Nitrogen Fixation Associated with Grasses Assessment of Opportunity

Summary

A review of the literature was requested by MLA to provide an opportunity for nitrogen fixing grasses in Australian production systems. The objectives were to: 

Describe the need or opportunity for non-symbiotic grasses in southern Australian pastures 

Review the literature to determine the nature and prospects for investing in non-symbiotic N fixation research relevant to grasses. 

Review a new project application based on scoping the potential for non-symbiotic N fixation in southern Australian perennial grasses and assess the likelihood of success of this project. 

Identify research areas required to develop the opportunity for non-symbiotic N fixation in association with grasses. 

A number of key Australian and international scientists with strong experience in non-symbiotic N fixation with crops and grasses were consulted and literature gathered which was reviewed in the context of the above objectives. In essence the opportunity for non-symbiotic N fixation are the perceived benefits for reducing reliance of pasture systems on fertiliser N inputs for dividends in reduced costs and environmental impacts through reduced green house gas emissions and leaching of N through the soil profile. An additional potential benefit may be in the replacement of pasture legumes with grasses which are more phosphorous efficient and so reduced fertiliser P costs may result. The process of biological N fixation as being mediated by the nitrogenase enzyme in bacteria and a range of non-symbiotic N fixation opportunities defined: Free-living N fixing bacteria that live in loose association with the rhizosphere of plant roots and in the surrounding soil in micro sites conducive to their function. 
Rhizo-diazotrophs (N fixing bacteria) which attach to or colonise plant roots and have a closer association with the plant then the free-living organisms. 
Endophytic diazotrophs which live inside the plant within cell structures in roots, stems and leaves A review of the literature revealed the following (described briefly): 
A diverse range of diazotrophs can exist in association with grasses and be shown to fix N. The N fixation benefit is complicated by other plant growth promoting effects they may have. 
They function best under conditions of high carbon, plentiful moisture, warm temperatures and low soil N. 
While there are examples of experiments under controlled conditions where agronomically significant amounts of N have been shown to be fixed by diazotrophs in association with plants, few of these have been translated reliably field situations. 
Most examples are based on tropical crop (eg sugarcane) and grass mono-cultures in systems with high carbon inputs and warm moist conditions. It is expected that in southern Australian conditions with low soil C and variability in soil type, temperature, pasture composition and management would make it unlikely for the benefits to be reliably captured. 
Diazotroph abundance and function are affected by many factors including oxygen (aneorobic conditions are required to preserve nitrogenase activity), carbon, temperature, moisture, plant genotype, soil characteristics (eg structure and pH), level of soil N (best at low soil N), other nutrients, soil disturbance etc 

The consensus of the literature and the scientists consulted was that there is little prospect for effective and significant non-symbiotic N fixation in association with grasses in the Australian environment, particularly with free-living diazotrophs. 
The case for free-living N fixing bacteria is least compelling as they have a high energy requirements, must compete with the diverse range of soil micro-organisms for root exudates and other energy sources, survive in sufficient numbers and remain active at N fixation to supply significant amounts of plant N demand. This seems remote. 
There may be more success through a focus on endophyte diazotrophs because thay have access to more protection and plant carbon and moisture. The methodologies to assess N fixation by diazotrophs is questioned for accuracy and sensitivity to measure small amounts fixed against significant soil N background and the likely contamination from rain and dust. However, modern molecular techniques using microarrays show promise. Claims for evidence of quantified non-symbiotic N fixation have been challenged in a number of examples on the basis of the methods used and the interpretation and extrapolation of data. An assessment of the submitted project proposal submitted to MLA to use molecular techniques to identify diazotroph diversity and abundance in rhizosphere soils associated with perennial grasses at prescribed research sites was review and it was concluded that:The likelihood of success of the project as described was high. 
The outcome would be to determine what diazotrophs were present and if they could fix N. It would not reveal the potential quantity of N fixed under field conditions nor the rates of N fixation at different times. 

On the basis the conclusions of the review and the scepticism among many researchers that free-living diazotrophs have potential to reliably and consistently supply fixed N to plants, it was questioned as to whether the work should be undertaken at all as the prospects, under Australian conditions, of translating knowledge on the diversity and presence of diazotrophs in the rhizosphere to translate to practical on-farm solutions is considered remote. 
However closer scrutiny of opportunities through N fixing endophytes was suggested. Some future research areas were suggested which included: Screen prospective perennial pasture plants for effective N fixing endophytes 
Investigate the characteristics of grass plants that survive and produce well in low N environments to identify the key features that favour non-symbiotic N fixation and evaluate their application to breeding objectives. 
Support basic science to better understand how N fixing endophytes and grass plants co-exist and optimise N fixation 
If endophytes are pursued, ways to utilize non-culturable forms need to be developed and they need to be assessed for any negative effects on animal health. 
Investigations into the potential of plant growth promoting effects which are complementary to the n fixing functions could be undertaken for additive effects. 
Methods for accurately and reliably assessing non-symbiotic N fixation at low levels under field conditions over time and space are required.