Draft COST action on ecological aspects of denitrification

Brussels, 14 March 2002

European Cooperation in the field of Scientific and Technical Research - COST Secretariat. Draft Memorandum of Understanding for the implementation of a European Concerted Research Action designated as COST Action 856 "Ecological Aspects of Denitrification, with emphasis on Agriculture". Brussels, 11 March 2002 (document COST 222/02) Full text

The Signatories to this Memorandum of Understanding, declaring their common intention to participate in the concerted Action referred to above and described in the Technical Annex to the Memorandum, have reached the following understanding:

1. The Action will be carried out in accordance with the provisions of document COST 400/01 "Rules and Procedures for Implementing COST Actions", the contents of which the Signatories are fully aware of.

2. The main objective of the Action is to better understand factors governing the loss of N- fertilisers in agriculture due to the microbial activities.

3. The overall cost of the activities carried out under the Action has been estimated, on the basis of information available during the planning of the Action, at about 30 Million Euro in 2001 prices.

4. The Memorandum of Understanding will take effect on being signed by at least five Signatories.

5. The Memorandum of Understanding will remain in force for a period of five years, calculated from the date of the first meetingof the Management Committee, unless the duration of the Action is modified according to the provisions of Chapter 6 of the document referred to in Point 1 above.

A. BACKGROUND

Denitrification, one of the main branches of the global nitrogen cycle, is an energy-yielding process in which microorganisms utilise nitrate as terminal respiratory electron acceptor under oxygen limited conditions. The overall reaction sequence employs the following intermediates in a pathway where nitrate is reduced via nitrite to gaseous products:

Nitrate Nitrite Nitric oxide Nitrous oxide Dinitrogen

Denitrification is also called nitrate respiration or dissimilatory nitrate reduction, where these terms stress different physiological roles of the process. For many years it was believed to be performed exclusively by eubacteria. However, there are indications that some fungi (e.g. the pathogenic species Fusarium oxysporum) and archaea are also able to denitrify. Bacteria of many different systematic groups can perform denitrification. However, some microorganisms can reduce nitrate only to nitrite and others only to nitrous oxide. Several bacteria utilize nitrite or nitrous oxide but not nitrate. Overall, approximately 50% of all known bacteria can perform denitrification (or at least some of its partial reactions). The occurrence of denitrification in organisms of totally unrelated affiliations suggests that denitrification has been distributed evolutionary by lateral gene transfer.

The single steps of the denitrification process are catalysed by specific reductases. The first reaction, the conversion of nitrate to nitrite, is catalysed by a Mo-containing nitrate reductase. All nitrate reductases have a molybdopterin-cofactor and contain FeS centres, and, in addition, some possess cytochrome b or c. Several forms of nitrate reductases exist (membrane-bound, periplasmic and also an assimilatory enzyme with similar characteristics to the dissimilatory enzymes) making the study of this step interesting but also complex, particularly since some organisms contain more than one type of enzyme.

The next step in the denitrification pathway is the reduction of nitrite to nitric oxide catalysed by nitrite reductase. Bacteria employ two different forms of nitrite reductase, containing either cytochrome cd1- or Cu in their prosthetic group. Organisms possess either of the two enzymes. The cytochrome cd1 containing nitrite reductase appears to be more widespread among bacteria, whereas the Cu-enzyme apparently is more conserved evolutionary. The product of the reaction, NO, in addition to being toxic and highly reactive, is an important signal molecule for plant or animal life. Intact denitrifying organisms evolve at best minute amounts of this gas which is effectively utilized by the cytochrome b and c containing nitric oxide reductase. The conversion of NO to N2O catalysed by this enzyme involves the formation of the dinitrogen double (triple) bond which is a biochemically extremely interesting reaction that is poorly understood currently. Finally, nitrous oxide is reduced to the dinitrogen molecule by nitrous oxide reductase which contains Cu-atoms in a novel tetra-nuclear cluster at the active site.

By the action of denitrifying microorganisms, the global dinitrogen content in the atmosphere is largely in balance due to the formation of the dinitrogen gas from terrestrial nitrate. Nitrate is the main N­source for the growth of plants in agriculture but can simultaneously by used also by microorganisms in soils. Denitrification is generally regarded as an anaerobic process, but there are indications that it may take place also in well-aerated soils. The conditions favouring denitrification in soils have not yet been elucidated in much detail. It is, however, clear that any use of nitrate by bacteria means a loss of N for the growth of plants. Thus, denitrification has severe impact on agriculture, and the COST Action will particularly put emphasis on these agricultural aspects. In addition, products of denitrification (nitrate respiration) have manifold other, mainly adverse effects on soils but also on the atmosphere and on waters. The major concern is thus agriculture, but it does not make sense to leave out environmental impacts of denitrification in this COST Action. Such impacts can be summarized as follows:

In contrast to ammonia, which is tightly bound in soil, nitrate is easily washed out and transported to the groundwater where it (and its reduction product nitrite) adversely affects water quality. On the other hand, denitrifying bacteria, together with nitrifying microbes, play a crucial role in the two-step process of biological sewage treatment. Denitrification also contributes in a beneficial way to the removal of nitrogen from polluted, eutrophic lakes. In addition, nitrogenous oxides released from soils and waters have several impacts on the atmosphere. Nitrous oxide is next to CO2 and CH4 in its importance as a potent greenhouse gas. Nitric acid and its chemical oxidation product NO2 are major constituents of acid rain, and NO and also N2O interact with ozone in complex reactions and are major causes of the destruction of the protective ozone layer in the stratosphere.

Thus the impacts of products of denitrification on agriculture as well as on waters and the atmosphere are complex, but are of extreme relevance for human welfare. In particular, the increasing N2O content of the atmosphere is giving cause for serious environmental concern.

However, many aspects of the denitrification processes are poorly understood at present. This COST Action will significantly contribute to increase the knowledge in the field, particularly to understand complex interactions of the biological N-cycle by bringing together scientists from different countries and different areas of competence. There are no series of International Congresses on denitrification in contrast to other fields of the N-cycle (e.g. dinitrogen fixation, nitrate assimilation). Therefore this COST Action is necessary to concentrate European activities and to stimulate the progress in the field. It should be stressed that such goals will not be achieved without the instalment of this COST Action....

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