rrn操纵元序列PCR鉴定术后龟分支杆菌暴发感染的研究

Study on the postoperative infection of M. chelonae subsp. abscessus by polymerase chain reaction of the rrn operons sequence

目的 本研究旨在采用分子生物学技术 ,从基因水平上确认医院内术后暴发感染的致病菌 ,并建立一套快速的PCR方法鉴定龟分支杆菌脓肿亚种。方法 根据分支杆菌的rrn操纵元序列即 16srDNA和 16s - 2 3srDNA间隔区序列 ,分别设计合成一对引物 ,采用聚合酶链反应技术 ,并以结核分支杆菌和常见速生长非结核分支杆菌作对照 ,对临床分离株进行PCR扩增 ,并根据DNA带的大小和数量鉴定分支杆菌。结果 5 3株龟分支杆菌脓肿亚种临床分离株和标准株 ,用16srDNA扩增 ,均被扩增出一条特异的 5 84bpDNA带 ,相应的 16s - 2 3srDNA扩增 ,为特异的380bpDNA带。而速生长的非结核分支杆菌均扩增出不同大小的 1或 2条DNA带。 结论 基因水平上确认此次引起医院内术后暴发感染的致病菌为龟分支杆菌脓肿亚种。rrn操纵元PCR扩增检测体系灵敏、特异 ,能鉴定龟分支杆菌脓肿亚种临床株 ,并与其它速生长分支杆菌区别开。

Objective To study the pathogen of the postoperative infection outbreak at gene level, using molecular-biological technology. On the basis of this study, the M. chelonae subsp. abscessus clincial strain by rapid polymerase chain reaction was established. Methods A single pair of primers of 16s rDNA or 16s-23s rDNA were designed, according to the rrn operon sequence of mycobacterium, which were 16s ribosomal DNA sequence and 16s-23s ribosomal DNA spacer sequences. All of the clincial strains were amplified by polymerase chain reaction with M. tuberculosis and usual rapid-growing mycobacterium as controls. As a result,mycobacterium was identified with different number & sizes of DNA fragments. Results For 53 clincial and standard strains,only one specific DNA fragment-584bp was amplified by 16s rDNA PCR and 380bp by 16s-23s rDNA PCR. 1 or 2 DNA fragments could be amplified with rapid-growing mycobacterium by PCR. Conclusion The pathogen of the post operative outbreak of infection was identified as M. chelonae subsp. abscessus.at gene level. The style of the rrn operons PCR was sensitive and specific. M. chelonae subsp. abscessus was identified from other rapid-growing mycobacterium.