Gut Microbiome Standardization in Control and Experimental Mice

Kathy D. McCoy1, Markus B. Geuking2, Francesca Ronchi3

1 Calvin, Phoebe, and Joan Snyder Institute for Chronic Diseases, Cumming School of Medicine, University of Calgary, Calgary, Alberta, 2 Department of Microbiology, Immunology, and Infectious Diseases, Cumming School of Medicine, University of Calgary, Calgary, Alberta, 3 Maurice Müller Laboratories, Department of Clinical Research (DKF), UVCM, University Hospital, Bern
Publication Name:  Current Protocols in Immunology
Unit Number:  Unit 23.1
DOI:  10.1002/cpim.25
Online Posting Date:  April, 2017
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Abstract

Mouse models are used extensively to study human health and to investigate the mechanisms underlying human disease. In the past, most animal studies were performed without taking into consideration the impact of the microbiota. However, the microbiota that colonizes all body surfaces, including the gastrointestinal tract, respiratory tract, genitourinary tract, and skin, heavily impacts nearly every aspect of host physiology. When performing studies utilizing mouse models it is critical to understand that the microbiome is heavily impacted by environmental factors, including (but not limited to) food, bedding, caging, and temperature. In addition, stochastic changes in the microbiota can occur over time that also play a role in shaping microbial composition. These factors lead to massive variability in the composition of the microbiota between animal facilities and research institutions, and even within a single facility. Lack of experimental reproducibility between research groups has highlighted the necessity for rigorously controlled experimental designs in order to standardize the microbiota between control and experimental animals. Well controlled experiments are mandatory in order to reduce variability and allow correct interpretation of experimental results, not just of host‐microbiome studies but of all mouse models of human disease. The protocols presented are aimed to design experiments that control the microbiota composition between different genetic strains of experimental mice within an animal unit. © 2017 by John Wiley & Sons, Inc.

Keywords: co‐housing; fecal transplants; germ‐free; littermate controls; litter swaps; microbiota standardization

     
 
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Table of Contents

  • Introduction
  • Basic Protocol 1: Littermate‐Controlled Experimental Setup to Ensure Standardization of Gut Microbiomes of Control and Experimental Mice
  • Alternate Protocol 1: Experimental Setup for Colonizing Germ‐Free Mice to Ensure Standardization of Gut Microbiomes of Control and Experimental Mice
  • Basic Protocol 2: Fecal Transplantation to Determine the Dominance of the Microbiota Versus the Host Genome on Driving a Phenotype
  • Alternate Protocol 2: Co‐Housing and/Or Litter Swap to Determine the Dominance of the Microbiota Versus the Host Genome on Driving a Phenotype
  • Commentary
  • Literature Cited
  • Figures
     
 
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Materials

Basic Protocol 1: Littermate‐Controlled Experimental Setup to Ensure Standardization of Gut Microbiomes of Control and Experimental Mice

  Materials
  • Adult male and female mice (6 to 8 weeks old that have reached sexual maturity) of the control background (e.g., C57BL/6) and mutant or genetically modified strain
  • Animal housing facility with appropriate sterile food, water, bedding, and environmental enrichments
  • 2‐ml microcentrifuge tubes with safe lock lid
  • Liquid nitrogen tank and container
  • −80°C freezer
  • Additional reagents and equipment for genotyping (see Conner, )

Alternate Protocol 1: Experimental Setup for Colonizing Germ‐Free Mice to Ensure Standardization of Gut Microbiomes of Control and Experimental Mice

  Additional Materials (also see protocol 1)
  • Germ‐free adult wild‐type male mice of the control background (e.g., C57BL/6)
  • SPF adult female mice of the genetically modified strain

Basic Protocol 2: Fecal Transplantation to Determine the Dominance of the Microbiota Versus the Host Genome on Driving a Phenotype

  Materials
  • Adult mice of the control and mutant mouse strains
  • Phosphate‐buffered saline, sterile (PBS; see appendix 2A)
  • 3‐week‐old pups taken at the time of weaning
  • 50‐ml conical tubes, sterile
  • 2‐ml microcentrifuge tubes with safe lock lid, sterile
  • Liquid nitrogen tank and container
  • −80°C freezer
  • Gavage feeding needles with 1‐ml syringe, sterile

Alternate Protocol 2: Co‐Housing and/Or Litter Swap to Determine the Dominance of the Microbiota Versus the Host Genome on Driving a Phenotype

  Materials
  • Adult male and female mice (6 to 8 weeks old that have reached sexual maturity) of the control background (e.g., C57BL/6) and mutant mouse strains
  • Animal housing facility with appropriate sterile food, water, bedding, and environmental enrichments
  • 2‐ml microcentrifuge tubes with safe lock lid
  • Liquid nitrogen tank and container
  • −80°C freezer
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Figures

Videos

Literature Cited

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Key References
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