Impact of Donor Individuality, Temporal Variation, and Culture Medium Type on Microbiota Composition and Metabolic Activity in Human Fecal Batch Culture

Fecal batch culture (FBC) studies often rely on a single fecal sample collection and the use of one type of medium for cultivation, bringing challenges to the interpretation of results and the comparison between studies. This study investigated the impact of donor individuality, temporal variation and culture medium type on microbiota composition and metabolic activity in an FBC setting with the fiber polydextrose (PDX) as carbon and energy source. FBCs were inoculated with fecal microbiota from three healthy donors sampled at three different days (day 1, 2 and 30), using either basal or rich culture medium with PDX as carbon source. Microbiota composition and metabolic activity were determined after 0, 6, 12, and 24 h of incubation. Microbiota composition variation explained by donor individuality dropped from 51% to 16% during incubation, while that explained by medium and PDX supplementation increased from 0% to 17% and 20%, respectively. Independent of the medium, the genera Erysipelotrichaceae UCG-003, Blautia and Fusicatenibacter were stimulated by PDX supplementation. In basal medium Bacteroides and Anaerostipes grew better, whereas Bifidobacterium, Faecalibacterium and Megasphaera grew better in rich medium. Metabolite variation was explained up to 50% by PDX supplementation during incubations, with butyrate being produced at the highest concentrations among all metabolites. Temporal variation explained less than 3% of the variation in both microbiota and metabolite composition. In conclusion, in this study donor individuality had the most profound impact on microbiota succession while medium and PDX supplementation had larger impacts on metabolic activity in FBCs. IMPORTANCEFBCs or other in vitro models are often chosen to assist in obtaining mechanistic insights complementing in vivo microbiome observations by mimicking the colonic fermentation. However, FBCs are prone to a variety of factors such as the individuality of feces donor, temporal variation in microbiota composition within the individual, and cultivation medium. The importance of our study is in reinforcing that both donor individuality and medium type have major impacts on PDX degradation, whilst the impact of temporal variation is limited. Of interest is that bifidobacterial growth was more stimulated in rich medium with PDX as carbon source, whereas growth of members of the Bacteroidetes were more stimulated in basal medium with PDX as carbon source. We recommend that variations in medium and donor samples should be considered when planning and interpreting in vitro incubation studies.