Multiplex PCR for Simultaneous Detection of Salmonella spp., Listeria monocytogenes, and Escherichia coli O157:H7 in Meat Samples

2005 ◽  
Vol 68 (3) ◽  
pp. 551-556 ◽  
Author(s):  
SUSUMU KAWASAKI ◽  
NAOKO HORIKOSHI ◽  
YUKIO OKADA ◽  
KAZUKO TAKESHITA ◽  
TAKASHI SAMESHIMA ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Listeria Monocytogenes ◽  
Multiplex Pcr ◽  
Simultaneous Detection ◽  
Detection Sensitivity ◽  
Rapid Screening ◽  
Escherichia Coli O157 ◽  
Salmonella Spp ◽  
E Coli ◽  
Meat Samples

A multiplex PCR method was developed for simultaneous detection of Salmonella spp., Listeria monocytogenes, and Escherichia coli O157:H7 in meat samples. DNA detection sensitivity for this method was 103 CFU/ml for each pathogen. When this protocol was used for the detection of each of the above pathogenic bacteria in spiked pork samples, 1 cell per 25 g of inoculated sample could be detected within 30 h. In the samples of naturally contaminated meat, Salmonella spp., L. monocytogenes, and E. coli O157:H7 were detected over the same time period. Excellent agreement was obtained for the results of multiplex PCR and the conventional culture method, which suggests that the multiplex PCR is a reliable and useful method for rapid screening of meat products for Salmonella spp., L. monocytogenes, and E. coli O157:H7 contamination.

scholarly journals Evaluation of Multiplex PCR System for Simultaneous Detection of Escherichia coli O157:H7, Listeria monocytogenes and Salmonella enteritidis in Shrimp Samples

1970 ◽  
Vol 24 (1) ◽  
pp. 42-46 ◽  
Author(s):  
Mahmuda Yasmin ◽  
Susumu Kawasaki ◽  
Shinichi Kawamoto
Keyword(s):  
Escherichia Coli ◽  
Listeria Monocytogenes ◽  
Multiplex Pcr ◽  
Salmonella Enteritidis ◽  
Simultaneous Detection ◽  
Dna Amplification ◽  
Rapid Screening ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Pcr Method

A multiplex polymerase chain reaction (PCR) method was evaluated for simultaneous detection of Escherichia coli O157:H7, Listeria monocytogenes and Salmonella enteritidis in shrimp samples. The sensitivity of DNA amplification by PCR in this method was found to be 103 cfu/ml for each pathogen. When this protocol was adopted for the detection of each of the above mentioned pathogen in spiked shrimp extract culture, similar sensitivity was observed. However, this method detected 1 bacterial cell for E. coli O157:H7 and S. enteritidis and 100 for L. monocytogenes per 25 g spiked shrimp samples after overnight enrichment. In the commercially imported shrimp samples, none was found to contain any of the three pathogens by multiplex PCR or by conventional method, which suggests that the multiplex PCR is a reliable and useful for rapid screening of shrimp samples for E. coli O157:H7, L. monocytogenes and S. enteritidis. This will save time and increase our ability to assure food safety. Keywords: Multiples PCR, Shrimp extract, Spiked sample, Escherichia coli O157:H7, Listeria monocytogenes, Salmonella enteritidisDOI: http://dx.doi.org/10.3329/bjm.v24i1.1236 Bangladesh J Microbiol, Volume 24, Number 1, June 2007, pp 42-46

scholarly journals Assessment of a multiplex detection method for Salmonella enterica, Escherichia coli O157:H7, and Listeria monocytogenes in cow milk

2019 ◽  
Vol 24 (1) ◽  
pp. 277-294
Author(s):  
Rocio Esperanza Patiño-Burbano ◽  
Ana Karina Carrascal ◽  
Jorge Luis Parra-Arango ◽  
José Luis Rodríguez-Bautista
Keyword(s):  
Escherichia Coli ◽  
Listeria Monocytogenes ◽  
Detection Rate ◽  
Pathogenic Bacteria ◽  
Simultaneous Detection ◽  
Cow Milk ◽  
Escherichia Coli O157 ◽  
Salmonella Spp ◽  
E Coli ◽  
Microbiological Methods

Raw cow milk is considered one of the most important vehicles for pathogenic bacteria like Salmonella spp., Escherichia coli O157:H7, and Listeria monocytogenes. These three bacteria are responsible for foodborne diseases. Routine microbiological methods to detect these microorganisms in cow milk can be complicated and time consuming. The aim of this work was to evaluate a method to simultaneously detect Salmonella spp., Escherichia coli O157:H7, and Listeria monocytogenes in experimentally contaminated cow milk. The assessed method combined a standard microbiological culture step, using a pre-enrichment medium that favors the growth of the three focal microorganisms: SEL broth, followed by a single PCR assay. A total of 43 interference bacterial strains were used to evaluate the method’s specificity. The detection rate for the microbiological method with standard culture media was 10 UFC/mL, and that of the PCR detection, following pre-enrichment in SEL broth, was 10 UFC/mL for S. enterica and L. monocytogenes and between 1 and 5 UFC/mL for E. coli O157:H7. The PCR method showed specificity for the reference strains. Simultaneous detection by multiple PCR using SEL broth was successful for the detection of S. enterica, E. coli O157:H7, and L. monocytogenes in samples of experimentally contaminated cow milk, featuring both a high detection rate and a high specificity. This approach promises to be a feasible routine procedure when testing milk samples in industry and public health control setups.

