Дисертації з теми "Nitrate use efficiency"

Nitrate (NO₃^-) leaching from fertilized land can be detrimental to aquatic ecosystems and human health. Although NO ₃^- leaching potential is generally found to be low in turfgrass, certain conditions can occur that result in increased leaching loss of nitrogen. Kentucky bluegrass (Poa pratensis L.) is the most widely used cool-season turfgrass species in the temperate and subarctic climate zones in the United States. Due to its popularity, many new cultivars are bred and released each year. Despite the ample amount of information characterizing the agronomic features of Kentucky bluegrass genotypes, little is available documenting their physiological characteristics related to nitrogen use efficiency and how these might affect the NO₃ ^- leaching potential of the genotypes. Such information would be of great value for both scientists and customers.

In this comprehensive study, a screening procedure was developed using nutrient solution culture to evaluate differences in NO₃ ^- uptake among sixty Kentucky bluegrass genotypes. Two cultivars were selected from the sixty to represent genotypes having high vs. low capacity for NO₃^- uptake. These two cultivars were then used to examine the relationship between NO₃^- uptake efficiency and competitiveness for soil nitrogen. Finally, these two cultivars were compared to determine if NO₃^- uptake efficiency affected NO₃^- leaching potential.

There were significant differences in NO₃^- uptake at both high (1 mM) and low (0.05 mM) N concentrations among sixty Kentucky bluegrass genotypes, with a strong correlation between uptake rates at high and low N concentrations. Julia and Midnight were selected as representing cultivars with efficient and inefficient nitrate uptake, respectively. In nutrient solution culture, Julia exhibited 56% higher NO₃ ^- uptake rates than Midnight. In a root competition study, in which the root systems of the two cultivars occupied the same soil volume, Julia was more competitive for nitrogen acquisition (absorbed 20 to 71% more NO ₃^- ) than Midnight. However, the higher NO₃ ^- uptake capacity and better competitiveness for soil N in Julia did not result in lower NO₃^- leaching potential. These results suggest that differences in root morphology and architecture may play a more important role than uptake capacity in determining nitrate leaching potential among Kentucky bluegrass genotypes.

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
A aplicação de amônio pode, em algumas condições, melhorar a eficiência de uso do fósforo (EUP) e o desenvolvimento do milho, especialmente em solos tropicais, normalmente ácidos e pobres em P disponível. O presente trabalho tem por objetivo estudar o efeito da adubação amoniacal sobre o aproveitamento do P, as condições bioquímicas da rizosfera, a morfologia radicular e o desenvolvimento da cultura do milho. Os dois experimentos iniciais foram conduzidos em casa de vegetação, o primeiro em vasos de 12 L e o segundo em Plant Growth Containers (PGCs), com solo previamente corrigido e adubado segundo os tratamentos: 3 (três formas de adubação nitrogenada - amoniacal, nítrica e mista) x cinco doses de fosfato (0, 40, 80, 120 e 160 mg P kg-1 de solo). Aos 35 dias após a emergência as raízes foram removidas dos containers e vasos e as amostras de planta e solo foram coletadas para análises. O solo rizosférico no experimento I foi obtido por “agitação-lavagem” e no experimento II por “fatiamento em camadas”, de forma que quatro das oito repetições foram congeladas frescas (análises: amônio, nitrato, C orgânico e P microbiano) e quatro foram transformadas em TFSE (análises: fracionamento químico do P e pH). As amostras de planta foram submetidas a pesagem e análise nutricional. O experimento III foi conduzido em um sistema de scanners em solução nutritiva de acordo com os tratamentos: 2 (amônio ou nitrato) x (alta dose ou baixa dose de P), com 10 repetições. Imagens foram obtidas diariamente para avaliação do desenvolvimento radicular. Ao final dos ensaios foram obtidos peso fresco e seco, análises de P em tecidos e, com dados de diferentes parâmetros radiculares, foi feito o desenho da planta média no RootBox. Os tratamentos com amônio e nitrato + amônio, na dose mais alta de P no experimento I ou em todas as doses no experimento II, resultaram em maior produção de matéria seca, menor relação raiz: parte aérea, desenvolvimento de raiz mais aequado à absorção de P, maior recuperação de P, maior acúmulo de amônio, nitrato e N-mineral nas camadas mais próximas ao rizoplano, menor pH, maior mobilização de formas menos lábeis de P, maior consumo de P geoquímico e P biológico. Houve diferença entre solo rizosférico e solo não rizosférico e entre as diferentes camadas da rizosfera, para todos os parâmetros, entre as formas de nitrogênio. O amônio e nitrato + amônio resultaram em maior produção de C orgânico solúvel em água e maior dose de superfosfato triplo resultou em maior quantidade de P na fração microbiana. O amônio resultou desenvolvimento radicular mais ramificado e com maior número de raízes laterais. O amônio melhora a eficiência de uso do P, devido às mudanças químicas e biológicas na rizosfera, e às mudanças na arquitetura radicular, que favorecem a solubilidade, a mineralização e a absorção de fosfato e, consequentemente, aumenta a produção de matéria seca nos estádios iniciais da cultura do milho.
Ammonium application can improve phosphorus use efficiency (PUE) and the development of maize plants, especially in tropical soils, usually acids and P-limiting. The aim of this study is evaluate how ammoniacal fertilization impacts the P use efficiency (PUE), the biochemical conditions of the rhizosphere, the root morphology and the development of maize plants. Two initial experiments were carried on greenhouse, the first one in pots of 20 L and the second one in Plant Growth Containers (PGCs), using soil after liming and fertilized according to the following treatments: 3 (nitrogen fertilization: ammonium only, nitrate only and both) x five doses of phosphate (0, 40, 80, 120 and 160 mg P kg-1 of soil). At 35 days after emergence the roots were removed from the containers and pots and the plant and soil samples were collected for analysis. The rhizosphere soil in experiment I was obtained by "shaking-washing" and in the experiment II by "layer slicing". Four of the eight soil replicates were fresh frozen (analyzes: ammonium, nitrate, organic C and microbial P) and four of them were dried (analyzes: chemical fractionation of P and pH). Plant samples were weighed and analyzed for P content. The experiment III was carried on a system of nutrient solution scanners according to the treatments: 2 (ammonium or nitrate) x (high or low dose of P), with 10 replicates. Images were obtained in daily basis for the evaluation of root development. At the end of the tests, fresh and dry weights and P content were obtained. Data of different root traits were used to draw the average plant on the RootBox. The treatments with ammonium and nitrate + ammonium, plus the highest dose of P in the experiment I or at all doses in the experiment II, resulted in higher dry matter yield, lower root ratio: aerial part, root development adequate for P absorption, higher P recovering, higher ammonium, nitrate and N-mineral accumulation in the nearest layersto the rhizoplane, lower pH, greater mobilization of less labile P forms, higher mobilization of geochemical and biological P. There were differences between rhizospheric and bulk soils and between the different layers of the rhizosphere, for all parameters and treatments. Ammonium and nitrate + ammonium resulted in higher organic C water soluble organic C, and higher dose of triple superphosphate resulted in higher amount of P in the microbial fraction. Ammonium resulted in a branched root with a greater number of lateral roots. Ammonium improves the use efficiency, solubility, mineralization and uptake of P, due to chemical and biological changes in the rhizosphere and root architecture in the early stages of corn development.
FAPESP: 13/18870-1
FAPESP: 15/12399-0

