Cyclic di-GMP (c-di-GMP), a universal second messenger in bacteria, orchestrates a wide array of essential life processes. Its intracellular dynamics are meticulously regulated by diguanylate cyclases (DGCs) and phosphodiesterases (PDEs), ensuring precise spatiotemporal control. The functional output of c-di-GMP signaling hinges on effector proteins—molecular decoders that translate c-di-GMP signals into specific cellular responses. This review systematically examines diverse classes of c-di-GMP effectors, using several representative bacterial species as model systems, to dissect their structural and mechanistic diversity. Particular emphasis is placed on their pivotal roles in bacterial pathogenicity, antibiotic tolerance, and host–pathogen interactions, offering fresh insights into the regulatory mechanisms underlying c-di-GMP signaling.

Cronobacter sakazakii is a food-borne pathogen that can thrive in various environments, including the human body. The human body’s physiological temperature exceeds that of the environment (22–30 °C), necessitating adaptations to heat stress during this transition. Managing heat stress is crucial when transitioning from the environment to the human body. In this study, we explored the effect of human body temperature on the growth of planktonic C. sakazakii, as well as its acid resistance, osmotic stress resistance, autoaggregation, and cell surface hydrophobicity. Our study demonstrated that human body temperature facilitated the growth, acid resistance, and osmotic resistance of C. sakazakii, compared to 28 °C. The relationship between human body temperature and phenotypes was studied by comparing gene expression at human and environmental temperatures (37 to 28 °C) using high-throughput sequencing. The results revealed up-regulation in the expression of 626 genes, including genes involved in arginine and proline metabolism, carbon fixation pathways, and nitrogen metabolism. Further analysis showed that human body temperature is essential for the environmental stress resistance of C. sakazakii. It boosts denitrification, betaine transport, and universal stress proteins, supporting membrane integrity and osmoprotectant transport. This study enhances our understanding of the strategies employed by C. sakazakii during its adaptation to the human body.

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.