Real time loop-mediated isothermal amplification using a portable fluorescence scanner for rapid and simple detection of Vibrio parahaemolyticus

We evaluated the usefulness of the loop-mediated isothermal amplification (LAMP) using a portable ESE Quant tube scanner as a rapid and simple method for the detection of Vibrio parahaemolyticus, an important pathogen causing seafood-borne gastroenteritis. The real time LAMP (RT-LAMP) assay using a hemolysin gene (tlh/ldh)-specific primers was verified using V. parahaemolyticus strains (n = 91) from different countries and other non-target strains to check the utility of this method. Both the sensitivity and specificity of the RT-LAMP using 3 pairs of tlh/ldh-specific primers developed in this study were excellent (100%). The detection limit of the RT-LAMP was as low as 7 Colony forming unit per reaction and detection time was only 20 min. Comparative evaluation of the target bacterial strains with the RT-LAMP using ESE Quant tube scanner, API 20E system and conventional RT-PCR method revealed that the RI-LAMP assay developed in this study is simpler and more rapid than the latter two methods. Therefore, the RI-LAMP method using the easily portable ESE Quant tube scanner can be considered as an effective tool for the rapid screening of V parahaemolyticus strains in environmental and clinical samples, especially, in remote areas of developing countries during epidemic periods. (C) 2014 Elsevier Ltd. All rights reserved.