We describe the architecture of nitrifying bacteria biofilms immobilized on a three-dimensional (3D) polyurethane foam that permits efficient water flow through a bioreactor. The 3D spatial organization of immobilized bacterial colonies is characterized on three resolution levels with X-ray tomography, light confocal microscopy, and scanning electron microscopy (SEM). Using these techniques we demonstrate biofilm distribution in the foam and the existence of several modes of binding of bacteria to the foam. Computed X-ray tomography permits observation of the distribution of the biofilm in the whole open cellular polyurethane material volume and estimation of biofilm volume. SEM and confocal laser scanning microscopy techniques permit 3D visualization of biofilm structure. Three distinct immobilization patterns could be observed in the open cellular polyurethane material: (1) large irregular aggregates of bacterial biofilm that exist as irregular biofilm fragments, rope-like structures, or biofilm layers on the foam surface; (2) spherical (pom-pom) aggregates of bacteria localized on the external surface of biofilm; and (3) biofilm threads adherent to the surface of polyurethane foam. Finally, we demonstrate that immobilized bacteria exhibit metabolic activity and growth.

Ammonium and nitrite oxidizing bacteria were enriched in separate cultures using a primary inoculum of water from an enclosed shrimp rearing facility. Each enrichment was continued by transfer to increasing volumes of medium up to 151. Seventy four strains of heterotrophic bacteria were isolated; seventeen from each primary inoculum, and twenty from each of the 151 enrichments at the time when all the ammonium or nitrite had been utilized. All the isolates, described by 101 characters, were subjected to numerical taxonomy. During the enrichment for ammonium oxidizers, astrong selective pressure led to the disappearance of some strains, mostly enterobacteria. Two taxonomic groups composed of pseudomonads and asporogenous Gram positive rods were associated with the ammonium oxidizers. The population was characterized by an inability to use amino acids as carbon and energy sources. In contrast, the strains associated with nitrite oxidizers were able to use all the carbon and energy sources tested. Many of these strains were Gram positive, asporogenous aerobic rods, others were Gram negative pseudomonads.