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Biological nitrogen fixation associated with sugar cane

A recent15N dilution/N balance study confirmed that certain sugar cane varieties are capable of obtaining large contributions of nitrogen from plant-associated N2 fixation. It was estimated that up to 60 to 80% of plant N could be derived from this source, and under good conditions of water and mineral nutrient supply, it may be possible to dispense with N fertilization of these varieties altogether. The recently discovered bacterium,Acetobacter diazotrophicus, apparently responsible for this N2 fixation associated with the plants, has unique physiological properties for a diazotroph, such as tolerance to low pH, and high sugar and salt concentrations, lack of nitrate reductase, and nitrogenase activity which tolerates short-term exposure to ammonium. Furthermore, it also behaves as an endophyte, in that it is unable to infect sugar cane plants unless through damaged tissue or by means of VA mycorrhizae and is propagated via the planting material (stem pieces).

The effect of inoculating endophytic N2-fixing bacteria on micropropagated sugarcane plants

The aim of this work was to evaluate the effect of the inoculation of endophytic N2-fixing bacteria on the development of micropropagated sugarcane plants. The endophytic population of each inoculated species was monitored during the growth period, and biological nitrogen fixation (BNF) contribution of each inoculation treatment was assessed using the 15N-isotope dilution technique. Seven different combinations of inoculum were used, using five endophytic diazotrophic species (Gluconacetobacter diazotrophicus, Herbaspirillum seropedicae, Herbaspirillum rubrisubalbicans, Azospirillum amazonense and Burkholderia sp.), originally isolated from sugarcane plants. The results showed a clear physiological effect on the development of the inoculated plants, resulting in alteration of the dry matter-partitioning pattern and increase on root dry matter as compared to uninoculated plants. Indeed, all inoculated diazotrophic species could be reisolated in high numbers from the rhizomes of the inoculated plants, even 400 days after inoculation (DAI), suggesting the establishment of the inoculated bacteria. However, a negative effect of the mixture of all five species on the survival of plantlets was observed 45 days after inoculation, just after acclimatization. The analysis of the BNF contribution using the 15N-isotope dilution technique showed that inoculation promoted some increase in the BNF contribution to the plant tissues. The best treatment was the mixture of all five strains, followed by the treatment with a mixture of Herbaspirillum spp. The contribution was much lower when the plants were inoculated with a mixture of G. diazotrophicus with A. amazonense and Burkholderia sp. A BNF contribution around 30% of total nitrogen accumulated was observed in micropropagated plants inoculated with the mixture of strains, suggesting that the combination of species in the inocula is the best strategy to improve sugarcane crops dependent on the biological nitrogen fixation process.

The success of BNF in soybean in Brazil

Approximately forty years after commercial cropping of soybean in Brazil began, the total area under this crop has reached over 13 M ha with a mean productivity of 2400 kg ha−1. Soybean varieties introduced from the USA and varieties rescued from early introductions in Brazilian territory were part of the Brazilian soybean-breeding programme which spread the crop from high to low latitudes. Disease-resistance, pest-resistance, tolerance to low fertility soils, as well as production of plants with pods sufficiently high above the ground for efficient mechanical harvesting, were all aims of the programme. Although BNF was not explicitly considered as a trait for selection in the breeding/selection programme, maximisation of biological nitrogen fixation (BNF) was favoured by conducting selection and breeding trials on soils low in N, in which the seeds were inoculated with efficient Bradyrhizobium inoculants but without N fertiliser application. Several efficient imported Bradyrhizobium strains were found to be unable to compete with native soil micro-flora and other previously-introduced Bradyrhizobium strains. Surprisingly, after being in the soil for many years one or two of these strains had become more competitive while maintaining their high BNF capacity. Today, these strains are included amongst the recommended Brazilian inoculants and have promoted significant improvements in grain yields. The breeding of soybeans in conditions that made grain yield highly dependent on BNF, and the continuous attention paid to the selection of Bradyrhizobium strains appropriate for the newly released varieties, have been the main contributors to todays high yields and their great benefit to the Brazilian economy. There seems to be no reason why this ongoing research programme should not serve as an appropriate model to improve BNF inputs to grain legumes in other countries of the world.

