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| Real-time recombinase polymerase amplification for detection of Vibrio parahaemolyticus |
| ZHAN Li, XU Chang-ping, ZHANG Yun-yi, CHEN Hong-hu, ZHANG Zheng, CHEN Jian-cai, ZHANG Jun-yan, MEI Ling-ling
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| Zhejiang Provincial Center for Disease Control and Prevention,Hangzhou,Zhejiang 310051,China |
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Abstract Objective To establish real-time recombinase polymerase amplification（RPA）for the rapid detection of Vibrio parahaemolyticus（VP）.Methods An exo probe and primers were designed according to the conserved sequence of thermolabile hemolysin（tlh）gene of VP and then RPA for detection of VP was established. The sensitivity of the assay was evaluated by detecting different concentration of VP;the specificity was evaluated by detecting different bacteria;the stability was evaluated by repeat trials;the application effect was evaluated by detecting food samples which were simultaneously tested with traditional culture method according to GB 4798.7-2013 Detection of VP.Results A real-time RPA was established to complete VP amplification within 20 min at a constant temperature of 39 ℃. The analytical sensitivity of the assay was five pg per reaction and no cross-reactivity with other pathogenic bacteria observed. The RPA detection results with different concentration of VP and E. coli DNA templates at three time points were consistent. The detection results of 51 food samples by RPA were the same as those by traditional culture method.Conclusion The established real-time RPA can qualitatively detect VP,with simple operation and interpretation of results,which is suitable for rapid detection of VP in public health emergencies and food safety supervision.
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Received: 14 February 2019
Revised: 15 March 2019
Published: 02 July 2019
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