The effect of c-di-GMP (3′–5′-cyclic diguanylic acid) on the biofilm formation and adherence of Streptococcus mutans

SummaryDepending on a biofilm lifestyle, Streptococcus mutans (S. mutans) is thought to be one of the primary causative agents of dental caries. Biofilm formation and adhesion are crucial physiological functions and virulence factors for S. mutans. Thus, attempts to control the development of dental caries only by inhibiting one of the several virulence factors are not effective. Cyclic diguanylate (c-di-GMP) [bis(3′,5′)-cyclic diguanylic acid] is a prokaryotic cyclic dinucleotide second messenger that has been implicated in determining the timing and amplitude of complex biological processes from biofilm formation and virulence to photosynthesis. Here, we demonstrate that this signaling molecule also plays a role in the ability of S. mutans to initiate biofilm formation and adhere to tooth surfaces. To test this hypothesis, S. mutans UA159 and its gcp gene knockout mutant were assayed for their ability to initiate biofilm formation and adherence. The spatial distribution and architecture of the biofilms were examined by scanning electron microscopy. These results show that inactivation of the gcp gene resulted in the formation of an abnormal biofilm. We confirmed that c-di-GMP was effective in preventing biofilm formation of S. mutans UA159. We also found that extracellular c-di-GMP inhibited the adherence of S. mutans to tooth surfaces and reduced (>50%) biofilm formation compared to the untreated control. These results indicate that c-di-GMP attenuates the caries-inducing virulence factors of S. mutans. This suggests that c-di-GMP may be used alone or in combination with other antimicrobial agents, and that such a treatment could be developed into a novel method to prevent tooth decay.