The diguanylate cyclase CrdA contributes to cell adhesion and gut colonization by switching motility and sessile state of Cronobacter malonaticus.

Cronobacter malonaticus is an emerging foodborne pathogen posing severe threats to vulnerable populations. Despite the established role of the ubiquitous second messenger cyclic diguanylate (c-di-GMP) in bacterial virulence, the specific diguanylate cyclases (DGCs) that orchestrate pathogenicity in C. malonaticus are unknown. Here, we systematically characterized 13 putative DGCs in C. malonaticus and identified CrdA as a pivotal regulator. We demonstrated that CrdA reciprocally inhibits swimming motility and promotes cellulose-dependent biofilm formation by modulating intracellular c-di-GMP levels. Functional and structural analyses revealed that the N-terminal transmembrane domains of CrdA are essential for its dimerization and enzymatic activity. Furthermore, CrdA enhances bacterial adhesion to intestinal epithelial cells and gut colonization in vivo, with its expression being induced upon host cell contact. Mechanistically, CrdA suppresses motility partially through the c-di-GMP effector YcgR and regulates biofilm formation in a BcsA-dependent manner. Collectively, this work elucidates a novel signaling pathway where CrdA integrates environmental cues to orchestrate a virulence-associated lifestyle switch, advancing our understanding of c-di-GMP signaling in Cronobacter pathogenicity and highlighting a potential target for disrupting biofilm-mediated persistence in food systems.