Free-living amoebae are described as potential reservoirs for pathogenic bacteria in the environment. It has been hypothesized that this might be the case for subsp. , the bacterium responsible for paratuberculosis. In a previous work, we isolated an amoeba from a water sample in the environment of infected cattle and showed that this amoeba was associated with subsp. . While a partial 18S rRNA gene has allowed us to suggest that this amoeba was -like, at that time we were not able to sub-cultivate it. In the present study, we succeeded in cultivating this strain at 20-25°C. This amoeba is among the smallest (5-7 μm) described. The sequencing of the whole genome allowed us to extract the full 18S rRNA gene and propose this strain as a new species of the genus, i.e., . Of note, the mitochondrial genome is particularly large (184,954 bp). Finally, we showed that this amoeba was able to phagocyte subsp. and that the bacterium was still observed within amoebae after at least 3 days. In conclusion, we characterized a new environmental amoeba species at the cellular and genome level that was able to interact with subsp. . As a result, is a potential candidate as environmental reservoir for subsp. but further experiments are needed to test this hypothesis.