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
Forrageiras do gênero Cynodon são conhecidas por sua capacidade de resposta a altas doses de nitrogênio (N). Em condições tropicais o N pode se tornar um problema ambiental e financeiro devido à sua baixa eficiência de uso pelas plantas, principalmente como resultado de perdas por volatilização e/ou lixiviação. Portanto, objetivou-se com este trabalho avaliar fontes e doses de N no crescimento e produção de matéria seca do Tifton 85, bem como mudanças nas frações do carbono (C) no solo. Um experimento foi conduzido de 2012 a 2014, e constituiu de um fatorial em um delineamento de blocos ao acaso, onde os tratamentos foram a associação de duas fontes de N (nitrato de amônio [NA] e ureia), e cinco doses do nutriente (0, 60, 120, 180 e 240 kg ha-1 N por corte) aplicadas a cada 30 dias. Neste experimento foram utilizados NA e ureia enriquecidos com 15N como uma ferramenta para quantificar a recuperação do N advindo do fertilizante, nas plantas e no sistema solo-planta. As plantas foram cortadas para avaliar a produção de matéria seca, concentração de N na parte aérea e recuperação. O índice de área foliar (LAI), radiação fotossinteticamente ativa interceptada (PARi) e índice de clorofila (CI) foram mensurados no dia anterior ao corte. As amostras de solo foram coletadas nas parcelas que receberam NA e foram fracionadas em carbono orgânico particulado (POC), fração leve livre (FLF) e fração mineral (C-min); O teor de C nas frações foi quantificado. Os resultados demonstraram que o NA pode ser uma fonte mais eficiente somente quando a quantidade de precipitação é insuficiente para incorporar o fertilizante ao solo, resultando em maior produção de forragem, na somatória dos dois anos ambas as fontes produziram a mesma quantidade de matéria seca onde a maior produtividade 37,2 Mg ha-1, foi atingida com a dose de 210 kg ha-1 N por corte. Por outro lado, a quantidade de N recuperado pelo sistema de planta+solo foi maior quando a ureia foi utilizada, com destaque para a quantidade de N no solo, onde a ureia foi capaz de manter 10% mais N que o NA, a recuperação do nutriente diminuiu à medida que as doses foram elevadas. Nenhuma alteração no conteúdo de C foi notada devido às diferentes doses de N utilizadas, no entanto o POC e o C-min foram mais sensíveis às mudanças na camada de 0-0,1 m do que na camada de 0,1-0,2 m.
Cynodon hybrids are known by their ability to respond to high rates of nitrogen (N). In tropical conditions, N may become an environmental and financial issue due to its low efficiency of use by plants, mainly as a result of losses by volatilization and/or leaching. Therefore, the aims with this work were to evaluate N sources and rates on growth and dry matter yield of Tifton 85 and to determine changes in soil carbon (C) fractions in response to N fertilization. The 2-yr field experiment was conducted from 2012 to 2014, and consisted of a factorial design. Treatments were a combination of two sources of N (ammonium nitrate [AN] and urea) and five rates of the nutrient (0, 60, 120, 180 and 240 kg ha-1 N per cut) applied broadcast every 30 days. This study used AN and urea enriched with 15N as a tool to quantify the recovery of N derived from fertilizer in plants and the soil-plant system. Forage was cut at 30-d intervals for dry matter yield, shoot N concentration determinations and N recovery. The leaf area index (LAI), photosynthetic active radiation intercepted (PARi) and chlorophyll index (CI) were evaluated on the day before clipping. Soil samples were collected in plots receiving AN and they were fractionated in to particulate organic carbon (POC), free light fraction (FLF) and mineral fraction (C-min); The C concentration of the various fractions was determined. Results showed that AN was a more efficient source only when the amount of precipitation is insufficient to incorporate the fertilizer to the soil, resulting in increased production, in the sum of the two years both sources produced the same amount of dry matter in which the highest productivity, 37.2 t ha-1 , was achieved at the rate of 210 kg ha-1 N per cutting. On the other hand, the amount of N recovered by the plant+soil system was higher when urea was used, especially the amount of N in the soil, where urea was able to maintain 10% more N than AN, the nutrient recovery decreased as the rates were increased. No change in soil C concentration was detected in response to the different N rates used, however the POC and the C-min were more sensitive to changes in the layer of 0-0.1 m than the layer from 0.1-0.2 m.

A utilização de inibidores de nitrificação pode ser uma alternativa interessante para aumentar a eficiência do uso do fertilizante nitrogenado em diversas culturas, porém, essa alternativa vem sendo pouco estudada em condições de solo e clima do Brasil. Assim, objetivou-se com esse trabalho avaliar a eficiência do inibidor de nitrificação fosfato de 3,4-dimetilpirazol (DMPP) em três solos com gradiente textural, bem como avaliar o destino do nitrogênio (N-NO3-, N-NH4+ e 15N) no solo, a absorção e o uso do nitrogênio (N-total e 15N) nas plantas de algodão. Três experimentos foram montados e desenvolvidos. No primeiro foi efetuada a incubação do solo em condições de laboratório para avaliar a inibição da nitrificação pelo DMPP aplicado na forma de ureia e sulfonitrato de amônio (SNA) em três solos (Neossolo Quartzarênico - NQ, Latossolo Vermelho Amarelo - LVA, Latossolo Vermelho - LV). No segundo experimento plantas de algodão foram cultivadas em colunas de lixiviação com os mesmos solos, recebendo ureia e sulfonitrato de amônio, com e sem DMPP como fonte de N. Foram avaliados a produção de matéria seca, o acúmulo de nutrientes nas plantas, a eficiência de uso do N pelo algodoeiro, a lixiviação e a quantidade de N mineral no solo após o cultivo do algodão por 60 dias. No terceiro experimento, plantas de algodão foram cultivadas também em colunas lixiviação, porém, com um solo de textura média (Latossolo Vermelho Amarelo), o qual recebeu três doses de N em cobertura (50, 100 e 150 kg ha-1 de N) na forma de ureia marcada no isótopo 15N, com e sem DMPP. Foram avaliados a produção de matéria seca, teor de N e recuperação do N aplicado na planta, lixiviação de N, teor de N-total, nítrico e amoniacal no solo, e recuperação do N aplicado no solo após o cultivo do algodão, por 90 dias. Melhores resultados foram obtidos com a aplicação do DMPP em ureia em relação ao SNA. No experimento de incubação, o DMPP foi capaz de manter menor o teor de nitrato nos três solos analisados. A nitrificação do N na ureia foi mais rápida comparado ao SNA, o que permitiu melhor desempenho do inibidor na ureia em dois dos três solos estudados. O efeito do DMPP aumentou seguindo a seguinte ordem: NQ>LVA>LV. O inibidor foi mais eficiente nos solos com menor teor de argila e matéria orgânica. O uso do DMPP em ureia aplicada no solo arenoso (NQ) reduziu significativamente a lixiviação de N e aumentou a produção de matéria seca, a eficiência do uso do N e a absorção de fósforo pela planta. No solo de textura média (LVA), sob irrigação intensa, o DMPP reduziu significativamente as perdas de N do sistema e aumentou a recuperação do 15N aplicado na planta e no solo, o que, porém, não se traduziu em maior produção de matéria seca provavelmente pelo N não ter sido limitante, devido à mineralização da matéria orgânica. As atividades das enzimas redutase do nitrato e urease não diferiram entre tratamentos com e sem DMPP
The use of nitrification inhibitors may be an interesting alternative to increase nitrogen fertilizer use efficiency in different crops, although it has been little studied in soil and climate conditions of Brazil. This study aimed to evaluate the efficiency of the nitrification inhibitor 3,4-dimethylpyrazole phosphate (DMPP) in three soils with textural gradient (represented by clay percentage), as well as to evaluate the fate of soil nitrogen (N-NO3-, N-NH4+ and 15N), nitrogen uptake, and N-use efficiency (total-N and 15N) by cotton plants. Three experiments were set and carried out in a controlled environment. In the first one, a laboratory incubation was performed to evaluate the inhibition of nitrification by DMPP applied to urea and ammonium sulfate nitrate (ASN) in three soils (Typic Quartzipsammnet - NQ, Typic Hapludox - LVA, Rhodic Hapludox - LV). In the second experiment, leaching columns with the same soils were planted with cotton receiving ammonium sulfate nitrate and urea with or without DMPP as nitrogen source. It were evaluated plant dry matter yield (shoot plus roots), nutrient uptake and nitrogen use efficiency by cotton plants, as well as the nitrogen leaching, and mineral nitrogen amount in the soil after cotton growth, for 60 days. In the third experiment, leaching columns with a medium textured soil (LVA) were planted with cotton and received three nitrogen rates in topdressing (50, 100 and 150 kg ha-1) as 15N-urea with and without DMPP. The production of dry matter, nitrogen content and recovery of applied N in the plant and soil, as well as the nitrogen leaching, total nitrogen, nitrate and ammonium in the soil were evaluated after cotton growth for 90 days. Best results were obtained with the application of DMPP to urea than in ASN. In the incubation experiment, the DMPP was able to maintain lower nitrate content in the three soils. The nitrification of nitrogen from urea was faster than the one from ASN, which allowed a better performance of the inhibitor in the urea in two out of three soils analyzed. The effect of DMPP increased in the following order: NQ> LVA> LV. The inhibitor was more effective in soils with lower clay and organic matter contents. The use of DMPP in urea applied on the sandy soil (NQ) significantly reduced N leaching and increased dry matter production, nitrogen use efficiency and phosphorus uptake by cotton plants. In a medium textured soil (LVA) under intense irrigation, DMPP significantly reduced N losses in the system and increased the recovery of applied 15N in plant and soil, which however did not translate into higher dry matter production because nitrogen was probably not limiting. The activity of the enzymes nitrate reductase and urease were not different between treatments with and without DMPP

