Inhibition, Gene Expression and Modeling of Ammonia Oxidation in Biofilms of Nitrosomonas europaea

This dissertation explores the physiology and gene expression of the ammonia-oxidizing bacterium Nitrosomonas europaea in surface-associated bacterial communities, or biofilms. Biofilms of N. europaea were cultivated in drip flow reactors for several weeks and gene expression microarrays were used to detect 240 genes differentially expressed between the mature biofilms and exponential batch cells. Using RT-qPCR, genes up-regulated in the biofilm microarray were detected in continuous cultures of suspended cells, and were increasingly up-regulated with decreasing dilution rates. The observations suggest a correlation between the biofilm-related gene expression and slow growth rates of the cells.;N. europaea cells in the biofilms upon exposure to the aromatic hydrocarbons phenol and toluene were more resistant to inhibition of ammonia oxidation than suspended cells. 50% inhibition was observed upon exposure of mature biofilms to 60 microM phenol and 100 microM toluene, compared to 10 microM phenol and 20 microM toluene with exponential batch (suspended) cells. However, the transcriptional response to the hydrocarbons was similar between suspended cells and biofilms, with two genes up-regulated in both growth states in response to phenol (NE1545-NE1546) while no transcriptional response was observed during toluene exposure.;To further explore the response to phenol exposure, cells were cultivated in continuous reactors at various growth rates and NH3 oxidation rates and exposed to phenol. The inhibition of ammonia oxidation by 20 microM phenol decreased with slower growth rates and NH3 oxidation rates and approached the inhibition level of the biofilms. Increasing the dissolved oxygen (DO) concentration in the biofilms resulted in higher NH3 oxidation rates and greater phenol inhibition, leading to the conclusion that the tolerance of biofilms is likely related to O2 limitation causing slow NH3 oxidation rates and slower growth rates.;DO and pH were measured in the biofilms with microsensors and the concentration profiles were simulated with 2-D reactive transport that combined advective and diffusive transport. The half-saturation coefficient for suspended cells fit the biofilm profiles, indicating that the kinetics of NH3 oxidation between the biofilm and suspended cells were the same. The model was also able to simultaneously predict the observed pH and DO biofilm profiles for tests conducted with limited buffering capacity, where pH was lowered by the cells, which resulted in NH3 limitations and reduced O2 consumption.