Biological nitrification inhibition (BNI) - is there potential for genetic interventions in the Triticeae

The natural ability of plants to release chemical substances from their roots that have a suppressing effect on nitrifier activity and soil nitrification, is termed ‘biological nitrification inhibition’ (BNI). Though nitrification is one of the critical processes in the nitrogen cycle, unrestricted and rapid nitrification in agricultural systems can result in major losses of nitrogen from the plant-soil system. This nitrogen loss is due to the leaching of nitrate out of the rooting zone and emission of gaseous oxides of nitrogen to the atmosphere, where it causes serious pollution problems. Using a newly developed assay system that quantifies the inhibitory activity of plant roots (i.e. BNI capacity), it has been shown that BNI capacity is widespread among crops and pastures. A tropical pasture grass, Brachiaria humidicola has been used as a model system to characterize BNI function, where it was shown that BNIs can provide sufficient inhibitory activity to suppress soil nitrification and nitrous oxide emissions. Given the wide-range of genetic diversity found among the Triticeae, and the current availability of genetic tools for moving traits/genes across members, there is great potential for introducing/improving the BNI capacity of economically important members of the Triticeae (i.e. wheat, barley and rye). This review outlines the current status of knowledge regarding the potential for genetic improvement in the BNI capacity of the Triticeae. Such approaches are critical to the development of the next-generation of crops and production systems where nitrification is biologically suppressed/regulated to reduce nitrogen leakage and protect the environment from nitrogen pollution

Developing Methods to Evaluate Phenotypic Variability in Biological Nitrification Inhibition (BNI) Capacity of \u3cem\u3eBrachiaria\u3c/em\u3e Grasses

As part of the nitrogen (N) cycle in the soil, nitrification is an oxidation process mediated by microorganisms that transform the relatively immobile ammonium (NH4+)to the water soluble nitrate (NO3-), enabling the production of nitrous oxide (N2O, a potent greenhouse gas) by denitrification as a by-product (Canfield et al. 2010). Researchers at CIAT-Colombia in collaboration with JIRCAS-Japan, reported that Brachiaria humidicola forage grasses have the ability to inhibit the nitrification process by exuding chemical compounds from its roots to the soil. A major hydrophobic compound was discovered and named brachial-actone (Subbarao et al. 2009). This capacity of Brachiaria grasses is known as biological nitrification inhibition (BNI) and it could contribute to better N use efficiency in crop-livestock systems by improving recovery of applied N while reducing NO3- leaching and N2O emission. The current methodologies for quantifying the BNI trait need further improvement to facilitate high throughput evaluation to quantify genotypic differences. In this paper, we aim to develop new (or improve the existing) phenotyping methods for this trait. Preliminary results were obtained using three different methods to quantify BNI: (1) a mass spectrometry method to quantify brachialactone; (2) a static chamber method to quantify N2O emission from soils under greenhouse conditions; and (3) an improved molecular method to quantify microbial populations by Real-Time PCR. Using these three methods we expect to score a bi-parental hybrid population (n=134) of two B. humidicola accessions differing in their BNI capacity CIAT26146 (medium to low BNI) x CIAT16888 (high BNI), in an attempt to identify QTLs associated with the BNI trait

Biological Nitrification Inhibition (BNI) in \u3cem\u3eBrachiaria\u3c/em\u3e Pastures: A Novel Strategy to Improve Eco-Efficiency of Crop-Livestock Systems and to Mitigate Climate Change

Up to 70% of the nitrogen (N) fertilizers applied to agricultural systems are lost due to nitrification and denitrification. Nitrification is a microbiological process that generates nitrate (NO3-) and promotes the losses of N fertilizers by leaching and denitrification. Nitrification and denitrification are the only known biological processes that generate nitrous oxide (N2O), a powerful greenhouse gas contributing to global warming. There is an urgent need to suppress nitrification process in soil to improve N-recovery and N use efficiency (NUE) of agricultural systems and to mitigate climate change (Subbarao et al. 2012). Certain Brachiaria grasses (B. humidicola) can suppress soil-nitrification by releasing biological nitrification inhibitors (BNIs) from roots, thereby reducing N2O emissions. This phenomenon, termed biological nitrification inhibition (BNI), has been the subject of recent research to characterize and validate the concept under field conditions (Subbarao et al. 2009). Advances on three aspects of BNI research are reported here: (1) gene quantification of soil nitrifying microorganisms to determine BNI activity in B. humidicola; (2) screening of B. humidicola breeding materials to identify hybrids with contrasting levels of BNI: and (3) quantification of the BNI-residual effect from B. humidicola on N-recovery and agronomic-NUE of the subsequent maize crop