O uso de determinadas fontes nitrogenadas, doses e formas de nitrogênio (N) pode aumentar a eficiência de uso de N (EUN) em soqueiras de cana-de-açúcar (Saccharum spp.), mantendo, em longo prazo, elevados índices de produtividade, lucro e proteção ao ambiente. Experimentos de campo e de casa-de-vegetação foram desenvolvidos com os seguintes objetivos: (i) avaliar o efeito do manejo do N-fertilizante e dos ciclos de soqueira da cultura na produtividade, parâmetros tecnológicos, acúmulo de biomassa, acúmulo e balanço de nutrientes e nas alterações dos atributos químicos do solo; (ii) avaliar a produção de biomassa seca, metabolismo e nutrição de plantas de cana-de-açúcar cultivadas em solução nutritiva, com suprimento exclusivo ou misto de nitrato (NO3-) e amônio (NH4+) como fontes de N; (iii) avaliar a EUN do 15N-nitrato (15NH4NO3), 15N-amônio (NH415NO3) e 15N-amida [(15NH2)2CO] aplicados na quarta soqueira de cana-de-açúcar em condições de campo; e avaliar a EUN do 15N-amônio e 15N-nitrato aplicados em mudas de cana-de-açúcar cultivadas em solução nutritiva. No campo, o N-fertilizante foi repetido nas mesmas parcelas experimentais (parcelas estáticas), em anos subsequentes (quatro ou cinco soqueiras) e o manejo de N-fertilizante, que incluiu fontes e doses de N, não influenciou na produtividade de colmos e de açúcar no local 1 (Latossolo Vermelho Amarelo distrófico - LVAd). Entretanto, no local 2 (também LVAd), o NA100 e o NAC100 [nitrato de amônio (NA) e nitrato de amônio e cálcio (NAC), ambos na dose de 100 kg ha-1 de N] foram os manejos mais vantajosos para a obtenção das maiores produtividades de colmo e de açúcar em soqueiras quando comparado aos demais tratamentos. No geral, nos dois locais, o NAC150 (150 kg ha-1 de N) proporcionou os maiores acúmulos de biomassa, N e K nas plantas. Além disso, enquanto o controle (sem adição de N) apresentou as menores produções de biomassa e acúmulos de nutrientes (com exceção ao P), o balanço de nutrientes do solo nesse tratamento foi relativamente neutro após anos de cultivo da cana-de-açúcar. Em condições controladas, plantas de cana-de-açúcar supridas exclusivamente com amônio apresentaram menor produção de biomassa, acúmulo de nutrientes e área foliar do que os outros tratamentos (supridos com combinações de NO3-/NH4+ e somente com nitrato), indicando que o uso exclusivo dessa forma de N não foi eficiente para sustentar o crescimento vegetal. A maior EUN foi obtida com o fornecimento exclusivo de nitrato, que também proporcionou maior produção de biomassa seca, área foliar, acúmulo de macronutrientes, atividade fotossintética e crescimento do sistema radicular. A utilização de 15N indicou que, no campo, não houve diferença na EUN pelas plantas devido à utilização do 15N-nitrato e 15N-amônio. Em contraste, a EUN do 15N-amônio foi maior que a do 15N-nitrato durante o 1º, 4º e 7º dia após a aplicação do fertilizante (DAAF), em condições controladas. Contudo, o uso do 15N-nitrato teve maior EUN nas raízes e planta inteira que o 15N-amônio no 15º DAAF. Isso indica que a cana-de-açúcar pode aproveitar eficientemente as duas formas minerais de N, embora o aproveitamento do amônio seja mais rápido.
The use of different fertilizers and rates of nitrogen (N) can increase the N use efficiency (NUE) in sugarcane ratoon (Saccharum spp.), sustaining high yield levels, profitability and environmental protection in long-term. Thus, field and greenhouse studies were performed with the following objectives: (i) evaluate the effect of fertilizer-N management and ratoon crop cycles on sugarcane yield, technological parameters, biomass accumulation, nutrient content, temporal variation in soil chemical properties and soil nutrient balance; (ii) evaluate the biomass production, metabolism and nutrient acquisition of sugarcane plants cropped in nutrient solution, with solely or combined supply of ammonium and nitrate as N sources; (iii) evaluate the NUE of 15N-ammonium (15NH4NO3), 15N-nitrate (NH415NO3), and 15N-amide [(15NH2)2CO] applied to sugarcane under field conditions and the NUE of 15N-ammonium and 15N-nitrate applied to seedlings sugarcane under nutrient solutions. In the field, N fertilizers were replicated in the same experimental plots (static plots) in subsequent years (four or five ratoons) and the management of N-fertilizer, which included fertilizers and N rates, resulted in lack of influence on cane and sucrose yield at Site 1 (Typic Hapludox). However, at Site 2 (also Typic Hapludox), the N treatments AN100 and CAN100 [ammonium nitrate (AN) and calcium ammonium nitrate (CAN), both at a rate of 100 kg N ha-1] provided the highest cane and sugar yield as compared to the remaining treatments. Overall, at both sites, CAN150 (150 kg N ha-1) resulted in the highest biomass accumulation and N and K content by sugarcane. In addition, while the control (no N added) had the lowest biomass accumulation and nutrient content (except for P), the soil nutrient balance in this treatment was relatively neutral, after years of sugarcane cultivation. Under controlled conditions, NH4+-fed plants presented lower biomass accumulation, nutrient content and leaf area than the other treatments (different NO3-/NH4+ ratios and NO3--fed plants), indicating that this N form, when applied solely, was not effective for the proper plant growth. The highest NUE was obtained with NO3--fed plants, which also had the highest production of dry biomass, leaf area, nutrient accumulation, photosynthetic activity, and growth of the root system. The use of 15N-nitrate and 15N-ammonium indicated that there was no difference in NUE by plants in the field. In contrast, the NUE of 15N-ammonium was greater than 15N-nitrate during the 1st, 4th and 7th day after fertilizer application (DAFA), under controlled conditions. However, the use of 15N-nitrate had greater NUE in roots and whole plant than 15N-ammonium in the 15th DAFA. It can be concluded that sugarcane can use both N forms although the recovery of NH4+ was faster than NO3-.