The Potential of C4 Perennial Grasses for Developing a Global BIOHEAT Industry

Unprecedented opportunities for biofuel development are occurring as a result of rising fossil fuel prices, the need to reduce greenhouse gases, and growing energy security concerns. An estimated 250 million hectares (ha) of farmland could be utilized globally to develop a bioenergy industry if efficient and economical perennial biomass crops and bioenergy conversion systems are employed. In temperate zones, C4 or warm-season grass research and development efforts have found switchgrass (Panicum virgatum) and Miscanthus capable of producing biomass yields of 10 to 20 oven dried tonnes (ODT)/ha/yr, while in tropical areas Erianthus and napier grass (Pennisetum purpureum) are producing 25 to 35 ODT/ha/yr. The potential to annually produce 100 barrels of oil energy equivalent/ha with a 25:1 energy output to input ratio appears achievable with high-yielding, N-fixing warm-season grasses grown on marginal lands in the tropics. Commercialization of densified herbaceous plant species has been slow because of the relatively high alkali and chlorine contents of the feedstocks, which leads to clinker formation and the fouling of boilers. This challenge can be overcome by improving biomass quality through advances in plant breeding and cultural management to reduce the chlorine, alkali, and silica content and through the use of new combustion technologies. Warm-season grasses can be readily densified provided suitable grinding and densification equipment and pressure are utilized. The major advantages of producing densified warm-season grasses for BIOHEAT include: it is the most efficient strategy to use marginal farmlands in most temperate and tropical climates to collect solar radiation; it has an excellent energy balance; the feedstocks can be used conveniently in a variety of energy applications; and it is relatively environmentally friendly. Densified warm-season grass biofuels are poised to become a major global fuel source because they can meet some heating requirements at less cost than all other alternatives available today.

Yield of micropropagated sugarcane varieties in different soil types following inoculation with diazotrophic bacteria

It is well described that the beneficial interactions between plants and bacteria are genotype and site specific. Brazilian sugarcane varieties can obtain up to 70% of their nitrogen requirement from biological nitrogen fixation (BNF), and this contribution is related to the Brazilian breeding and selection processes, by example of the variety SP70-1143. In this study the effect of two inoculation mixtures containing diazotrophic bacteria in our earlier pot experiment was evaluated with two sugarcane varieties, a known responder, SP70-1143, and a newly selected variety, SP81-3250, to investigate the sugarcane genotype effect and the role of the mixtures. The sugarcane varieties SP70-1143 and SP81-3250 were grown under commercial field conditions at three sites with contrasting soil types: an Alfisol, an Oxisol and an Ultisol that means a low, medium and high natural fertility respectively. The stem yield and BNF contribution in response to bacterial inoculation were influenced by the strain combinations in the inoculum, the plant genotype, and the soil type and nitrogen fertilization, confirming the genetic and environmental influence in PGP-bacteria interactions. Inoculation effects on the BNF contribution and stem yield increased in the variety SP70-1143 grown in the Alfisol without nitrogen fertilization for three consecutive crops, and it was equivalent to the annual nitrogen fertilization. The plants grown in the Oxisol showed small increases in the productivity of the variety SP70-1143, and in the Ultisol the sugarcane plants presented even decreases in the stem productivity due to inoculation with diazotrophic bacteria mixtures. The results demonstrate the feasibility of the inoculation technology using diazotrophic bacteria in micropropagated sugarcane varieties grown in soils with low to medium levels of fertility. In addition, the results also indicated that specific plant – bacteria – environment combinations are needed to harness the full benefits of BNF.

The contribution of biological nitrogen fixation for sustainable agricultural systems in the tropics

Abstract The pressing need to increase food production in the tropics to feed the burgeoning population of the Third World requires that crop yields ha−1 must be increased without prejudicing the resource base for future generations. Biological nitrogen fixation (BNF), especially that associated with legumes, has great potential to contribute to productive and sustainable agricultural systems for the tropics, but more research is required to investigate how biologically fixed N, and the increased BNF contributions resulting from research innovations, can be incorporated into viable agricultural systems to increase crop or pasture yields and to substitute N fertiliser inputs. A majority of the soils of the humid and semi-humid tropics have mineral fractions composed of 1:1 lattice clays or sesquioxides of relatively low capacity to retain nutrients (CEC) and water (WHC). It is the soil organic matter (SOM) which has high CEC (after liming) and WHC, and soils under undisturbed climax vegetation are usually high in organic matter which is responsible for their fertility. The key to the long term fertility of such soils is to maintain their soil organic matter by the preservation of crop residues and the selection of suitable crop rotations or fallows. In this review we examine several types of agricultural systems utilised in the tropics ranging from pastures, ley cropping, zero-till rotations as well as green manuring and discuss the management options that can be adopted to preserve their agricultural productivity through the strategic use of legumes in these systems, and their effects on pasture and crop yields. The introduction of forage legumes into tropical pastures can increase and sustain their productivity, with only modest inputs of lime and P and K fertilisers. Similarly, crop and pasture rotations (ley cropping) maintain SOM and soil fertility and crop yields can benefit greatly from the introduction of pasture legumes into the ley. Continuous cropping under minimum or zero tillage can maintain soil cover, and stimulate the retention of SOM, such that nutrient losses are often minimal, and legume derived N can be efficiently transferred to subsequent crops. The options for the resource-poor small-holder to efficiently utilise biologically fixed N as a N supply for cereal grains are more limited and need more attention from researchers as well as less neglect from government organisations. The addition of lime and P fertiliser in modest quantities in many under-developed regions could make large contributions to increased crop yields. If such modest fertiliser inputs were to be combined with suitable crop rotations including green manure or grain legume crops, larger increases in crops yields could be achieved on a sustainable basis, but in many regions agricultural extension services are non-existent and poor farmers have little access to even these basic chemical inputs.

