Grains Research and Development

Date: 04.05.2015

Nitrogen genetics open a new efficiency frontier

Author: Dr Gio Braidotti

Key points

  • Around the world cereals, oilseeds and pulses that make better use of applied nitrogen fertiliser are under development
  • Australian research in nitrogen use efficiency (NUE) has drawn international interest and investment
  • Despite the trait’s complexity, especially in water-limited conditions, there are promising signs that genetics can deliver NUE gains 

Efforts are underway to use plant genetics to help growers retain nitrogen fertiliser’s yield and grain-quality benefits while reducing its impact on farm profits and the environment

Diagram of Nitrogen cycles

The nitrogen cycle

The world has a nitrogen budget that is essential to food and economic security, but that budget is just as skewed as many nations’ balance of trade and debt levels.

Update through the GRDC

A review of overall progress on developing nitrogen use efficiency (NUE) traits within Australia took place in 2014 when the GRDC brought together Australia’s leading NUE researchers for a workshop in Melbourne. The GRDC has also commissioned a report, Pre-breeding Opportunities for Enhanced NUE in Wheat, Barley, Legumes and Oilseeds, co-authored by Dr Trevor Garnett, Dr Darren Plett and Dr Mamoru Okamoto of the Australian Centre for Plant Functional Genomics and the University of Adelaide.

Since the Green Revolution of the 1960s, the planet has relied on nitrogen fertiliser to ensure crop yields keep pace with rapid bursts of population growth and economic development. This strategy, however, is incurring a cost to both farm profits – with nitrogen inputs now one of the largest single costs for many cereal growers – and to natural resources essential to food production more broadly.

Rather than maintaining this status quo of achieving short-term gains at long-term costs, researchers are attempting to better balance the world’s nitrogen budget in ways that benefit growers directly.

The idea looks simple on paper – endow all the major cereals, legumes and oilseeds with an enhanced potential to take up, assimilate, store or move nitrogen into grain thereby maximising yield and grain quality while reducing costs and waste.

Collectively, this multifaceted trait is called nitrogen use efficiency (NUE).

NUE is highly desirable to commercial breeding companies, both domestically and internationally, and has resulted in significant private-sector investment in Australian pre-breeding laboratories.

Among the companies working with Australian researchers are Arcadia Biosciences, Bayer CropScience, Pioneer DuPont and Vilmorin & Cie. The GRDC has also funded research in this area.

While a gene-driven approach that is unabashedly GM-centric dominates research efforts internationally, to date this approach has failed to deliver a commercially viable trait. In the meantime, Australian scientists are mounting a more calculated, paddock-based campaign. In fact, the strategy to crack NUE traits resembles Australia’s pioneering approach to water use efficiency (WUE) and there is a reason for this similarity.

The genetics underlying both NUE and WUE tend not to be hardwired into plants but rather cause the plant to respond to variation in environmental conditions. The result is a dynamic gene-by-environment effect that can confuse GM research strategies, especially if undertaken in pots that limit root growth.

Fast facts: Nitrogen’s modern-day inefficiencies

  • In 2011, global nitrogen fertiliser applications reached an annual total of about 100 million tonnes, with Australian agriculture using one million tonnes.
  • Wheat and other crops typically absorb less than half of applied nitrogen.
  • Unabsorbed nitrogen fertiliser can volatilise as nitrous oxide (N2O), a greenhouse gas that is 300 times more damaging to ozone depletion and climate change than carbon dioxide (CO2).
  • Agriculture is the second-largest industrial contributor to global greenhouse gases.
  • Oceanic dead zones result from the overgrowth of algae blooms fed by excessive nutrient runoff. The decomposition of the blooms depletes oxygen from the water. More than 146 dead zones around the world were identified in a 2004 United Nations Environment Programme report, the largest exceeding 70,000 square kilometres.

