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Research Feature: Nitrogen cycling and microbial communities in forest soils
Stephanie Boyle completed her Ph.D. in Soil Science in March of 2007. For her dissertation, she used modern molecular techniques to study forest soil bacterial and fungal communities. Her primary advisor was Dave Myrold, professor of Soil Science, and her co-advisor was Peter Bottomley, professor of Microbiology and Soil Science.
In this short Web interview, Stephanie describes her research project.
What was your project about?
My project focused on understanding the contribution of bacteria and fungi in nitrogen cycling. I worked with soils beneath alder and Douglas fir dominated tree stands in two different forests – The Cascade Head Experimental Forest, a Coast Range forest north of Lincoln City, and the H.J. Andrews Experimental Forest, a Cascade Mountains forest near Blue River.
My goal was to understand more about the processes and biological drivers of nitrogen cycling by comparing microbial communities and nitrogen transformations at the different sites. For example, Cascade Head is extremely nitrogen rich and I wanted to know whether the microbial community there were different than in the forest soils at the H.J. Andrews where nitrogen is a limiting nutrient.
What were some of your research methods?
I looked at nitrogen transformations using stable isotope techniques and monitoring 15N. I used antibiotics to selectively block fungal or bacterial protein synthesis, while running isotope dilution experiments.
I also used DNA techniques to look at the composition of the microbial communities in the soils. I generated T-RFLP community fingerprints, clone libraries, and sequences. I also did some work with fatty acid extractions and quantitative PCR. These techniques allowed me to examine the size and composition of microbial populations of bacteria, fungi, archaea, and nitrifiers.
One of the fun things about this project was getting to work with the molecular techniques. I ended up making contacts with many other labs and was always impressed with the willingness of others to help and take time out to teach me about a particular research technique.
What were some of your most important findings?
I found that bacteria were the dominant contributors to nitrogen cycling in the Cascade Head soils but that fungi were the dominant contributors in the H.J. Andrews forest. I also found that there was a unique “community fingerprint” for the fungal, archaeal, and bacterial communities in the soils from the two forests. As for the role of tree type, I found that the fungal and archaeal communities were different beneath the alder and Douglas fir stands within a site, but that the bacterial community was not.
This project was some of the first sequencing of bacteria, fungi, and archaea from Cascade Head. That information will now be available to others through GenBank, a Web site for distributing genetic information.
My isotope dilution experiments also showed that nitrification was predominately carried out by prokaryotes. These nitrifiers include bacteria at both sites, but there may be some archaeal nitrification going on in Cascade Head soils. The Cascade Head soils are pretty acidic – they have a pH of 3 – and it raises questions about whether the presence of archaea in this environment is related to pH. Archaea are a kingdom of living things that have some of the characteristics of bacteria and some of the characteristics of Eukaryotes. They make up a relatively small proportion of the microbes in soils (5% compared with 95% bacteria) so they have not been studied very much in that environment – instead, they have mostly been studied in extreme environments. In recent years however, people have found that some archaea contain the gene to carry out ammonia oxidation and can play a role in nitrification and this has led to more studies of archaea in soils. This is interesting and an area for future research.
My work also suggests that dissolved organic nitrogen (DON) may be an important source of nitrogen for both fungi and bacteria. When antibiotics were added to soils from both sites, gross ammonification tended to increase, suggesting that organic N containing molecules continued to be broken down even when protein synthesis was blocked.
Working on a dissertation is a research project, but also a learning experience, what were some of the most valuable lessons for you?
An important lesson was how to write proposals and design experiments. I developed the idea for my project by writing an application for an EPA STAR fellowship during my second year in the ESB IGERT program. I didn’t end up receiving the fellowship, but I was able to pursue the project with funding from a different source. Later, I wrote a small grant to the Andrews LTER fund and received the money that allowed me to create clone libraries.
Two other important lessons were about learning how to take a piece of a project and make it my own and about persistence. I know this last one is a cliché, but it’s true. When you see my final project, it looks like everything that worked out, but there are always bumps in the road. I spent nine months working on a primer for a molecular analysis and in the end it didn’t work – there are always times like that and it takes persistence to keep going.
Now that you have completed your Ph.D., what are your future plans?
Since I finished in March, I’ve been working for Peter Bottomley as a research assistant. It’s giving me a chance to write up my dissertation research for publication and train another graduate student in some of the molecular work. I’m also job hunting – I really enjoy teaching and research and my ideal would be to be a professor at a smaller college.
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