| Summary: | One of the obstacles to eradicating <i>paratuberculosis</i> or Johne’s Disease (JD) seems to be the persistence of <i>Mycobacterium avium</i> subsp. <i>paratuberculosis</i> (<i>Map</i>) in the environment due to its ability to survive alone or vectorized. It has been shown that <i>Map</i> is widely distributed in soils and water. Previously, we isolated amoebae associated with <i>Map</i> strains in the environment of bovines from an infected herd. This work aims to verify our working hypothesis, which suggests that amoebae may play a role in the transmission of JD. In this study, we sampled water in the vicinity of herds infected with <i>Map</i> or <i>Mycobacterium bovis</i> (<i>M. bovis</i>) and searched for amoebae and mycobacteria. Live amoebae were recovered from all samples. Among these amoebae, four isolates associated with the presence of mycobacteria were identified and characterized. <i>Map</i> and other mycobacterial species were detected by qPCR and, in some cases, by culture. This study suggests that amoebae and <i>Map</i> may be found in the same environment and might represent a risk of exposure of animals to pathogenic mycobacteria. These data open up new perspectives on the control measures to be put in place to prevent contamination by <i>Map</i>.
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