Original Article

A rapid method for separating and concentration of food-borne pathogens using elution from ready-to-eat vegetables

Abstract

Background and Objectives: Traditional culture methods for detection of food-borne pathogens, a major public health problem, are simple, easily adaptable and very practical, but they can be laborious and time consuming. In this study, we eliminated culturing steps by developing a new separation method and therefore, decreased the detection time of food-borne pathogens (Salmonella enterica serovar Typhimurium, Escherichia coli O157:H7 and Listeria monocytogenes) to a few hours.
Materials and Methods: We used alkaline water and different alkaline buffers to elute bacteria from the lettuce surface as a model for ready-to-eat vegetables. Buffers used were as follows: 1) 0.05 M glycine; 2) 0.05 M glycine -100 mM Tris base -1% (w/v) beef extract; 3) buffer peptone water; 4) buffer phosphate saline. Buffers were adjusted to pH of 9, 9.5 and 10. In order to elute the bacteria, the lettuce pieces were suspended into buffers and shacked for 30, 45 and 60 min. Moreover, a multiplex PCR method for the simultaneous detection of food-borne pathogens was performed.
Results: The results showed that buffer peptone water at pH 9.5 for 45 min have high ability to elute bacteria from the lettuce surface and the bacteria can be detected using multiplex PCR.
Conclusion: We developed a new rapid and efficient method for simultaneous separation of food-borne pathogens. This method eliminates culturing stages and permits the detection and identification of target pathogens in a few hours.

References

Elizaquıvel P, Aznar R. A multiplex RTi-PCR reaction for simultaneous detection of Escherichia coli O157:H7, Salmonella spp. and Staphylococcus aureus on fresh, minimally processed vegetables. Food Microbiol 2008; 25: 705-713.

Yu Q, Zhai L, Bie X, Lu Z, Zhang C, Tao T, Li J, Lv F, Zhao H. Survey of five food-borne pathogens in commercial cold food dishes and their detection by multiplex PCR, Food Control 2016, 59: 862-869.

Ryu J, Park SH, Yeom YS, Shrivastav A, Lee SH, Kim YR, Kim HY. Simultaneous detection of Listeria species isolated from meat processed foods using multiplex PCR. Food Control 2013; 32(2): 659-664.

Jasson V, Jacxsens L, Luning P, Rajkovic A, Uyttendaele M. Alternative microbial methods: An overview and selection criteria. Food Microbiol 2010; 27: 710-730.

Thapa SP, Han R, Cho JM, Hur JH. Multiplex PCR and DNA array for the detection of Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes, Escherichia coli O157:H7 and Salmonella spp. Targeting virulence-related genes. Annu Microbiol 2013; 63: 725-731.

Zhang QY, Zhou WW, Zhou Y, Wang XF, Xu JF. Response surface methodology to design a selective co-enrichment broth of Escherichia coli, Salmonella spp. and Staphylococcus aureus for simultaneous detection by multiplex PCR. Microbiol Res 2012; 167: 405-412.

Ohk SH, Bhunia AK. Multiplex fiber optic biosensor for detection of Listeria monocytogenes, Escherichia coli O157:H7 and Salmonella entrica from ready-to-eat meat samples. Food Microbiol 2013; 33: 166-171.

Yang Y, Xu F, Xu H, Aguilar ZP, Niu R, Yuan Y, Sun J, You X, Lai W, Xiong Y, Wan C, Wei H. Magnetic nano-beads based separation combined with propidium monoazide treatment and multiplex PCR assay for simultaneous detection of viable Salmonella Typhimurium, Escherichia coli O157:H7 and Listeria monocytogenes in food products. Food Microbiol 2013; 34: 418-424.

Donhauser SC, Niessner R, Seidel M. Sensitive quantification of Escherichia coli O157:H7, Salmonella enterica, and Campylobacter jejuni by combining stopped polymerase chain reaction with chemiluminescence flow-through DNA microarray analysis. Anal Chem 2011; 83: 3153-3160.

Vasanth D, Pugazhenthi G, Uppaluri R. Fabrication and properties of low cost ceramic microfiltration membranes for separation of oil and bacteria from its solution. J Membrane Sci 2011; 379: 154-163.

Coklin T, Farber JM, Parrington LJ, Bin Kingombe CI, Ross WH, Dixon BR. Immunomagnetic separation significantly improves the sensitivity of polymerase chain reaction in detecting Giardia duodenalis and Cryptosporidium spp. in dairy cattle. J Vet Diagn Invest 2011; 23: 260-267.

Hagren V, Von Lode P, Syrjala A, Korpimaki T, Tuomola M, Kauko O, Nurmi J. An 8-hour system for Salmonella detection with immunomagnetic separation and homogeneous time-resolved fluorescence PCR. Int J Food Microbiol 2008; 125: 158-161.

Brandao D, Liebana S, Pividori MI. Multiplexed detection of food-borne pathogens based on magnetic particles. New Biotechnology 2015; 32(5): 511-520.

Lim MC, Park JY, Park K, Ok G, Jang HJ, Choi SW. An automated system for separation and concentration of food-borne pathogens using immunomagnetic separation. Food control 2017; 73(B): 1541-1547.

Croci L, Dubois E, Cook N, Medici D de, Schultz AC, China B, Rutjes SA, Hoorfar J, Van der Poel WHM. Current methods for extraction and concentration of enteric viruses from fresh fruit and vegetables: Towards international standards. Food Anal Method 2008; 1: 73-84.

Butot S, Putallaz T, Sanchez G. Procedure for rapid concentration and detection of enteric viruses from berries and vegetables. Appl Environ Microb 2007; 73(1): 186-192.

