Many but not all ammonia-oxidizing bacteria (AOB) produce urease (urea amidohydrolase, EC 3.5.1.5) and are capable of using urea for chemolithotrophic growth. We sequenced the urease operons from two AOB, the ␤-proteobacterium Nitrosospira sp. strain NpAV and the ␥-proteobacterium Nitrosococcus oceani. In both organisms, all seven urease genes were contiguous: the three structural urease genes ureABC were preceded and succeeded by the accessory genes ureD and ureEFG, respectively. Green fluorescent protein reporter gene fusions revealed that the ure genes were under control of a single operon promoter upstream of the ureD gene in Nitrosococcus oceani. Southern analyses revealed two copies of ureC in the Nitrosospira sp. strain NpAV genome, while a single copy of the ure operon was detected in the genome of Nitrosococcus oceani. The ureC gene encodes the alpha subunit protein containing the active site and conserved nickel binding ligands; these conserved regions were suitable primer targets for obtaining further ureC sequences from additional AOB. In order to develop molecular tools for detecting the ureolytic ecotype of AOB, ureC genes were sequenced from several ␤-proteobacterial AOB. Pairwise identity values ranged from 80 to 90% for the UreC peptides of AOB within a subdivision. UreC sequences deduced from AOB urease genes and available UreC sequences in the public databases were used to construct alignments and make phylogenetic inferences. The UreC proteins from ␤-proteobacterial AOB formed a distinct monophyletic group. Unexpectedly, the peptides from AOB did not group most closely with the UreC proteins from other ␤-proteobacteria. Instead, it appears that urease in ␤-proteobacterial autotrophic ammonia oxidizers is the product of divergent evolution in the common ancestor of ␥-and ␤-proteobacteria that was initiated before their divergence during speciation. Sequence motifs conserved for the proteobacteria and variable regions possibly discriminatory for ureC from ␤-proteobacterial AOB were identified for future use in environmental analysis of ureolytic AOB. These gene sequences are the first publicly available for ure genes from autotrophic AOB.We are interested in the metabolic diversity of ammoniaoxidizing bacteria (AOB) and the selective advantage that individual traits may afford these specialized bacteria. AOB obtain usable energy and reductant solely from ammonia and fix carbon autotrophically (5, 19). The abilities to hydrolyze urea as a source of ammonia and carbon dioxide and to use the products of ureolysis for modification of the pH in the vicinity of the cell (4) appear to be important ecologically selected traits provided by the urease enzyme. AOB isolated from acidic soils are generally ureolytic (6), and ureolytic AOB can grow at lower pH with urea as an ammonia source (4). Ureolytic AOB may also have an advantage in soils receiving animal wastes or urea fertilizers. Molecular tools for examining functional genes involved in N metabolism in AOB have previously been developed for the gen...

An agricultural soil was treated with dairy-waste compost, ammonium-sulfate fertilizer or no added nitrogen (control) and planted to silage corn for 6 years. The kinetics of nitrification were determined in laboratory-shaken slurry assays with a range of substrate concentrations (0-20 mM NH(4)(+)) over a 24-h period for soils from the three treatments. Determined concentrations of substrate and product were fit to Michaelis-Menten and Haldane models. For all the treatments, the Haldane model was a better fit, suggesting that significant nitrification inhibition may occur in soils under high ammonium conditions similar to those found immediately after fertilization or waste applications. The maximum rate of nitrification (V(max)) was significantly higher for the fertilized and compost-treated soils (1.74 and 1.50 mmol N kg(-1) soil day(-1)) vs. control soil (0.98 mmol kg(-1) soil day(-1)). The K(m) and K(i) values were not significantly different, with average values of 0.02 and 27 mM NH(4)(+), respectively. Our results suggest that both N sources increased nitrifier community size, but did not shift the nitrifier community structure in ways that influenced enzyme affinity or sensitivity to ammonium. The K(m) values are comparable to those determined directly in other soils, but are substantially lower than those from most pure cultures of ammonia-oxidizing bacteria.