Nitrogen Management in Grasslands and Forage-Based Production Systems–Role of Biological Nitrification Inhibition (BNI)

Nitrogen (N), being the most critical and essential nutrient for plant growth, largely determines the productivity in both extensive- and intensive- grassland systems. Nitrification and denitrification processes in the soil are the primary drivers generating reactive-N: NO3-, N2O, and NO, and is largely responsible for N-loss and degradation of grasslands. Suppressing nitrification can thus facilitate the retention of soil-N to sustain long-term productivity of grasslands and forage-based production systems. Certain plants can suppress soil nitrification by releasing inhibitors from roots, a phenomenon termed ‘biological nitrification inhibition’ (BNI). Recent methodological developments (e.g. bioluminescence assay to detect BNIs from plant-root systems) led to significant advances in our ability to quantify and characterize BNI function in pasture grasses. Among grass-pastures, BNI-capacity is strongest in low-N adapted grasses such as Brachiaria humidicola and weakest in high-N environment grasses such as Italian ryegrass (Lolium perenne) and B. brizantha. The chemical identity of some of the BNIs produced in plant tissues and released from roots has now been established and their mode of inhibitory action determined on nitrifying bacteria Nitrosomonas. Synthesis and release of BNIs is a highly regulated and localized process, triggered by the presence of NH4+ in the rhizosphere, which facilitates the release of BNIs close to soil-nitrifier sites. Substantial genotypic variation is found for BNI-capacity in B. humidicola, which opens the way for its geneticmanipulation. Field studies suggest that Brachiaria grasses suppress nitrification and N2O emissions from soil. The potential for exploiting BNI function (from a genetic improvement and a system perspective) to develop production systems that are low-nitrifying, low N2O-emitting, economically efficient and ecologically sustainable, will be the subject of discussion

Phenotyping of Urochloa humidicola grass hybrids for agronomic and environmental performance in the Piedmont region of the Orinoquian savannas of Colombia

In the low fertility acid soils of the Orinoquian savannas of Colombia, Urochloa humidicola cv. Tully or Humidicola is one of the most widely planted tropical forage grasses for improving livestock productivity. Low nutritional quality of this grass limits sustainable livestock production in this region. In this study, we conducted a phenotypic evaluation under field and greenhouse conditions of one of the first hybrid populations of U. humidicola generated from the forage breeding program of CIAT. Our objective was to identify a set of new hybrids of U. humidicola that combine improved productivity and nutritional quality plus the biological nitrification inhibition (BNI) trait/ability to reduce nitrogen (N) losses via leaching and nitrous oxide (N2O) emissions. To this end, we tested 118 hybrids (planted in pots) in the greenhouse for over 6 months and measured potential nitrification rates (NR) using soil microcosm incubation. NR values observed ranged from 0.27 to 5.75 mg N-NO3− kg soil−1 day−1. Later, 12 hybrids with different levels of NR were selected and field-tested in the Orinoquia region over a 4 years period (2013–2017) for dry matter production, nutrition quality (crude protein, in vitro digestibility and fibres content) and NR in each year. In the rainy season of 2018, two hybrids with superior agronomic performance and contrasting field level NR (Uh08/1149 and 0450) were subjected to analysis of soil-borne N2O emissions after fertilization during 13 days. The NR values recorded were not directly correlated with the forage quality parameters evaluated, however, the two grasses with the lowest NR values were among those with the highest biomass production, crude protein content, and N uptake. The grass hybrid Uh08/1149 and the germplasm accession CIAT 16888 were found as materials with superior forage value, with production of 14.1 and 14.6 tons dry matter ha−1 year−1 (up to 8% higher than the cv. Tully), crude protein of 11.5 and 9.1% per cut (up to 20% higher than the cv. Tully), and N uptake of 31.6 and 25.7 kg N ha−1 cut−1 (up to 30% higher than the cv. Tully). Additionally, these two grasses are likely to exhibit high-BNI ability, with potential to improve N use efficiency in managed pastures

