Browsing by Subject "Biological nitrification inhibition"

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Biochemical and ecophysiological characterization of BNI (Biological Nitrification Inhibition) by Brachiaria humidicola
(2021) Egenolf, Konrad; Rasche, Frank
In perennial grasslands, especially Brachiaria humidicola (syn. Urochloa humidicola) dominated swards, one hypothesized mechanism of high N efficiency is the plant exerted control of nitrification via the synthesis and release of nitrification inhibitors (NI) into the soil. This phenomenon has been conceptualized as Biological Nitrification Inhibition (BNI). This doctoral thesis was conducted with the aim of broadening our fundamental understanding on BNI ecophysiology, with a special emphasis on the edaphic parameters soil pH and soil texture as factors shaping the soil microbial community composition. The overarching objectives were to (1) screen root exudates of B. humidicola for major bioactive secondary metabolites with nitrification inhibiting activity, to (2) proof the significance of rhizosphere pH and nutritional N form for NI release and understand the underlying exudation mechanism, and to (3) elucidate the influence of soil pH and soil texture on the ammonia oxidizer community composition and BNI performance of B. humidicola. Root exudate screening via LC-MS and NMR techniques revealed several novel NI compounds with significantly higher NI activity compared to previously described brachialactone, i.e. the brachialactone isomers/derivatives 3-epi-brachialactone (ED50 ~ 20 µg ml-1, ED80 ~ 40 µg ml-1) and 3,18-epoxy-9-hydroxy-4,7-seco-brachialactone (ED50 ~ 40 µg ml-1) as well as the phenol aldehyde vanillin (ED50 ~ 12.5 µg ml-1, ED80 ~ 20 µg ml-1). In the case of the described brachialactone derivatives, internal tissue concentrations were extremely low (2-8 µg g-1 root DM), suggesting so far undiscovered cytosolic precursors. In the case of vanillin, its chemical proximity to other phenolic compounds previously described as NI, i.e. methyl-coumarate, methyl-ferulate and methyl 3-(4-hydroxyphenyl) propionate, drew the attention to phenylalanine and coumaric acid as common precursors and possible BNI breeding target. With regard to the NI exudation, the hypothesized positive effect of low rhizosphere pH and NH4+ nutrition was confirmed for both the brachialactone isomers/derivatives and vanillin. However, for 3-epi-brachialactone it was demonstrated that NH4+ did not constitute an essential prerequisite for NI synthesis and release. In contrast, NI release correlated with the transmembrane proton gradient, which in turn depends on soil pH and is favored by rhizosphere acidification occurring under cation-dominant nutrition (e.g. NH4+). These findings were considered as evidence for an active NI release via secondary transporters (possibly MATE transporters). The effects of soil pH and soil texture on the ammonia oxidizer community and BNI performance of B. humidicola were investigated through a three-factorial pot trial including liming and different soil types as experimental factors. No clear conclusion could be drawn with regard to the hypothesized effects of soil pH, soil texture and the ammonia oxidizer community composition on BNI performance of B. humidicola. In the presented pot trial, B. humidicola reduced net nitrification rates by 50-85% compared to the non-planted control, but this reduction was observable irrespective of soil pH, soil texture and the ammonia oxidizer community composition. Furthermore, the reduction of net nitrification was largely dependent on microbial N immobilization and efficient plant N (probably NO3-) uptake rather than BNI. This absence of a clear BNI effect was mainly attributed to high N inputs, which is in accordance with previous studies indicating that BNI was impaired in high nitrifying environments. The argument was underlined by theoretical enzyme-kinetic calculations, revealing a strong influence of substrate (NH4+) availability on soil nitrification dynamics, but as well BNI performance: Assuming soil NI concentrations at ED50 (~ effective dose 50% inhibition), it could be shown that – with the only exception of AOA populations suppressed by non-competitive inhibitors – the efficacy of NI is severely disrupted by increasing soil NH4+ availability. Besides contrasting AOA and AOB sensitivities towards NI, the inter-domain differences of ammonia-monooxygenase (AMO) kinetics probably delivers an additional explanation for the observation, that under field conditions BNI has mainly been confirmed for AOA, and to a lesser extent for AOB. Based on the findings of the presented doctoral thesis, it was concluded that BNI may play an important role in extensive B. humidicola pasture systems, especially on acid, coarse textured and AOA dominated soils. Intensification, especially increasing N amendments, will most likely disrupt the nitrification inhibiting effect and under these circumstances, N immobilization and efficient plant N uptake may display the dominant factors controlling net nitrification rates.