O presente trabalho foi desenvolvido na área experimental da ESALQ-USP, localizada no município de Piracicaba/SP, em um Latossolo Vermelho Amarelo, textura areno-argiloso. O experimento teve como objetivo estudar a dinâmica do N no sistema solo-planta da cultura do milho, sob implantação do sistema plantio direto, e, pela técnica do 15N, avaliar a lixiviação de nitrato à profundidade de 0,80 m, a eficiência de utilização do N e o efeito residual do fertilizante nas culturas subseqüentes. O projeto envolveu dois cultivos de milho e um de aveia preta, cultivada na entressafra. O delineamento experimental foi o de blocos casualizados (3x2), com 4 repetições. Os tratamentos consistiram de doses de nitrogênio, na forma de sulfato de amônio (60, 120 e 180 kg ha-1 de N) e uma testemunha, além dos parcelamentos (30 kg ha-1 de N na semeadura e o restante em cobertura; 60 kg ha-1 de N na semeadura e o restante em cobertura). A aplicação de sulfato de amônio enriquecido com 15N foi feita, na dose de 120 kg ha-1 de N, em subparcelas, previamente definidas, apenas no primeiro cultivo do milho. Foram feitas as seguintes avaliações: densidade de fluxo de água e de nitrato na profundidade de 0,80 m do solo (dose de 120 kg ha-1 de N); massa de 100 grãos; massa de matéria seca; teor de N; rendimento de grãos de milho; átomos % 15N nas plantas e solo; fração do N fertilizante na solução do solo na profundidade de 0,80 m, e no perfil do solo; balanço do N na sucessão de culturas; efeito residual do N fertilizante (aplicado no primeiro cultivo de milho) na aveia e no segundo cultivo de milho. Pelos resultados obtidos, concluiu-se que: a) a perda de nitrato total por lixiviação, a 0,80 m de profundidade, no 1º cultivo de milho, na dose de 120 kg ha-1 de N, foi de aproximadamente 96 e 68 kg ha-1, para os parcelamentos com 60 kg ha-1 e 30 kg ha-1 de N na semeadura, respectivamente, dos quais apenas 3 e 1 kg ha-1 foram provenientes do fertilizante nitrogenado; b) na dose 120 kg ha-1 de N, o tratamento com maior quantidade de N aplicado na semeadura (60 kg ha-1), no cultivo de milho do ano agrícola 2003/2004, proporcionou um maior aproveitamento do N do fertilizante (65%), em relação ao tratamento com a dose de 30 kg ha-1 na semeadura (45%) e que, para esse ano, a recuperação do N fertilizante pelo solo, na camada 0-0,80 m, para ambos os tratamentos, 60-60 e 30-90 kg ha-1, foi de 40 e 49%, respectivamente; c) Ainda referindo-se aos tratamentos 60-60 e 30-90 kg ha-1, do N remanescente do sulfato de amônio (120 kg ha-1), aplicado no primeiro ano agrícola (efeito residual), 3,72 e 1,79% foram utilizados pela aveia preta e 2,84 e 2,06% pelo milho cultivado no ano subseqüente (2004/2005), permanecendo no solo (camada 0-0,80 m), 30,19 e 33,47% após o cultivo de aveia preta e 16,79 e 17,91% após o segundo cultivo de milho, respectivamente, para os referidos tratamentos.
The general objective of this work was to study the dynamics of nitrogen in the soil-plant system of two maize crops with a black oats one in between, introducing the no-tillage system. Besides, fertilizing the first maize crop with 15N enriched nitrogen fertilizer, it was also objective of this work to assess the total and the derived from fertilizer nitrate leached at the depth of 0.8 m, the nitrogen use efficiency by the crops and the residual effect of the labeled nitrogen fertilizer. The experiment was carried in the experimental areas of ESALQ-USP in the country of Piracicaba (SP), in a yellow red latossol, sandy-loamy texture. The first maize crop was in 2003/2004 and the second one in 2004/2005. The experimental design was randomized blocks (3 x 2), with four replications. Treatments consisted of nitrogen (as ammonium sulphate) levels (60, 120 and 180 kg ha-1 of nitrogen) and the control, besides the splittings (30 and 60 kg ha-1 of nitrogen in the sowing and the rest in covering). The fertilization with 15N enriched ammonium sulphate in the first maize crop was made only for the level of 120 kg ha-1 of nitrogen, in subplots previously defined. The following measurements were carried out: soil-water and nitrate flux densities at the depth of 0.8 m (120 kg ha-1 of N); weight of 100 grains; weight of plant dry matter; nitrogen content; grain yield; total nitrogen; plant nitrogen use efficiency; fraction of the applied nitrogen in the soil solution at the depth of 0.8 m and in the soil profile; nitrogen balance in the crops succession; nitrogen residual effect in the black oat and the second maize crop. From the results, it could be concluded that: a) the leaching losses of total N, at the depth of 0,80 m, in the first maize crop, at the fertilization level of 120 kg ha-1 of N, were of 96 and 68 kg ha-1 for the splittings 60 and 30 kg ha-1 of N at sowing, respectively, from which only 3 and 1 kg ha-1 were derived from the nitrogen fertilizer; b) at the N fertilization level of 120 kg ha-1, the treatment with higher amount of applied N at sowing (60 kg ha-1) of the first maize crop (2003/2004) led to a higher plant N fertilizer recovery (65%) than the 30 kg ha-1of applied N at the sowing treatment (45%) and that, at the end of this crop cycle, the N fertilizer recovery by the 0-0,80 m soil layer, for 60-60 and 30-90 kg ha-1 treatments, were 40 and 49%, respectively; c) still for treatments 60-60 and 30-90 kg ha-1, from the total N applied in the first maize crop (120 kg ha-1), 3.72 and 1.79% were used by the black oats and 2.84 and 2.06% by the second maize crop, remaining in the soil (0-0.8 m layer), 30.19 and 33.47% after the black oats crop and 16.79 and 17.91% after the second maize crop, respectively for 60-60 and 30-90 kg ha-1 of N treatments.

Les biocarburants de 2ème génération pourraient fournir une énergie renouvelable au secteur des transports et ainsi permettre de lutter contre le changement climatique. Toutefois, leurs bilans gaz à effet de serre, énergétiques et environnementaux seront probablement très dépendants des ressources utilisées. Les cultures lignocellulosiques candidates à la production de biocarburant 2G devront ainsi concilier forte productivité, faibles besoins en intrants et faibles impacts environnementaux. L’objectif de la thèse a été de quantifier les bilans d’eau, d’azote et de carbone à l’échelle de la parcelle, pour différentes cultures candidates et différentes pratiques culturales. Nous nous sommes appuyés sur le dispositif expérimental de long terme « Biomasse & Environnement », mis en place en 2006 à Estrées-Mons, en Picardie. Il compare deux cultures pérennes en C4 (Miscanthus × giganteus et switchgrass), deux cultures pluriannuelles fourragères en C3 (fétuque et luzerne) et deux cultures annuelles récoltées en plante entière (sorgho fibre et triticale). Il inclut deux niveaux de fertilisation et deux dates de récolte pour les cultures pérennes : récolte précoce (octobre) ou récolte tardive (février). Les mesures effectuées ont porté sur : i) la production de biomasse, ii) l’évolution des stocks d’eau du sol en continu pendant 7 ans, iii) la profondeur et la densité des systèmes racinaires, iv) le drainage et la concentration en nitrate de l’eau drainée, évalués avec le modèle STICS à partir des stocks d’eau et d’azote minéral du sol mesurés en milieu d’automne et fin d’hiver, v) les stocks de carbone organique du sol en 2006 et 2011-2012, vi) le devenir de l’engrais azoté, suivi par marquage isotopique 15N de l'engrais pendant 4 ou 5 années successives.Grâce à leur enracinement profond, les cultures pérennes et pluriannuelles ont prélevé davantage d’eau que les cultures annuelles, notamment en profondeur. Le drainage sous les cultures pluriannuelles a été plus faible que sous les cultures annuelles (64 contre 133 mm an-1 en moyenne sur 7 ans), malgré une production de biomasse équivalente. Il a été intermédiaire pour les cultures pérennes (56-137 mm an-1) et très fortement lié à la production (elle-même fonction de l’espèce et de la fertilisation azotée). La concentration en nitrate a varié de 2 à 23 mg l-1. Elle a été en général plus faible sous les cultures pérennes, sauf pour le miscanthus lors de la première année de mesure. Les stocks de carbone du sol ont augmenté fortement sous les cultures pluriannuelles (+0.93 t C ha-1 an-1) mais n'ont pas varié significativement pour les autres cultures. Le 15N retrouvé dans la biomasse récoltée a été plus faible pour les cultures pérennes, particulièrement lorsqu’elles sont récoltées tardivement, mais cela est compensé par une plus forte proportion de 15N dans leurs organes souterrains et dans le sol. Le 15N retrouvé dans le système sol-plante a été de 69% de l’azote apporté pour les cultures pérennes, 61% pour les cultures pluriannuelles et 56% pour les cultures annuelles, ce qui suggère que des pertes importantes ont eu lieu par volatilisation et dénitrification. Dans nos conditions pédoclimatiques, les cultures pérennes en C4 sont les plus intéressantes pour concilier forte production de biomasse, forte efficience d’utilisation de l’eau et de l’azote et faibles pertes d’azote vers l’hydrosphère et l’atmosphère. En revanche, seules les cultures pluriannuelles permettent de stocker du carbone à court terme
Second generation biofuels could provide renewable energy to the transportation sector while mitigating climate change. However, their greenhouse gas, energy and environmental balances will probably depend on the feedstock used for their production. Bioenergy crops that could be used for second generation biofuels will have to fulfil several requirements, including high productivity, low input requirements, and low environmental impacts. The objective of this work was to assess the water, N and C balances at the plot scale for various bioenergy crops with different management. The study is based on a long term field experiment, called “Biomass & Environment”, established at the INRA experimental station in Estrées-Mons, northern France. This experiment includes two perennial C4 crops (Miscanthus × giganteus and switchgrass), two semi-perennial forage C3 crops (fescue and alfalfa) and two annual C4/C3 crops (fibre sorghum and triticale). It compares two nitrogen treatments and two dates of harvest of perennial crops: early (October) or late harvest (February). Measurements have been carried out on: i) biomass production; ii) soil water stocks, monitored continuously during 7 years; iii) root depth and density; iv) drainage and nitrate concentration in drained water, assessed from soil water and mineral N content measurements (in mid-autumn and late winter) and using the STICS model; v) soil organic carbon (SOC) stocks in 2006 and 2011-2012; vi) the fate of 15N-labelled fertiliser applied during 4 or 5 successive years.Thanks to their deep rooting system, perennial and semi-perennial crops consumed more water than annual crops. The amount of drained water was lower under semi-perennial than annual crops (64 vs. 133 mm yr-1 average over 7 years), despite an equivalent biomass production. It was intermediate under perennial crops (56-137 mm yr-1) and negatively correlated to biomass production, itself depending on crop species and N rate. Nitrate concentration in drained water varied between 2 and 23 mg l-1. It was generally lower under perennial than other crops, except for miscanthus on the first year of measurement. SOC stocks increased markedly over time under semi-perennial crops (+0.93 t C ha-1 yr-1), whereas no significant change occurred under perennial and annual crops. The 15N recovery in the harvested biomass was lower for perennial than other crops, particularly when harvested late, but compensated by a higher 15N recovery in belowground organs and soil. The overall 15N recovery in the soil-plant system was 69% in perennials, 61% in semi-perennials and 56% in annual crops, suggesting that important fertiliser losses occurred through volatilisation and denitrification. In our pedo-climatic conditions, the C4 perennial crops performed best in terms of production, water and nitrogen use efficiency, and nitrogen losses towards the groundwater and the atmosphere. However, only semi-perennial crops yielded in SOC sequestration

