CAJ | 학술논문

目的建立基质辅助激光解析/电离飞行时间质谱(MALDI-TOF MS)鉴定非结核分枝杆菌(NTM)的方法,评估其快速鉴定临床和环境分离NTM的可行性、准确性和重复性.方法回顾性收集2012年1至3月上海复旦大学附属中山医院微生物实验室临床分离NTM 15株及2011至2012年医院供水系统环境分离NTM 68株.比较3种样本前处理和蛋白抽提方案,采用MALDI-TOFMS技术检测临床和环境分离NTM,并以16S rRNA测序结果为对照.结果 3种方法均鉴定出相同细菌,但方法1的鉴定值高于其他2种方法.对生长于Middle brook 7H10琼脂平板的83株共10个种的NTM进行鉴定,与16S rRNA测序结果比较,所有菌株均能鉴定至属水平,67株(80.7％)鉴定至种,6株鉴定到复合群水平,鉴定值大于1.700的NTM占81.9％ (68/83),且重复性好.培养后获得的菌落从前处理至鉴定完成仅需1.5h.结论 MALDI-TOF MS可快速准确地鉴定临床和环境分离的NTM,在临床实验室常规鉴定方面有着较好的应用前景.
목적 건립기질보조격광해석/전리비행시간질보(MALDI-TOF MS)감정비결핵분지간균(NTM)적방법,평고기쾌속감정림상화배경분리NTM적가행성、준학성화중복성.방법 회고성수집2012년1지3월상해복단대학부속중산의원미생물실험실림상분리NTM 15주급2011지2012년의원공수계통배경분리NTM 68주.비교3충양본전처리화단백추제방안,채용MALDI-TOFMS기술검측림상화배경분리NTM,병이16S rRNA측서결과위대조.결과 3충방법균감정출상동세균,단방법1적감정치고우기타2충방법.대생장우Middle brook 7H10경지평판적83주공10개충적NTM진행감정,여16S rRNA측서결과비교,소유균주균능감정지속수평,67주(80.7％)감정지충,6주감정도복합군수평,감정치대우1.700적NTM점81.9％ (68/83),차중복성호.배양후획득적균락종전처리지감정완성부수1.5h.결론 MALDI-TOF MS가쾌속준학지감정림상화배경분리적NTM,재림상실험실상규감정방면유착교호적응용전경.
Objective The purpose of this study was to compare and develop the method for identification of Non-tuberculous Mycobacteria (NTM) using matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS),and evaluate the feasibility,accuracy and repeatability of MALDI-TOF MS to discriminate NTM.Methods Fifteen clinical strains were collected from January to March in 2012 and 68 environmental strains were retrospectively collected from 2011 to 2012.A protocol for sample pre-treatment and protein extraction was developed and utilized it to identify clinical and environmental isolates.The results from 16 s rRNA sequencing were served as control.Results Method A was more effective in protein extraction.Although all the three methods got the same species result,a total of 83 strains belonging to 10 distinct species grown in Middle brook 7H10 media were analyzed.All members of the NTM were identified accurately at the genus level and 80.7％ (67/83) of strains could be identified at the species level.Six strains were identified at the complex level.81.9％ (68/83) of NTM got high spectral scores.The identification of cultured colony could be completed in 1.5 hours.And it had good reproducibility.Conclusion MALDI-TOF MS can be used as a rapid and accurate method for identification of Mycobacteria in clinical microbiology laboratories,implying its good application prospects.