Plant-associated Biological Nitrogen Fixation Research Articles

Sugarcane Genetic Controls Involved in the Association with Beneficial Endophytic Nitrogen Fixing Bacteria

Sugarcane is an economically important culture in Brazil, being the world most important source of sugar and ethanol production. Brazilian sugarcane culture is able to obtain large and significant contributions of nitrogen from plant-associated Biological Nitrogen Fixation (BNF), reducing the use of nitrogen fertilizers and leading to an increase in energy balance of the culture. Different populations of endophytic N2-fixing bacteria associated with sugarcane were identified. Besides BNF, these bacteria exhibit growth promoting traits by mechanisms involving nutrient solubilization, plant hormone production and pathogens antagonistic activity. BNF contribution efficiency to sugarcane is controlled by plant and bacteria genotypes as well as environmental conditions. Here, there will be reviewed the advances made in studies on molecular, cellular and physiological mechanisms by which sugarcane controls establishment and efficiency of endophytic association. Transcriptome profile analysis indentified several differentially expressed sugarcane genes during early stages of the association. Regulatory networks involved in various plant processes such as plant-microorganism recognition, defense, plant hormone signaling, plant growth and nitrogen metabolism were reported to be responsive to endophytic diazotrophic bacteria colonization. The data indicate that sugarcane actively participates in the association with these bacteria. Comprehension of how various sugarcane regulatory mechanisms are coordinated and connected to genotype and environmental signals, in order to control the establishment of a beneficial and endophytic type of association is still a big challenge. Nevertheless, the knowledge being accumulated may guide studies to improve plant association with endophytic diazotrophic bacteria and, possibly, to extend it to other crops.

Long-term Effects of Pre-harvest Burning and Nitrogen and Vinasse Applications on Yield of Sugar Cane and Soil Carbon and Nitrogen Stocks on a Plantation in Pernambuco, N.E. Brazil

Since the 1970s the area under sugarcane in Brazil has increased from 2 million to over 5 million ha (M ha), and it is expected to pass the 7 M ha mark in 2007. More than half of the cane is harvested to produce bioethanol as a fuel for light vehicles. The distilleries produce approximately 13 L of distillery waste (vinasse) for each litre of ethanol produced. In the 1980s there was considerable concern over the long-term effects of the disposal of this material (containing about 1% carbon and high in K) on cane yields if it was applied to the field. At the same time there was a growing movement to abandon the practice of pre-harvest burning and some research was showing that some Brazilian varieties of sugar cane were able to obtain significant contributions of N from plant-associated biological nitrogen fixation (BNF). For these reasons an experiment was installed on a cane plantation in the state of Pernambuco, NE Brazil to investigate the long-term effects of vinasse and N fertiliser additions and the practice of pre-harvest burning on crop and sugar yield, soil fertility parameters, N balance and soil C stocks. The results showed that over a 16-year period, trash conservation (abandonment of burning) increased cane yields by 25% from a mean of 46 to 58 Mg ha−1. Vinasse applications (80 m3 ha−1 crop−1) increased mean cane and sugar yield by 12 to 13% and the application of 80 kg N ha−1 as urea increased cane yields by 9%, but total sugar yield by less than 6% (from 7.0 to 7.4 Mg ha−1 crop−1). The total N balance for the soil/plant system when only the surface 20 cm of the soil was considered was positive in plots where no N fertiliser was added. However, the data indicated that during the 16 years of the study considerable quantities of soil organic matter were accumulated below 20 cm depth such that the N balance considering the soil to 60 cm depth was strongly positive, except where N fertiliser was added. The data indicated that there were considerable BNF inputs to the system, which was consistent with its low response to N fertiliser and low N fertiliser-use-efficiency. There were no significant effects of vinasse or urea addition, or trash conservation on soil C stocks, although the higher yields proportioned by trash conservation had potentially significant benefits for increased mitigation of CO2 emissions where the main use of the cane was for bioethanol production.