Biological nitrogen fixation in Azospirillum strain-maize genotype associations as evaluated by the 15N isotope dilution technique

Few studies of the inoculation of cereal crops with N2-fixing bacteria have included more than one or two plant genotypes. In a recent study performed in Argentina using 12 different maize genotypes, it was found in 2 consecutive field experiments that several of them responded consistently, either negatively or positively, to inoculation with a mixture of strains of Azospirillum spp. The present study in post was performed to investigate the effect of inoculation of individual strains (and a mixture) of Azospirillum spp., and their nitrate reductase negative (NR-) mutants, on the growth of four of these maize genotypes. Two of these genotypes were grown in 15N-labelled soil with the aim of quantifying any contributions of biological N2 fixation. Two genotypes (Morgan 318 and Dekalb 4D-70) produced similar increases in grain yield when they were inoculated with a mixture of Azospirillum spp. strains or fertilized with the equivalent of 100 kg N ha-1. The other genotypes (Dekalb 2F-11 and CMS 22) showed little response to inoculation or N fertilization. The Morgan 318 and Dekalb 4D-70 genotypes showed a large increase in total N accumulation, suggesting that the response was due to increased N acquisition, but not due to bacterial nitrate reductase as the NR- mutants generally caused plant responses similar to those of the parent strains. Despite problems with the stabilization of the 15N enrichment in the soil, the 15N isotope dilution results indicated that there were very significant biological nitrogen fixation (BNF) contributions to the Dekalb 4D-70 and CMS 22 maize genotypes.

Carbon turnover (δ13C) and nitrogen mineralization potential of particulate light soil organic matter after rainforest clearing

Abstract Soil samples from under a rainforest, a papaya plantation, a pure Brachiaria humidicola pasture and a B. humidicola/Desmodium ovalifolium sward from the humid tropical Bahia, Brazil, were analyzed for fertility parameters, litter quality, particulate light fraction [ PLF > 100 53 -100 μ m , 100 μm) PLF was of younger age than the finer PLF or whole soil samples. The age of the 53–100 μm PLF was very close to that of stabilized organic matter as indicated by 13C data, its low C-to-N ratio (15–17:1) and its turnover time was about 30 y. The PLF appeared not to consist of one uniform pool and thus may have to be subdivided to be useful for modelling purposes (e.g. active, passive pools and charcoal). The contribution of PLF and above ground litter to total soil N mineralization (anaerobic incubation) was highest in the undisturbed rainforest ( 12 16 % for the PLF/litter, respectively, in the 0–2 cm layer), but much less in the other systems. However, mineralization of PLF from different sources was unpredictably altered by the density agent. Introducing D. ovalifolium into these pastures increased, significantly, the amounts of N in PLF, decreased the C-to-N ratio of PLF and tended to increase, the mineralization potential despite its high lignin and polyphenol content. The combination of δ13C analysis and size/density separations, of surface samples in particular, allowed sensitive detection of changes in soil organic matter dynamics and soil fertility.

Quantification of the contribution of N2 fixation to field-grown grain legumes-a strategy for the practical application of the 15N isotope dilution technique.

Abstract Four experiments were performed at two sites in Brazil to evaluate the use of the 15 N isotope dilution technique for the quantification of N 2 fixation to inoculated and non-inoculated soybean, cowpea and groundnut using three independent control crops in each experiment. The data showed that the 15 N enrichment of the N derived from the soil by the different control crops was different even though slow-release forms of labelled N had been applied. These results illustrate the principal problem associated with the use of the isotope dilution technique in that its application theoretically requires that all test (“N 2 -fixing”) and control crops accumulate N from the soil with the same 15 N enrichment. It was also shown that the three control crops behaved differently at the different sites, so that the idea that “appropriate” controls can be universally recommended for use with any particular legume crop is fallacious. The use of several control crops, in combination with slow-release labelled fertilizer, was shown to be a successful strategy for the application of this technique to quantify the contribution of N 2 fixation to field-grown grain legumes.

Quantification of the contribution of biological nitrogen fixation to tropical green manure crops and the residual benefit to a subsequent maize crop using 15N-isotope techniques.

In this study the contribution of biological N2 fixation (BNF) to leguminous green manures was quantified in the field at different sites with different 15N methodologies. In the first experiment, conducted on a Terra Roxa soil in Cuba, the BNF contribution to three legumes (Crotalaria juncea, Mucuna aterrima and Canavalia ensiformis) was quantified by applying 15N-labelled ammonium sulphate to the soil. The second experiment was planted in a very low fertility sandy soil near Rio de Janeiro, and the 15N natural abundance technique was applied to quantify BNF in C. juncea, M. niveum and soybean. In both studies the advantages of using several non-N2-fixing reference plants was apparent and despite the much greater accumulation of the C. juncea in the experiment performed on the fertile soil of Cuba, the above ground contributions of BNF at both sites were similar (40-80 kg N x ha(-1)) and greater than for the other legumes. In a further experiment the possible contribution of root-derived N to the soil/plant system of two of the legumes was quantified using a 15N-leaf-labelling technique performed in pots. The results of this study suggested that total below-ground N could constitute as much as 39 to 49% of the total N accumulated by the legume crops.