Understanding and Exploiting Biological Nitrogen Fixation for improvement of Brazilian Agriculture

After water, available nitrogen is the most limiting nutrient for plant growth. Although the demand for fixed nitrogen can be met by the use of chemical fertilisers, their use in agriculture has resulted in major environmental and economic impacts. At present less than half of the nitrogen used by farmers is assimilated by crops. Excess nitrogen leaks into the environment, leading to significant effects on soil and water quality, biodiversity, and atmospheric pollution. However, some bacteria can perform a process known as biological nitrogen fixation, which converts nitrogen gas from the atmosphere into ammonia, a source of nitrogen that can be assimilated by animals and plants. This provides a more sustainable alternative to the use of synthetic fertilisers.

Although soil-dwelling nitrogen fixing bacteria can associate with food and energy crops, they do not readily release their fixed nitrogen for the benefit of the plant, although they can have beneficial effects on plant growth. In Brazil, significant increases in crop yield have been observed in response to inoculation with nitrogen fixing bacteria, some of which have an endophytic lifestyle and can gain access to intercellular spaces in plant roots and shoots. This provides an opportunity to engineer closer associations between nitrogen-fixing bacteria and food and energy crops, in which fixed nitrogen is delivered more effectively to the plant. This proposal for a Virtual Joint Centre (VJC) brings together a team of Brazilian and UK investigators focused towards understanding and exploiting plant-diazotroph interactions as a means to enhance agricultural productivity in Brazil. Using fundamental knowledge of nitrogen regulation in endophytic nitrogen fixing bacteria, we will engineer strains, which excrete ammonium to benefit crop growth, and examine the potential of these modified strains as inoculants. We will also design selection strategies to isolate associative diazotrophs that are competitive in the rhizosphere in relation to plant varieties that respond well and have a growth advantage in the presence of these endophytes. We will identify genes required for efficient plant-microbe interactions and quantitate the level of biological, nitrogen fixation in relation to crop yield. Finally, we will we examine the potential for the use of native and engineered strains as inoculants. To achieve these objectives, we will exploit strong synergies and world-leading expertise of UK and Brazilian researchers in biological nitrogen fixation, microbial population dynamics, bacterial and plant genetics, genomics and synthetic biology, in order to engineer efficient cultivar-endophyte combinations and develop improved inoculant technologies. Enhancing nitrogen fixation in our target crops promises substantial benefits for Brazilian agriculture, while decreasing the use and environmental impact of industrial nitrogen fertilisers.

Although we are only midway through the grant period, we have made considerable progress on each of the work packages. Diazotrophic bacteria of importance to Brazilian agriculture have been manipulated to excrete ammonia and a rhizopine biosensor has been designed to enable plant-inducible nitrogen fixation in Azospirillum brasilense. Field trails have been initiated with Pennisetum and sugar cane to assess the influence of inoculation with diazotrophic bacteria on crop yield. We are also evaluating a recently isolated diazotrophic endophyte, Azoarcus olearius DQS-4, for plant growth promotion and the role of nitrogen fixation. Preliminary analysis of RNA-seq data on sugar cane, suggests that plant nitrogen metabolism is reprogrammed when beneficial plant-microbe associations are established.