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Tiedje Lab - Projects

DENITRIFICATION AND NITROGEN CYCLING

Oceanic N Cycling

Oceanic primary production is widely thought to be limited by the availability of fixed nitrogen (N). The local and global extent to which N removal processes occur can therefore affect both ecosystem function and global biogeochemical cycles (2). The current marine N budget appears to be unbalanced with a seemingly greater removal of N through denitrification than N inputs through in situ N fixation, precipitation, and riverine sources (1, 7). A net loss of N by the world's oceans could result in reduced photosynthesis and concurrent oceanic carbon dioxide (CO2) fixation thereby further elevating atmospheric CO2 (1). Although rates of sedimentary denitrification vary widely (3), which can serve to explain the unbalanced budget, it is important to characterize the processes and related organisms that may contribute to net N loss.

The Role of Anammox in Marine Sediments: Distribution, Abundance, and Activity (Ryan Penton)

In 1994, a novel N removal process was discovered in which the disappearance of nitrate in a fluidized bed reactor treating effluent from a methanogenic reactor with the concurrent consumption of ammonium and N2 formation was observed. This process was termed anammox (http://www.anammox.com) for anaerobic ammonium oxidation (10). The anammox process produces twice the amount of N2 as denitrification per molecule of nitrite consumed (24) and is thus a potentially important global oceanic N sink. The anammox process is now known to be an autotrophic, CO2-fixing process with a direct anaerobic biological conversion of ammonium and nitrite to dinitrogen gas, thereby consuming 100% less reducing agent (5).

Anammox has been proposed to be responsible for the consumption of more than 40% of the fixed N that sinks to the anoxic water in the Black Sea where FISH anammox cell densities of ~1,900 800 cells ml-1 were reported (6). In the anoxic waters of Golfo Dulce, a coastal bay in Costa Rica, anammox was found responsible for 19 to 35% of the total N2 formation in the water column based on 15N tracer studies (2). Additionally, anaerobic ammonium oxidation accounted for between 24 and 67% of total N2 production in continental shelf sediments from Baltic-North Sea transition zones (9). Furthermore, one-half to one-third of global N removal is believed to occur in the oxygen depleted zones of the world's oceans (4) and thus the anammox reaction is expected to be a globally important oceanic N sink. By understanding this novel piece in the N puzzle, the distribution, abundance, and contribution to N removal by 'anammox' bacteria can serve to further enhance current oceanic N models.

The Anammox Project

Marine sediments were collected from deep sea, shelf, and shallow sites in the Pacific Northwest and Arctic Ocean. Molecular and biochemical techniques are being utilized for the identification of anammox bacteria in marine sediments. Novel 16S rRNA gene based PCR primers were developed to identify the presence of Anammox bacteria with a greater degree of accuracy than previously attained while allowing flexibility for the capture of novel sequences in constructed libraries (8). Due to mismatches to "universal" primers, it is thought that the use of these primers, such as 27F, 1392R, etc. in previous biodiversity studies have resulted in PCR bias against anammox members. Thus, the development of these primers may show that anammox bacteria are ubiquitous in nature and that the breadth of the anammox group may be much greater than previously thought. Real-time PCR is being used to quantify these bacteria and relate their numbers to biogeochemical data. Fluorescence In-Situ Hybridization (FISH) is also being used as a method to identify specific anammox groups and to determine spatial relationships with other bacteria. Labeled 15N studies will be carried out by Allan Devol (http://www.ocean.washington.edu/2004/academics/options/chemical/faculty/DevolA/DevolA.html) at the University of Washington to confirm the in-situ activity of these organisms.

References:

  1. Codispoti, L.A. 1995. Is the ocean losing nitrate? Nature. 376:724.
  2. Dalsgaard T., D.E. Canfield, J. Petersen, B. Thamdrup, J. Acuna-Gonzalez. 2003. N2 production by the anammox reaction in the anoxic water column of Golfo Dulce, Costa Rica. Nature 422:606-608.
  3. Devol A.H. 1991. Direct measurement of nitrogen gas fluxes from continental shelf sediments. Nature. 349:319-321.
  4. Gruber N., J.L. Sarmiento. 1997. Global patterns of marine nitrogen fixation and denitrification, Global Biogeochemical Cycles 11:235-266.
  5. Kuai L., W. Verstraete. 1998. Ammonium removal by the oxygen-limited autotrophic nitrification-denitrification system. Appl. Env. Microb. 64(11):4500-4506.
  6. Kuypers M.M.M., A.O. Sliekers, G. Lavik, M. Schmid, B.B. Jergensen, J.G. Kuenen, J.S.S. Damste, M. Strous, M.S.M. Jetten. 2003. Anaerobic ammonium oxidation by anammox bacteria in the Black Sea. Nature 422:608-611.
  7. Middleburg J.J., K. Soetaert, P.M.J. Herman, C.H.R Heip. 1996. Denitrification in marine sediments: a model study. Global Biogeochem. Cycles 10:661-673.
  8. Penton, C. Ryan, A.H. Devol and J.M. Tiedje. 2006. Molecular evidence for the broad distribution of anammox in freshwater and marine sediments. Appl. Environ. Microbiol.
  9. Thamdrup B., T. Dalsgaard. 2002. Production of N2 through anaerobic ammonium oxidation coupled to nitrate reduction in marine sediments. Appl. Env. Microb. 68(3):1312-1318.
  10. Van de Graaf, A.A., A. Mulder, P. de Bruijn, M.S.M. Jetten, L.A. Robertson, J.G. Kuenen. 1995. Anaerobic oxidation of ammonium is a biologically mediated process. Appl. Env. Microb. 61(4):1246-1251.

