张慧芳, 龚杰, 张炳华, 张建中, 肖迪. 基于MALDI-TOF MS预提取样本制备法废弃成分识别病原菌的方法建立及识别能力评价[J]. 疾病监测, 2019, 34(11): 964-968. DOI: 10.3784/j.issn.1003-9961.2019.11.005
引用本文: 张慧芳, 龚杰, 张炳华, 张建中, 肖迪. 基于MALDI-TOF MS预提取样本制备法废弃成分识别病原菌的方法建立及识别能力评价[J]. 疾病监测, 2019, 34(11): 964-968. DOI: 10.3784/j.issn.1003-9961.2019.11.005
Huifang Zhang, Jie Gong, Binghua Zhang, Jianzhong Zhang, Di Xiao. Establishment and evaluation of a pathogen identification assay based on waste components of MALDI-TOF MS pre-extraction sample preparation[J]. Disease Surveillance, 2019, 34(11): 964-968. DOI: 10.3784/j.issn.1003-9961.2019.11.005
Citation: Huifang Zhang, Jie Gong, Binghua Zhang, Jianzhong Zhang, Di Xiao. Establishment and evaluation of a pathogen identification assay based on waste components of MALDI-TOF MS pre-extraction sample preparation[J]. Disease Surveillance, 2019, 34(11): 964-968. DOI: 10.3784/j.issn.1003-9961.2019.11.005

基于MALDI-TOF MS预提取样本制备法废弃成分识别病原菌的方法建立及识别能力评价

Establishment and evaluation of a pathogen identification assay based on waste components of MALDI-TOF MS pre-extraction sample preparation

  • 摘要: 目的为提高基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)鉴定病原菌的能力,尤其是对微量病原菌的鉴定能力,建立一种基于预提取样本制备法的废弃成分进行病原菌识别鉴定的新方法。方法选择63株病原菌(23株细菌、40株真菌),培养后采用常规乙醇/甲酸法提取病原菌蛋白后,利用废弃沉淀物涂抹样本靶,与基质结合后进行MALDI-TOF MS数据采集及分析,并与常规分析成分进行病原菌识别能力比较。结果常规乙醇/甲酸法将50株菌(79.37%)识别到种水平,将10株菌(15.87%)识别到属水平,3株菌(4.76%)未被识别;沉淀涂抹法将49株菌(77.78%)识别到种水平,将14株菌(22.22%)识别到属水平。 在同样采集条件下,沉淀涂抹法的数据采集能力略强于常规法;在同样信噪比下,2种方法所获得的质谱峰数目没有统计学差异。结论新的基于MALDI-TOF MS进行病原菌鉴定的样本采用方法,一次实验即可获得2种样本,达到了样本资源的最大限度利用,可提高MALDI-TOF MS鉴定病原菌尤其是单菌落病原菌的能力,为病原菌质谱高通量快速识别鉴定能力的增强开辟了新的途径。

     

    Abstract: ObjectiveTo improve the pathogen identification power, especially the identification of trace pathogen, of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), a new sample adoption assay was established by using the waste components of its pre-extraction sample preparation.MethodsIn this study, 63 freshly cultured pathogen strains (including 23 strains of bacteria and 40 strains of fungi) were used. After ethanol/formic acid extraction of protein of the pathogen strains, the waste sediment was used to smear the sample target, then the MALDI-TOF MS data collection and analysis were conducted after the combination with the matrix. The identification power for the pathogens was compared with conventional analysis components.ResultsUsing the conventional ethanol/formic acid assay, 50 strains (79.37%) were identified at species level, 10 strains (15.87%) were identified at genus level and 3 strains (4.76%) were not identified. Using precipitation smear assay, 49 strains (77.78%) were identified at species level, 14 strains (22.22%) were identified at genus level. Under the same collection condition, the data collection ability of precipitation smear assay was slightly better than that of conventional assay, and there was no statistical difference in the number of mass spectra peaks obtained by two methods under the same signal-to-noise ratio.ConclusionIn this study, a new pathogen identification assay based on waste components of MALDI-TOF MS pre-extraction sample preparation was established. Two samples can be obtained in one experiment, maximizing the use of sample resources. This method can improve the identification power of MALDI-TOF MS for pathogens, especially for single colony pathogen, providing us a new way for the enhancement of high-throughput and rapid identification of pathogens by mass spectrometry.

     

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