Department of Hygienic Inspection, School of Public Health, Jilin University, 1163 Xinmin Street, Changchun, 130021, Jilin, China.
Research Laboratory, Changchun Children's Hospital, Changchun, 130061, Jilin, China.
Anal Bioanal Chem. 2020 Jan;412(1):93-101. doi: 10.1007/s00216-019-02209-y. Epub 2019 Dec 3.
The aim of this study was to develop an effective and specific visual method to rapidly detect and identify Vibrio parahaemolyticus (V. parahaemolyticus) based on the polymerase spiral reaction (PSR). The method utilized only two pairs of primers designed specifically to target the conserved tlh gene sequence of V. parahaemolyticus. Nucleic acid amplification can be achieved under isothermal conditions using DNA polymerase. The reaction could be accomplished in < 40 min with high specificity and sensitivity. The limits of detection of V. parahaemolyticus in purified genomic DNA and pure culture were 300 fg/μL and 2.4 CFU/mL per reaction, respectively, which were 100-fold more sensitive than with conventional PCR. The model food samples showed consistent specificity and sensitivity to the pure bacterial culture. With these encouraging results, it is expected that the novel, effortless and reliable isothermal nucleic acid testing assay developed in this study has potential to be applied to screening for V. parahaemolyticus in seafood samples.
本研究旨在开发一种基于聚合酶螺旋反应（PSR）的有效且特异的可视化方法，用于快速检测和鉴定副溶血性弧菌（V. parahaemolyticus）。该方法仅使用两对专门针对副溶血性弧菌保守 tlh 基因序列设计的引物。在等温条件下，使用 DNA 聚合酶即可实现核酸扩增。该反应可在 < 40 分钟内完成，具有高特异性和灵敏度。在纯化基因组 DNA 和纯培养物中的副溶血性弧菌检测限分别为 300 fg/μL 和每反应 2.4 CFU/mL，比常规 PCR 灵敏 100 倍。模型食品样品对纯细菌培养物表现出一致的特异性和灵敏度。有了这些令人鼓舞的结果，预计本研究中开发的新型、轻松和可靠的等温核酸检测方法有望应用于海产品样本中副溶血性弧菌的筛选。
