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Ampicillin is applied in rodents to induce a temporarily depleted microbiota. To elucidate whether bacteria are just temporarily suppressed or fully eliminated, and how this affects the re-colonisation process, we compared the microbiota and immune system in conventionally housed untreated mice with newly weaned ampicillin treated mice subsequently housed in either a microbe containing environment or in an isolator with only host associated suppressed bacteria to recolonize the gut. Two weeks ampicillin treatment induced a seemingly germ-free state with no bacterial DNA to reveal. Four weeks after treatment caeca were still significantly enlarged in both treated groups, but bacteria re-appeared even in isolator housed mice. While some suppressed bacteria were able to recover and even dominate the community, the abundances and composition were far from the untreated mice and differed between isolator …