Their phenotypie eharaeteristies were similar to those of L. pneumophila strains. Results of PCR identifica- tion by using Legiortella-speeifie primer were positive. Enzymatic digestion analysis showed that the 226 bp PCR products of two isolates were not digested by Taa I. The two strains were classified as Legionella bus- anens/s as indicated by gene sequencing analysis of 16S rRNA, mip and rpoB gene. Conclusion Two L. busanensis strains were first isolated from environmental and air conditioning cooling water systems in Chi- na. Due to their biochemical characteristics, L. busanensis strains were commonly misidentified as L. pneu- mophila, but could be effectively identified by PCR-enzymatie digestion analysis and multiple genes identifi- cation.

关 键 词：釜山军团菌 PCR-酶切分型 基因序列分析 

分 类 号：R378[医药卫生—病原生物学]