 

Shewanella project

Visit the Shewanella oneidensis MR-1 page

 

Stable isotope probing for biostimulated metal, uranium and nitrate reducing bacteria (Mary Beth Leigh and Erick Cardenas, with Terence Marsh (http://www.mmg.msu.edu/faculty/marsh.htm) and Nathaniel Ostrom (http://www.msu.edu/user/ostrom/ostrom%20lab/index.htm)

See summary under Bioremediation

 

Assessment of bacterial diversity in soils under different land use systems in Amazon (Ederson da Conceicao Jesus)

The Amazonian forest is known for its large biological diversity and recent studies indicate that a large soil bacterial diversity is also present in Amazonian soils (Moreira et al., 1993; Borneman and Triplett, 1997). Agriculture is also a reality in Amazon and its effects on soil organism communities are not well understood yet. Anthropic activities such as agriculture have a great potential to change the soil environment and consequently affect microbial communities which are responsible for several environment functions and consequently influence the ecosystem health. Comprehending how diversity is changed by soil utilization is important to delineate management practices that help to preserve microbial diversity and achieve benefit from it.

The current work is part of the Conservation and Sustainable Management of Below Ground Biodiversity (BGBD) project implemented by the United Nations Program which has been carried out in seven countries including Brazil. Several groups of soil organisms have been studied including soil fauna, mycorrhizal fungi, bacterial communities and pathogenic fungi as well as soil processes carried out by these organisms. Our objective is to study the diversity of overall bacterial communities as well as the diversity of bacterial communities involved in the N cycle in Amazon soils and to assess the effect of land utilization over these communities. We chose to focus on the nitrogen cycle because of its importance for agricultural systems and to complement the study of microbial processes including N2O emissions which will be made by other research group participating in the project.

In order to achieve our objective we are exploring genes as the 16S rDNA, amoA, nirK, nirS and nifH with culture-independent techniques, such as the T-RFLP, and constructing some clone libraries to have a finer view of the bacterial communities. The AmoA, nifH and nir genes code for key enzymes in the nitrogen cycle and have been used as genetic markers to study bacteria communities (Brakere et al., 1998; Avrahami et al. 2002; Tan et al., 2003). Diversity indices and recent techniques for the study of microbial communities will be applied in order to evaluate the ecology of these communities.

References:

Avrahami, S, Conrad, R., and G. Braker. 2002. Effect of soil ammonium concentration on n2o release and on the community structure of ammonia oxidizers and denitrifiers. Applied and Environmental Microbiology, 68:5685-5692.

Borneman, J.; Triplett, E. W. 1997. Molecular microbial diversity in soils from Eastern Amazonia: Evidence for unusual microorganisms and microbial population shifts associated with deforestation. Applied and Environmental Microbiology 63:2647-2653.

Braker, G., Fesefeldt, A. and Witzel, K. P. 1998. Development of PCR Primer Systems for Amplification of Nitrite Reductase Genes ( nirK and nirS ) To Detect Denitrifying Bacteria in Environmental Samples. Environmental Microbiology 64:3769-3775.

Moreira, F. M. S.; Gillis, M.; Pot, B.; Kerskers, K.; Franco, A. A. 1993. Characterization of rhizobia isolated from different divergence groups of tropical leguminosae by comparative polyacrylamide gel eletrophoresis of their total proteins. Systematic and Applied Microbiology 17:135-146.

Tan, Z., Hurek, T., Reinhold-Hurek, B. 2003. Effect of N-fertilization, plant genotype and environmental conditions on nifH gene pools in roots of rice. Applied and Environmental Microbiology 5:1009-1015.

 

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