基质辅助激光解析电离飞行时间质谱技术快速鉴定耐万古霉素屎肠球菌

Rapid identification of vancomycin-resistant Enterococcus faecium by MALDI-TOF MS

目的:利用全自动生物质谱检测系统即基质辅助激光解析电离飞行时间质谱仪(matrix-assisted laser desorption/ionization time-of-flight mass spectrometry,MALDI-TOF MS)快速鉴别耐万古霉素屎肠球菌(vancomycin resistant Enterococcus faecium,VRE)和万古霉素敏感屎肠球菌(vancomycin susceptible Enterococcus faecium,VSE)。方法:收集临床分离的屎肠球菌,用仪器法和纸片扩散法(Kirby-Bauer,K-B)进行万古霉素耐药性确认。用MALDI-TOF MS对菌株进行再鉴定,采集图谱。用Clin Pro Tools 3.0软件对质谱峰图进行分析:随机选取25株VRE和50株VSE的质谱峰图用于模型的建立,用建立的模型验证剩余的15株VRE和35株VSE。结果:用Clin Pro Tools 3.0软件建立了遗传算法(GA)、快速分类算法(QC)、监督式神经网络算法(SNN),3种模型的交叉验证和识别能力分别为95.65%、100%,87.62%、94.98%,62.54%、100%。质荷比为2 194.94、3 022.15、3 167.47和4 058.08的质谱峰是用于区分VRE和VSE的最为明显的特征性峰,即两类菌株间存在的差异蛋白质谱峰(分别是峰6、峰27、峰32、峰52)。受试者工作特征曲线显示,这4个特征性峰的曲线下面积(AUC)均约为0.9。用3种模型分别进行的外部验证显示,GA算法、QC算法、SNN算法对VRE的敏感性和特异性分别为86.7%、82.8%,73.3%、85.7%,66.7%、80.0%。结论:在严格控制实验条件的情况下,MALDI-TOF MS可以在鉴定屎肠球菌的同时大致区分VRE和VSE,具有耗时短,敏感性、特异性相对较高的优势。

Objective: To identify vancomycin resistant Enterococcus faecium( VRE) and vancomycin susceptible Enterococcus faecium( VSE) by using the matrix-assisted laser desorption/ionization time-offlight mass spectrometry( MALDI-TOF MS). Methods: The clinically isolated Enterococcus faecium were collected,and the resistance of vancomycin was confirmed by instrumental method and Kirby-Bauer( K-B)method. The strain was identified by MALDI-TOF MS and the map was collected. The mass spectra were analyzed by Clin Pro Tools 3. 0 software. The mass spectra of 25 VRE and 50 VSE were randomly selected for the establishment of the model. The remaining 15 VRE and 35 VSE were validated by the established model. Results: The genetic algorithm( GA),fast classification algorithm( QC),supervised neural network algorithm( SNN) were established by Clin Pro Tools 3. 0 software. The cross validation andrecognition capabilities of the three models were 95. 65%/100%,87. 62%/94. 98%,62. 54%/100%.Mass spectral ratios of 2 194. 94,3 022. 15,3 167. 47 and 4 058. 08 m/z were the most significant characteristic peaks for distinguishing VRE and VSE,i. e.,differential protein peaks between the two strains( peak 6,peak 27,peak 32,peak 52). The receiver operator characteristic curve showed that the area under curve of these four characteristic peaks was about 0. 9. Three types of models were used for external validation: the sensitivity and specificity of the GA,QC,SNN algorithm for the VRE were 86. 7%/82. 8%,73. 3%/85. 7%,66. 7%/80. 0%. Conclusion: MALDI-TOF MS can be used for rapid identification of VRE and VSE with strict control of experimental conditions,which is time-saving,sensitive and specific.