Multiplex real-time polymerase chain reaction assay for simultaneous detection and quantification of Salmonella species, Listeria monocytogenes, and Escherichia coli O157:H7 in ground pork samples

Foodborne Pathogens and Disease
S KawasakiShinichi Kawamoto

Abstract

Salmonella sp., Listeria monocytogenes, and Escherichia coli O157:H7 are foodborne pathogens capable of causing serious gastrointestinal illness. We previously described simultaneous detection of these pathogens by multiplex polymerase chain reaction (PCR) in 44 types of spiked food samples, including meat, produce, fish, and dairy products, targeting genes specific for each pathogen. Based on the previous work, a multiplex real-time PCR assay using fluorescent probes was developed to detect and accurately quantify Salmonella sp., L. monocytogenes, and E. coli O157:H7 in ground pork samples. The detection sensitivity for this method was 2.0 x 10(2) CFU/mL for each pathogen, and the quantification range was 10(2)-10(7) CFU/mL with a high correlation coefficient (R(2) > 0.99) and high PCR efficiency (84.2% to 99.2%). When this protocol was used for the detection of each of the pathogens in spiked pork samples, one cell per 25 g of inoculated sample after enrichment for 20 h could be detected within 24 h. As a result, this multiplex real-time PCR assay will be valuable as a screening method for foods contaminated with these pathogens.

References

Jul 27, 1999·Journal of Food Protection·B KimuraS J Flood
Oct 8, 1999·Emerging Infectious Diseases·P S MeadR V Tauxe
May 5, 2007·Journal of Food Protection·Benjamin R WarrenKeith R Schneider
Aug 19, 2008·Applied and Environmental Microbiology·S Thisted LambertzM Lindblad

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Citations

Feb 4, 2011·Foodborne Pathogens and Disease·Beatriz QuiñonesErica D Dawson
Apr 5, 2012·Foodborne Pathogens and Disease·Giulia AmaglianiGiorgio Brandi
Jan 9, 2016·Critical Reviews in Food Science and Nutrition·S Umesha, H M Manukumar
Dec 6, 2014·Food Microbiology·Alejandro Garrido-MaestuAna G Cabado
Apr 1, 2014·Journal of Food Protection·Martin WiedmannKendra Nightingale
Nov 13, 2013·Journal of Food Protection·Abdela WoubitTemesgen Samuel
May 20, 2018·Biotechnology and Bioengineering·Quang Nghia PhamNae Yoon Lee
Jan 17, 2019·Critical Reviews in Analytical Chemistry·Shalini MuniandyBey Fen Leo
Mar 23, 2013·The Journal of General and Applied Microbiology·Ashwani KumarVirender Kumar Batish

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Methods Mentioned

BETA
PCR
electrophoresis
Assay

Software Mentioned

ABI PRISM 7900
Sequence Detection

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