Combination of Immunomagnetic Separation and Polymerase Chain Reaction for the Simultaneous Detection of Listeria monocytogenes and Salmonella spp. in Food Samples

Hsih, Hsien-Yee; Tsen, Hau‐Yang

doi:10.4315/0362-028x-64.11.1744

Cited by 71 publications

(37 citation statements)

References 30 publications

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“…Immunomagnetic separation (IMS) has been previously applied to isolation of microorganisms (Grant et al, 1998;Hsih and Tsen, 2001;Pinto and Tantillo, 2002). Madonna et al (2001) also reported isolation of Salmonella choleraesuis from a variety of matrices including river water, urine and chicken blood.…”

Section: Immunomagnetic Separationmentioning

confidence: 97%

Analysis of Bacillus anthracis Spores in Milk Using Mass Spectrometry

Whiteaker

Karns²,

Fenselau

et al. 2004

Foodborne Pathogens and Disease

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New approaches for identifying biological threat agents in raw milk using spectroscopy were tested using Bacillus anthracis (BA) Sterne strain spores seeded into unpasteurized bulk tank milk. Direct filtration onto Tyvek membranes provided the optimal filtration approach from raw milk, but detection limits were not ideal. When beads coated with anti-BA antibodies were mixed with spores in raw milk, the beads were capable of concentrating the spores that could be later detected and characterized by MALDI spectroscopy based on presence of previously characterized small acid-soluble proteins (SASP's). This approach could provide a very rapid assessment of whether milk or milk products have been purposefully contaminated with BA spores. This work was fundamentally a proof-of-concept study demonstrating feasibility of the approach in milk. Other parameters could be changed to potentially lower detection limits, and additional studies are currently underway to improve the approach.

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Section: Immunomagnetic Separationmentioning

confidence: 97%

Analysis of Bacillus anthracis Spores in Milk Using Mass Spectrometry

Whiteaker

Karns²,

Fenselau

et al. 2004

Foodborne Pathogens and Disease

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“…Magnetic separation (MS) has become a routine method in food and veterinary microbiology laboratories and is commonly used in combination with culture or molecular methods for the detection and isolation of pathogenic bacteria such as Listeria monocytogenes (13,31), Salmonella spp. (22,25), and Escherichia coli O157:H7 in both the food (15) and veterinary (20) clinical sample testing context.…”

mentioning

confidence: 99%

Maximizing Capture Efficiency and Specificity of Magnetic Separation for Mycobacterium avium subsp. paratuberculosis Cells

Foddai

Elliott

Grant

2010

Appl Environ Microbiol

108

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In order to introduce specificity for Mycobacterium avium subsp. paratuberculosis prior to a phage amplification assay, various magnetic-separation approaches, involving either antibodies or peptides, were evaluated in terms of the efficiency of capture (expressed as a percentage) of M. avium subsp. paratuberculosis cells and the percentage of nonspecific binding by other Mycobacterium spp. A 50:50 mixture of MyOne Tosylactivated Dynabeads coated with the chemically synthesized M. avium subsp. paratuberculosis-specific peptides biotinylated aMp3 and biotinylated aMptD (i.e., peptide-mediated magnetic separation [PMS]) proved to be the best magnetic-separation approach for achieving 85 to 100% capture of M. avium subsp. paratuberculosis and minimal (<1%) nonspecific recovery of other Mycobacterium spp. (particularly if beads were blocked with 1% skim milk before use) from broth samples containing 10 3 to 10 4 CFU/ml. When PMS was coupled with a recently optimized phage amplification assay and used to detect M. avium subsp. paratuberculosis in 50-ml volumes of spiked milk, the mean 50% limit of detection (LOD 50 ) was 14.4 PFU/50 ml of milk (equivalent to 0.3 PFU/ml). This PMS-phage assay represents a novel, rapid method for the detection and enumeration of viable M. avium subsp. paratuberculosis organisms in milk, and potentially other sample matrices, with results available within 48 h.The prospect of being able to detect viable Mycobacterium avium subsp. paratuberculosis organisms in food or veterinary samples within 48 h using a commercially available phage amplification assay (FASTPlaqueTB assay; Biotec Laboratories Limited, Ipswich, United Kingdom), rather than waiting weeks for conventional culture results, is an exciting recent development (7,8,26). However, the mycobacteriophage used in the phage amplification assay has a broader mycobacterial host range than M. avium subsp. paratuberculosis alone (23). Consequently, plaques obtained when naturally infected, rather than artificially spiked, samples are tested may not necessarily emanate from M. avium subsp. paratuberculosis alone if other Mycobacterium spp. are also present in the sample. Some additional selective step prior to phage infection, such as magnetic separation (12), is needed to introduce selectivity for M. avium subsp. paratuberculosis.Magnetic separation (MS) has become a routine method in food and veterinary microbiology laboratories and is commonly used in combination with culture or molecular methods for the detection and isolation of pathogenic bacteria such as Listeria monocytogenes (13, 31), Salmonella spp. (22, 25), and Escherichia coli O157:H7 in both the food (15) and veterinary (20) clinical sample testing context. Magnetic-separation methods selectively separate the target bacterium from other, nontarget microorganisms and inhibitory sample components while concentrating the target bacterial cells into a smaller volume. Collectively, these properties of magnetic separation enhance the analytical specificity and sensitivity of the ...