So while Australia is pursuing some GM gene-driven projects, there is a broader awareness among researchers – particularly at the University of Western Australia (UWA), the University of Adelaide (UA), the Australian Centre for Plant Functional Genomics (ACPFG) and CSIRO Plant Industry – that due consideration is needed of environmental realities and management systems within growers’ paddocks.

Of particular interest is the way a variety’s NUE status changes with root diseases, soil constraints such as salinity, and as water becomes scarce with dry finishes.

“With NUE, it is not just about genetics,” says CSIRO’s Dr Greg Rebetzke, a researcher with a strong track record running field trials that helped crack complex traits such as WUE and tolerance to dryland salinity in cereals.

“How genes interact with the soil, agronomy and nitrogen-management regimes is also really important. Growers are developing tools to predict the nitrogen requirements of their paddocks so we thought long and hard about the germplasm and genotypes needed to further enhance these efficiencies,” Dr Rebetzke says.

In giving growing conditions due consideration, local pre-breeders have acquired a broader – and in many ways deeper and more sophisticated – set of research priorities as they avoid committing exclusively to a GM approach.

That means as progress stalls worldwide, Australia’s strategy is making impressive headway. The main finding is that vast diversity in NUE capabilities exist within the gene pool of Australian cereals, particularly wheat, that appear exploitable under real growing conditions.

Pre-breeders are now focused on developing the tools to make use of this NUE biodiversity. Research underway includes devising cheap and easy measurements of NUE characteristics, including DNA markers.

Paddock realities

Of the NUE field trials recently completed, two projects were public–private ventures and the third was a GRDC project headed by Winthrop Professor Zed Rengel from UWA. He tested 1377 wheat and 551 barley lines at locations in WA, Victoria and New South Wales, but with an emphasis on WA and InterGrain varieties.

Image of Professor Zed Rengal

Professor Zed Rengel: many modern wheat cultivars have genetic potential for improved nitrogen use efficiency.

PHOTO: Evan Collis

“In general, many current commercial cultivars have good NUE, probably as a result of inadvertent selection during screening for yield under conditions that were frequently challenging regarding soils or climate,” Professor Rengel says. However, he notes that breeding companies are working with genotypes that differ widely for this trait, meaning continuous evaluation of that material is required to avoid perpetuating poor NUE in their germplasm.

Another project involving field trials, but with a national focus, was led by Dr Rebetzke in a partnership between CSIRO Plant Industry and Bayer CropScience.

For four years, across the Australian wheatbelt, variation in nitrogen uptake and movement of nitrogen into grain was assessed in both water-limited and irrigated conditions and using optimal (120 per cent) and sub-optimal (80 per cent) nitrogen levels. Management practices were used that sought to achieve Australian Hard grain quality standards (greater than 11.5 per cent protein).

“The really important thing for us was emulating growers’ management of nitrogen,” Dr Rebetzke says. “Our idea was to do a baseline assessment of commercial wheat germplasm that is not just about genetics but looks at the interaction between genetics and management. So in the trials, nitrogen was delivered in the same way as growers would and at around the same crop nitrogen requirements.”

Researchers took into account the trial site’s paddock history and rotation, measured starting nitrogen in the soil across the field and soil moisture, measured how much nitrogen was mineralised through the season, and controlled for factors that affect nitrogen capacity independently of genetics, such as root diseases and salinity.

Image of Dr Greg Rebetzke

Dr Greg Rebetzke, CSIRO Plant Industry.

Dr Rebetzke found there are outstanding genotypes in the Australian commercial wheat gene pool to increase the capacity for nitrogen uptake and remobilisation to the grain in water-limited finishes typical of the Australian wheatbelt.

“We also saw a huge amount of variation,” he says. “There are lines out there that take up a lot of nitrogen and lines that take up very little nitrogen. And there are lines that make more nitrogen available for grain filling.

“That’s under favourable conditions. As soon as you constrain water or temperature, they all do different things in terms of uptake and remobilisation of nitrogen.

