Inflammatory bowel disease (IBD) and colorectal cancer (CRC) are the major diseases of the lower gastrointestinal tract. The intestinal epithelium plays a critical role in the host's interactions with the large communities of resident luminal bacteria. Epithelial cells recognize the bacterial components via pattern-recognition receptors. Toll-like receptors (TLRs) are a major class of pattern-recognition receptors that are present on intestinal epithelial cells, including putative stem cells. Stem cells are responsible for tissue homeostasis and regeneration after injury including IBD. Stem cells are also implicated in the pathogenesis of CRC. In susceptible individuals, disruption of normal homeostatic balance between the host's mucosal cells and enteric microflora is believed to result in aberrant immune responses against the resident commensal bacteria, leading to IBD. Microbiological analyses have revealed that the composition and localization of microbiota is altered in CRC and IBD. It is plausible that stem cells directly sense and respond to microbiota. This review aims to summarize the current knowledge on the effect of microbiota and TLR signaling on intestinal stem cells. It also describes how TLR signaling could affect the stem cell regulatory pathways.