Zimmermann Jennifer, Schmidt Herbert, Loessner Martin J, Weiss Agnes
Institute of Food Science and Biotechnology, Department of Food Microbiology, University of Hohenheim, Garbenstrasse 28, 70599 Stuttgart, Germany.
Institute of Food, Nutrition and Health, ETH Zurich, Schmelzbergstrasse 7, 8092 Zurich, Switzerland.
Food Microbiol. 2014 Sep;42:19-25. doi: 10.1016/j.fm.2014.02.010. Epub 2014 Feb 24.
Certain species of the genus Cronobacter are considered opportunistic pathogens, but their detection in milk products according to ISO/TS 22964 may take up to six days. The aim of this study was to develop a fast and sensitive PCR-based detection system for these species including enrichment, DNA-isolation and detection by real-time PCR, using the outer membrane protein gene ompA as a target. The assay was successfully validated using type strains of the genus Cronobacter, as well as 18 strains of closely related genera as controls. A total of 40 Cronobacter spp. food isolates yielded positive results, while the food matrix itself did not influence the PCR reaction. An equal detection limit as achieved with the ISO/TS 22964 method was established in this study, when 0.01 CFU Cronobacter sakazakii DSM 4485(T) per gram powdered infant formula were successfully detected after 28 days of storage at ambient temperature. In comparison to the ISO/TS 22964 method, the method described here has an equal detection limit, but offers a specific detection at the genus level in an analysis time of 24 h.
克罗诺杆菌属的某些菌种被视为机会致病菌，但根据ISO/TS 22964标准在乳制品中检测它们可能需要长达6天的时间。本研究的目的是开发一种基于PCR的快速灵敏检测系统，用于检测这些菌种，包括富集、DNA提取以及通过实时PCR进行检测，以外膜蛋白基因ompA作为靶标。该检测方法使用克罗诺杆菌属的模式菌株以及18株密切相关属的菌株作为对照进行了成功验证。总共40株克罗诺杆菌属食品分离株得到了阳性结果，而食品基质本身并未影响PCR反应。在本研究中，当每克婴儿配方奶粉中含有0.01 CFU阪崎克罗诺杆菌DSM 4485(T)在室温下储存28天后成功检测到时，确立了与ISO/TS 22964方法相同的检测限。与ISO/TS 22964方法相比，此处描述的方法具有相同的检测限，但能在24小时的分析时间内实现属水平的特异性检测。
