Advancements have been
made in elucidating the composition of ammonium oxidizing bacteria (AOB)
communities in soil, which produce NO2-/NO3-.,
by taking advantage of variations in gene sequences of 16S ribosomal DNA (rDNA)
or the catalytic subunit of NH3 monooxygenase gene (amoA)( Mintie et al. 2003). Analysis of these functional genes provides a
vigorous, culture-independent method of examining ammonia-oxidizing bacterial
diversity and community composition in agroecosystems (Yeager et al. 2005). Stable-isotope
probing is a method used in microbial ecology that provides a means by which
specific functional groups of organisms that incorporate particular substrates
are identified without cultivation. Stable-isotope-labeled nitrogen (15N)
sources are essentially assimilated into the microbial biomass of environmental
samples (Neufeld, 2007). Molecular analysis will reveal phylogenetic and
functional information about the ammonia-oxidizing prokaryotes in agricultural systems here in the United States (Huntsville,
AL) and in China (Nanjing). In addition, bacterial genome diversity has
been found to be significantly higher with pyrosequencing analysis as compared
to the usual culture-based analysis (Price et al. 2009) and microarray
technology (Gilbert et al. 2008).
These
techniques have limitations when exploring the diversity of natural
environments. Contributions of most
organisms are missed through cloning because only a select number of clones are
chosen for analysis, and microarray technology only gives information about
those sequences for which it was designed. This proposed research involving
pyrosequencing methods will provide substantial information of both microbial
species, such as Nitrosospira and genes responsible for ammonium
oxidation and thus increase the knowledge of the diversity and distribution of
that group of microorganisms. We
hypothesize that ammonia-oxidizers play an important role in N2
cycling in soil-water systems, and ammonia monooxygenase (amoA) genes may not only be phylogenetically diverse, but also
geographically widespread. The REU students selected should have a strong
interest in environmental microbiology. They will become knowledgeable of the
role the nitrogen cycle plays in the environment, more specifically ammonium
oxidizers. The students will also become
skilled in basic molecular techniques such as DNA extraction, polymerase chain
reaction (PCR), and more advanced high-throughput techniques such as
pyrosequencing.