African Journal of
Microbiology Research

  • Abbreviation: Afr. J. Microbiol. Res.
  • Language: English
  • ISSN: 1996-0808
  • DOI: 10.5897/AJMR
  • Start Year: 2007
  • Published Articles: 5232

Full Length Research Paper

Application of an improved loop-mediated isothermal amplification detection of Vibrio parahaemolyticus from various seafood samples

Li Wang1, Xihong Zhao2, Jin Chu3, Yue Li1, Yanyan Li4, Chunhou Li5*, Zhenbo Xu2,6*and Qingping Zhong1
  1Food Safety Key Laboratory of Guangdong Province, College of Food Science, South China Agricultural University, Guangzhou 510642, China. 2College of Light Industry and Food Sciences,South China University of Technology, Guangzhou 510640, China. 3School of Food Science and Nutrition, Faculty of Mathematics and Physical Sciences, University of Leeds, Leeds LS2 9JT, United Kingdom. 4State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China. 5South China Sea Fisheries Research Institute, CAFS; Key Laboratory of South China Sea Fishery Resources Exploitation, Ministry of Agriculture, Guangzhou 510300, China. 6Department of Microbial Pathogenesis, Dental School, University of Maryland-Baltimore, Baltimore, MD 21201, USA.
Email: [email protected], [email protected]

  •  Accepted: 16 November 2011
  •  Published: 23 December 2011

Abstract

 

The specificity and sensitivity of an improved loop-mediated isothermal amplification (LAMP) method for rapid detection of Vibrio parahaemolyticus strains from various seafood samples had been developed and evaluated in this study. Six primers, including outer primers and inner primers, were specially designed for recognizing six distinct sequences on the target gene of tlh. The optimal reaction condition was found to be 65°C for 45 min, with the detection limit as 10 CFU/ml. Application of LAMP assays was performed on 416 food borne V. parahaemolyticus strains isolated from various seafood samples, and the total detection rate for LAMP and polymerase chain reaction (PCR) assay was found to be 96.2% (400/416) and 85.6% (356/416). This is the first report of an improved and simple LAMP detection assay on V. parahaemolyticus employed procedures of simple template deoxyribonucleic acid (DNA) preparation, equipment for LAMP reaction (water bath) and direct result determination via observation of color change. In addition, this is also the first application of LAMP detection on this considerable amount of V. parahaemolyticus isolates (416 strains for application together with 105 reference strains for establishment) with the total identification rate as 96.2%, as well as its extensive application to marine fish, shrimp, oyster, mussel, jellyfish, cuttlefish and seaweed samples, with detection rate ranging from 89.5 to 100%.

 

Key words: Loop-mediated isothermal amplification (LAMP), Vibrio parahaemolyticus, seafood samples.