ABSTRACTIntestinal epithelial cells (IECs), including secretory goblet cells, form essential physiochemical barriers that separate luminal bacteria from underlying immune cells in the intestinal mucosa. IECs are common targets for enteric bacterial pathogens, with hosts responding to these microbes through innate toll-like receptors that predominantly signal through the MyD88 adaptor protein. In fact, MyD88 signaling confers protection against several enteric bacterial pathogens, includingSalmonella entericaserovar Typhimurium andCitrobacter rodentium. Since IECs are considered innately hyporesponsive, it is unclear whether MyD88 signaling within IECs contributes to this protection. We infected mice lacking MyD88 solely in their IECs (IEC-Myd88−/−) withS.Typhimurium. Compared to wild-type (WT) mice, infectedIEC-Myd88−/−mice suffered accelerated tissue damage, exaggerated barrier disruption, and impaired goblet cell responses (Muc2 and RELMβ). Immunostaining revealedS.Typhimurium penetrated the IECs ofIEC-Myd88−/−mice, unlike in WT mice, where they were sequestered to the lumen. When isolated crypts were assayed for their antimicrobial actions, crypts fromIEC-Myd88−/−mice were severely impaired in their antimicrobial activity againstS.Typhimurium. We also examined whether MyD88 signaling in IECs impacted host defense againstC. rodentium, withIEC-Myd88−/−mice again suffering exaggerated tissue damage, impaired goblet cell responses, and reduced antimicrobial activity againstC. rodentium. These results demonstrate that MyD88 signaling within IECs plays an important protective role at early stages of infection, influencing host susceptibility to infection by controlling the ability of the pathogen to reach and survive at the intestinal mucosal surface.