应用MALDI-TOF MS技术快速鉴定布鲁菌  

Rapid identification of Brucella by MALDI-TOF MS

作　　者：俞静[1] 陈峰[1] 李媛睿[1] 刘瑛[1] YU Jing;CHEN Feng;LI Yuanrui;LIU Ying(Department of Clinical Laboratory,Xinhua Hospital Affiliated to School ofMedicine of Shanghai Jiaotong University,Shanghai 200092,China)

机构地区：[1]上海交通大学医学院附属新华医院检验科,上海200092

出　　处：《国际检验医学杂志》2018年第12期1443-1447,共5页International Journal of Laboratory Medicine

基　　金：上海申康医院发展中心市级医院临床辅助科室能力建设项目(SHDC22014003)

摘　　要：目的利用MicroflexTMLT型MALDI-TOF质谱仪建立布鲁菌蛋白质量指纹图谱数据库,并评估其在布鲁菌临床分离株鉴定中的应用价值。方法 2013年10月该院微生物实验室分离到1株疑似布鲁菌,经VITEK~2 Compact全自动微生物分析系统及16SrDNA基因测序确认为马耳他布鲁菌,按照厂家推荐方法自建了布鲁菌蛋白质量指纹图谱数据库。对2015年4月及2016年6月该实验室分离到的2株疑似布鲁菌进行革兰染色、柯氏染色及氧化酶、触酶、快速尿素酶等生化反应试验。使用Microflex^(TM)LT型质谱仪收集这2株及外院3株布鲁菌疑似菌株的质量图谱,比对分析图谱时在MALDI Biotyper数据库基础上添加自建数据库,同时使用16SrDNA基因测序法鉴定上述细菌。结果按照自建库推荐操作流程成功获得了基于BRUxh1的自建布鲁菌质量指纹图谱数据库。2015年4月及2016年6月分离到的布鲁菌疑似菌株,涂片染色镜检均为革兰阴性短小球杆菌,氧化酶阳性,触酶阳性,尿素酶阳性(5min内)。这5株疑似菌株的质谱鉴定结果均为布鲁菌(质谱评分分别为2.407、2.475、2.436、2.466、2.397),16SrDNA基因测序确认这5株细菌为布鲁菌。其中来自外院的2株细菌于外院VITEK~2 Compact系统鉴定时均错误地鉴定为吉氏玫瑰单胞菌。结论联合MALDI Biotyper数据库与自建布鲁菌数据库对临床分离株进行质量图谱比对分析,可以实现对布鲁菌属细菌的准确鉴定,该方法快速、灵敏,对布鲁菌病的临床快速诊断具有较高的应用价值。Objective To establish Brucellapeptide mass fingerprint database by MicroflexTMLT MALDITOF mass spectrometer,and evaluate its application value in the identification of clinical isolates of Brucella.Methods In October 2013,a suspected Brucellastrain was isolated in the microbiology laboratory of the hospital.It was confirmed as Brucella meltensis by VITEK~2 Compact automatic microbial analysis system and 16 SrDNA gene sequencing.Brucella peptide mass fingerprint database was established with this strain according to the manufacturer′s recommended method.Two suspected Brucellastrains isolated in the laboratory in April 2015 and June 2016 were tested for Gram stain,Kozlowski stain and biochemical reactions such as oxidase,catalase and rapid urease activity.MicroflexTMLT mass spectrometer was used to collect the mass spectrums of these two strains and three suspected Brucella strains isolated from other hospitals.When these spectrums were comparatively analyzed,self-built database was added on the basis of the MALDI Biotyper database.The 16 SrDNA gene sequencing was also used to identify the above-mentioned bacteria.Results According to the recommended self-built database construction process,Brucella peptide mass fingerprint database based on BRUxh1 was obtained successfully.The suspected Brucella strains isolated in April 2015 and June 2016 showed small Gram-negative coccobacillus under microscopic examination of stained smear,and were positive for oxidase,catalase,urease activity（within 5 min）.The identification results of mass spectrometry for the 5 suspected strains were all Brucella（mass spectrometry score：2.407,2.475,2.436,2.466 and2.397）.The 5 strains were confirmed as Brucellaby 16 SrDNA gene sequencing.Two strains among bacteria from other hospitals were falsely identified as Roseomonas gilardii by the VITEK~2 Compact system.Conclusion The combination of MALDI Biotyper database and self-built Brucelladatabase can be used to comparatively analyze the mass spectrometry of clinical isolates.

关 键 词：基质辅助激光解析电离飞行时间质谱 自建库 布鲁菌 鉴定 

分 类 号：R446.5[医药卫生—诊断学]