Type 1 Diabetes (T1D) is a chronic autoimmune disease caused by the destruction of insulin-producing pancreatic β-cells due to T-cell autoreactivity. The insulinB:9-23 peptide is recognized as a critical epitope triggering T1D. In our previous study, we discovered that a gut bacteria Parabacteroides distasonis contains a hypoxanthine phosphoribosyltransferase (hprt) 4-18 peptide, exhibiting 60% homology to insB:9–23 and stimulates human T-cell clones and nonobese diabetic (NOD) mice T-cell hybridomas specific to insB:9-23. Colonization of P.distasonis in NOD mice elevated immune cell infiltration (insulitis) in the pancreatic islets and accelerated T1D onset compared to the controls. Notably, A reduction in helper T-cells, effector T-cellsand B-cell populations in the intestines of colonized mice was observed. The data indicated that P. distasonis does not stimulate an unspecific inflammatory response. To delineate the specific impacts of P. distasonis and nullify the effect of other microbes, we utilized the NOD Germ-Free (GF) mice model. Notably, P. distasonis colonization in GF mice also induced insulitis in 12 weeks of age. Next, to understand the mechanism by which P. distasonis induces inflammation, we evaluated serum metabolite changes and intestinal permeability. Quantitative PCR analyses unveiled unaltered mRNA levels of the proteins, indicative of preserved gut barrier function following colonization in both GF and Specific Pathogen-Free (SPF) NOD mice. Altogether, the findings support our previous hypothesis regarding the molecular mimicry mechanism by P. distasonis in inducing T1D as colonization resulted in specific T-cell infiltration in the pancreas of NOD mice, rather than in the intestine.