”Having built up a picture of the characteristics that contribute to NUE under realistic growing conditions, Dr Rebetzke is now keen to pinpoint the sites within the wheat genome where genetic variation has the most beneficial impact for growers’ yields.

“We are thinking a lot about what influences growth and what we want to achieve in terms of genetics that we could sensibly put into a grower’s paddock,” Dr Rebetzke says. “With NUE, as with the water-efficiency traits CSIRO developed, we design the science with a view to trait delivery.”

New partnerships

NUE is also a trait subject to a suite of research projects at the ACPFG and UA overseen by Associate Professor Sigrid Heuer.

The NUE genes

Many genes thought to play a pivotal role in nitrogen metabolism have been trialled using a GM approach in which the gene is inserted and over-expressed in various crop varieties.

None have shown promise, with the exception of the barley alanine aminotransferase (AlaAT) gene, which has nitrogen use efficiency (NUE) properties discovered accidentally during drought tolerance research in Canada by Dr Allen Good of the University of Alberta.

However, the NUE effect is only observed if the transgene’s expression is restricted to the roots.

Over-express the gene throughout the plant – as typically occurs in most GM research – and the NUE effect disappears.

AlaAT is under development in the US by Arcadia Biosciences for use in wheat and many other crops. In Australian, AlaAT wheat and barley varieties have been under development since 2007 in a partnership between Arcadia Biosciences, the Australian Centre for Plant Functional Genomics and CSIRO Plant Industry.

In 2012, CSIRO obtained a regulatory licence to trial 17 wheat lines and 10 barley lines genetically modified to express AlaAT.

Included were field trials in South Australia that detected lines that perform well under low nitrogen. These included Australian wheat cultivars and exotic germplasm from the International Maize and Wheat Improvement Center (CIMMYT).

Additionally, there is a longstanding partnership with Pioneer DuPont that involves manipulating nitrogen uptake and transport genes in maize, with expertise gained now being turned to wheat.

“Different transporters are needed for nitrogen uptake and storage, and for its transportation to leaves,” Associate Professor Heuer says.

“So far very little is known about those genes in wheat, and pioneering work on the wheat nitrogen genes is now ongoing at the ACPFG, UA and elsewhere.”

There are also two new collaborative Australian Research Council (ARC) Linkage grants: one headed by Dr Trevor Garnett looking at nitrogen uptake and the other led by Associate Professor Heuer focusing on preventing loss of nitrogen in drought-stressed plants.

“The ACPFG has now brought together three Australian universities and all major Australian wheat breeding companies to study nitrogen remobilisation during grain filling and nitrogen loading into grains – in the lab and in the field,” Associate Professor Heuer says.

“The GRDC is a partner in this ARC-funded Industrial Transformation Research Hub project launched in early 2015. The research on nitrogen will be tightly linked with other activities within the ‘wheat hub’, such as development of wheat with combined heat and drought tolerance, which are essential traits for yield improvement.”

Dr Garnett will undertake controlled-environment experiments at the Plant Accelerator including establishing methods to more easily measure or ‘phenotype’ NUE in cereals. This project is a collaboration with Pioneer DuPont and Australian Grain Technologies (AGT) as the commercial partners, and UWA and the University of Melbourne as the investigative partners.

The robotised glasshouse at the Plant Accelerator allows Dr Garnett to non-destructively measure biomass by just taking images on a conveyor belt. The Smart House there can automatically apply water and nitrogen stress to varying degrees.

“We are doing experiments looking at both drought and nitrogen,” Dr Garnett says. “We then sit down with AGT breeders and work out how our screens relate to the results they have from their extensive field trials. In three years we want to have a method that is the gold standard for phenotyping cereals for their NUE. Ideally that will let us find plants from which we can breed.”

Overall, the signs are promising that NUE traits can be developed, allowing nitrogen’s yield-enhancing value to be retained while diminishing its spiralling costs. The outcome for growers could be varieties that make better use of applied nitrogen. 

 

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