Mouse atonal homolog 1 (Math1/Atoh1) is a basic helix-loop-helix transcription factor important for the differentiation of secretory cells within the intestinal epithelium. The analysis of Paneth depletion efficiency upon Math1^lox/loxVilCreER^T2 ( Math1^∆IEC) mice treatment with Tamoxifen in the presence or absence of intestinal microbiota, showed a failure on Paneth cell depletion in germ-free mice as compared to SPF mice. However, goblet cells were efficiently depleted in Math1^∆IEC germ-free mice. The gene expression of Math1 was significantly reduced in the ileum of germ-free Math1^∆IEC mice 5 days post tamoxifen injection as compared to germ-free control, but its protein expression was still detectable in the nuclei of epithelial cells in the crypts. Germ-free mice showed low proliferative ileal crypts as well as apoptotic cells that were mainly detected in the tip of the villus, consistent with a slow turnover rate of epithelial cells. Although Paneth cells were not depleted in germ-free Math1^∆IEC mice for the first 7 weeks after the last tamoxifen injection - far already from the 5 days timelaps observed in SPF conditions- but an incomplete depletion of Paneth cells was observed 14 weeks after last tamoxifen injection. Colonization of germ-free mice restored the phenotype observed in SPF mice, highlighting the regulatory role of gut microbes in our model. We conclude that absence of intestinal microbiota in Math1^∆IEC mice is associated with reduced epithelial cell renewal and delays the depletion of preexisting Paneth cells.