Detection of Vibrio parahaemolyticus virulence gene by cross primer thermostatic amplification technology
10.3760/cma.j.issn.1003-9279.2018.01.018
- VernacularTitle: 交叉引物恒温扩增技术在副溶血性弧菌毒力基因检测中的应用
- Author:
Xiaofang WU
1
;
Lei JI
1
;
Wei YAN
1
;
Shiwang HUANG
2
;
Xuejun MA
3
Author Information
1. Huzhou Center for Disease Control and Prevention Microbiology Laboralory, Huzhou 313000, Zhejiang, China
2. Jianggan District Center for Disease Control and Prevention Laboroatory, Hangzhou 310000, China
3. National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China
- Publication Type:Journal Article
- Keywords:
Vibrio parahaemolyticus;
Cross-primer amplification technique;
Virulence gene
- From:
Chinese Journal of Experimental and Clinical Virology
2018;32(1):85-88
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish a CPA-nucleic acid test strip method for rapid detection of Vibrio parahaemolyticus thermostable direct hemolysin (TDH) and heat-labile hemolysin (TLH).
Methods:Primers and probes were designed according to the sequence of TDH and TLH virulence genes of Vibrio parahaemolyticus, The optimal reaction temperature and time were established by optimizing the reaction system. The CPA method was compared with the fluorescence quantitative PCR.
Results:The optimal temperature and time for the CPA-nucleic acid test strip method established in this study were 60 ℃ and 40 min, which were highly specific to the TDH and TLH virulence genes of Vibrio parahaemolyticus. The TLH And TDH detection limit of 1.8 cfu/mL and 16.0 cfu/mL; consistent with the method of fluorescence quantitative PCR, and has good stability. The positive rates of TLH and TDH genes were 100% (464/464) and 72.6% (337/464) in 464 isolates, respectively. The positive rate of TDH virulence genes in isolates of aquatic products was 7.81 % (10/128). The positive rate of TDH virulence gene in clinical isolates was 97.32% (327/336).
Conclusions:The CPA-nucleic acid test strip method for detection of Vibrio parahaemolyticus TDH and TLH genes is rapid, efficient, sensitive, specific and easy to operate. It can be used in the rapid field detection of Vibrio parahaemolyticus.