Orientador: Edson Luiz Mendes Coutinho Coutinho
Coorientador: Takashi Muraoka
Banca: Henrique Coutinho Junqueira Franco
Banca: Valdeci Orioli Júnior
Banca: José Eduardo Corá
Banca: Carolina Fernandes
Resumo: Forrageiras do gênero Cynodon são conhecidas por sua capacidade de resposta a altas doses de nitrogênio (N). Em condições tropicais o N pode se tornar um problema ambiental e financeiro devido à sua baixa eficiência de uso pelas plantas, principalmente como resultado de perdas por volatilização e/ou lixiviação. Portanto, objetivou-se com este trabalho avaliar fontes e doses de N no crescimento e produção de matéria seca do Tifton 85, bem como mudanças nas frações do carbono (C) no solo. Um experimento foi conduzido de 2012 a 2014, e constituiu de um fatorial em um delineamento de blocos ao acaso, onde os tratamentos foram a associação de duas fontes de N (nitrato de amônio [NA] e ureia), e cinco doses do nutriente (0, 60, 120, 180 e 240 kg ha-1 N por corte) aplicadas a cada 30 dias. Neste experimento foram utilizados NA e ureia enriquecidos com 15N como uma ferramenta para quantificar a recuperação do N advindo do fertilizante, nas plantas e no sistema solo-planta. As plantas foram cortadas para avaliar a produção de matéria seca, concentração de N na parte aérea e recuperação. O índice de área foliar (LAI), radiação fotossinteticamente ativa interceptada (PARi) e índice de clorofila (CI) foram mensurados no dia anterior ao corte. As amostras de solo foram coletadas nas parcelas que receberam NA e foram fracionadas em carbono orgânico particulado (POC), fração leve livre (FLF) e fração mineral (C-min); O teor de C nas frações foi quantificado. Os resultados demonstraram que o... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: Cynodon hybrids are known by their ability to respond to high rates of nitrogen (N). In tropical conditions, N may become an environmental and financial issue due to its low efficiency of use by plants, mainly as a result of losses by volatilization and/or leaching. Therefore, the aims with this work were to evaluate N sources and rates on growth and dry matter yield of Tifton 85 and to determine changes in soil carbon (C) fractions in response to N fertilization. The 2-yr field experiment was conducted from 2012 to 2014, and consisted of a factorial design. Treatments were a combination of two sources of N (ammonium nitrate [AN] and urea) and five rates of the nutrient (0, 60, 120, 180 and 240 kg ha-1 N per cut) applied broadcast every 30 days. This study used AN and urea enriched with 15N as a tool to quantify the recovery of N derived from fertilizer in plants and the soil-plant system. Forage was cut at 30-d intervals for dry matter yield, shoot N concentration determinations and N recovery. The leaf area index (LAI), photosynthetic active radiation intercepted (PARi) and chlorophyll index (CI) were evaluated on the day before clipping. Soil samples were collected in plots receiving AN and they were fractionated in to particulate organic carbon (POC), free light fraction (FLF) and mineral fraction (C-min); The C concentration of the various fractions was determined. Results showed that AN was a more efficient source only when the amount of precipitation is insufficient to... (Complete abstract click electronic access below)
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Funda??o Carlos Chagas Filho de Amparo ? Pesquisa do Estado do RJ - FAPERJ
Nitrogen is one of the nutrient elements most limiting for plant growth. Thus, increasing plant nitrogen usage efficiency (NUE) is an essential factor for sustainable agriculture, leading to an increased food production with less fertilizer input and less environment impact. NUE in plant involves the uptake and accumulation efficiency, and N use efficiency for grain production. Understanding the mechanisms regulating these processes is crucial for the improvement of NUE in crops. Therefore, is essential to comprehend plants response to different N regimes, mainly N limitation. The usage of plants or varieties with differences in nutrient use efficiency is another important factor to determine the main mechanisms involved in these processes. The objectives of this study were to evaluate N metabolism and expression of genes involved in N uptake and assimilitation of two rice varieties (Piau? and IAC-47), under different N supply conditions. The N metabolism was evaluated by quantification of nitrogen fractions in plant tissue and enzymatic activity, expression of genes that code for high and low affinity N transporters, and N assimilation enzymes. Two experiments were conducted: the first, to investigate the plants behavior when grown under high and low N supply; and the second to verify plants response to NO3 - resupply. The IAC-47 variety showed greatest mass accumulation in all treatments. When plants were grown for a long period under low N supply it was observed a higher nitrate (NO3 -) content in the tissue of Piau? variety, which is adapted to low fertility conditions, as well as a high activity of nitrate reductase and glutamine synthetase in the roots of these plants. When under high NO3 - supply for a long period, Piau? variety had higher NO3 - concentrations in roots and sheaths, but no differences were observed in enzymes activity. The nitrate transporters expression was higher in roots of Piau? plants under low supply of NO3 - when compared to Piau? plants under high N supply. Nitrate transporters expression in roots of IAC-47 plants showed a pattern opposite to the one observed for Piau? plants. When plants were resupplied with NO3 -, after a 72 hours period without N, there was a rapid increase, after 6 hours of resupply, in the expression of high affinity nitrate transporters genes (OsNRT2.1) and the plastidial glutamine synthetase (GS2) in the Piau? variety roots, and a higher expression of these in IAC-47 roots after a 24 hours of N resupply. The efficiency of Piau? variety is related to the differentiated control of N absorption and assimilation mechanisms, which allow bigger expression of high affinity nitrate transporters in tissue with high levels of soluble N, high N status, indicating that the control of nitrogen uptake system may be essential for the nitrogen usage efficient.
O nitrog?nio ? um dos elementos minerais que mais limita o desenvolvimento das plantas. Assim, aumentar a efici?ncia de uso de nitrog?nio (EUN) ? um fator ? essencial para uma agricultura sustent?vel, levando a um aumento da produ??o de alimentos com menor uso de insumos e menos impactos ao ambiente. A EUN envolve a efici?ncia de absor??o e ac?mulo e a efici?ncia de utiliza??o de N para produ??o. Compreender os mecanismos envolvidos nesses processos e como eles s?o controlados ? fundamental para a melhoria da EUN nas plantas cultivadas. Para isso, ? essencial entender a resposta e comportamento das plantas a diferentes regimes de N, principalmente ? limita??o de N. O uso de plantas ou variedades com diferen?as na EUN ? outro fator importante para verificar os principais mecanismos envolvidos. Este trabalho teve por objetivo desse trabalho foi verificar o metabolismo e a express?o de genes envolvidos na absor??o e assimila??o de nitrog?nio nas variedades de arroz Piau? e IAC-47 sob diferentes condi??es de disponibilidade desse nutriente. O metabolismo de N foi avaliado atrav?s da quantifica??o das fra??es nitrogenadas no tecido e atividade enzim?tica, a express?o de genes que codificam para transportadores de N de alta e baixa afinidade e enzimas de assimila??o de N. Foram realizados dois experimentos: o primeiro para verificar o comportamento das plantas quando cultivadas sob alto e baixo suprimento de N; e o segundo, para verificar as repostas ao ressuprimento de N. O ac?mulo de massa foi maior na variedade IAC-47 em todos os tratamentos. Quando as plantas foram cultivadas por um longo per?odo sob baixo suprimento de N foi observado maior teor de nitrato (NO3 -) no tecido da variedade Piau?, adaptada a condi??es de baixa fertilidade, assim como uma alta atividade da nitrato redutase e glutamina sintetase nas ra?zes dessas plantas. Sob alto suprimento de NO3 -, por um longo per?odo, a variedade Piau? apresentou maiores teores de NO3 - nas bainhas e ra?zes, mas n?o foram observas diferen?as na atividade de enzimas. A express?o dos transportadores de nitrato na variedade Piau? foi mais elevada nas ra?zes das plantas sob baixo suprimento de NO3 - quando comparado as plantas sob alto suprimento de N. Na variedade IAC-47 a express?o dos transportadores de NO3 - teve comportamento oposto ao observado para a Piau?. Quando as plantas receberam NO3 -, ap?s um per?odo de 72 horas sem N, houve um r?pido aumento, ap?s 6 horas do ressuprimento, na express?o do gene que codifica para o transportador de NO3 - de alta afinidade OsNRT2.1 e glutamina sintetase plastidial (GS2) nas ra?zes da variedade Piau?, e 24 horas ap?s o ressuprimento a express?o foi maior na variedade IAC-47. A efici?ncia da variedade Piau? esta relacionada ao controle diferenciado dos mecanismos de absor??o e assimila??o de N, que permite a maior express?o dos transportadores de nitrato de alta afinidade em tecido com altos teores de N sol?vel, alto status de N, indicando que o controle do sistema de absor??o de N pode ser essencial para a efici?ncia no uso de nitrog?nio.

