GcrR in <i>Streptococcus mutans</i> Inhibits the Morphological Transition of <i>Candida albicans</i> Through Ras1–cAMP/PKA Pathway in the Cross‐Kingdom Biofilms

The dual-species biofilms formed by Streptococcus mutans and Candida albicans exhibit enhanced cariogenic potential due to the production of extracellular polysaccharides (EPSs). The response regulator GcrR in S. mutans negatively regulates EPS synthesis, but its impact on dual-species biofilms remains unclear. The objective of the present study was to investigate the effect of GcrR on the cariogenicity of cross-kingdom biofilms established by S. mutans and C. albicans. The crystal violet (CV) staining was conducted to evaluate the dual-species biofilm formation. The anthrone-sulfuric acid assay was employed to assess EPS formation. The biofilm morphology was visualized through scanning electron microscopy (SEM), and the structure was examined by confocal laser scanning microscopy (CLSM). The expression levels of genes were determined by quantitative real-time PCR (qRT-PCR). The animal assay was conducted to elucidate the anti-cariogenic effect of GcrR. The overexpression of GcrR suppressed dual-species biofilm formation, which was accompanied by a reduction in both water-soluble glucans (WSGs) and water-insoluble glucans (WIGs). The morphological transformation of C. albicans from yeast to hyphal form was inhibited by GcrR. Expression of gtfB/D and gbpB/C genes in S. mutans and Ras1-cAMP/PKA pathway genes in C. albicans was downregulated. Furthermore, we found that GcrR attenuated the virulence of dual-species biofilms. This study suggests that GcrR in S. mutans plays a critical role in modulating the cariogenic potential of bacterial-fungal biofilms.