摘要
目的将16SrRNA基因序列分析技术应用于传统方法鉴定可靠率低或无法鉴定细菌的分类鉴定,以提高临床诊断的准确性。方法以本院微生物实验室保存的5株标准菌株验证实验方法的准确性后,收集实验室常规方法不能鉴定或鉴定可靠性低的临床分离菌15株(包括革兰阳性球菌,革兰阳性杆菌及革兰阴性杆菌),提取DNA模板,以通用引物PCR扩增16SrRNA目的片段后测序,将测序结果在GenBank数据库中比对分析以确定菌种。结果 15株实验细菌中有14株(占93.3%)鉴定到种,包括鼻疽诺卡菌,大肠埃希菌,阿尔莱特葡萄球菌,脆弱拟杆菌,弗氏柠檬酸杆菌,短小芽孢杆菌,河流漫游球菌,极小短小杆菌,伴放线凝聚杆菌,毗邻颗粒球菌;1株菌鉴定到属(占6.7%),归属于短状杆菌属。结论 16SrRNA基因序列分析技术具有准确、快速和方法简便的特点,适用于对临床非典型菌、少见菌以及新型细菌的鉴定,可作为细菌常规鉴定手段的必要补充。
Objective To use 16S rRNA gene sequence analysis to identify bacteria that cannot be identified accurately by conventional methods in a clinical laboratory in order to improve the accuracy of clinical diagnosis. Methods Experi mental accuracy was first verified by analyzing 5 ATCC standard strains in a clinical laboratory. Then, 15 clinical isolates （including Gram-positive cocci, Gram-positive bacilli, and Gram-negative bacilli）that were identified inaccurately by con ventional methods were selected. Bacterial DNA was extracted and 16S rRNA PCR amplification was performed with a u niversal primer. The PCR products were sequenced and compared to the GenBank database to determine bacterial species. Results The species of 14 （93.3%）of the 15 strains was successfully identified, including Nocardia farcinica , Esche richia coli, Staphylococcus arlettae, Bacteroides fragilis, Citrobacter freundii, Bacillus pumilus, Vagococcus flu＇via lis, Curtobacterium pusillum, Aggregatibacter actinomyceterncomitans, and Granulicatella adiacens. The genus of 1 strain（6.7 % ）was identified as Brachybacterium. Conclusion Bacterial identification based on 16S rRNA gene sequcnce analysis is accurate, simple, and fast. This technique can be used to identify clinically atypical bacteria, rare bacteria, and even new forms of bacteria. This technique must be used to supplement routine bacterial identification.
出处
《中国病原生物学杂志》
CSCD
北大核心
2013年第11期972-974,992,共4页
Journal of Pathogen Biology
基金
国家临床重点专科建设项目[财社(2010)305号]
