Maternal Cafeteria Diet and Methyl Donor Supplementation Modulate Gut Microbiota and Anxiety-Like Behavior in Male Offspring

Maternal cafeteria diet and methyl donor supplementation modulate gut microbiota and behavioral outcomes in male offspring. Background: Maternal hypercaloric diets rich in saturated fats and refined sugars are associated with metabolic alterations, gut microbiota dysbiosis, and behavioral disturbances in offspring. Methyl donor supplementation has been proposed as a potential strategy to modulate these effects. Objective: To evaluate the effect of maternal cafeteria diet consumption and methyl donor supplementation on gut microbiota composition and behavioral outcomes in male offspring in a murine model. Design: Female C57BL/6 mice were assigned to four dietary groups: control, cafeteria diet, control supplemented with methyl donors, and cafeteria diet supplemented with methyl donors. Diet exposure occurred during pre-gestation, gestation, and lactation. Male offspring were evaluated at 8 weeks of age using behavioral tests and gut microbiota analysis based on 16S rRNA gene sequencing. Associations between gut microbiota and behavioral parameters were evaluated using adjusted linear regression models controlling for maternal diet and methyl donor supplementation. Results: Offspring from cafeteria-fed dams supplemented with methyl donors showed higher microbial diversity compared to the non-supplemented cafeteria group. Cafeteria diet increased the abundance of Deferribacteres, Mucispirillum, Adlercreutzia, Butyricicoccus, and Prevotella, whereas methyl donor supplementation reduced Deferribacteres and modified the abundance of Paraprevotella and Ruminococcus_1. At the species level, Mucispirillum schaedleri and Lactobacillus reuteri were increased under cafeteria dietary conditions. No significant effects were observed in sociability-related variables after adjustment. However, Butyricicoccus remained associated with central and peripheral zone behavior, whereas Paraprevotella remained positively associated with wall-leaning behavior after adjustment for maternal diet and supplementation status. Discussion: Maternal cafeteria diet modulated offspring gut microbiota composition and was associated with anxiety-related behavioral parameters. Methyl donor supplementation showed differential effects depending on maternal dietary context, reducing specific bacterial taxa associated with inflammatory and metabolic alterations. Conclusions: Maternal methyl donor supplementation attenuated specific microbiota alterations induced by cafeteria diets and was associated with microbiota–behavior relationships related to anxiety-like responses in offspring.