The Robogut: A Bioreactor Model of the Human Colon for Evaluation of Gut Microbial Community Ecology and Function

Abstract

AbstractThe human colon is inhabited by a complex community of microbes. These microbes are integral to host health and physiology. Understanding how and when the microbiome causally influences host health will require microbiome models that can be tightly controlled and manipulated. While in vivo models are unrivalled in their ability to study host‐microbial interplay, in vitro models are gaining in popularity as methods to study the ecology and function of the gut microbiota, and benefit from tight controllability and reproducibility, as well as reduced ethical constraints. In this set of protocols, we describe the Robogut, a single‐stage bioreactor system designed to replicate the conditions of the distal human colon, to culture whole microbial communities derived from stool and/or colonic biopsy samples, with consideration of methods to create culture medium formulations and to build, run, and sample the bioreactor apparatus. Cleaning and maintenance of the bioreactor system are also described. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC.Basic Protocol 1: Growth medium preparationSupport Protocol 1: Preparing medium supplementsBasic Protocol 2: Preparing the bioreactor vesselsSupport Protocol 2: Making acid and base bottlesSupport Protocol 3: Preparing the effluent bottlesSupport Protocol 4: Making acid solutionSupport Protocol 5: Making base solutionBasic Protocol 3: Preparing inoculum and inoculating bioreactorsAlternate Protocol 1: Preparing inoculum less than 0.5% (w/v) of vessel volumeAlternate Protocol 2: Preparing synthetic community aliquots and inoculation via the septumAlternate Protocol 3: Preparing inoculum from a tissue sampleBasic Protocol 4: Sampling the bioreactor vesselBasic Protocol 5: Harvesting bioreactor vessel contents at end of experimentSupport Protocol 6: Cleaning and sterilizing sampling needlesBasic Protocol 6: Cleaning the bioreactor vesselSupport Protocol 7: Cleaning bioreactor support bottles