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Inoculation of Gut Fermentation Models

The inoculation and colonization of in vitro fermentation systems influences the reproducibility of the studies and constitute a challenge of the models (Payne et al. 2012). Fermenters are usually inoculated with a liquid fecal suspension from an individual or pooling stools from several subjects. In the last case, the fecal samples are used to inoculate a fed-batch fermenter to produce a standardized inoculum that is stored frozen. Cinquin et al. (2004) developed an immobilization process for the entrapment of fecal microbiota in mixed xanthan–gellan gum gel beads to reach high cell densities and to maintain the microbial diversity over long time continuous colonic fermentations. The model has recently been updated into the model PolyFermS where it is possible to stably and reproducibly cultivate complex intestinal communities in multiple reactors allowing studying in parallel the impact of different treatments compared to a control reactor (Zihler Berner et al. 2013). In order to facilitate the reproducibility of experiments, recent developments are addressing the inoculation of fermentation models with defined populations of human gut microorganisms represented by common saccharolytic and amino acid-fermenting populations in the large intestine (Newton et al. 2013).

Host-Gut Microbiota Interaction

A major drawback of gut fermentation models is the limitations for simulating the host functionality. The combination of fermentation models and intestinal cell cultures represent a common approach to reproduce in vitro the host responses. These studies have been usually performed with colon epithelial cell cultures and/or immune cells to evaluate adherence, cytokine production and gene expression, among others (Venema and Van den Abbeele 2013). Additional tools that have improved modeling of the physiological colonic conditions are the incorporation in the reactors of a mucosal environment able to differentiate between the luminal microbiota and the microbial biofilms adhering to the colonic epithelium (Macfarlane et al. 2005; Van den Abbeele et al. 2012). The Host-Microbiota Interaction (HMI) module is a recently developed device adapted to the SHIME model that allows long-term studies of a complex microbial community colonizing a mucus layer, while being co-cultured microaerophilically (up to 48 h) in the presence of shear forces and a monolayer of enterocyte human cells (Marzorati et al. 2014). This combination of in vitro models represents an approximation to the conceptualized ideal experimental model for the study of host-gut microbiota interactions described by Fritz et al. (2013). These authors claim that the ideal model should include epithelial cells, complex gut microbiota, anaerobic/microaerophilic conditions, a mucus layer and physiological conditions of pH, fluid retention times and dissolved O2 concentrations.

The progress made in developing in vitro fermentation models able to closely mimic the gut microbial environment and the increasing knowledge regarding microbial populations and host-gut microbiota interactions can offer remarkable insights into gut microbiota functions. Moreover, ongoing studies would allow the development of well-defined mixtures of microorganisms that retain a high level of diversity and encompass key functions required for healthy intestinal homeostasis in an approach coined “microbial ecosystem therapeutics” by Petrof et al. (2013).

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