分支杆菌临床分离株的16S rRNA基因序列分析

The sequence analyses of 16S rRNA genes of mycobacteria

目的 建立分支杆菌的 16SrDNA序列分析的方法 ,采用该方法鉴定临床结核分支杆菌分离株。方法 用PCR从重庆某医院的肺部感染病人的痰标本中分得的 2 9株分支杆菌菌株中扩增 16SrRNA基因 (16SrDNA)的 5′端片段 (～5 0 0bp) ,并测定所得到片段的DNA序列。用DNA分析软件将所获得的序列与GenBank的分支杆菌序列相比较 ,计算种间相似性 ,并用序列构建分支杆菌菌株的系统发育树 ,对分离株进行分类与鉴定。结果 有 14株的 16SrDNA序列分别与已知结核分支杆菌复合群 (Mycobacteriatuberculosiscomplex ,MTC)、戈登分支杆菌、新金色分支杆菌、氯酚红分支杆菌、龟分支杆菌或脓肿分支杆菌的序列完全相同。依据完全一致的序列可以准确鉴定这些临床分离株为相应的种 ;部分分离株的 16SrDNA在GenBank序列检索中无完全一致的匹配序列。用临床分离株的 16SrDNA序列与已知的分支杆菌 16SrDNA序列构建的系统发育树 ,结果提示某些菌株可能是与已知分支杆菌密切相关的新种或是它们的亚种。结论 16SrDNA序列分析鉴定分支杆菌是一种快速、可靠、成本较低的方法 ;

To establish the sequence analysis of 16S rDNA of mycobacteria for the identification of mycobacterial strains isolated from patients.The 5' end fragments (～500bp) of 16S rRNA genes were amplified from 29 mycobacterial strains isolated from sputa of the patients with lung infection in a hospital of Chongqing city by polymerase chain reaction (PCR) and the DNA fragments were directly sequenced. The sequences obtained in this study were compared with the 16S rDNA sequneces reported in GenBank by software of DNA analyses. Results showed the 16S rDNA sequences from 14 strains were identical to that of MTC, M.gordonae, M.neoaurum, M.chlorophenolicum, and M.abscessus/ M.chelonae. Therefore, these strains may correctly be identified based on the identical sequences. Some of the 16S rDNA sequences of the isolated strains were not identical to any sequence in the GenBank,but similar to sequences of certain mycobacterial species. The sequence comparison of 16S rDNA sequences between the isolates and the members of mycobacterial genus revealed that certain strains isolated in Chongqing are the new species or subspecies. It is concluded that 16S rDNA sequence analyses may be a reliable, rapid, and low costs way for identification of mycobacterial strains and the species of mycobacteria causing lung infection in Chongqing are different from that reported previously.