摘要
目的:利用膜芯片技术建立一种快速、简便的分枝杆菌分子菌种鉴定方法。方法:以DNA直接测序法为对照,通过PCR-SSCP和膜芯片技术分析11种分枝杆菌标准菌株、9种非分枝杆菌和199株分枝杆菌临床分离株的菌种。结果:应用膜芯片技术分析11种分枝杆菌标准菌株和7种非分枝杆菌菌株,特异性100%。199株分枝杆菌临床分离株中,经16S rRNA PCR-SSCP初步菌种鉴定,30株为结核分枝杆菌复合群,应用膜芯片分析,显示与分枝杆菌属探针分枝杆菌和结核分枝杆菌复合群探针a杂交阳性,两种鉴定方法结果一致;169株PCR-SSCP初步鉴定为非结核分枝杆菌的分离株,经芯片分析,58株为龟分枝杆菌,46株为胞内分枝杆菌,33株为堪萨斯、瘰疬、胃和猿猴分枝杆菌复合群,6株为偶然分枝杆菌,15株为戈登分枝杆菌,3株为鸟分枝杆菌,2株为海和溃疡分枝杆菌复合群,另6株只与探针分枝杆菌杂交,经测序显示2株为胞内分枝杆菌,但其基因序列与标准菌株不完全相同,1株为土分枝杆菌,1株为迪氏分枝杆菌,1株为草分枝杆菌,1株为新金色分枝杆菌,芯片上无鉴定该菌种的探针。结论:应用膜芯片技术可简便、快速、灵敏、特异地将大多数分枝杆菌鉴定到种,指导临床合理治疗。
Objective To develop a rapid and simple method for identification of mycobacterial（M） species with the membrane chip technique, and to provide a thereunder for the correct diagnosis of mycobacterium disease. Methods DNA sequencing was used as the control, the target DNA fragments of 11 mycobacterial reference strains,9 non-mycobacterial strains and 199 mycobacterial isolates were analyzed by PCR-SSCP and membrane chip. Results The reference strains of 11 mycobacteria and 7 non-mycobacteria were analyzed by membrane chips, the results showed that the membrane chip was specific. Of 199 mycobacterial clinical isolates, 30 strains were identified as mycobacterium tuberculosis complex（MTBC） with PCR-SSCP, and were positive hybridization with mycobacterium genus probe M and MTBC-specific probe with membrane chip. One hundred and sixty-nine strains were identified as non -tuberculous mycobacterium（NTM）with PCR-SSCP, and their species were identified with membrane chips, including 58 strains of M. chelonae,46 of M. intracellulare, 33 of M. kansasii, M. gastri, M. scrofulaceum and M. simiae complex, 6 of M. fortuitum, 15 of M. gordonae, 3 of M. avium, 2 of M. marinum and M. abscess complex. Six strains that were only positive hybridization with probe M were sequenced and showed that 2 strains were M. intracellulare that had different DNA sequences with the reference strain, 1 was M. terrae, 1 was M .diernhoferi, 1 was M. phlei and 1 was M. neoaurum. There were not specific probes for them on membrane chip. Conclusion It may be a simple, rapid, sensitive and specific method for identification of most mycobacterial species by membrane chip, and it can raise correct diagnosis rate of mycobacterial disease, and instruct the physicians to perform rational chemotherapy.
出处
《实用医学杂志》
CAS
2007年第5期616-618,共3页
The Journal of Practical Medicine
基金
军队医学杰出中青年人才科研基金项目(编号:01J020)
关键词
分枝杆菌属
聚合酶链反应
单链构象多态性
菌种鉴定
基因芯片
Mycobacterium Polymerase chain reaction Single-stranded conformation polymorphism Species identification Membrane chip