Kim H, Kwak I, Hwang I, Ko G. Optimization of methods for detecting norovirus on various fruit. J Virol Methods 2008; 153: 104-110.

Dubois E, Hennechart C, Deboosere N, Merle G, Legeay O, Burger C, et al. Intra-laboratory validation of a concentration method adapted for the enumeration of infectious F-specific RNA coliphage, entrovirus and hepatitis A virus from inoculated leaves of salad vegetables. Int J Food Microbiol 2006; 108: 164-171.

Bahreini M, Habibi Najafi MB, Bassami MR, Yavarmanesh M. Optimization of extraction and concentration methods of enteric viruses from the surface of ready to eat vegetables using MS2 coliphage as a model. J Food Sci Tech 2014; 11(43): 1-10.

Sapers GM. Efficacy of Washing and Sanitizing Methods for Disinfection of Fresh Fruit and Vegetable Products. Food Technol Biotech 2001; 39: 305-311.

Love DC, Casteel MJ, Meschke JS, Sobsey MD. Methods for recovery of hepatitis A virus (HAV) and other viruses from processed foods and detection of HAV by nested RT-PCR and TaqMan RT-PCR. Int J Food Microbiol 2008; 126: 221-226.

ISO, 11290-2: Microbiology of food and animal feeding stuffs – Horizontal method for the detection and enumeration of Listeria monocytogenes– Part 2: Enumeration method. 1998.

ISO. 16654: Microbiology of food and animal feeding stuffs – Horizontal method for the detection and enumeration of Escherichia coli O157:H7. 2002.

ISO. 6579: Microbiology of food and animal feeding stuffs – Horizontal method for the detection of Salmonella spp. 2002.

Coudray-Meunier C, Fraisse A, Martin-Latil S, Guillier L, Delannoy S, Fach P, Perelle S. A comparative study of digital RT-PCR and RT-qPCR for quantification of Hepatitis A virus and Norovirus in lettuce and water samples. Int J Food Microbiol 2015; 201: 17-26.

Casteel MG, Schmidt CE, Sobsey MD. Chlorine disinfection of produce to inactivate hepatitis A virus and coliphage MS2. Int J Food Microbiol 2008; 125: 267-273.

Maurer JJ, Schmidt D, Petrosko P, Sanchez S, Bolton L, Lee MD. Development of primers to O-antigen biosynthesis genes for specific detection of Escherichia coli O157:H7 by PCR. Appl Environ Microbiol 1999; 65: 2954-2960.

Rahn K, De Grandis SA, Clarke RC, McEwen SA, Galán JE, Ginocchio C, Curtiss R, Gyles CL. Amplification of an invA gene sequence of Salmonella typhimurium by polymerase chain reaction as a specific method of detection of Salmonella. Mol Cell Probes 1992; 6: 271-279.

Germini A, Masola A, Carnevali P, Marhelli R. Simultaneous detection of Escherichia coli O157:H7, Salmonella spp. and Listeria monocytogenes by multiplex PCR. Food Control 2009; 20(8): 733-738.

Chiang YC, Yang CY, Li C, Ho YC, Lin CK, Tsen HY. Identification of Bacillus spp., Escherichia coli, Salmonella spp., Staphylococcus spp. and Vibrio spp. with 16S ribosomal DNA-based oligonucleotide array hybridization. Int J Food Microbiol 2006; 107(2): 131-137.

Wang L, Li Y, Mustaphai A. Rapid and simultaneous quantitation of Escherichia coli 0157:H7, Salmonella, and Shigella in ground beef by multiplex real-time PCR and immunomagnetic separation. J Food Protect 2007; 70: 1366-1372.

Schrader KN, Fernandez-Castro A, Cheung WKW, Crandall CM, Abbott SL. Evaluation of commercial antisera for Salmonella serotyping. J Clin Microbiol 2008; 46: 685-688.

Kim TH, Park J, Kim CJ, ChoYK. Fully integrated lab-on-a-disk for nucleic acid analysis of food-borne pathogenes. Anal Chem 2014; 86(8): 3841-3848.

Lee N, Kwon KY, Oh SK, Chang HJ, Chun SH, Choi SW. A multiplex PCR assay for simultaneous detection of Escherichia coli O157:H7, Bacillus cereus, Vibrio parahaemolyticus, Salmonella spp., Listeria monocytogenes, and Staphylococcus aureus in Korean ready-to-eat food. Food-borne Pathog Dis 2014; 11(7): 1-7.

Guan ZP, Jiang Y, Gao F, Zhang L, Zhou GH, Guan ZJ. Rapid and simultaneous analysis of five food-borne pathogenic bacteria using multiplex PCR. Eur. Food Res Technol 2013; 237: 627-637.

Garrido A, Chapela MJ, Román B, Fajardo P, Vieites JM, Cabado G. In-house validation of a multiplex real-time PCR method for simultaneous detection of Salmonella spp., Escherichia coli O157:H7 and Listeria monocytogenes. Int J Food Microbiol 2013; 164: 92-98

Files
IssueVol 10 No 6 (2018) QRcode
SectionOriginal Article(s)
Keywords
Rapid detection Elution Multiplex polymerase chain reaction Food-borne pathogens Ready-to-eat vegetables

Rights and permissions
Creative Commons License This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.
How to Cite
1.
Rajabzadeh S, Bahreini M, Sharifmoghadam MR. A rapid method for separating and concentration of food-borne pathogens using elution from ready-to-eat vegetables. Iran J Microbiol. 2019;10(6):385-393.