Biological nitrogen fixation associated with tropical pasture grasses

This paper originates from an address at the 8th International Symposium on Nitrogen Fixation with Non-Legumes, Sydney, NSW, December 2000 The semi-humid or humid tropics are ideal for the production of large quantities of biomass from fast-growing C4 grasses, but high yields normally require large quantities of fertiliser, especially N, which has a very high input from fossil fuels (natural gas). A program has been started recently to use elephant grass (Pennisetum purpureum Schum.) to substitute firewood as a fuel and also to make charcoal for iron production. In this case, any large N fertiliser additions would mean that the yield of bio fuel per unit of fossil fuel invested would be detrimentally affected. In this study, we report on the potential for the selection of genotypes of fast-growing C4 tropical grasses of the genera Pennisetum and Brachiaria for their capacity to obtain N inputs from plant-associated biological nitrogen fixation (BNF). Fourteen genotypes each of Brachiaria and Pennisetum were screened for BNF contributions by growing them in 15N-labelled soil. In the case of the Pennisetum, after a suitable cutting height for the crop had been selected, there were large differences in dry matter production, N accumulation and 15N enrichment. The differences in 15N enrichment between genotypes were statistically significant and BNF inputs were estimated as high as 41% of accumulated N. In the study on Brachiaria genotypes, potential inputs of BNF seemed lower. Only one or two genotypes of B. brizantha and B. ruziziensis obtained more then 20% of their N from BNF. The N2-fixing bacteria that were most commonly associated with shoots and roots the Pennisetum genotypes were of the genus Herbaspirillum, but predominantly of a recently described new species. The Brachiaria spp. from three different sites (Rio de Janeiro, Goânia, Bahia) were predominately colonised by Azospirillum spp., most of the isolates being of the species Azospirillum amazonense. Very few Herbaspirilla were isolated from these plants.

Field application of the15N isotope dilution technique for the reliable quantification of plant-associated biological nitrogen fixation

Field application of the15N isotope dilution technique for the reliable quantification of plant-associated biological nitrogen fixation

Calculations and assumptions involved in the use of the ‘A-value’ and 15N isotope dilution techniques for the estimation of the contribution of plant-associated biological N2 fixation

In a paper published recently in this Journal a new equation was presented for the calculation of the proportion of plant nitrogen derived from plant-associated biological nitrogen fixation (BNF) using the ‘A-value’ technique. To apply this technique it is assumed that the specific availability (‘A-value’) of soil N is constant regardless of the quantity of added labelled N fertilizer. The authors stated that it follows from this assumption that the proportion of plant N derived from the 15N-labelled fertilizer (%Ndff) is proportional to the rate of addition of the labelled fertilizer. In this present paper we show that this is not the case, and furthermore data in the literature indicate that the ‘A-value’ of soil N is often not constant at different applied N levels. This technique, in common with the isotope dilution (ID) technique, relies on the assumption that the specific availability of soil N is equal for different plants (both ‘N2-fixing’ and ‘non-N2-fixing’). However, as in addition the assumption is made that the ‘A-value’ of soil N is constant at different rates of applied N, we argue that the ID technique is more reliable, particularly when BNF contributions are small.

Quantification of biological nitrogen fixation associated with sugar cane using a 15N aided nitrogen balance

A nitrogen balance study was performed on four commercial sugar cane cultivars grown in pots containing 64 kg of soil from a sugar cane growing area, with the objective of quantifying possible contributions of biological nitrogen fixation (BNF) to the plants. Distillery (stillage) waste from an alcohol distillery was added to half of the pots, and all treatments were replicated 10 times. The pots were maintained in the field and amended with the equivalent of 80 kg N ha −1 of 15N-labelled urea fertilizer. After 12 months of growth the plants accumulated the equivalent of between 10 and 24% of the total N in the soil and added fertilizer. In the next 9 months of growth, with no more N additions, the plants further accumulated the equivalent of between 8.5 and 19% of the original soil + fertilizer N. The cultivar CB 47–89 accumulated significantly more N than the other cultivars. This difference was greatest in the absence of distillery waste, and in this treatment the 15N enrichment of this cultivar was almost half that of the others, which suggests that there was a contribution of unlabelled N from the air to this cultivar via BNF. After 21 months of growth the N content of the soil and roots was determined on 5 replicates. These analyses revealed that the planted plots lost between 7.5 and 12.5 g of N from soil + fertilizer, but all accumulated more than this in plant tissue. In view of the fact that unplanted plots lost between 10 and 14 g of N during this period, the data suggest that all of the cultivars received some N from associated BNF. In the case of the cultivar CB 47–89, in the absence of distillary waste the plants removed approx. 10 g of N from the soil and fertilizer but accumulated almost 35 g of N. These data constitute the first direct evidence of very significant contributions of plant-associated BNF to a sugar cane cultivar although further experiments must be performed to determine whether such large contributions would be encountered under field conditions.