Probiotic microorganisms are able to survive during transit through the gastrointestinal tract, and among other beneficial health effects, may help in the reduction of inflammatory conditions and promote the normalization of the intestinal microbiota. Meanwhile, vitamin D has its biological functions mainly through the mediation of the vitamin D receptor (VDR), acting on the antibacterial mechanism of the innate immune system. Vitamin D/VDR deficiency may be associated with the development or aggravation of certain illnesses, including inflammatory bowel diseases. The VDR is encountered in several sites of the body, which may explain this connection. Besides, probiotics may have a protective effect against induced colitis depending on the VDR status. The administration of butyrate, a microbial metabolite, raises the VDR expression, suggesting a direct relation between the potential health benefits of probiotics and the VDR expression. Furthermore, vitamin D and VDR expression are linked to the composition of the intestinal microbiota, as well as the susceptibility for the development of autoimmune diseases. Thus, this chapter aims to discuss the interaction of probiotics, vitamin D, and VDR regarding the intestinal microbiota and the possible immunomodulatory effects.