Animal Models of Listeria Infection

Didier Cabanes1, Marc Lecuit2, Pascale Cossart3

1 Group of Molecular Microbiology, Instituto de Biologia Molecular e Celular (IBMC), Universidade do Porto, Portugal, 2 Service des Maladies Infectieuses et Tropicales, Hopital Necker‐Enfants Malades, Faculté de Médecine, Paris, France, 3 Unité des Interactions Bactéries Cellules, Institut Pasteur, Paris, France
Publication Name:  Current Protocols in Microbiology
Unit Number:  Unit 9B.1
DOI:  10.1002/9780471729259.mc09b01s10
Online Posting Date:  August, 2008
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Abstract

Listeria monocytogenes is an intracellular foodborne pathogen that causes listeriosis, an infection characterized by gastroenteritis, meningitis, encephalitis, and maternofetal infections in humans. L. monocytogenes enters the host via contaminated foods, invades the small intestine, translocates to mesenteric lymph nodes, and spreads to the liver, spleen, brain and, in pregnant women, the fetoplacental unit. Many pathogenicity tests for studying L. monocytogenes have been developed, including tests using laboratory animals. A number of small animal species can be experimentally infected with Listeria. Mice and guinea pigs can be infected either intragastrically or intravenously, and virulence evaluated either by enumerating bacteria within infected target organs or by evaluating the 50% lethal dose (LD50). Although mice and guinea pigs can be infected with Listeria by a variety of routes, the intragastric route is the most relevant to the human foodborne listeriosis. Curr. Protoc. Microbiol. 10:9B.1.1‐9B.1.17. © 2008 by John Wiley & Sons, Inc.

Keywords: Listeria monocytogenes; listeriosis; foodborne pathogen; infection; animal model

     
 
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Table of Contents

  • Introduction
  • Basic Protocol 1: Mouse Intragastric (IG) Infection
  • Basic Protocol 2: Mouse Intravenous (IV) Infection
  • Alternate Protocol 1: Guinea Pig Intragastric (IG) Infection
  • Basic Protocol 3: Organ Recovery and Bacterial Enumeration
  • Basic Protocol 4: Determination of Mortality
  • Support Protocol 1: Preparation of Bacterial Inoculum
  • Commentary
  • Literature Cited
  • Figures
     
 
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Materials

Basic Protocol 1: Mouse Intragastric (IG) Infection

  Materials
  • 6‐ to 8‐week‐old sex‐matched iFABP‐hEcad mice
  • Bacterial inoculum ( protocol 6)
  • Phosphate‐buffered saline (PBS), sterile
  • Brain heart infusion (BHI) agar plates
  • 150 mg/ml CaCO 3 in PBS, prepare fresh
  • Housing cages for the mice
  • Personal protective equipment (e.g., laboratory coat, gloves, and eye protection)
  • 37°C incubator
  • Sterile 1‐ml syringe with a long, bulbous‐ended needle (e.g., animal feeding needle 20‐G × ½ in. with silicon tip)
  • Additional reagents and equipment for the anesthesia of rodents (Donovan and Brown, )

Basic Protocol 2: Mouse Intravenous (IV) Infection

  Materials
  • 6‐ to 8‐week‐old female BALB/c mice
  • Bacterial inoculum ( protocol 6)
  • Phosphate‐buffered saline (PBS), sterile
  • BHI agar plates
  • Personal protective equipment (e.g., laboratory coat, gloves, and eye protection)
  • 37°C incubator
  • Housing cages for the mice
  • Heat lamp
  • Restraining device for mice (Harvard Apparatus)
  • Sterile 1‐ml syringe with a small diameter needle (e.g., 26‐G × ⅜ in.)
  • Sterile gauze

Alternate Protocol 1: Guinea Pig Intragastric (IG) Infection

  Materials
  • 300‐g, sex‐matched Hartley guinea pigs
  • Bacterial inoculum ( protocol 6)
  • Phosphate‐buffered saline (PBS), saline
  • BHI agar plates, sterile
  • 15 mg/ml ketamine in sterile PBS, prepare fresh
  • 25 mg/ml CaCO 3 in PBS, prepare fresh
  • Personal protective equipment (e.g., laboratory coat, gloves, and eye protection)
  • 37°C incubator
  • 1‐ml syringe with a small diameter needle (e.g., 26‐G × ⅜ in.), sterile
  • 20‐ml syringe
  • Sterile suction catheter (1.7‐mm × 270‐mm; Vygon, ref. no. 530.05)
  • Animal cages
  • Additional reagents and equipment for the anesthesia of rodents (Donovan and Brown, )

Basic Protocol 3: Organ Recovery and Bacterial Enumeration

  Materials
  • Infected 6‐ to 8‐week‐old sex‐matched mice ( protocol 1or protocol 2)
  • 70% ethanol in a spray bottle
  • Phosphate‐buffered saline (PBS), sterile
  • Dulbecco's minimum essential medium (DMEM)
  • 100 mg/liter gentamicin in sterile PBS, prepare fresh
  • Sterile Brain Heart Infusion (BHI) medium
  • BHI agar petri plates, sterile
  • Personal protective equipment (e.g., laboratory coat, gloves, and eye protection)
  • 15‐ or 50‐ml conical polypropylene centrifuge tubes, sterile
  • Sterile dissection instruments including:
    • Medium dissection scissors
    • Tissue forceps
    • Small iris scissors
    • Iris forceps
    • Small‐toothed forceps
    • Surgical scissors
    • Dissecting pins
  • Styrofoam or cork cutting board and pins
  • Paper towels
  • 20°C incubator
  • Rotor‐stator homogenizer (IKA UltraTurrax T25, or similar)
  • Additional reagents and equipment for the euthanasia of rodents (Donovan and Brown, )

Basic Protocol 4: Determination of Mortality

  Materials
  • Bacterial inoculum ( protocol 6)
  • Phosphate‐buffered saline (PBS), sterile
  • BHI agar plates, sterile
  • 37°C incubator
  • Additional reagents and equipment for the euthanasia of rodents (Donovan and Brown, )

Support Protocol 1: Preparation of Bacterial Inoculum

  Materials
  • Single colony of Listeria cells
  • Sterile Brain Heart Infusion (BHI) medium
  • Phosphate‐buffered saline (PBS), sterile
  • BHI agar plates
  • 250‐ml Erlenmeyer flask
  • 37°C shaking incubator
  • 2‐liter flask
  • 50‐ml polypropylene conical tubes, prechilled
  • Microtubes, prechilled
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Figures

Videos

Literature Cited

Literature Cited
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