Exploring the effect of dietary components on gut microbiota "in vitro"obtention, maintenance and characterization of complex microbial communities

Abstract

Gut microbiota is a complex and dynamic ecosystem comprising the myriads of microorganisms that inhabit the human intesnal tract. Gut microbiota plays relevant funcons that are crucial to human health. Diet is the major factor by which the gut microbiota can be modulated. Dietary components that resist digeson in the upper gastrointesnal tract reach the large intesne, which harbors the most numerous and diverse microbial populaons of the intesne, where they constute a substrate for microbial growth. This Doctoral Thesis hypothesizes that the human colon funcons as an enrichment culture and that rests of undigested foods reaching the large intesne promote the proliferaon of microorganisms able to degrade them, or microorganisms directly or indirectly benefing from the metabolism or acvity of the former. Addionally, complex relaonships are established among microbiota members, which may impact both communies composion and acvity. This research addresses the study of the gut microbiota from a community perspecve and proposes that microbial communies of the human gut microbiota derived from the metabolism of different difficult-to-digest nutrients can be obtained and maintained in vitro for their study and characterizaon, based on a sequenal batch culture approach. The objecve of this study was to obtain, maintain and characterize microbial communies from the human gut microbiota derived from the metabolism of food components resistant to digeson by human enzymes. Part I of this Doctoral Thesis focused on the study of gluten, a difficult-to-digest protein. Part II addressed the analysis of communies derived from the metabolism of non-digesble carbohydrates classified as dietary fibers, including guar gum, locust-bean gum, inulin, pecn and cellulose. For this purpose, enrichment culture media containing gluten (pepsin-digested gluten and/or Non-Digested Gluten (NDG)), or the different dietary fibers were inoculated with the fecal microbiota of healthy volunteers. Microbial communies were maintained in vitro by daily subculturing in the enrichment media and communies composion, microbial growth and metabolic acvity were analyzed along me and across the different condions. Findings from Part I showed that, through the developed methodology, microbial communies from the human gut microbiota derived from the metabolism of gluten were obtained and maintained in vitro with a stable composion and metabolic acvity along me. Compared to only digested gluten, the presence of digested gluten plus NDG in the culture media did not affect microbial growth. Nevertheless, NDG presence resulted in communies significantly enriched in microorganisms and taxonomic groups with potenal implicaons in human health. Similarly, communies cultured in NDG presence or absence showed different producon or consumpon raons for several metabolites, many of which are involved in human metabolic pathways, physiological processes or for which altered levels in feces have been associated with various disorders. Results obtained in Part II indicated that the proposed methodology enabled the obtenon of microbial communies from the human gut microbiota derived from the metabolism of different dietary fibers and their successful maintenance along me with a predominantly stable composion and metabolic acvity. The presence of different fibers resulted in variable microbial growth rates and disncvely affected communies composion and metabolic acvity. Notable variaons in different taxonomic groups and metabolites regarded as relevant for human physiology and health were detected. Altogether, the results in this Doctoral Thesis indicate that microbial communies from the human gut derived from the metabolism of difficult-to-digest nutrients can be obtained and stably maintained in vitro for their further characterizaon by performing sequenal batch cultures in appropriate enrichment culture media. Findings presented in this study suggest that dietary components parally or totally resistant to digeson by human enzymes may influence the composion, the growth and the metabolic acvity of the gut microbiota and, consequently, potenally affect human physiology and health at both intesnal and systemic levels. Globally, the results in this study support the relevance of studying the gut microbiota from a community perspecve, considering the importance of the interacons among community members in the final community outcomes.