College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, PR China.
College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, PR China.
J Microbiol Methods. 2021 Jan;180:106100. doi: 10.1016/j.mimet.2020.106100. Epub 2020 Nov 27.
To identify the main spoilage bacterium on fresh-cut leafy vegetables and establish a multiplex PCR assay.
Based on physiological-biochemical, molecular identification, and artificial contamination tests, the main bacterium to spoil fresh-cut leafy vegetables was identified as Exiguobacterium spp. and Exiguobacterium acetylicum. Comparative genomics showed that P401_RS0117025 and oxi_50,582,462 genes are specific to Exiguobacterium spp. and E. acetylicum. Based on this, three pairs of primer sets to EaG-291, EaS-2B, and Ea16S-12 genes were designed and used to develop a multiplex PCR assay, which exhibited 100% specificity among 16 Exiguobacterium and 10 non-Exiguobacterium strains. Finally, 84 fresh-cut leafy vegetable samples were analyzed by multiplex PCR assay and standard physiological-biochemical experiments, the results showed multiplex PCR assay reached a detection rate of 96%.
The main spoilage bacterium was identified as Exiguobacterium spp. and E. acetylicum on fresh-cut leafy vegetables based on the novel specific genes explored in this study.
A rapid, specific, and sensitive PCR assay was developed for the detection of Exiguobacterium spp. and E. acetylicum.
鉴定鲜切叶类蔬菜中的主要腐败菌，并建立多重 PCR 检测方法。
基于生理生化、分子鉴定和人工污染试验，确定鲜切叶类蔬菜的主要腐败菌为节杆菌属和醋酸钙不动杆菌。比较基因组学表明，P401_RS0117025 和 oxi_50、582、462 基因是节杆菌属和醋酸钙不动杆菌特有的。基于此，设计了三对针对 EaG-291、EaS-2B 和 Ea16S-12 基因的引物对，建立了多重 PCR 检测方法，该方法在 16 株节杆菌和 10 株非节杆菌菌株中具有 100%的特异性。最后，用多重 PCR 检测方法和标准生理生化实验对 84 份鲜切叶类蔬菜样品进行了分析，结果表明，该方法的检测率达到 96%。
基于本研究中探索的新的特异性基因，鉴定出鲜切叶类蔬菜中的主要腐败菌为节杆菌属和醋酸钙不动杆菌。
建立了一种快速、特异、敏感的检测节杆菌属和醋酸钙不动杆菌的 PCR 检测方法。
