1. Diversity and Functional Variation of Denitrifying Bacterial Communities in the Cape Fear River Estuary Brian Shirey Marine Biology

2. Project Goals Provide insight into denitrifying bacterial community structure Examine relationships between bacteria and their environment Provide answers to questions

3. What’s The Big Deal About Bacteria? The good The bad And the ugly Salmonella typhimurium

4. Denitrifying Bacteria Anaerobic –Utilize nitrate during respiration Comprise over 50 different genera Ubiquitous

5. Ecological Importance Vital role in global nitrogen cycle

6. Ecological Importance cont. Contribution to global warming –Production of greenhouse gasses NO 3 - NO 2 - N 2 O NO N 2 Nitrogen oxides

7. Ecological Importance cont. Effective in bioremediation –Reduction of nitrate entering ground and surface waters

8. Global Significance Understanding the mechanisms of denitrification is critical to our understanding of the global nitrogen cycle Applied methodologies promote a healthy, balanced, captive ecosystem

9. Experimental Design Collect samples Perform bacterial diversity and phylogenetic analysis Perform community functionality analysis

10. Sediment Sampling Sediments collected from 5 sites along the cape fear river estuary

12. Measuring Bacterial Diversity DNA sequencing –Isolation of functional genes encoding for nitrate reductase Comparative analysis –Along environmental gradients T-RFLP analysis –Profile of community structure

13. Measuring Functional Diversity Incubation experiments –Designed to monitor denitrifying activity Control NaClNitrate Anaerobic Enrichment Cultures

14. Research Hypothesis Different components of a community will respond to different environmental inputs to maintain community functionality

15. Questions To Be Answered Do denitrifying bacteria show the capability to readily adapt to sudden changes in their environment? Is survivorship of denitrifying bacteria greatly reduced in sub optimal conditions? To what degree do environmental conditions determine community structure?

16. Schedule Phase 1 –Sequencing all DNA, setting up and initialize incubation experiment Phase 2 –Completing incubation experiment and performing T-RFLP analysis Phase 3 –Analyze data, write paper Jan. Feb. March April May June July Aug. Sep. Oct. Nov. Dec. Phase 1 Phase 2 Phase 3

17. Conclusion Denitrifying bacteria have a tremendous impact on the biosphere Our biosphere is fueled by an underlying microbial world The global community and the microbial community are eternally linked Bacterial community as a window into past, present, and future

18. QUESTIONS?