Simultaneous detection of Vibrio cholerae, Vibrio alginolyticus, Vibrio parahaemolyticus and Vibrio vulnificus in seafood using dual priming oligonucleotide (DPO) system-based multiplex PCR assay
Simultaneous detection of Vibrio cholerae, Vibrio alginolyticus, Vibrio parahaemolyticus and Vibrio vulnificus in seafood using dual priming oligonucleotide (DPO) system-based multiplex PCR assay
复制标题
DOI：
10.1016/j.foodcont.2016.06.024
复制
发表时间：
2017
期刊：
影响因子：
6
通讯作者：
Yigang Xu;Liu-Mei Sun;Yusai Wang;Peipei Chen;Zhongmei Liu;Yi-jing Li;Lijie Tang
中科院分区：
文献类型：
--
作者：
Yigang Xu;Liu-Mei Sun;Yusai Wang;Peipei Chen;Zhongmei Liu;Yi-jing Li;Lijie Tang
In this study, a rapid and reliable multiplex PCR assay for simultaneous detection of Vibriocholerae, Vibrio parahaemolyticus, Vibrio vulnificus and Vibrio alginolyticus in seafood was developed using the dual priming oligonucleotide (DPO) system. Species-specific DPO primers were designed targeting themdh,vvhA,colHandtoxRgenes for the discrimination ofV. cholerae,V.vulnificus,V.alginolyticusandV.parahaemolyticus, respectively. Compared to conventional PCR assay, the DPO system-based multiplex PCR assay allowed a wider annealing temperature at 48 °C–68 °C to effectively amplify target genes followed an analytical detection limit of <1.5 × 102CFU/mL (or g) for each Vibrio species in pure cultures or artificially contaminated food matrix. A total of 396 bacterial strains including 209 targets and 187 other bacterial strains were used to test the specificity of the DPO system-based multiplex PCR assay, and results showed that specific PCR product was only observed in target bacterial strains without nonspecific priming. Practical application of the assay indicated that targetVibriospecies in seafood, clinical samples and foodborne outbreaks can be accurately detected. This DPO system-based multiplex PCR assay developed in this study would be a powerful tool for the rapid and reliable detection of the targetVibriospecies.
