<b>Persistence mechanisms of Crohn's disease-associated adherent invasive</b> <i> <b>E</b> scherichia <b>coli</b> </i> <b>within macrophages</b>

Patients with Crohn's disease exhibit abnormal intestinal colonization by <i>Proteobacteria</i>, particularly adherent-invasive <i>Escherichia coli</i> (AIEC). These bacteria predominate in the mucus, adhere to epithelial cells, colonize them, and survive inside macrophages. We recently demonstrated that the AIEC strain LF82 adapts to phagolysosomal stress through a two-step process: initial replication arrest generating stress-tolerant persisters, followed by the resumption of replication, leading to the formation of intracellular bacterial communities (IBCs) embedded in a biofilm-like matrix. Given the significant genomic diversity among strains with the AIEC phenotype, we performed a comparative genomic and functional analysis of 13 AIEC isolates from Crohn's disease patients in France and Spain. Our results demonstrate that IBCs are replicative niches for all AIEC strains within THP-1 macrophages, yet their formation relies on distinct mechanisms, including variations in phagosome detoxification, biofilm architecture, and macrophage responses. Our study identifies a strong positive correlation between vacuole acidification and persister induction, which underlies the intracellular survival of the different strains. Furthermore, we revealed distinct AIEC dissemination strategies outside macrophages, potentially contributing to the propagation of inflammation in the human host. These findings highlight that research on pathogens and pathobionts with dynamic genomes should extend beyond classical bacterial models.