Date of Award:

5-1-2005

Document Type:

Thesis

Degree Name:

Master of Science (MS)

Department:

Biology

Department name when degree awarded

Life Sciences: Biology

Committee Chair(s)

John M. Stark

Committee

John M. Stark

Committee

Jeanette M. Norton

Committee

Bradley R. Kropp

Abstract

Ammonia-oxidizing bacteria carry out the first step of nitrification, producing nitrite that is subsequently converted to nitrate, which may contribute to environmental pollution. The objective of this study was to gain insight into the functional diversity of ammonia-oxidizing bacteria in agricultural soils. We investigated the urease encoding gene sequences of several nitrifying bacteria and the kinetics of nitrification of soils treated with different nitrogen sources. Many, but not all, ammonia-oxidizing bacteria produce urease and are capable of hydrolyzing urea for their source of ammonium. The urease operons were sequenced from the betaproteobacterial Nitrosospira sp. NpAV and the gammaproteobacterial Nitrosococcus oceani. In both organisms, all seven ure genes were contiguous: ureDABCEFG. Southern analyses revealed two copies of ureC in Nitrosospira sp. NpAV and one copy in the N. oceani genome. Several regions were suitable primer targets for obtaining further ureC sequences from additional ammonia-oxidizing bacteria. The deduced peptide sequences were used to construct alignments and make phylogenetic inferences. UreC proteins from betaproteobacterial ammonia oxidizers formed a distinct monophyletic group. This phylogeny suggests that urease in betaproteobacterial ammonia-oxidizers is the product of divergent evolution in the common ancestor of gamma- and betaproteobacteria that was initiated before their divergence. The ure gene sequences revealed in this study are the first publicly available from autotrophic ammonia-oxidizing bacteria. In soil slurry laboratory experiments, the kinetics and NH4+ inhibition of nitrification were determined for soils treated with (NH4)2SO4 and dairy compost over six years. The rate of nitrification was determined for a range of NH4+ substrate concentrations (0-20 mM NH4+). The rate data were fit to the Michaelis-Menten and Haldane models. The Haldane model, which included the effect of substrate inhibition at high concentrations, better represented the observed kinetic relationship, indicating that these soils exhibited NH4+ inhibition. Vmax was the only parameter that was significantly different and was lower for control soils than in soils that had received either (NH4)2SO4 or compost. This research suggests that the Haldane model may be more applicable than the Michaelis-Menten model for representing the kinetics of ammonia-oxidizing bacteria in agricultural soils.

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Biology Commons

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