The metabolic processes supported in a biofilm are often determined by a biofilm’s mesoscale structure (10-4-10-2 meter scale), and mesoscale structure is in turn shaped by the interplay between microbial metabolism and detachment. Despite the widespread adoption of biofilm reactors for water pollution control, much is unknown regarding the relationship between mesoscale biofilm structure and process performance. Optical coherence tomography (OCT) has recently emerged as a promising technique to non-destructively capture mesoscale three-dimensional images of environmentally relevant biofilms. For this study, OCT was used to image the three dimensional structure of mixed culture biofilms enriched in a synthetic wastewater. Distinct morphological features such as pores and microcolonies were detectable as deep as 1mm within the biofilm matrix. Quantitative structural parameter values (e.g. porosity, thickness, diffusion distance) were consistent across multiple fields of view and compared favorably with images acquired by confocal laser scanning microscopy.