Akkermansia muciniphila (A. muciniphila) is an intestinal bacterium that specializes in degrading mucins, which are heavily O-glycosylated proteins that form the mucus lining of the intestine. Despite its ability to adhere to mucins, the molecular mechanisms underlying its binding to mucins remain poorly understood. This study demonstrates that A. muciniphila binds to mucins independently of oxygen levels and is not affected by pasteurization. Using a novel cell-based mucin array, the researchers identified that A. muciniphila selectively binds to the unsialylated LacNAc disaccharide on core2 and core3 O-glycans. This epitope is abundant on human colonic mucins, and the bacterium's endogenous neuraminidase activity can uncover this epitope, facilitating binding. The study highlights the importance of specific O-glycan structures in A. muciniphila's ability to colonize the gut, as it preferentially binds to core3 O-glycans on MUC2 mucins. The findings suggest that A. muciniphila's mucin-binding properties are crucial for its role in the gut microbiome, as it can utilize mucins for energy and support the growth of other bacteria. The study also shows that A. muciniphila's neuraminidase activity is essential for exposing the LacNAc epitope on mucins, enabling binding. These results provide insights into the molecular mechanisms that allow A. muciniphila to interact with mucins and highlight the importance of O-glycan structures in bacterial-mucin interactions. The study underscores the potential of A. muciniphila as a probiotic due to its beneficial effects on gut health and metabolic disorders.Akkermansia muciniphila (A. muciniphila) is an intestinal bacterium that specializes in degrading mucins, which are heavily O-glycosylated proteins that form the mucus lining of the intestine. Despite its ability to adhere to mucins, the molecular mechanisms underlying its binding to mucins remain poorly understood. This study demonstrates that A. muciniphila binds to mucins independently of oxygen levels and is not affected by pasteurization. Using a novel cell-based mucin array, the researchers identified that A. muciniphila selectively binds to the unsialylated LacNAc disaccharide on core2 and core3 O-glycans. This epitope is abundant on human colonic mucins, and the bacterium's endogenous neuraminidase activity can uncover this epitope, facilitating binding. The study highlights the importance of specific O-glycan structures in A. muciniphila's ability to colonize the gut, as it preferentially binds to core3 O-glycans on MUC2 mucins. The findings suggest that A. muciniphila's mucin-binding properties are crucial for its role in the gut microbiome, as it can utilize mucins for energy and support the growth of other bacteria. The study also shows that A. muciniphila's neuraminidase activity is essential for exposing the LacNAc epitope on mucins, enabling binding. These results provide insights into the molecular mechanisms that allow A. muciniphila to interact with mucins and highlight the importance of O-glycan structures in bacterial-mucin interactions. The study underscores the potential of A. muciniphila as a probiotic due to its beneficial effects on gut health and metabolic disorders.