Distinct Gut Microbiota Signatures Characterize Glucoregulatory Effects of Prenatal Choline and Betaine in Wistar Rat Offspring

BACKGROUND: Prenatal supplementation with the essential nutrient choline improves insulin sensitivity in Wistar rat offspring. Whether these benefits extend to choline's oxidized derivative, betaine, and how they relate to gut microbiota composition and function remains unclear. OBJECTIVES: We investigated the effects of prenatal choline or betaine supplementation on metabolic phenotypes and whether gut microbiota features predict functional outcomes in offspring. METHODS: Pregnant Wistar rats (n = 11/group) were fed an AIN-93G diet and randomly assigned to receive either 0.25% choline, 0.25% betaine, or no supplementation (control) in their drinking water during pregnancy. One female and one male offspring from each dam (n = 11/group) were weaned to a high-fat diet for 12 wk. Metabolic measures were analyzed using analysis of variance models, gut microbiota profiles were evaluated with Analysis of Composition of Microbiomes with Bias Correction, and predictive capacity was tested using nested cross-validation. RESULTS: Offspring of choline- and betaine-supplemented dams showed lower body weight (8% in females, P < 0.0001; 7% in males, P < 0.01) and food intake (7% in females, 10% in males; both P < 0.05) compared with control, with variations in fasting blood glucose, plasma glucagon, and insulin. Colon total glucagon-like peptide-1 (GLP-1) concentrations were higher in both supplemented groups (50% in females, 40% in males; P < 0.0001), with betaine exerting 80% higher circulating total GLP-1 in males (P < 0.001). Prenatal choline and betaine produced distinct, sex-specific gut microbiota signatures, with 40% higher fecal butyrate concentrations (P < 0.0001). Machine learning identified Akkermansia and Adlercreutzia, overlapping with betaine exposure, as predictors of fecal butyrate (r = 0.48, P < 0.001) and colon GLP-1 concentrations (r = 0.34, P < 0.05). CONCLUSIONS: Prenatal supplementation of choline or betaine enhances features of the glucoregulatory system in offspring in concert with shifts in gut microbiota composition that were predictive of metabolic function.