克罗诺杆菌属细菌数值鉴定系统的研制与效果评价

Development and effectiveness evaluation of a numerical identification system for Cronobacter

【背景】克罗诺杆菌属(Cronobacter)细菌的分离鉴定是临床医学中精准治疗的理论基础,然而,目前的生化鉴定方法仅可将其鉴定至属水平。【目的】开发一种简便、可靠的克罗诺杆菌属生化分种鉴定方法,建立克罗诺杆菌属细菌数值鉴定系统并对其鉴定性能进行评价。【方法】将鉴定系统所需的生化反应配方浓缩整合到一组注塑件中制得鉴定条,辅以数值鉴定理论构建克罗诺杆菌属细菌数值鉴定系统,利用克罗诺杆菌属标准株及分离株对其鉴定准确度进行验证。分离株的鉴定以fusA基因序列分析方法的鉴定结果作为参考标准,与数值鉴定法的检测结果进行比较。【结果】创建了用于鉴别克罗诺杆菌属内9个种/亚种的生化反应阳性概率数据库,同时确定了数值法中的计算方法和结果评价标准,并完成了数值鉴定系统软件的开发。5株克罗诺杆菌属标准株通过细菌数值鉴定系统均得到了正确鉴定,结果评价为极好的鉴定结果;其中,DSM 18707被准确鉴定为都柏林克罗诺杆菌奶粉亚种(C.dublinensis subsp.lactaridi)。测试集的84株克罗诺杆菌属分离株的数值鉴定法与fusA基因测序法的总鉴定一致率为100%;除C.sakazakii IV型(1株)及C.dublinensis III型(1株)的数值鉴定为不可接受的鉴定结果外,其余菌株均得到了明确的鉴定。【结论】本研究开发的克罗诺杆菌属细菌数值鉴定系统操作简便、价格低廉、准确率高,可为临床诊断及相关食品安全检测提供实验依据,具有良好的应用推广前景。

[Background]The isolation and identification of Cronobacter forms the theoretical basis for precise treatment in clinical medicine.However,the current biochemical identification methods can only identify them at the genus level.[Objective]To develop a simple and reliable method for species identification of Cronobacter,we established a numerical identification system and evaluated its identification performance.[Methods]The biochemical reaction formulations required for the identification system were concentrated and integrated into a set of plastic containers to prepare identification strips.The numerical identification system of Cronobacter was developed based on the numerical identification theory,and the identification accuracy was assessed with the standard strains and isolates of Cronobacter.The identification results based on the sequencing of fusA were taken as the reference and compared with those obtained with the numerical identification method.[Results]We established a biochemical reaction positive probability database for distinguishing the 9 species/subspecies of Cronobacter.Simultaneously,we defined the calculation methods and result evaluation criteria for the numerical method and developed a numerical identification system.Five standard strains of Cronobacter were correctly identified by the numerical identification system,and the results were evaluated as excellent.Among them,strain DSM 18707 was accurately identified as C.dublinensis subsp.lactaridi.The total identification agreement between the numerical identification method and the fusA sequencing method of the 84 Cronobacter isolates in the test set was 100%.The numerical identification method clearly identified other strains except C.sakazakii type IV(one strain)and C.dublinensis type III(one strain)with unacceptable identification results.[Conclusion]The numerical identification system developed in this study for Cronobacter.has simple operation,low cost,and high accuracy,which can provide an experimental basis for clinical diagnosis and food safety detection and demonstrates a promising prospect of application and popularization.