Background: Rare plants that contain corrinoid compounds mostly comprise cobalamin analogues, which may compete with cobalamin (vitamin B₁₂ (B₁₂)) metabolism. We examined the presence of B₁₂ in a cultivated strain of an aquatic plant: Wolffia globosa (Mankai), and predicted functional pathways using gut-bioreactor, and the effects of long-term Mankai consumption as a partial meat substitute, on serum B₁₂ concentrations. Methods: We used microbiological assay, liquid-chromatography/electrospray-ionization-tandem-mass-spectrometry (LC-MS/MS), and anoxic bioreactors for the B₁₂ experiments. We explored the effect of a green Mediterranean/low-meat diet, containing 100 g of frozen Mankai shake/day, on serum B₁₂ levels during the 18-month DIRECT-PLUS (ID:NCT03020186) weight-loss trial, compared with control and Mediterranean diet groups. Results: The B₁₂ content of Mankai was consistent at different seasons (p = 0.76). Several cobalamin congeners (Hydroxocobalamin(OH-B₁₂); 5-deoxyadenosylcobalamin(Ado-B₁₂); methylcobalamin(Me-B₁₂); cyanocobalamin(CN-B₁₂)) were identified in Mankai extracts, whereas no pseudo B₁₂ was detected. A higher abundance of 16S-rRNA gene amplicon sequences associated with a genome containing a KEGG ortholog involved in microbial B₁₂ metabolism were observed, compared with control bioreactors that lacked Mankai. Following the DIRECT-PLUS intervention (n = 294 participants; retention-rate = 89%; baseline B₁₂ = 420.5 ± 187.8 pg/mL), serum B₁₂ increased by 5.2% in control, 9.9% in Mediterranean, and 15.4% in Mankai-containing green Mediterranean/low-meat diets (p = 0.025 between extreme groups). Conclusions: Mankai plant contains bioactive B₁₂ compounds and could serve as a B₁₂ plant-based food source.

Background: Polyphenols are secondary metabolites produced by plants to defend themselves from environmental stressors. We explored the effect of Wolffia globosa ‘Mankai’, a novel cultivated strain of a polyphenol-rich aquatic plant, on the metabolomic-gut clinical axis in vitro, in-vivo and in a clinical trial. Methods: We used mass-spectrometry-based metabolomics methods from three laboratories to detect Mankai phenolic metabolites and examined predicted functional pathways in a Mankai artificial-gut bioreactor. Plasma and urine polyphenols were assessed among the 294 DIRECT-PLUS 18-month trial participants, comparing the effect of a polyphenol-rich green-Mediterranean diet (+1240 mg/polyphenols/day, provided by Mankai, green tea and walnuts) to a walnuts-enriched (+440 mg/polyphenols/day) Mediterranean diet and a healthy controlled diet. Results: Approximately 200 different phenolic compounds were specifically detected in the Mankai plant. The Mankai-supplemented bioreactor artificial gut displayed a significantly higher relative-abundance of 16S-rRNA bacterial gene sequences encoding for enzymes involved in phenolic compound degradation. In humans, several Mankai-related plasma and urine polyphenols were differentially elevated in the green Mediterranean group compared with the other groups (p < 0.05) after six and 18 months of intervention (e.g., urine hydroxy-phenyl-acetic-acid and urolithin-A; plasma Naringenin and 2,5-diOH-benzoic-acid). Specific polyphenols, such as urolithin-A and 4-ethylphenol, were directly involved with clinical weight-related changes. Conclusions: The Mankai new plant is rich in various unique potent polyphenols, potentially affecting the metabolomic-gut-clinical axis.

Many higher and lower animal gut ecosystems have complex resident microbial communities. In contrast, ingested soil is the primary source of the gut microbial diversity of earthworms, invertebrates of fundamental importance to the terrestrial biosphere. Earthworms also harbor a few endemic bacteria including Tenericutes-affiliated Candidatus Lumbricincola of unknown function. Gut microbes are subject to nutrient fluctuations due to dilution effects during gut passage, the nutrient richness of the anoxic gut, and dietary organic carbon, factors that could alter their activity/detection. This study's objective was to assess the potential impact of these factors on the occurrence and activity of ingested and endemic bacteria in gut content of Lumbricus terrestris. Fermentation product profiles of anoxic undiluted and diluted gut content treatments were similar, suggesting that experimental increase in water content and nutrient dilution had marginal impact on fermentation. However, 16S ribosomal Ribonucleic Acid (16S rRNA) sequence abundances indicated that stimulated bacterial taxa were not identical in undiluted and diluted treatments, with dominate potentially functionally redundant phylotypes being affiliated to the Firmicutes, Fusobacteria and Proteobacteria. Although the earthworm-associated Tenericutes were not stimulated in these treatments, the occurrence of three Tenericutes-affiliated phylotypes varied with the organic carbon richness of the earthworm diet, with two phylotypes being associated with high organic carbon richness. 16S rRNA sequence abundances indicated that other dominant gut taxa also varied with dietary organic carbon richness. These findings illustrate that functionally redundant ingested bacteria and earthworm-associated Tenericutes might be influenced by nutrient fluctuations in the gut and organic carbon richness of the earthworm diet.