Development of the in vitro Cecal Chicken ALIMEntary tRact mOdel-2 to Study Microbiota Composition and Function

Abstract

The digestive system of the chicken plays an important role in metabolism, immunity, and chicken health and production performance. The chicken ceca harbor a diverse microbial community and play a crucial role in the microbial fermentation and production of energy-rich short-chain fatty acids (SCFA). For humans, dogs, and piglets in vitro digestive system models have been developed and are used to study the microbiota composition and metabolism after intervention studies. For chickens, most research on the cecal microbiota has been performed in in vivo experiments or in static in vitro models that may not accurately resemble the in vivo situations. This paper introduces an optimized digestive system model that simulates the conditions in the ceca of the chicken, i.e., the Chicken ALIMEntary tRact mOdel-2 (CALIMERO-2). The system is based on the well-validated TNO in vitro model of the colon-2 (TIM-2) and is the first dynamic in vitro digestion model for chickens species. To validate this model, the pH, temperature, and different types of microbial feeding were compared and analyzed, to best mimic the conditions in the chicken ceca. The bacterial composition, as well as the metabolite production at 72 h, showed no significant difference between the different microbial feedings. Moreover, we compared the CALIMERO-2 digestive samples to the original inoculum and found some significant shifts in bacterial composition after the fermentation started. Over time the bacterial diversity increased and became more similar to the original inoculum. We can conclude that CALIMERO-2 is reproducible and can be used as a digestive system model for the chicken ceca, in which the microbial composition and activity can be maintained and shows similar results to the in vivo cecum. CALIMERO-2 can be used to study effects on composition and activity of the chicken cecum microbiota in response to in-feed interventions.

In vitro Modeling of Chicken Cecal Microbiota Ecology and Metabolism Using the PolyFermS Platform

“…Here, there was an overall shift in microbial composition and a decrease in diversity in vitro compared with in vivo cecal inoculum as measured by qPCR and 16S rRNA sequencing. Such decrease was also observed in other in vitro fermentation models from chicken, humans (infants, adult, and elderly), and swine (Van Den Abbeele et al, 2010;Tanner et al, 2014b;McDonald et al, 2015;Card et al, 2017;Oost et al, 2021). Indeed, in vitro models cannot simulate all conditions encountered in the host, which are highly individual and not well known or microbiotahost complex interactions that cannot be mimicked, such as the immune response, hormone and digestive secretions, adsorption, and peristaltic movement, all of which influences microbial diversity (Lacroix et al, 2015).…”

“…Until now, in vitro chicken cecal continuous fermentation models have been little investigated and applied, whereas most research was done in vivo (Mota de Carvalho et al, 2021;Oost et al, 2021). Card et al (2017) and Gong et al (2019) have described and implemented a continuous culture of the chicken ceca microbiota inoculated with free-cell suspension in single-stage chemostat fermentation.…”

“…Low cell density can highly impact competition and cross-feeding among bacteria, leading to a significant difference in both the α and β diversity over time (Card et al, 2017). Very recently, Oost et al (2021) reported a new in vitro cecal chicken dynamic model developed on the TIM 2 platform. The model used a pooled frozen inoculum that was cryopreserved with addition of 15% glycerol.…”

Characteristics of carbohydrates determine the shape of the gut microbiota in a chicken cecal in-vitro model

“…The cecal contents of broiler chickens (Ross 308) were obtained from slaughterhouse van der Linden Poultry Products B.V. (Beringe, the Netherlands), as previously reported (Oost et al, 2021). Brie y, the days before slaughter, the broiler chickens were fed a coccidiostat-free diet and were not treated with antibiotics.…”

The Interplay between Salmonella and Intestinal Innate Immune Cells in Chickens

“…Targeting the innate immune cells using oligosaccharide compounds, either in-ovo or post-hatch through feed formulations, could be an alternative options to strengthen immune mediated disease resistance in chickens. The recent development of novel in vitro chicken intestine models [231,232] can be used to study immune response of novel carbohydrate compounds against intestinal pathogens including Salmonella. This would help us to identify compounds that can enhance the immune fitness of innate immune cells, which can be used as alternatives to antibiotics.…”