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Immunology and Infection

Oral Transmission of Listeria monocytogenes in Mice via Ingestion of Contaminated Food

Published: May 6th, 2013

DOI:

10.3791/50381

1Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky

This paper describes a novel method for oral infection of mice using Listeria monocytogenes-contaminated food. The protocol can readily be adapted for use with other food borne bacterial pathogens.

L. monocytogenes are facultative intracellular bacterial pathogens that cause food borne infections in humans. Very little is known about the gastrointestinal phase of listeriosis due to the lack of a small animal model that closely mimics human disease. This paper describes a novel mouse model for oral transmission of L. monocytogenes. Using this model, mice fed L. monocytogenes-contaminated bread have a discrete phase of gastrointestinal infection, followed by varying degrees of systemic spread in susceptible (BALB/c/By/J) or resistant (C57BL/6) mouse strains. During the later stages of the infection, dissemination to the gall bladder and brain is observed. The food borne model of listeriosis is highly reproducible, does not require specialized skills, and can be used with a wide variety of bacterial isolates and laboratory mouse strains. As such, it is the ideal model to study both virulence strategies used by L. monocytogenes to promote intestinal colonization, as well as the host response to invasive food borne bacterial infection.

Listeria monocytogenes are facultative intracellular bacterial pathogens that cause food borne infections in humans. The bacteria are resistant to many of the processes used to protect our food supply, such as drying, salting, or refrigeration 1,2 and infections are typically linked to processed, "ready-to-eat" foods that are not heated prior to consumption. In several previous outbreaks, the source of L. monocytogenes-contaminated food was identified, and the restricted group of exposed individuals was monitored closely 3-6. In those examples, clinical disease in otherwise healthy individuals varied from a mild, self-limiting g....

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1. Preparation of Selective Agar Media (BHI/L+G) to Inhibit Intestinal Microbiota

  1. Weigh out 26 g Brain Heart Infusion Agar (Difco), 7.5 g LiCl and 5.0 g glycine and place in a 1 liter flask. Add 500 ml of deionized water.
  2. Heat on a magnetic stirrer until the agar boils, stirring continuously. Take the flask off the heat, let the bubbles settle briefly, and then bring back to a boil again.
  3. Autoclave for 30 min at 121 °C under 16-19 psi on a liquid cycle. Note: leave the.......

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L. monocytogenes colonies will be visible on BHI/L+G plates after 36-48 hr incubation at 37 °C. The colonies have a smooth, dome-shaped creamy white appearance (Figure 1A). Growth will be inhibited for the majority of the intestinal microbiota, but it is common to see some colonies that are not L. monocytogenes, particularly when plating small intestine or colon directly without significant dilution (Figure 1B). Suspect colonies can be confirmed by plating on ChromA.......

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Inbred mice are not uniformly receptive to feeding at all times of the day, and their willingness to eat the contaminated bread will depend both on the strain type and the age of the mice 17. In our experience, 6-9 week old B6 mice are receptive to feeding at any time of day, but BALB mice will not consistently eat the bread piece unless it is offered during their dark cycle. The light cycle of the room used to house the animals can be altered so the dark phase coincides with the normal working day for laborat.......

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This work was supported by grants from the National Institutes of Health (AI079442 and AI091918) awarded to S.E.F.D.

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Name Company Catalog Number Comments
Name of Reagent/Material Company Catalog Number Comments
Brain Heart Infusion Agar Difco BD-241830
Lithium chloride Sigma L9650
Glycine Omnipur 4840
EDTA Gibco 15575-038
DTT Sigma D5545
Collagenase, type IV Worthington LS004089
DNAse I Worthington LS002007
Diff-Quik Dade-Behring B4132-1A
PowerGen 1000 homogenizer Fisher 14-261-06
stainless steel type 304 mesh #80 Small Parts, Inc. CX-0080-C
Cytospin Statspin M801-22

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