Vibrio parahaemolyticus, the leading cause of seafood-associated gastroenteritis, often exists in a biofilm state. Biofilm formation in this bacterium is regulated by a complex network of factors, including the second messenger c-di-GMP. CalR, a LysR-type transcriptional regulator, has been demonstrated to play a role in regulating the expression of virulence genes and swarming motility of V. parahaemolyticus. In this study, we demonstrate that CalR positively regulates biofilm formation by enhancing the production of exopolysaccharides (EPSs), extracellular proteins, extracellular DNA (eDNA), and c-di-GMP. RNA sequencing reveals that CalR regulates the transcription of 167 genes, including those potentially involved in c-di-GMP metabolism (e.g., vpa0184, vpa0198, and vpa1429) and biofilm-associated regulators (e.g., scrP). CalR directly suppresses the transcription of scrP, vpa0184, and vpa0198 while activating vpa1429 transcription in a direct manner. Additionally, CalR directly activates the transcription of cpsA but indirectly activates that of scvE, both of which are involved in EPS production. In summary, these findings elucidate the regulatory mechanisms by which CalR promotes biofilm formation in V. parahaemolyticus and highlight its role in modulating c-di-GMP homeostasis and biofilm matrix production. This work enhances our understanding of the regulatory network governing biofilm formation by V. parahaemolyticus.IMPORTANCEBiofilm formation is a critical survival strategy for V. parahaemolyticus, allowing it to persist in adverse environments. Understanding the regulatory mechanisms underlying biofilm formation is essential for developing strategies to control infections caused by this pathogen. This work demonstrates that CalR positively regulates the production of EPS, extracellular proteins, and eDNA, as well as biofilm formation by V. parahaemolyticus. CalR regulates the global gene expression in V. parahaemolyticus, including genes such as scrP, vpa0184, vpa0198, and vpa1429. ScrP negatively impacts V. parahaemolyticus biofilm formation, whereas vpa0184, vpa0198, and vpa1429 contribute to c-di-GMP metabolism. CalR represses scrP, vpa0184, and vpa0198 while activating vpa1429 and also promotes the production of c-di-GMP. These findings indicate how CalR positively regulates biofilm formation by V. parahaemolyticus.
Chang et al. (Thu,) studied this question.