Zhejiang Provincial Key Laboratory of Biometrology and Inspection & Quarantine, China Jiliang University, Xueyuan Street, Xiasha Higher Education Zone, Hangzhou 310018, China.
World J Microbiol Biotechnol. 2012 Mar;28(3):1013-20. doi: 10.1007/s11274-011-0899-8. Epub 2011 Oct 8.
Contamination of Cronobacter spp. (Enterobacter sakazakii) in infant formulas and other food products is a severe problem. Here a loop-mediated isothermal amplification (LAMP) assay was developed for rapidly detecting Cronobacter spp. in powdered infant formula. Sequences of 16S/23S rDNA internal intergenic spacer of Cronobacter spp. were used as the target template to design LAMP primers. The detection outcome can be evaluated by the white precipitate or the fluorescence intensity under ultraviolet irradiation, both visible to naked eyes. The sensitivity and specificity of the LAMP assay was further analyzed in comparison with that of regular PCR and real time quantitative PCR. The results showed that all of Cronobacter spp. strains display positive reaction to the detections while all of the non-Cronobacter spp. strains were negative, and that the LAMP assay exhibits a high sensitivity of 9.1 fg/μL (The sensitivity of regular PCR and real time quantitative PCR is 91 and 9.1 pg/μL, respectively.). The amplified reaction could be accomplished in about 1 h, with the results visible to naked eyes. Hence, the LAMP assay developed by this study can provide a rapid and simple approach for the detection of Cronobacter spp. in infant formula.
阪崎克罗诺杆菌（Enterobacter sakazakii）污染婴儿配方奶粉和其他食品是一个严重的问题。本研究开发了一种环介导等温扩增（LAMP）检测方法，用于快速检测婴儿配方奶粉中的阪崎克罗诺杆菌。以阪崎克罗诺杆菌 16S/23S rDNA 内基因间隔区序列为靶标模板设计 LAMP 引物。检测结果可通过肉眼可见的白色沉淀或紫外照射下的荧光强度进行评估。与常规 PCR 和实时定量 PCR 相比，进一步分析了 LAMP 检测方法的灵敏度和特异性。结果表明，所有阪崎克罗诺杆菌菌株均对检测呈阳性反应，而所有非阪崎克罗诺杆菌菌株均为阴性，LAMP 检测具有很高的灵敏度（9.1 fg/μL，常规 PCR 和实时定量 PCR 的灵敏度分别为 91 和 9.1 pg/μL）。扩增反应可在约 1 小时内完成，结果肉眼可见。因此，本研究开发的 LAMP 检测方法可提供一种快速简便的方法来检测婴儿配方奶粉中的阪崎克罗诺杆菌。