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“…The results described here indicate that the MAAB not only has similar or better detection limits than other sensors, but also requires only 15 min for analysis of prepared samples. Sample processing is simple, requiring only homogenization of the foodstuff or excreta, followed by coarse filtration or centrifugation; this ease of sample preparation contrasts greatly with the large numbers of manipulations typically required for nucleic acid-based techniques (6,10,31), even with immunomagnetic separation.…”

Section: Discussionmentioning

confidence: 99%

“…Thus, a pre-enrichment step is typically required to increase the number of viable cells while effectively diluting inhibitory substances present in the sample. A large number of papers describing PCR-based detection of Salmonella from spiked or naturally contaminated foods have claimed LODs ranging from 0.1 to 100 CFU/g (6,10,31,37,40). However, many of these LODs represent the concentrations of inoculum used rather than actual bacterial counts after enrichment; postenrichment concentrations can reach 10 6 to 10 9 CFU/ml (6,7,17,23).…”

Section: Discussionmentioning

confidence: 99%

Detection ofSalmonella entericaSerovar Typhimurium by Using a Rapid, Array-Based Immunosensor

Taitt

Shubin

Angel

et al. 2004

Appl Environ Microbiol

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The multianalyte array biosensor (MAAB) is a rapid analysis instrument capable of detecting multiple analytes simultaneously. Rapid (15-min), single-analyte sandwich immunoassays were developed for the detection of Salmonella enterica serovar Typhimurium, with a detection limit of 8 ؋ 10 4 CFU/ml; the limit of detection was improved 10-fold by lengthening the assay protocol to 1 h. S. enterica serovar Typhimurium was also detected in the following spiked foodstuffs, with minimal sample preparation: sausage, cantaloupe, whole liquid egg, alfalfa sprouts, and chicken carcass rinse. Cross-reactivity tests were performed with Escherichia coli and Campylobacter jejuni. To determine whether the MAAB has potential as a screening tool for the diagnosis of asymptomatic Salmonella infection of poultry, chicken excretal samples from a private, noncommercial farm and from university poultry facilities were tested. While the private farm excreta gave rise to signals significantly above the buffer blanks, none of the university samples tested positive for S. enterica serovar Typhimurium without spiking; dose-response curves of spiked excretal samples from university-raised poultry gave limits of detection of 8 ؋ 10 3 CFU/g.

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Combination of Immunomagnetic Separation and Polymerase Chain Reaction for the Simultaneous Detection of Listeria monocytogenes and Salmonella spp. in Food Samples

Cited by 71 publications

References 30 publications

Analysis of Bacillus anthracis Spores in Milk Using Mass Spectrometry

Analysis of Bacillus anthracis Spores in Milk Using Mass Spectrometry

Maximizing Capture Efficiency and Specificity of Magnetic Separation for Mycobacterium avium subsp. paratuberculosis Cells

Detection ofSalmonella entericaSerovar Typhimurium by Using a Rapid, Array-Based Immunosensor

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