The gut microbiome and Vitamin B12 status in humans
Creators: name: Rittmann, Marijke
Publisher: name: No Publisher Supplied
Year: 2024

description: Background: Vitamin B12 is an essential nutrient required for hematological and neurological function. B12 deficiency results in megaloblastic anemia and neurodegenerative disease and may contribute to cognitive decline and dementia risk in older adults. Some bacteria among human gut microbiota can synthesize B12 while others require B12 from the diet of the host. Consequently, variation in gut microbiota may influence B12 status in humans. Aims: The purpose of this dissertation is to assess the associations between gut microbiota composition and host B12 status as assessed by serum total B12, holotranscobalamin (HoloTC), methylmalonic acid (MMA), and total homocysteine (Hcy) concentrations.Methods: A pilot study was first conducted consisting of a secondary analysis of a cohort of 90 adults (44 males/46 females; 20-75y) in Sydney, Australia. A second study was conducted with a cohort of 100 healthy adults (76 females/24 males; 18-81y) recruited in New Brunswick, New Jersey. 16S rRNA gene V3-V4 amplicon sequencing was used to profile gut microbiota at the amplicon sequence variant (ASV) level. Serum B12 was measured by automated chemiluminescence assay, serum MMA by high pressure liquid chromatograhy-tandem mass spectrometry, serum HoloTC by active B12 ELISA kit assay, plasma Hcy by HPLC with post-column fluorescence detection, and plasma short chain fatty acids (propionate, butyrate, and acetate) by gas chromatography-tandem mass spectrometry. In addition, a composite score called combined B12 (cB12) was calculated using a published formula that includes all four blood B12 biomarkers. Microbial guilds were identified based on co-abundance clustering using the Ward clustering algorithm. Random forest regression with feature selection by the Boruta algorithm was used to identify microbial co-abundance guilds that are predictive of measured B12 status biomarker concentrations adjusting for age, sex, body mass index (BMI), and fiber intake in the Australian cohort, and adjusting for age, sex, BMI, race/ethnicity, creatinine, alcohol intake, caffeine intake, fiber intake, and supplemental B12 in the New Jersey cohort. Linear regression was used to assess the associations between the guilds and B12 status biomarkers, and short chain fatty acids (New Jersey cohort only) in univariate analyses and in multivariate analyses adjusting for the covariates listed above for the two cohorts. Mediation analysis was conducted to determine if propionate mediated the associations between guilds and MMA in the New Jersey cohort.Results: In the Australian cohort, mean ± SD serum B12 and MMA were 389 ± 177 pmol/L and 210 ± 80 nmol/L. Thirty microbial co-abundance guilds were identified. Random forest regression with controlling for age, sex, BMI, and fiber intake, revealed that age and guilds 3 and 15 were predictors of B12 (p=0.027), and guilds 4, 27, and 29 were predictors of MMA (p=0.003). Multivariate linear regression revealed that little of the variance in B12 (adjusted R2=0.038) was associated with the guilds, while a higher amount of variance in MMA (adjusted R2=0.196) was associated with the guilds. In the New Jersey cohort mean ± SD serum B12, MMA, HoloTC, Hcy, cB12 and propionate were 349 ± 143 pmol/L, 161 ± 119 nmol/L, 92 ± 49 pmol/L, 7.75 ± 2.36 µmol/L, 0.65 ± 0.05 and 1.34 ± 0.76 µmol/L, respectively. Twenty-eight microbial co-abundance guilds were identified. Random forest regression with controlling for covariates revealed that supplemental B12 and guilds 14, 15, and 26 were predictors of B12 (p&lt;0.0001); guilds 2, 6, and 14, sex, creatinine and BMI were predictive of Hcy (p&lt;0.00001), and guild 13 was a predictor of MMA and cB12 (p=0.01 and p=0.001). No guilds were predictive of HoloTC. In the multilinear regression, serum B12, HoloTC and cB12 models did not reach significance. The amount of variance in Hcy explained by the model was strong (adjusted R2=0.116) and was strengthened by the addition of covariates (adjusted R2=0.201). Likewise, the amount of variance in MMA explained by the model also was strong (adjusted R2=0.218). Propionate was directly associated with MMA and explained 12.6% of the study sample variance in MMA (R2=0.126; p=0.0003). After controlling for age, sex, B12, creatinine, BMI, and fiber intake, the association with propionate remained significant (p=0.002) with the full model including all covariates explaining 31.1% of the variance in MMA (R2=0.311). No significant associations between butyrate or acetate and MMA were found. The mediation analysis revealed that the mediation effect of propionate of the association between one of the guilds (guild 6) and MMA was 20%, but this mediation effect did not reach statistical significance (p=0.14).Conclusion: These results indicate that in these cohorts of healthy adults different microbial co-abundance guilds are predictive of total B12, MMA, HoloTC, Hcy and cB12. Moreover, guilds account for a larger percentage of the variance in MMA and Hcy compared with total B12 and HoloTC. Propionate appears to partially mediate the relationship between MMA and at least one guild. This suggests that some of the variance in serum MMA and Hcy in adults is not specific to B12 status. MMA is generally considered to be a specific biomarker of B12 status, but is also influenced by renal function and age, and as we show here variance in the gut microbiome. This variance is mediated by propionate. This has ramifications for interpreting measures of B12 status in healthy adults and may provide insight into the metabolic connections between gut dysbiosis and diseases such as cancer, cardiovascular disease and neurodegenerative disease.