Developing indicators and characterizing direct and residual effects of biological nitrification inhibition (BNI) by the tropical forage grass Brachiaria humidicola
(2018) Karwat, Hannes; Cadisch, Georg
Nitrogen (N) losses from agroecosystems harm the environment via increased nitrate (NO3-) amounts in water-bodies and nitrous oxide (N2O) emissions to the atmosphere. Bacteria and archaea oxidize ammonium (NH4+) to NO3- under aerobic conditions. Furthermore, under mainly anaerobic conditions, microbial denitrification reduces NO3- to gaseous N forms. The tropical forage grass Brachiaria humidicola (Rendle) Schweick (Bh) has been shown to reduce soil microbial nitrification via root derived substances. Therefore, biological nitrification inhibition (BNI) by Bh might contribute to reduction of N losses from agroecosystems. The present doctoral thesis aimed at assessing the potential of the actual BNI by Bh, as well as the residual BNI effect with new developed methodologies. The overall research was based on the following major objectives: (1) characterization of the residual BNI effect by Bh on recovery of N by subsequent cropped maize (Zea mays L.) under different N fertilization rates; (2) investigate if low enzymatic nitrate reductase activity (NRA) in leaves of Bh is linked to reduced NO3- nutrition by effective BNI; (3) identify a possible link between plant delta 15N of Bh and the BNI effect of different Bh genotypes on nitrification, plant N uptake and NO3- leaching losses. The overall objective was to use and test new methodologies with a minimum of disturbance of the plant-soil system, to characterize BNI of different Bh genotypes in greenhouse and field studies. The first research study focused on the investigation of a potential residual BNI effect of a converted long-term Bh pasture on subsequent maize cropping, where a long-term maize monocrop field served as control. The residual BNI effect was characterized in terms of enhanced maize grain yield, total N uptake and 15N (labeled) fertilizer recovery. Furthermore, the impact of residual BNI effect on soil N dynamics was investigated. The residual BNI effect was confirmed for the first maize crop season after pasture conversion on the basis of lower nitrification in incubation soil, higher total N uptake and higher maize grain yields. However, the residual BNI effect did not result in higher 15N fertilizer uptake or reduced 15N fertilizer losses, nor in reduced N20 emissions. Applied N was strongly immobilized due to long-term root turnover effects, while a significant residual BNI effect from Bh prevented re-mineralized N from rapid nitrification resulting in improved maize performance. A significant residual Bh BNI effect was evident for less than one year only. In the second research study it was the aim to verify the potential of nitrate reductase activity (NRA) as a proxy for the detection of in vivo performance of BNI by selected Bh accessions and genotypes grown under contrasting fertilization regimes. NRA was detected in Bh leaves rather than in roots, regardless of NO3- availability. Leaf NRA correlated with NO3- contents in soils and stem sap of contrasting Bh genotypes substantiating its use as a proxy of in vivo performance of BNI. The leaf NRA assay facilitated a rapid screening of contrasting Bh genotypes for their differences in in vivo performance of BNI under field and greenhouse conditions; but inconsistency of the BNI potential by selected Bh genotypes was observed. The third research study emphasized to link the natural abundance of delta 15N in Bh plants with reduced NO3- losses and enhanced N uptake due to BNI. Increased leached NO3- was positively correlated to rising delta 15N in Bh grass, whereas the correlation between plant N uptake and plant delta 15N was inverse. Long-term field cultivation of Bh decreased nitrification in incubated soil, whereas delta 15N of Bh declined and plant N% rose over time. Delta 15N of Bh correlated positively with assessed nitrification rates in incubated soil. It was concluded that decreasing delta 15N of Bh over time reflects the long-term effect of BNI linked to lower NO3- formation and reduced NO3- leaching, and that generally higher BNI activity of Bh is indicated by lower delta 15N plant values. Within the framework of this thesis, a residual BNI effect by Bh on maize cropping could be confirmed for one season due to the combined methodological approaches of soil incubation and 15N recovery. The development of the NRA assay for sampled Bh leaves was validated as a rapid and reliable method linked to the actual soil nitrification after NH4+ fertilizer supply. Consequently, the assay could be used for both greenhouse and field studies as BNI proxy. The gathered data from the third study indicated that decreasing delta 15N of Bh over time reflects the long-term effect of BNI linked to lower NO3- formation and reduced NO3- leaching, and that generally higher BNI activity of Bh is indicated by lower delta 15N plant values. Consequently, it was suggested that delta 15N of Bh could serve as an indicator of cumulative NO3- losses. Overall, this doctoral thesis suggests the depressing effect on nitrification by Bh might be a combined effect by BNI and fostered N immobilization. Furthermore, BNI by Bh might be altered by different factors such as soil type, plant age and root morphology of the genotypes. Finally, future studies should consider that Bh genotypes express their respective BNI potential differently under contrasting conditions.