MALDI-TOF MS用于副溶血性弧菌检测初探
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摘要
目的评价基质辅助激光解析电离飞行时间质谱(MALDI-TOF MS)对副溶血性弧菌的识别能力,并将该方法用于副溶血性弧菌的同源性分析。方法运用标准菌株判断MALDI-TOF MS方法用于检测副溶血性弧菌的重复性及特异性,将从5种贝类中收集到的65株菌株运用MALDI-TOF MS做鉴定,PCA同源性分析。结果 MALDI-TOFMS对65株副溶血性弧菌的质谱鉴定结果与生化鉴定及PCR鉴定结果一致,即能有效区分副溶血性弧菌、创伤弧菌、蜡样芽胞杆菌、金黄色葡萄球菌、大肠埃希菌等5种细菌,其中在副溶血性弧菌,3%NaCl TSA培养24h后的结果鉴定效果更好。10次重复检测副溶血性弧菌的分值均在2.3分以上,PCA聚类分析将65株菌株被分成了4个大类,大多数菌株的PC1、PC2、PC3相对集中靠拢。结论 MALDI-TOF MS检测副溶血性弧菌具有较好的稳定性、特异性,对不同型别的细菌具有较强的分辨能力,且重复性好,同源性分析具有高通量、快速、简单、低成本等优点。
Objective To evaluate the ability of MALDI-TOF MS to recognize Vibrio parahaemolyticus and to use this technique to analyze its homology. Methods Standard strains tested with MALDI-TOF MS were used to evaluate the repeatability and specificity of the technique.Sixty-five strains collected from 5 species of shellfish were identified using MALDI-TOF MS and homology was analyzed using PCA. Results The results of mass spectrometry identification of65 strains were consistent with biochemical identification and PCR identification,and flight time mass spectrometry effectively distinguished five bacteria:Vibrio parahaemolyticus,V.traumatica,Bacillus cereus,Staphylococcus aureus,and Escherichia coli.The most appropriate conditions for MALDI-TOF MS identification of V.parahaemolyticus were after culturing on TSA with 3% NaCl for 24 h.The scores on 10 repetitions of testing for V.parahaemolyticus were all above2.3.The 65 strains were divided into 4 groups according to PCA with cluster analysis.PC1,PC2,and PC3 were relatively concentrated for most strains. Conclusion The MALDI-TOF MS technique has good consistency and specificity,and it is able to distinguish different types of bacteria.Analysis of homology with MALDI-TOF MS has the advantages of a high throughput,rapidity,simplicity,and a low cost.
Objective To evaluate the ability of MALDI-TOF MS to recognize Vibrio parahaemolyticus and to use this technique to analyze its homology. Methods Standard strains tested with MALDI-TOF MS were used to evaluate the repeatability and specificity of the technique.Sixty-five strains collected from 5 species of shellfish were identified using MALDI-TOF MS and homology was analyzed using PCA. Results The results of mass spectrometry identification of65 strains were consistent with biochemical identification and PCR identification,and flight time mass spectrometry effectively distinguished five bacteria:Vibrio parahaemolyticus,V.traumatica,Bacillus cereus,Staphylococcus aureus,and Escherichia coli.The most appropriate conditions for MALDI-TOF MS identification of V.parahaemolyticus were after culturing on TSA with 3% NaCl for 24 h.The scores on 10 repetitions of testing for V.parahaemolyticus were all above2.3.The 65 strains were divided into 4 groups according to PCA with cluster analysis.PC1,PC2,and PC3 were relatively concentrated for most strains. Conclusion The MALDI-TOF MS technique has good consistency and specificity,and it is able to distinguish different types of bacteria.Analysis of homology with MALDI-TOF MS has the advantages of a high throughput,rapidity,simplicity,and a low cost.
引文
[1] 龚艳清,陈信忠.MALDI-TOF-MS方法检测、鉴定副溶血性弧菌[J].食品安全质量检测学报,2016,4(2):521-7.
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[3] 刘瑛,俞静,陈峰,等.MicroflexTM MALDI-TOF MS和Vitek2Compact微生物分系统对肠杆菌科细菌鉴定能力的比较[J].检验医学,2015,30(2):122-7.
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[5] Suh MJ,Hamburg DM,Gregory ST,et al.Extending ribosomalprotein identifications to unsequenced bacterial strains using ma-trix-assisted laser desorption/inoization mass spectrometry[J].Proteomics,2005,5(18):4818-31.
[6] Christensen JJ,Dargis R,Hammer M,et al.Matrix-assisted laserdesorption/inoization-time of flight mass spectrometry analysis ofGram-positive,catalase-negative cocci not belonging to the strepto-coccus or Enterococcus genus and benefits of database ex-tention[J].J Clin Microbio,2012,50(5):1787-91.
[7] Wolters M,Rohde H,Maier T.MALDI-TOF MS fingerprintingal-lows for discrimination of major methicillin-resistant Staphylo-coccus aureus lineages[J].Int J Med Microbiol,2011(301):64-8.
[8] 郑秋月,战小微,肖珊珊,等.沙门氏菌MALDI-TOF MS分型方法建立[J].中国公共卫生,2014,10(30):1344-7.
[9] 王哗茹,崔生辉,李凤琴.基质辅助激光解吸/电离飞行时间质谱在沙门菌检测和鉴定分型中的应用研究[J].卫生研究,2008,37(6):685-9.
[2] 张慧芳,李莉,麻丽丹,等.基于MALDI-TOF MS的弧菌识别能力分析[J].中国病原生物学杂志,2016,6(11):522-34.
[3] 刘瑛,俞静,陈峰,等.MicroflexTM MALDI-TOF MS和Vitek2Compact微生物分系统对肠杆菌科细菌鉴定能力的比较[J].检验医学,2015,30(2):122-7.
[4] 屠博文,史伟峰,韩晓冬,等.MALDI-TOF质谱在院内感染致病菌检测中的应用[J].重庆医学,2015,11(33):4665-70.
[5] Suh MJ,Hamburg DM,Gregory ST,et al.Extending ribosomalprotein identifications to unsequenced bacterial strains using ma-trix-assisted laser desorption/inoization mass spectrometry[J].Proteomics,2005,5(18):4818-31.
[6] Christensen JJ,Dargis R,Hammer M,et al.Matrix-assisted laserdesorption/inoization-time of flight mass spectrometry analysis ofGram-positive,catalase-negative cocci not belonging to the strepto-coccus or Enterococcus genus and benefits of database ex-tention[J].J Clin Microbio,2012,50(5):1787-91.
[7] Wolters M,Rohde H,Maier T.MALDI-TOF MS fingerprintingal-lows for discrimination of major methicillin-resistant Staphylo-coccus aureus lineages[J].Int J Med Microbiol,2011(301):64-8.
[8] 郑秋月,战小微,肖珊珊,等.沙门氏菌MALDI-TOF MS分型方法建立[J].中国公共卫生,2014,10(30):1344-7.
[9] 王哗茹,崔生辉,李凤琴.基质辅助激光解吸/电离飞行时间质谱在沙门菌检测和鉴定分型中的应用研究[J].卫生研究,2008,37(6):685-9.