Quantifying diffusion in a biofilm of Streptococcus mutans.

In biofilms, diffusion may limit the chemical activity of nutrients, toxic compounds, and medicines. This study provides direct, noninvasive insight into the factors that will most effectively limit the transport of antibiotics and biocides in biofilms. Self-diffusion coefficients have been determined for a number of fluorescent probes in biofilms of Streptococcus mutans using fluorescence correlation spectroscopy. The effects of probe size and charge and the roles of biofilm pH, ionic strength, and heterogeneity were studied systematically. The relative diffusion coefficients (D in the biofilm divided by that in water) decreased with increasing probe size (3,000-molecular-weight [3K], 10K, 40K, 70K, and 2,000K dextrans). Studies using variably charged substrates (tetramethylrhodamine, Oregon Green, rhodamine B, and rhodamine 6G) showed that the self-diffusion coefficients decreased with an increasing negative charge of the fluorescent probes. No significant effect was observed for changes to the ionic strength (10⁻⁴ to 10⁻¹ M) or pH (4 to 9) of the biofilm. Biofilm heterogeneity was responsible for variations of ca. one order of magnitude in the diffusion coefficients.