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Rapid and sensitive enumeration of total coliforms and Escherichia coli in water and foods by most-probable-number loop-mediated isothermal amplification (MPN-LAMP) method

Chonnikan Tongphrom^a, Sutima Preeprem^a, Mitsuaki Nishibushi^b, Varaporn Vuddhakul^a, Pimonsri Mittraparp-arthorn^a,*

 
ABSTRACT:     Total coliforms and Escherichia coli are bacterial indicators used to assess the microbiological qualities of water and foods. In this study, we developed a most-probable-number loop-mediated isothermal amplification (most probable number (MPN-LAMP)) method for the enumeration of total coliforms and E. coli in drinking water and vegetables. The LAMP primers were designed based on lacZ (lacZ-LAMP) and uidA (uidA-LAMP) genes for the detection of total coliforms and E. coli, respectively. In pure culture, the lacZ-LAMP and the uidA-LAMP were able to detect all of the 37 coliforms and 30 E. coli strains, respectively. In the artificially contaminated drinking water and vegetables, the combination of MPN and LAMP techniques (MPN-LAMP) was able to detect E. coli at 1 colony-forming unit (CFU)/100 ml and 5 CFU/g, respectively. From analysis of 33 drinking water and 46 vegetable samples, the total coliform detection obtained by the MPN-LAMP method was in agreement with the MPN detection technique. However, MPN-LAMP was more sensitive than the MPN technique for E. coli detection. Our findings revealed that the MPN-LAMP assay was more rapid and highly sensitive than MPN method. Thus this method could be considered for detection and enumeration of total coliforms and E. coli in the food industry

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* Corresponding author, E-mail: pimonsri.m@psu.ac.th

Received 6 Jul 2018, Accepted 1 Oct 2018