Dans un contexte de développement durable, les enzymes sont des outils biologiques puissants pour catalyser des réactions avec de très grandes efficacités et spécificités. Inspirée des enzymes et de la catalyse organométallique, l’élaboration de métalloenzymes artificielles émerge depuis plusieurs années comme une stratégie de choix pour fournir aux chimistes de nouveaux biocatalyseurs, en accord avec les principes de la chimie verte. Elles sont construites par l’insertion par interactions supramoléculaires ou couplage covalent, d’un ion ou d’un complexe métallique au sein d’une protéine, qui leur apporte un environnement hydrophobe protecteur et chiral. Lors de cette thèse, plusieurs métalloenzymes artificielles ont été construites par couplage covalent de complexes métalliques dans deux protéines hôtes, qui sont la Xylanase A (Xln) et les protéines artificielles de la famille des Reps. Dans un premier temps, une hydrogénase artificielle a été construite dans le mutant XlnS212C par ancrage covalent d’un complexe de fer appelé complexe de Knölker. L’hydrogénase artificielle obtenue, XlnS212CK, s’est avérée capable de catalyser l’hydrogénation par transfert d’hydrure de la trifluoroacétophénone, TFAC, sans excès énantiomérique. Dans un second temps, quatre Diels-Alderases artificielles ont été construites à partir de la protéine bidomaine (A3_A3’) de la famille des αReps. Les deux meilleures Diels-Alderases, qui ont conduit respectivement au meilleur rendement et la meilleure énantiosélectivité dans la réaction de l’azachalcone sur le cyclopentadiène, ont été élaborées respectivement par fixation covalente de complexe de cuivre de ligands phénanthroline et terpyridine dans un mutant F119C de A3_A3’ : (A3_A3’)F119Phen-Cu(II) et (A3_A3’)F119Terpy-Cu(II). Finalement, une nouvelle hémoprotéine artificielle a été construite par couplage covalent de la méso-tétraphénylporphyrine de manganèse Mn(III)TPP-NHMal dans le mutant (A3_A3’)Y26C. L’hémoprotéine artificielle formée BH MnTPP seule ne montre aucune activité catalytique pour l’oxydation de co-substrats par H2O2. Cependant, de manière inattendue, l’addition d’imidazole et d’une autre protéine αRep, bA3-2, qui se fixe de manière spécifique sur A3_A3 et provoque son ouverture, permet non seulement de déclencher l’activité peroxydase de BH MnTPP, mais également une activité monooxygénase qui catalyse la sulfoxydation du thioanisole par H2O2. Il s’agit du premier exemple décrit à ce jour de métalloenzyme artificielle dont l’activité peut être induite par la fixation d’une protéine partenaire
In a context of sustainable development, enzymes are powerful biological tools to catalyze reactions with very high efficiencies and specificities. Inspired by enzymes and organometallic catalysis, the development of artificial metalloenzymes has emerged for several years as a strategy of choice to provide the chemists with new biocatalysts, in accordance with the principles of green chemistry. They are constructed by the insertion by supramolecular interactions or covalent coupling of an ion or a metal complex within a protein, which provides them with a protective and chiral hydrophobic environment. In this thesis, several artificial metalloenzymes were constructed by covalent coupling of metal complexes into two host proteins, Xylanase A (Xln) and artificial proteins of the Reps family. Initially, an artificial hydrogenase was constructed in the XlnS212C mutant by covalent anchoring of an iron complex known as the Knölker complex. The artificial hydrogenase obtained, XlnS212CK, was found to be capable of catalyzing hydride hydrogenation of trifluoroacetophenone, TFAC, without enantiomeric excess. In a second time, four artificial Diel-Alderases were constructed from the bidomain protein (A3_A3') of the αReps family. The two best Diels-Alderases which led respectively to the best yield and the best enantioselectivity in the reaction of azachalcone on cyclopentadiene were developed respectively by covalent attachment of copper complex of phenanthroline and terpyridine ligands in a mutant F119C of A3_A3' (A3_A3')F119Phen-Cu (II) and (A3_A3')F119 Terpy-Cu (II). Finally, a new artificial hemoprotein was constructed by covalent coupling of the manganese meso-tetraphenylporphyrin Mn(III)TPP-NHMal in the (A3_A3')Y26C mutant. The artificial hemoprotein formed BH-MnTPP alone shows no catalytic activity for the oxidation of co-substrates by H2O2. However, unexpectedly, the addition of imidazole and another αRep protein, bA3-2, which binds specifically to A3_A3’ and causes it to be opened, not only triggers the BH-MnTPP peroxidase activity but also a monooxygenase activity which catalyzes the sulfoxidation of thioanisole by H2O2. This is the first example described to date of artificial metalloenzyme whose activity can be induced by the attachment of a partner protein

