摘要
目的建立水产品中溶藻弧菌的基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)的快速鉴定方法。方法以溶藻弧菌标准菌株(CICC 10889)为研究对象,采集不同样品处理方法 (直接涂抹法和甲酸提取法)、激光强度和菌株传代次数的MALDI-TOF MS图谱并加以分析比较,探讨上述因素对方法准确性的影响。将溶藻弧菌标准菌株(CICC 10889)的MALDI-TOF MS图谱主产物(MSP)添加到数据库中,使用36株分离菌株及8株与溶藻弧菌近缘的弧菌标准菌株验证数据库的补充对鉴定结果可信度的影响。结果甲酸提取法处理所得图谱特征峰多、基线平滑、信噪比高,图谱质量优于直接涂抹法;激光强度对MALDI-TOF MS图谱有影响,过高或过低均会导致图谱质量的下降;标准菌株补充数据库后可提高鉴定可信度,分离株的鉴定分值均达到2.300以上;传代次数对鉴定结果没有影响。结论 MALDI-TOF MS方法可将溶藻弧菌准确鉴定到种水平。该方法准确、可靠,可用于溶藻弧菌的快速鉴定。
Objective To establish a rapid identification method of Vibrio alginolyticus using matrix assisted laser desorption/ionization time of flight mass spectrometry(MALDI-TOF MS).Methods The effect of different sample pretreat method(direct smear method and formic acid extraction method),laser intensity and times of subculture on the quality of spectrum was investigate using the reference strain CICC 10889.The impact of database on identification result was evaluate by comparing the identification result(species and Logscore)of 36 isolated strains before and after adding the main spectra product(MSP)of CICC 10889 to the database.Results The formic acid extraction method was better than the direct smear method.Spectra of ethanol/formic acid treatment got more characteristic peak of protein with smooth baseline,less noise and higher signal-noise ratio.Laser intensity could affect spectra.High quality spectra could be obtained when the laser intensity was appropriate.Database was the most important impact factor.The Logscore of identification result was improved greatly by adding the in-house database,score of isolated strains were all above 2.300.Times of subculture had no effect on spectra.Conclusion MALDI-TOF MS could accurately identify Vibrio alginolyticus to the species level.The established MALDI-TOF MS method was accurate and constant and could be used to identify Vibrio alginolyticus rapidly.
作者
赵晓娟
汪琦
王紫薇
徐蕾蕊
刘莉
韩笑
曾静
ZHAO Xiaojuan;WANG Qi;WANG Ziwei;XU Leirui;LIU Li;HAN Xiao;ZENG Jing(Beijing Customs,Beijing 100026,China)
出处
《中国食品卫生杂志》
北大核心
2018年第6期586-591,共6页
Chinese Journal of Food Hygiene
基金
"十三五"国家重点研发计划(2016YFD0401102)
国家质检总局科技计划项目(2017IK145)
北京检验检疫技术中心自立科研课题(2017JK008)
关键词
溶藻弧菌
基质辅助激光解吸电离飞行时间质谱
鉴定
影响因素
水产品
Vibrio alginolyticus
matrix assisted laser desorption/ionization time of flight mass spectrometry
identification
influencing factors
aquatic products
作者简介
赵晓娟,女,工程师,研究方向为食品安全与微生物检测,E-mail:zhaoxj@bjciq.gov.cn;通信作者:曾静,女,研究员,研究方向为食品安全与微生物检测,E-mail:zengj@bjciq.gov.cn.
