J. Agric. Food Chem.,
Web Release Date: October 23,
Copyright © 2004 American Chemical Society
Simultaneous and Sensitive Detection of Three Foodborne Pathogens by Multiplex PCR, Capillary Gel Electrophoresis, and Laser-Induced Fluorescence
and
Departamento de Microbiología y Ecología, Universitat València, Burjassot E-46100, Valencia, Spain, Instituto de Agroquímica y Tecnología de Alimentos (IATA), Consejo Superior de Investigaciones Científicas (CSIC), P.O. Box 73, Burjassot E-46100, Valencia, Spain, and Institute of Industrial Fermentations (CSIC), Juan de la Cierva 3, 28006 Madrid, Spain
Received for review June 15, 2004. Revised manuscript received September 10, 2004. Accepted September 13, 2004. This work has been supported by a CICYT grant (1FD97-0549) and a CAM project (7B- 0021-2002). V.G.-C. thanks Consejería de Educación y Cultura (Comunidad de Madrid) for a fellowship.
Abstract:
The simultaneous detection of Staphylococcus aureus, Listeria monocytogenes, and Salmonella spp. has been approached by a new multiplex PCR-based procedure followed by capillary gel electrophoresis with laser-induced fluorescence detection (multiplex-PCR-CGE-LIF). As compared to slab gel electrophoresis, the use of CGE-LIF improved from 10- to 1000-fold the sensitivity of the multiplex PCR analysis, allowing the detection of 2.6 × 103 cfu mL-1 of S. aureus, 570 cfu mL-1 of L. monocytogenes, and 790 cfu mL-1 of Salmonella in artificially inoculated food, without enrichment. Following 6 h of enrichment, as low as 260, 79, and 57 cfu mL-1 of S. aureus, L. monocytogenes, and Salmonella, respectively, were detected. The CGE-LIF method is shown to be reproducible, providing relative standard deviation (RSD) values lower than 0.8% for analysis time and lower than 5.8% for peak areas. The multiplex-PCR-CGE-LIF proved a powerful analytical tool to detect various food pathogens simultaneously in a fast, reproducible, and sensitive way.
Keywords: Multiplex PCR; CGE; LIF; food analysis; foodborne pathogens; DNA analysis; Salmonella spp.; Staphylococcus aureus; Listeria monocytogenes