Cultivation of Akkermansia muciniphila

A. muciniphila MucT (ATCC BAA-835) was obtained from the American Type Culture Collection (ATCC). The strain was cultured as described previously [21 (link)]. The bacterial culture medium was brain–heart infusion medium (Qingdao Hope Bio-Technology Company, Qingdao, China) supplemented with hog gastric mucin (0.25% w/v Type II; Sigma-Aldrich, St. Louis, MO, USA). The bacterial cells were grown under anaerobic conditions at 37 °C for 18 h, and were enriched by centrifugation (6000× g, 10 min) and washed with normal saline. Finally, the bacterial sludge was resuspended in normal saline at the required final concentration.