Nitrate is an essential nutrient for plant growth. Nitrate acquisition by roots and its intercellular translocation is mediated by nitrate permeable transport proteins. Nitrate transporters have been extensively studied in the model plant, Arabidopsis thaliana. Nitrate transporters belong to three protein families: NPF (Nitrate Transporter 1/Peptide Transporter), NRT2 (Nitrate Transporter 2) and CLC (Chloride Channel) (Miller et al., 2007; Wang et al., 2012). However, there is little known about how these proteins orchestrate nitrate transport in maize. Four putative nitrate transporter genes (ZmNPF6.4, ZmNPF6.5, ZmNPF6.6, and ZmNPF7.10) were cloned from a maize root cDNA population. Preliminary localization studies using C-terminal YFP-fusions showed maize NPF proteins targeting to the plasma membrane, with the exception of ZmNPF7.10, where targeting could not be resolved. Gene expression studies indicated ZmNPF6.6 was induced strongly in roots by nitrate. Its shoot expression was mostly absent. In contrast, ZmNPF6.4 exhibited a constitutive expression pattern in both root and shoot tissues and was not sensitive to nitrate. Both ZmNPF6.5 and ZmNPF7.10 showed little expression in either root or shoot tissues. Functional characterization studies were conducted on ZmNPF6.4 and ZmNPF6.6 as there was no nitrate transport activity measured with ZmNPF6.5 and ZmNPF7.10 using a preliminary screening experiment in Xenopus laevis oocytes. Combining electrophysiology and chemical flux analysis, ZmNPF6.4 was characterized as a pH-dependent, low-affinity, non-selective nitrate and chloride transporter. On the other hand, ZmNPF6.6 encoded a pH-dependent, dual-affinity, nitrate specific transporter, which was also permeable to chloride in the absence of nitrate. The functional differences between ZmNPF6.4 and ZmNPF6.6 were explored using site-directed mutagenesis experiments. The “affinity switch” Thr101 within the nitrate transceptor, AtNPF6.3, is conserved in ZmNPF6.6 (Thr104) (Liu, 2003). However, mutating ZmNPF6.6:Thr104 to alanine or aspartate (dephosphorylation and phosphorylation mimics, respectively), did not transform the dual-affinity transporter into either a high- or low-affinity monophasic transporter. Instead, both HATS and, predominantly, the LATS activities of ZmNPF6.6 were repressed by both T104A and T104D mutations. The equivalent of the predicted nitrate-binding residue in AtNPF6.3 (His356) was investigated in ZmNPF6.4 and ZmNPF6.6. In ZmNPF6.4, a tyrosine residue (Tyr370) is present instead of a histidine. Replacement of Y370 with histidine (ZmNPF6.4:Y370H) conferred dual-affinity nitrate transport and enhanced nitrate specificity over chloride. However, replacing His362 in ZmNPF6.6 with Tyr362 made the transporter non-functional. A preliminary analysis of the high-affinity nitrate transport system was conducted by functionally characterizing ZmNRT2.1 and ZmNRT3.1A. The plasma membrane targeting of ZmNRT2.1 required the presence of ZmNRT3.1A. This was confirmed using a C-terminal fusion of NRT2.1 with YFP. Signal was only detected in onion epidermal cells that were co-transformed with both ZmNRT2.1 and ZmNRT3.1A. Gene expression analysis identified both a N-starvation induced expression and a nitrate induced expression pattern for ZmNRT2.1. In contrast, ZmNRT3.1A exhibited a constitutive expression in both roots and shoots. When ZmNRT2.1 and ZmNRT3.1A were co-injected into Xenopus laevis oocytes, high-affinity nitrate transport activity was measured. Single injections of either cRNA failed to elicit a nitrate transport phenotype.
Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food & Wine, 2015

碩士
國立中興大學
園藝學系
84
Genotypial difference in nitrate content was found in 28 lettuce cultivars, with the maximum difference cultivar reached 100%. Variation of round tip was found higher than sharp tip type in leafnitrate content. The nitrate concentration of cultivars R8, R13, R43 increased with nitrogen application and rate of enhancement is greater than R14, R24, S25, R37 which identified as low nitrate cultivars. Leaf nitrate contents of 6 cultivars were not found positively correlated with nitrate reductase acticity. Leaf nitrate content and variation in 3mM NH4NO3 solution was lower than 6mM NO3-N treated plants at different time of a day. Six mM nitrogen treatment together with 20% or 50% ammonium-N effectively reduced leaf nitrate content; to the leavels of 24.6%, 33.9% under 50% shading condition with the addition of 20% and 50% of ammonium-N in the culture solution. Fifty percent ammonium-N also increased the dry weight of lettuce by 17%. There were variety difference in nitrogen use efficiency(NUE) with R37 the highest. The NUE decreased with increasing fertilizer application while nitrogen concentration in the culture solution was raised from 2mM to 15mM. R37, R14 were found to be the high NUE and lower nitrate content cultivars.

This thesis tested the hypothesis that nitrogen use efficiency of subirrigated potted chrysanthemum (Chrysanthemum morifolium Ramat.) can be improved by managing the NO3- status of the plant. Replacement of NO3- with water one week prior to flower development was the most effective method of reducing tissue NO3- content and improving the nitrogen usage index, as compared to various combinations of NO3- and Cl-. Shoot or flower dry mass and N content were unaffected and the medium electrical conductivity was reduced. Supplying N in the NH4+ form in combination with NO3- did not affect nitrogen use indices. Reducing N supply improved various indices of nitrogen use efficiency, with a slight loss of quality. Elimination of NO3- prior to flower development and reduction in N supplied are effective strategies for improving nitrogen use efficiency in subirrigated potted chrysanthemums without sacrificing quality.
Cecil Delworth Foundation

Cereal crops supply a major proportion of the world’s food and their production capacity is tightly linked to nitrogen (N) fertiliser use. With on average less than half of the applied N being captured by crops, there is scope and need to improve N uptake in cereals. With nitrate (NO₃⁻) being the main form of N available to cereal crops there has been a significant global research effort to understand plant NO₃⁻ uptake. Despite this, our understanding of how the NO₃⁻ uptake system is regulated remains limited. To advance our understanding of the NO₃⁻ uptake system and its regulation, three knowledge gaps were identified and explored in this thesis. Firstly, there is an identified need to better understand the NO₃⁻ uptake system and the signalling molecules which modulate it. Secondly, with the literature containing alternative approaches to studying NO₃⁻ uptake, there is a need to appreciate how these studies relate to better leverage the existing literature. And finally, with strong transcriptional control governing the NO₃⁻ uptake system, new leads were sought for modulating transcription of NO₃⁻ transporter genes. To explore these knowledge gaps, dwarf maize (Zea mays L. var. Gaspe Flint) was grown hydroponically with either sufficient or limiting NO₃⁻ availability. During the vegetative growth period a subset of plants grown were moved from sufficient to limiting NO₃⁻ conditions and a range of physiological parameters were measured. The results showed: the high affinity NO₃⁻ uptake system (HATS) appears to contribute a major proportion of total NO₃⁻ uptake capacity and responds to N demand at external concentrations where it was previously thought to be saturated; NO₃⁻ itself appears to play a key role in modulating the NO₃⁻ uptake system, and; temporal variation of NRT transcripts are more variable than previously understood. The observed responses to reduction in NO₃⁻ revealed a series of responses leading to a new model for the control of the NO₃⁻ uptake system. Using the same growth system, plants were grown under steady state NO₃⁻ conditions and a starvation and re-supply (primary nitrate response – PNR) response was explored in parallel. The information generated provided data to relate the PNR literature to longer term steady state studies. The ZmNRT2.5 gene was highlighted as an interesting candidate for revealing cis-trans regulatory elements associated with low N responses. To explore this, a combined phylogenomics and co-expressed gene promoter analysis was undertaken. A number of evolutionarily and functionally conserved regions were identified in the ZmNRT2.5 promoter with six regions showing no resemblance to known transcription factor binding sites. These sequences provide a new resource for the discovery of cis-trans regulatory mechanisms associated with the low N expression of ZmNRT2.5. The findings in this thesis have identified key time points for future transcriptome analysis, and revealed putative cis-elements as new leads for discovering novel cis-trans regulatory elements associated with the regulation of NO₃⁻ uptake. Ultimately, further research may lead to the identification of key regulatory genes as candidates for the improvement of N uptake efficiency and overall N use efficiency in cereal crops.
Thesis (Ph.D.) (Research by Publication) -- University of Adelaide, School of Agriculture, Food and Wine, 2015.