A gold nanoparticles-assisted multiplex PCR assay for simultaneous detection of Salmonella typhimurium, Listeria monocytogenes and Escherichia coli O157:H7

2020 ◽  
Vol 12 (2) ◽  
pp. 212-217 ◽  
Author(s):  
Juan Du ◽  
Shujing Wu ◽  
Liyuan Niu ◽  
Junguang Li ◽  
Dianbo Zhao ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Gold Nanoparticles ◽  
Listeria Monocytogenes ◽  
Salmonella Typhimurium ◽  
Multiplex Pcr ◽  
Simultaneous Detection ◽  
Escherichia Coli O157 ◽  
Pcr Assay ◽  
Multiplex Pcr Assay ◽  
Pcr Product

Unfunctionalized flower-shaped AuNPs is used as colorimetric sensor for PCR product detection by naked eyes.

Primers with 5′ Flaps Improve the Efficiency and Sensitivity of Multiplex PCR Assays for the Detection of Salmonella and Escherichia coli O157:H7

2013 ◽  
Vol 76 (4) ◽  
pp. 668-673 ◽  
Author(s):  
CHRIS TIMMONS ◽  
SHEFALI DOBHAL ◽  
JACQUELINE FLETCHER ◽  
LI MARIA MA
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Fold Increase ◽  
Detection Sensitivity ◽  
Escherichia Coli O157 ◽  
Bacterial Cells ◽  
Robust Detection ◽  
E Coli ◽  
Pcr Multiplex ◽  
Pcr Assays

Foodborne illnesses caused by Salmonella enterica and Escherichia coli O157:H7 are worldwide health concerns. Rapid, sensitive, and robust detection of these pathogens in foods and in clinical and environmental samples is essential for routine food quality testing, effective surveillance, and outbreak investigations. The aim of this study was to evaluate the effect on PCR sensitivity of adding a short, AT-rich overhanging nucleotide sequence (flap) to the 5′ end of PCR primers specific for the detection of Salmonella and E. coli O157:H7. Primers targeting the invA gene of Salmonella and the rfbE gene of E. coli O157:H7 were synthesized with or without a 12-bp, AT-rich 5′ flap (5′-AATAAATCATAA-3′). Singleplex PCR, multiplex PCR, and real-time PCR sensitivity assays were conducted using purified bacterial genomic DNA and crude cell lysates of bacterial cells. The effect of background flora on detection was evaluated by spiking tomato and jalapeno pepper surface washes with E. coli O157:H7 and Salmonella Saintpaul. When targeting individual pathogens, end-point PCR assays using flap-amended primers were more efficient than nonamended primers, with 20.4 and 23.5% increases in amplicon yield for Salmonella and E. coli O157:H7, respectively. In multiplex PCR assays, a 10- to 100-fold increase in detection sensitivity was observed when the primer flap sequence was incorporated. This improvement in both singleplex and multiplex PCR efficiency and sensitivity can lead to improved Salmonella and E. coli O157:H7 detection.

Validation of Apple Cider Pasteurization Treatments against Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes

2001 ◽  
Vol 64 (11) ◽  
pp. 1679-1689 ◽  
Author(s):  
PEGGY P. MAK ◽  
BARBARA H. INGHAM ◽  
STEVEN C. INGHAM
Keyword(s):  
Escherichia Coli ◽  
New York ◽  
Listeria Monocytogenes ◽  
Fluorescent Protein ◽  
Escherichia Coli O157 ◽  
Salmonella Spp ◽  
Apple Cider ◽  
E Coli ◽  
Log Reduction ◽  
Green Fluorescent

Time and temperature pasteurization conditions common in the Wisconsin cider industry were validated using a six-strain cocktail of Escherichia coli O157:H7 and acid-adapted E. coli O157:H7 in pH- and ∘Brix-adjusted apple cider. Strains employed were linked to outbreaks (ATCC 43894 and 43895, C7927, and USDA-FSIS-380–94) or strains engineered to contain the gene for green fluorescent protein (pGFP ATCC 43894 and pGFP ATCC 43889) for differential enumeration. Survival of Salmonella spp. (CDC 0778, CDC F2833, and CDC HO662) and Listeria monocytogenes (H0222, F8027, and F8369) was also evaluated. Inoculated cider of pH 3.3 or 4.1 and 11 or 14°Brix was heated under conditions ranging from 60°C for 14 s to 71.1°C for 14 s. A 5-log reduction of nonadapted and acid-adapted E. coli O157:H7 was obtained at 68.1°C for 14 s. Lower temperatures, or less time at 68.1°C, did not ensure a 5-log reduction in E. coli O157:H7. A 5-log reduction was obtained at 65.6°C for 14 s for Salmonella spp. L. monocytogenes survived 68.1°C for 14 s, but survivors died in cider within 24 h at 4°C. Laboratory results were validated with a surrogate E. coli using a bench-top plate heat-exchange pasteurizer. Results were further validated using fresh unpasteurized commercial ciders. Consumer acceptance of cider pasteurized at 68.1°C for 14 s (Wisconsin recommendations) and at 71.1°C for 6 s (New York recommendations) was not significantly different. Hence, we conclude that 68.1°C for 14 s is a validated treatment for ensuring adequate destruction of E. coli O157:H7, Salmonella spp., and L. monocytogenes in apple cider.

Simultaneous detection of Escherichia coli O175:H7, Salmonella spp., and Listeria monocytogenes by multiplex PCR

Food Control ◽  
2009 ◽  
Vol 20 (8) ◽  
pp. 733-738 ◽  
Author(s):  
Andrea Germini ◽  
Annalisa Masola ◽  
Paola Carnevali ◽  
Rosangela Marchelli
Keyword(s):  
Escherichia Coli ◽  
Listeria Monocytogenes ◽  
Multiplex Pcr ◽  
Simultaneous Detection ◽  
Salmonella Spp
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