In situ effective diffusion coefficient profiles in live biofilms using pulsed-field gradient nuclear magnetic resonance.

Diffusive mass transfer in biofilms is characterized by the effective diffusion coefficient. It is well documented that the effective diffusion coefficient can vary by location in a biofilm. The current literature is dominated by effective diffusion coefficient measurements for distinct cell clusters and stratified biofilms showing this spatial variation. Regardless of whether distinct cell clusters or surface-averaging methods are used, position-dependent measurements of the effective diffusion coefficient are currently: (1) invasive to the biofilm, (2) performed under unnatural conditions, (3) lethal to cells, and/or (4) spatially restricted to only certain regions of the biofilm. Invasive measurements can lead to inaccurate results and prohibit further (time-dependent) measurements which are important for the mathematical modeling of biofilms. In this study our goals were to: (1) measure the effective diffusion coefficient for water in live biofilms, (2) monitor how the effective diffusion coefficient changes over time under growth conditions, and (3) correlate the effective diffusion coefficient with depth in the biofilm. We measured in situ two-dimensional effective diffusion coefficient maps within Shewanella oneidensis MR-1 biofilms using pulsed-field gradient nuclear magnetic resonance methods, and used them to calculate surface-averaged relative effective diffusion coefficient (D(rs)) profiles. We found that (1) D(rs) decreased from the top of the biofilm to the bottom, (2) D(rs) profiles differed for biofilms of different ages, (3) D(rs) profiles changed over time and generally decreased with time, (4) all the biofilms showed very similar D(rs) profiles near the top of the biofilm, and (5) the D(rs) profile near the bottom of the biofilm was different for each biofilm. Practically, our results demonstrate that advanced biofilm models should use a variable effective diffusivity which changes with time and location in the biofilm.