Healthy adult gut microbiota sustains its own vitamin B12 requirement in an in vitro batch fermentation model

Abstract

Vitamin B12 (cobalamin) is present in the human lower gastrointestinal tract either coming from the unabsorbed dietary fraction or from in situ production of the gut microbiota. However, it is unclear whether the gut microbial communities need exogenous B12 for growth and metabolism, or whether B12 in low and high levels could affect gut community composition and metabolite production. Here, we investigated in vitro B12 production of human fecal microbiota and the effects of different levels of B12 (as cyanocobalamin) on composition and activity. Eight fecal communities from healthy human adults distributed over three enterotypes, dominated by Firmicutes (n = 5), Bacteroides (n = 1) or Prevotella (n = 2) were used to perform batch fermentations in Macfarlane medium supplemented with low B12 medium (Control, 5 ng/ml, within the tested fecal range), no B12 addition (NB12), and high B12 addition (ExtraB12, 2500 ng/ml). The microbiota community composition (qPCR, 16S rRNA metabarcoding), metabolic activity (HPLC-RI), and B12 levels (UHPLC-DAD) were measured after 24 h incubation at 37°C under strict anaerobic conditions. All fecal microbial communities produced B12 in the NB12 condition after 24 h, in the range from 152 ± 4 to 564 ± 25 ng/ml. None of the B12 treatments had an impact on total bacterial growth, community richness, diversity and total metabolite production, compared to the low B12 control. However, a significant increase of propionate was measured in ExtraB12 compared to NB12. Most taxonomic and metabolite changes compared to control incubations were donor-dependent, implying donor-microbiota-specific changes upon B12 treatments. Our in vitro data suggest that healthy human adult gut microbial communities have the capacity to produce B12 at levels fulfilling their own requirements, independently of the initial B12 content tested in the donor’s feces. Further, supplementation of exogenous dietary B12 may have limited impact on the healthy human gut microbial community composition and function.

Veillonella and Bacteroides are associated with gestational diabetes mellitus exposure and gut microbiota immaturity

“…Considering this information and our results, one hypothesis is that those prototrophic taxa, such as Veillonella , would help supply cobalamin to auxotrophic taxa during the first months of life. Kundra et al demonstrated that even in the absence of B12, the presence of prototrophic bacteria could supply sufficient cobalamin for the remaining 80% of auxotrophic taxa [ 46 ]. Thus, the presence of Veillonella could help modulate the configuration of the microbiota in the early stages of life.…”

Vitamin B12 analogues from gut microbes and diet differentially impact commensal propionate producers of the human gut

“…These analogues of B12 are used in diet and dietary supplements and also found in human feces ( 21 , 35 ). B12 forms were tested at two doses, 10 μg/L (1x) and a 20-fold higher dose of 200 μg/L (20x), which are, respectively, two times and 40 times higher than in the medium used for human colon microbiota cultivation ( 16 , 36 ). These concentrations were based on suggested concentrations for cultivating single gut microbes (10 μg/L) ( 33 ).…”

“…These concentrations were based on suggested concentrations for cultivating single gut microbes (10 μg/L) ( 33 ). In comparison, under normal dietary conditions of 4.5 μg/day, approximately 3.75 μg/L of B12 is expected to reach the colon from dietary sources, which increases to 2,500 μg/L when taking supplements equivalent to a single dose of 1,500 μg/day per tablet ( 16 ). Strains were cultivated in monocultures in BMA and inoculated after cell washing as described above.…”

“…For production of propionate via the succinate and the propanediol pathways, vitamin B12 (B12) is required as a cofactor for methylmalonyl-CoA mutase and glycerol/diol dehydratases, respectively ( 14 ). In vitro studies have shown that B12 supplementation can enhance propionate production by single microbes such as Akkermansia muciniphila , and also by complex human gut microbiota in fecal batch fermentation ( 15 , 16 ). However, the effect of exogenous B12 on propionate production is not yet fully understood, as it varies depending on the donor microbiota ( 16 ).…”

“…In vitro studies have shown that B12 supplementation can enhance propionate production by single microbes such as Akkermansia muciniphila , and also by complex human gut microbiota in fecal batch fermentation ( 15 , 16 ). However, the effect of exogenous B12 on propionate production is not yet fully understood, as it varies depending on the donor microbiota ( 16 ).…”