The expansion of livestock husbandry and biogas production in large parts of northwestern Germany during the last two decades increased the amount of accruing manure, as well as the demand for maize as fodder crop and substrate for biogas plants. To overcome phosphorus deficiency symptoms during early growth of maize, farmers commonly apply mineral starter fertilizers containing ammonium-nitrogen and phosphorus on top of the usual manure applications required to meet crop nutrient demand. This practice typically leads to overfertilization of N and P and the excess nutrients are then prone to be lost into the environment. Recent developments of agricultural machinery allow for the injection of slurry bands into the soil prior to maize planting. Due to high concentrations of ammonium and phosphorus in the manure band, chemical transformation and translocation of these nutrients is reduced. When the bands are placed near the seeds, even the radicles can access the applied nutrients. Hence, application of mineral starter fertilizers might be obviated. Earlier investigations showed insufficient knowledge of nutrient transformations in manure bands and their consequences on crop growth. To resolve these problems a research project at the University of Applied Sciences Osnabrück was conducted in close cooperation with the local agricultural extension services, machinery producers and farmers. In a series of field trials, broadcasting of liquid manure was compared to injection with and without a nitrification inhibitor in three consecutive growing seasons (2013 to 2015). The trials were conducted in a split-plot design, where all liquid manure treatments were divided in subplots with and without a mineral starter fertilizer. Biomass samplings at eight leaves stage and harvest gave insight into the performance of the treatments. Compared to broadcast application with starter fertilizer, manure injection showed slightly retarded early growth in some trials. However, yields and nitrogen uptake at harvest were similar. When a nitrification inhibitor was added to the injected manure, early growth was not retarded, yields were alike broadcast and injection treatments, but nitrogen uptake was higher in all seasons (on average ~7%). To further investigate nitrogen dynamics and crop growth, another field trial was conducted on a sandy soil close to Osnabrück in 2014 and 2015. Manure injection with and without a nitrification inhibitor was compared to broadcast application with mineral starter fertilizer and an unfertilized control treatment. Plant samplings were taken at regular intervals. Major precipitation events in May and June 2014 led to significant nitrate leaching, especially in the broadcast treatment. Manure injection delayed the nitrification of slurry ammonium and consequently the translocation out of the root zone. Thus, plants in injection treatments could accumulate more nitrogen in their biomass and showed less nitrogen deficiency symptoms. This led to increased yield (+16.5%) and nitrogen uptake (+9.6%) for injection treatment with nitrification inhibitor compared to broadcast treatment. In 2015, low temperatures impaired seminal root growth and phosphorus availability. The mineral starter fertilizer in the broadcast treatment led to better early growth than injected slurry. When a nitrification inhibitor was added to the injected manure, less P deficiency symptoms were observed, and the crop growth was only slightly retarded. Due to the high compensation potential of silage maize, these differences were equalized until harvest. Nevertheless, the mean apparent nitrogen recovery efficiency of both seasons was higher in injection treatments with and without nitrification inhibitor, compared to broadcast with mineral starter fertilizer (48%, 56% and 43%, respectively). To ease the handling of field trial series by decreasing the number of tissue samplings, the use of a handheld sensor was tested during vegetative growth of maize. In the series of field trials with the local extension service, the derived vegetation index showed significant correlations to biomass and nitrogen uptake at eight leaves stage. Measurements of the vegetative growth observed during the nitrogen dynamics trial showed that the sensor needs sufficient leaf area to deliver precise data, but also tends to saturate when maize tassels evolve. The best estimates were found between six and ten leaves. Thus, the sensor can be a valuable tool to reduce numbers of tissue samples and, thus, time and effort needed in fertilization trials. Altogether, these results should encourage farmers to obviate mineral starter fertilizers by using manure injection when cropping maize on sandy soils. The advantages that come along with manure injection based on the present research indicate higher shares of manure nutrients find their way into the plants due to delayed biochemical transformations. These nutrients are consequently not lost into the environment. Nitrification inhibitors have shown a positive effect on crop performance and led to a further reduction of nitrogen losses. However, further knowledge of their decomposition with special regard to the ecological impact of their compounds and metabolites need to be thoroughly evaluated.

Maize is the dominant crop in northwestern Germany and is mostly cultivated on sandy soils. Additionally, due to intensive livestock husbandry and biogas production, large amounts of liquid manures are produced. The current farm practice leads to high N and P surpluses at field level accompanied by environmental pollution, like nitrate leaching, eutrophication of non-agricultural ecosystems, and N2O emissions. The accruing liquid manures are often used for maize fertilization. Thereby, slurries are mainly broadcast applied using trailing hose applicators followed by incorporation into the topsoil. In addition, a mineral N P starter fertilizer (MSF) is band-applied below the seed-corn at planting to overcome the limited nutrient availability during the early growth stages. Using a slurry injection technique below the maize row before planting might serve a substitute for MSF. Addition of a nitrification inhibitor (NI) into the slurry before injection seems to be an option to further decrease N losses. The objectives of this thesis were to compare the current and novel fertilizing strategies with a special focus on soil mineral nitrogen (SMN) dynamics and plant P, zinc (Zn) and manganese (Mn) status. For both issues the effect of adding a NI into the slurry was investigated. To characterize the SMN dynamics after slurry injection an appropriate soil sampling strategy had to be developed. Therefore, three consecutive field trials were conducted. The first testing of the new soil sampling approach was implemented in an existing experiment where the slurry was injected at a depth of 12 cm (upper rim) below the soil surface. The soil profile (75 cm wide) centered below the maize row was sampled using a grid-like approach to a depth of 90 cm. Around the injection zone, soil monoliths (SM) were sampled using a purpose-built soil shovel. Below the SMs and in the interrow space (15 and 30 cm distance to the row) a standardized auger procedure was used. The second experiment aimed to improve the sampling strategy with focus on sample homogenization quality and necessary sample sizes per pooled sample. In the third experiment this improved sampling strategy was validated. Results from the first testing of the sampling procedure showed that the strategy is suitable, although some problems occurred. Especially the high spread in values among the replications caused high coefficients of variation (CV; mostly 40 – 60%). The improvement trial revealed that for the SM, which contains the slurry band, an intensive homogenization is required. In addition, suitable sample sizes (twelve auger samples and six soil monolith samples per pooled sample) have to be collected to obtain reliable SMN values. Following this enhanced sampling strategy in the final validation trial, the spread in values was considerably reduced and resulted in CV values of mostly < 20%. The method can be adapted to other fertilizer placement strategies and further row crops. To compare both fertilizing strategies with respect to the spatial and temporal SMN dynamics as well as to the plant nutrient status two field trials were conducted using pig slurry on sandy soils in 2014 and 2015. Four treatments were tested: unfertilized control, broadcast application + MSF, injection, and injection + NI. Soil samples were taken using the new sampling strategy at several dates during the growing season. Plant samples were simultaneously collected to evaluate the plant P, Zn, and Mn status at different growth stages. In 2014, all fertilized N was displaced from the top soil layer of the broadcast treatment until the 6-leaf stage due to heavy rainfall, while N displacement was significantly smaller after slurry injection. The lateral movement of injected slurry N was negligible. In 2015, almost no displacement of fertilized N out of the top soil layer occurred independently of treatments, due to distinctly lower rainfall. The release of slurry N was delayed following broadcast application and large SMN concentrations were detected in the injection zones until the 10-leaf stage. The addition of a NI resulted in significantly increased NH4-N shares in the injection zone throughout the early growth stages (+ 46% in 2014 and + 12% in 2015 at 6-leaf stage). Thus, in 2014 SMN displacement was delayed, and in 2015 increased SMN concentrations were found around the slurry band, most probably due to lower N losses via denitrification. Furthermore, NI addition significantly increased the nutrient uptake by maize during early growth in both years. With P deficiency due to cold weather conditions in 2015, broadcast application showed higher P uptake until the 6-leaf stage (36 – 58%), while it was lower at the 8- (32%) and 10- (19%) leaf stages compared to slurry injection (+ NI). Zn availability was enhanced during early growth after slurry injection (+ NI) and Zn as well as Mn uptake were higher at harvest. Furthermore, dry matter yields were higher (2014) or equal (2015) compared to broadcast application. The P balances were decreased by 10 – 14 kg P ha-1, while Zn and Mn balances were excessive independent of treatments. The field trials showed that after slurry injection, especially when combined with a NI, the applied nitrogen is located in a soil zone with better spatial availability for plant roots compared to broadcast application. Furthermore, the MSF can be substituted without affecting early growth of maize. In conclusion, slurry injection leads to equal (or even higher) yields and enables farmers in northwestern Germany to reduce the P and N surpluses. This would support several goals concerning sustainable land use: Lower pollution of ground and surface waters, reduced emission of NH3, more efficient use of the limited rock P reserves, and less need of transporting organic manures out of regions with intensive animal husbandry and/or biogas production. However, slurry injection enhances the risk of N2O emissions, which contributes to climate change. Thus, for a final evaluation of the environmental impact a life cycle assessment would be worthwhile.