Biological nitrification inhibitor-trait enhances nitrogen uptake by suppressing nitrifier activity and improves ammonium assimilation in two elite wheat varieties

Synthetic nitrification inhibitors (SNI) and biological nitrification inhibitors (BNI) are promising tools to limit nitrogen (N) pollution derived from agriculture. Modern wheat cultivars lack sufficient capacity to exude BNIs, but, fortunately, the chromosome region (Lr#n-SA) controlling BNI production in Leymus racemosus, a wild relative of wheat, was introduced into two elite wheat cultivars, ROELFS and MUNAL. Using BNI-isogenic-lines could become a cost-effective, farmer-friendly, and globally scalable technology that incentivizes more sustainable and environmentally friendly agronomic practices. We studied how BNI-trait improves N-uptake, and N-use, both with ammonium and nitrate fertilization, analysing representative indicators of soil nitrification inhibition, and plant metabolism. Synthesizing BNI molecules did not mean a metabolic cost since Control and BNI-isogenic-lines from ROELFS and MUNAL presented similar agronomic performance and plant development. In the soil, ROELFS-BNI and MUNAL-BNI plants decreased ammonia-oxidizing bacteria (AOB) abundance by 60% and 45% respectively, delaying ammonium oxidation without reducing the total abundance of bacteria or archaea. Interestingly, BNI-trait presented a synergistic effect with SNIs since made it also possible to decrease the AOA abundance. ROELFS-BNI and MUNAL-BNI plants showed a reduced leaf nitrate reductase (NR) activity as a consequence of lower soil NO3- formation and a higher amino acid content compared to BNI-trait lacking lines, indicating that the transfer of Lr#-SA was able to induce a higher capacity to assimilate ammonium. Moreover, the impact of the BNI-trait in wheat cultivars was also noticeable for nitrate fertilization, with improved N absorption, and therefore, reducing soil nitrate content.This project was funded by the Spanish Government (RTI2018-094623-B-C21 MCIU/AEI/FEDER, UE) and by the Basque Government (IT932-16; IT1560-22; 00048-ID2021-45). AB-L and LU held grants from the Basque Government (PRE-2020-2-0142 and PRE-2020-1-0127)

Enlisting wild grass genes to combat nitrification in wheat farming: A nature-based solution

Active nitrifiers and rapid nitrification are major contributing factors to nitrogen losses in global wheat production. Suppressing nitrifier activity is an effective strategy to limit N losses from agriculture. Production and release of nitrification inhibitors from plant roots is termed "biological nitrification inhibition" (BNI). Here, we report the discovery of a chromosome region that controls BNI production in "wheat grass" Leymus racemosus (Lam.) Tzvelev, located on the short arm of the "Lr#3Ns(b)" (Lr#n), which can be transferred to wheat as T3BL.3Ns(b)S (denoted Lr#n-SA), where 3BS arm of chromosome 3B of wheat was replaced by 3Ns(b)S of L. racemosus. We successfully introduced T3BL.3Ns(b)S into the wheat cultivar "Chinese Spring" (CS-Lr#n-SA, referred to as "BNI-CS"), which resulted in the doubling of its BNI capacity. T3BL.3Ns(b)S from BNI-CS was then transferred to several elite high-yielding hexaploid wheat cultivars, leading to near doubling of BNI production in "BNI-MUNAL" and "BNI-ROELFS." Laboratory incubation studies with root-zone soil from field-grown BNI-MUNAL confirmed BNI trait expression, evident from suppression of soil nitrifier activity, reduced nitrification potential, and N2O emissions. Changes in N metabolism included reductions in both leaf nitrate, nitrate reductase activity, and enhanced glutamine synthetase activity, indicating a shift toward ammonium nutrition. Nitrogen uptake from soil organic matter mineralization improved under low N conditions. Biomass production, grain yields, and N uptake were significantly higher in BNI-MUNAL across N treatments. Grain protein levels and breadmaking attributes were not negatively impacted. Wide use of BNI functions in wheat breeding may combat nitrification in high N input-intensive farming but also can improve adaptation to low N input marginal areas.We gratefully acknowledge funding support from Japanese Ministry of Agriculture, Forestry and Fisheries, CGIAR Research Program on WHEAT during the execution of the research presented in this study

