基质辅助激光解吸电离飞行时间质谱对阪崎肠杆菌的鉴定

Identification of Enterobacter sakazakii by matrix-assisted laser desorption ionization-time of flight mass spectrometry

目的利用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)法对阪崎肠杆菌进行鉴定,建立一种高效检测阪崎肠杆菌的方法,并为该技术的推广使用及阪崎肠杆菌的进一步研究提供科学依据。方法用MALDI-TOF-MS法检测38株野生阪崎肠杆菌、2株标准菌株和1株阴沟肠杆菌,结果与常规生化鉴定结果对比;同时对在不同培养基上培养的阪崎肠杆菌进行质谱分析比较,对比不同培养基对质谱结果是否有影响;对38株野生菌株质谱图进行聚类分析。结果 38株菌株鉴定结果均为阪崎肠杆菌,与生化鉴定结果一致,且质谱鉴定分值大多在2.0以上。通过MALDI-TOF-MS鉴定方法可以很明显地将阴沟肠杆菌与阪崎肠杆菌两种菌分开。4种培养基对MALDI-TOF-MS鉴定结果的影响不是很明显,TSA比较适合作为阪崎肠杆菌MALDI-TOF-MS鉴定的培养基。通过质谱图谱和离子峰值比较得出,所有菌株在5745 m/z附近均出现高的离子峰,在2871、4740、8288、6260和9488 m/z附近出现离子峰的实验菌株达95%以上;在差异水平在0.5时,MALDI-TOF-MS的聚类分析结果可将所有实验菌株分成5个类型,结合菌株对应的来源和种类分析表明本研究所用菌株与来源和种类之间并无明显关系。结论MALDI-TOF-MS方法具有准确且精确鉴定阪崎肠杆菌的能力;离子峰5745m/z具有作为阪崎肠杆菌的标记性离子峰的可能;差异水平为0.5进行MALDI-TOF-MS聚类分析,未发现5个类型与来源等具有一定关系,需要进一步研究。

Objective To establish a highly effective method for identification of Enterobacter sakazakii and provide scientific basis for the promotion of the technique and further studies on the pathogen.Methods 38 wild strains of Enterobacter sakazakii,2 reference strains and 1 strain of Enterobacter cloacae were identified with matrix-assisted laser desorption ionization-time of flight mass spectrometry（MALDI-TOF MS）.The results of MALDI-TOF-MS were compared to biochemical results.Identifications of Enterobacter sakazakii and Enterobacter cloacae cultivated on different media were performed with MALDI-TOF-MS;the results were compared to see whether different media could affect the MALDI-TOF-MS results.Cluster analysis was performed on all strains.Results The identification results by MALDI-TOF-MS of 40 strains of Enterobacter sakazakii were of very high accuracy and consitent with biochemical results.MALDI-TOF-MS had differentiated Enterobacter sakazakii from Enterobacter cloacae distinctively.The4 culture media didn＇t show obvious effect on the MALDI-TOF-MS results.The quasi-molecular ions of the 40 strains were very high at 5745 m/z.About 95 percent of the 40 strains had a quasi-molecular ion at 2871,4740,8288,6260,and 9488 m/z.From the dendrogram,5 kinds of MALDI-TOF-MS types appeared at distance level of 0.5.Conclusion MALDI-TOF-MS has ability to identify Enterobacter sakazakii,reference strains and Enterobacter cloacae correctly and exactly in this study.The character quasi-molecular ions on 5745 m/z could be a signal one for Enterobacter sakazakii.40 strains contained 5 kinds of MALDI-TOF-MS types were appeared at distance level 0.5 for the dendrogram.The correlation between the dendrogram and the source and kinds of all strains of Enterobacter sakazakii is not obviously,and more study about this aspect is necessary.