Role of Tad pili during the transition from planktonic to biofilm state in <i>Bradyrhizobium diazoefficiens</i> USDA 110

Free-living soil bacteria can exist in two main states: planktonic, as motile single cells, or sessile, within biofilms. In biofilms, bacterial cells are embedded in an extracellular matrix that provides protection from environmental stresses and enhances long-term survival. The transition from planktonic to biofilm states sometimes involves surface sensing and attachment, processes commonly mediated by flagella and pili. In this study, we investigated the role of Type IVc Tad pili in surface sensing, adhesion, and biofilm formation in <i>Bradyrhizobium diazoefficiens</i>, a nitrogen-fixing symbiont of soybean. Bioinformatic analyses revealed that Tad pili are widely distributed and highly conserved within the <i>Bradyrhizobium</i> genus. While pili deletion in other model organisms typically reduces biofilm formation, we found that deletion of the most conserved genomic cluster encoding Tad pili in <i>B. diazoefficiens</i> led to increased adhesion to abiotic surfaces and impaired motility-indicative of a physiological shift toward a biofilm-associated state. These findings suggest that Tad pili may play a sensory or regulatory role, potentially influencing cell-cell or cell-matrix interactions. Furthermore, we identified a link between Tad pili and intracellular c-di-GMP levels. Together, these results highlight the critical role of Tad pili in the physiology of <i>B. diazoefficiens</i> and offer new insights into bacterial surface adaptation, with potential applications in agriculture and biotechnology. Understanding these mechanisms is essential for improving biofilm management strategies and developing new approaches to enhance bacterial survival in soil and inoculant formulations, ultimately optimizing legume symbiosis.IMPORTANCEBiofilm formation is essential for bacterial survival in soil environments. In this study, we investigated the role of Tad pili in the biofilm-forming capacity of <i>Bradyrhizobium diazoefficiens</i> and their connection to the second messenger c-di-GMP, a key regulator of the transition between planktonic and sessile states. Bacteria used in agricultural inoculants are typically in the planktonic state, yet survival and persistence are optimized in the sessile state. Our findings may contribute to the development of strategies that promote the transition to the biofilm lifestyle in inoculant formulations, thereby enhancing bacterial viability in storage and soil and improving symbiotic performance with host plants.