Challenges and Opportunities for Improving Eco-Efficiency of Tropical Forage-Based Systems to Mitigate Greenhouse Gas Emissions

Forage-based livestock production plays a key role in national and regional economies, for food security and poverty alleviation. Livestock production is also considered as a major contributor to agricultural GHG emissions, however. While demand for livestock products is predicted to continue to increase, there is political and societal pressure both to reduce environmental impacts and to convert some of the pasture area to alternative uses such as crop production and environmental conservation. Thus it is essential to develop approaches for sustainable intensification of livestock systems to mitigate GHG emissions, addressing biophysical, socioeconomic and policy challenges. This paper highlights the potential of improved tropical forages in crop-livestock systems, and linked with policy incentives, to enhance livestock production while reducing its environmental footprint. We give examples for sustainable intensification to mitigate GHG emissions based on improved forages in Brazil and Colombia and suggest future perspectives

Accelerating the development of biological nitrification inhibition as a viable nitrous oxide mitigation strategy in grazed livestock systems

This position paper summarizes the current understanding of biological nitrification inhibition (BNI) to identify research needs for accelerating the development of BNI as a N2O mitigation strategy for grazed livestock systems. We propose that the initial research focus should be on the systematic screening of agronomically desirable plants for their BNI potency and N2O reduction potential. This requires the development of in situ screening methods that can be combined with reliable N2O emission measurements and microbial and metabolomic analyses to confirm the selective inhibition of nitrification. As BNI-induced reductions in N2O emissions can occur by directly inhibiting nitrification, or via indirect effects on other N transformations, it is also important to measure gross N transformation rates to disentangle these direct and indirect effects. However, an equally important challenge will be to discern the apparent influence of soil N fertility status on the release of BNIs, particularly for more intensively managed grazing systems

Climate-Smart Crop-Livestock Systems for Smallholders in the Tropics: Integration of New Forage Hybrids to Intensify Agriculture and to Mitigate Climate Change through Regulation of Nitrification in Soil

It is widely recognized that less than 50% of applied nitrogen (N) fertilizer is recovered by crops, and based on current fertilizer prices the economic value of this “wasted N” globally is currently estimated as US$81 billion annually. Worse still, this wasted N has major effects on the environment (Subbarao et al. 2012). CIAT researchers and their collaborators in Japan reported a major breakthrough in managing N to benefit both agriculture and the environment (Subbarao et al. 2009). Termed Biological Nitrification Inhibition (BNI), this natural phenomenon has been the subject of long-term collaborative research that revealed the mechanism by which certain plants (and in particular the tropical pasture grass B. humidicola) naturally inhibit the conversion of N in the soil from a stable form to forms subject to leaching loss (NO3) or to the potent greenhouse gas N2O (Subbarao et al. 2012). Brachiaria humidicola which is well adapted to the low-nitrogen soils of South American savannas has shown high BNI-capacity among the tropical grasses tested (Subbarao et al. 2007). The major nitrification inhibitor in Brachiaria forage grasses is brachialactone, a cyclic diterpene (Subbarao et al. 2009). Reduction of N loss from the soil under a B. humidicola pasture has a direct and beneficial environmental effect. We hypothesize that this conservation of soil N will have an additional positive impact on a subsequent crop (e.g. maize). At present, recovery of fertilizer N and the impact on crop yield is not known. The main purpose of our inter-institutional and multi-disciplinary project, targeting small-scale farmers, is to develop the innovative approach of BNI using B. humidicola forage grass hybrids to realize sustainable economic and environmental benefits from integrated crop-livestock production systems