Perinatal Gut Dysbiosis Reduces Milk Oligosaccharides via LPS‐Mediated Gut–Mammary Signaling in Mice

ABSTRACT Human milk oligosaccharides (HMOs) are bioactive components pivotal for infant health, whose biosynthesis is modulated by intrinsic (genetic, metabolic) and extrinsic (diet, microbiota) maternal factors. Emerging evidence suggests the gut microbiota regulates HMO production through the gut–mammary axis, but mechanistic understanding remains fragmentary. Here, we investigated how gestational and postpartum gut dysbiosis induced by dextran sulfate sodium (DSS) disrupts milk oligosaccharide (MO) synthesis in mice and explored whether targeted probiotic supplementation could counteract lactation insufficiency under dysbiotic conditions. These results revealed that DSS‐exposed dams exhibited gut barrier disruption and systemic inflammation, accompanied by a significant reduction in mammary sialylated oligosaccharides (particularly 3′‐sialyllactose, 3′‐SL) and compromised blood–milk barrier integrity. Through integrated analysis of gut/milk microbiota and host parameters, we identified gut‐derived lipopolysaccharide (LPS) as a key mediator suppressing mammary glycosyltransferase genes critical for lactose synthesis and sialylation. LPS treatment of human mammary epithelial cells (MCF‐10A) recapitulated these effects, confirming direct transcriptional repression. Systematic in vitro screening of 15 probiotic strains identified three candidates ( Lactobacillus reuteri DM24MUS05, Lactobacillus plantarum DM23M20, and Lactobacillus rhamnosus MP108) that upregulated oligosaccharide‐synthesis genes. Probiotic consortium administration restored MO levels in DSS‐treated dams by mitigating enteric inflammation and reconstructing Lactobacillus ‐dominant microbiota. This study explores possible mechanisms linking gut microbiota dysbiosis to impaired MO biosynthesis. Our findings suggest that synergistic probiotics may represent a targeted intervention to counteract gut dysbiosis–associated impaired MO biosynthesis. These results position microbiota modulation as a promising, translatable approach for improving maternal‐infant health outcomes.