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Title: DETECTION OF LISTERIA MONOCYTOGENES IN FRESH PRODUCE USING MOLECULAR BEACON - REAL-TIME PCR TECHNOLOGY

Author
item LIMING, SAMANTHA - NGEE ANN POLYT, SINGAPORE
item ZHANG, YIFAN - UMD, COLLEGE PK, MD
item MENG, JIANGHONG - UMD, COLLEGE PK, MD
item Bhagwat, Arvind

Submitted to: Journal of Food Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/9/2005
Publication Date: 8/1/2005
Citation: Liming, S.H., Zhang, Y., Meng, J., Bhagwat, A.A. 2005. Detection of listeria monocytogenes in fresh produce using molecular beacon - real-time pcr technology. Journal of Food Science. 69:240-245.

Interpretive Summary: Conventional methods to detect the foodborne pathogen Listeria monocytogenes take up to one and a half weeks. Considering the limited shelf-life of produce, rapid methods for pathogen detection are required. Real-time detection of Listeria monocytogenes strains will broaden our ability to a screen large number of samples in a short time. In this study, a DNA-based detection method for Listeria monocytogenes, based on polymerase chain reaction (PCR) was developed to enable rapid detection. The modified protocol requires approximately 26 hours and is able to handle high sample volumes. Detection of human pathogens from fresh produce is a crucial step in implementing food safety. Both the fresh produce industry and consumers will benefit from the results of this research.

Technical Abstract: The capability of an assay to detect Listeria monocytogenes from artificially-inoculated fresh-cut produce such as cantaloupe and mixed-salad was demonstrated. An oligonucleotide probe that becomes fluorescent upon hybridization to the target DNA (Molecular Beacon; MB) was used in a real-time polymerase chain reaction (PCR) assay. As few as 4 to 7 colony-forming units (CFU) of L. monocytogenes per 25 g of artificially contaminated produce could be detected. A comparison of two commercially available kits utilizing MB-PCR (iQ-Check, Bio-Rad Laboratories) and conventional PCR (BAX, Dupont Qualicon) was performed on artificially inoculated produce. The time required to detect L. monocytogenes (from produce to PCR data) was considerably less for the iQ-Check protocol (approximately 26 h) compared to the BAX- PCR (approximately 52 h). The iQ-Check protocol was also used to confirm the identity of the L. monocytogenes isolates obtained during a microbiological screen of conventional and organic leaf lettuce and alfalfa sprout samples from local supermarkets. The iQ check protocol was successful in differentiating L. monocytogenes isolates from rest of the Listeria spp. such as, L. welshimeri, L. innocua, and L. ivanovii. This is the first report of the application of the MB probe being used for real-time detection of L. monocytogenes in whole and fresh-cut fruits and vegetables.