Fluorescence imaging of the spatial distribution of ferric ions over biofilms formed by streptococcus mutans under microfluidic conditions

Streptococcus mutans, which is an initial dental plaque colonizer on the mucosal surface of oral cavities, utilize exogenous sucrose as a nutrient source to produce extracellular polysaccharides (EPS). EPS are highly sticky and allow bacterial cells to attach to the tooth surface. Bacteria, including S. mutans, acquire iron from the human body to survive. However, little is known about the mechanism of iron acquisition during the formation of S. mutans bio-films. In this study, we utilized a microfluidic device that generated the microfluidic condition often found in teeth and imaged the spatial distribution of ferric ions over S. mutans biofilms formed in the device using a ferric chemosensor. Our results showed that under iron-repleted conditions S. mutans produced a lower amount of EPS and thus formed smaller bio-films compared to under iron-depleted conditions, indicating that biofilm formation is not required for the survival of the microorganism in the presence of a sufficient amount of iron. Spatial imaging of the biofilms using a ferric chemosensor revealed that higher amounts of ferric ions accumulated in the inner EPS layer of the biofilm formed under both iron-depleted and iron-repleted conditions, suggesting that the EPS layer in the biofilm is responsible for acquiring and delivering ferric ions to the cell body.