Free-living mammals carry complex microbiota that co-evolved with their hosts over eons of years. The transfer of such microbiota from wild mice to genetically tractable laboratory mice has been shown to enhance modeling of human immune responses in preclinical studies. Here, we assessed the long-term stability of microbiota and immune phenotype of the first C57BL/6 mouse colony with natural microbiota (wildling mice). The bacterial gut microbiota of wildling mice maintained its increased α-diversity and richness over 5 years, with significantly greater stability than the gut microbiota of laboratory mice. Wildling mice had increased myeloid cell numbers across organs and increased activation and function of natural killer, B, and T cells, which was transferable to laboratory mice via co-housing. Immunological readouts in two preclinical models remained stable throughout the follow-up. These results demonstrate the feasibility of maintaining mouse colonies with natural, wild-derived microbiota as a sharable resource for basic and preclinical research.