Multiplex real-time polymerase chain reaction for rapid detection of Staphylococcus aureus, Vibrio parahaemolyticus, and Salmonella typhimurium in milk and kimbap

This study presented a multiplex, single-tube, realtime polymerase chain reaction (RTi-PCR) approach for detecting Staphylococcus aureus, Vibrio parahaemolyticus, and Salmonella typhimurium, three of the more frequent foodborne pathogenic bacteria typically investigated in a variety of foods. New pr...

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Bibliographic Details
Published in:Applied biological chemistry 2013, 56(6), , pp.715-721
Main Authors: Jeong, Yoo Seok, Jung, Hee Kyoung, Joo-Heon, Hong
Format: Article
Language:English
Subjects:
Amplification
Bacteria
Deoxyribonucleic acid
DNA
Food industry
Food processing industry
Foodborne pathogens
Fragments
Melting curve
Microbiological analysis
Milk
Multiplexing
Pathogens
Polymerase chain reaction
Real time
Replication origins
rRNA
rRNA 23S
Salmonella
Sensitivity analysis
Staphylococcus aureus
Target detection
Transcription
Vibrio parahaemolyticus
Waterborne diseases
농학
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Summary:This study presented a multiplex, single-tube, realtime polymerase chain reaction (RTi-PCR) approach for detecting Staphylococcus aureus, Vibrio parahaemolyticus, and Salmonella typhimurium, three of the more frequent foodborne pathogenic bacteria typically investigated in a variety of foods. New primer sequences were designed for detection of specific gene fragments in the 23s ribosomal RNA, transmembrane transcription regulator, and replication origin sequences of S. aureus, V. parahaemolyticus, and S. typhimurium. Simultaneous amplifications were performed under the optimized reaction conditions. Melting curve analysis using SYBR Green I RTi-PCR analysis produced characteristic Tm values for each target amplicon, demonstrating specific and efficient amplification of the three fragments. Addition of an internal amplification control did not affect detection sensitivity for the target pathogen. The analysis of frequent foodborne pathogenic bacteria in artificially inoculated food demonstrated analytical sensitivity for direct detection of each pathogen using the Chelex method rather than a commercial DNA extraction kit. The assay was sensitive to 103 colony-forming units (CFU)/reaction. With enrichment (2 or 4 h), each species could be detected at 101 CFU/g. These results provided that RTi-PCR is a rapid and costeffective procedure to detect foodborne pathogens. This assay could become a valuable tool for routine microbiological analysis in the food industry.
ISSN:2468-0834
2468-0842
DOI:10.1007/s13765-013-3194-6