Role and regulation of chemotaxis and motility in the Rhizobium-legume symbiosis

Thesis

Fertilisers are used to provide crop plants with essential nutrients such as nitrogen but with a high energy cost and detrimental environmental effects. The symbiosis of legumes with rhizobia exhibits none of these problems since the bacteria provide fixed nitrogen directly to the plant. Further understanding of the initiation of symbiosis may enable improvements to the farming of crop legumes and enable the same processes to be mimicked in engineered systems of other crops. Although motility and chemotaxis are known to be crucial for competitive plant root colonisation (the first stage of symbiosis), little is known about their importance after this stage. In this thesis, I aimed to describe the importance of chemotaxis and motility in sequential stages of symbiosis up to the generation of root nodules filled with nitrogen-fixing rhizobia. To this end, the chemotaxis and motility systems of the model rhizobia Rhizobium leguminosarum biovar viciae 3841 (Rlv3841) were characterised, in both free-living and plant-associated states.

Rlv3841 motility is driven by flagella rotation modulated by two chemotaxis systems, Che1 and Che2, that respond to signals sensed by chemoreceptors. In this work we found that Rlv3841 has a diverse set of chemoreceptors that enable the bacteria to sense and respond to a variety of chemical gradients, by both externally sensing in the periplasm and internally sensing metabolic state. We also found a novel system that halts the bacteria in response to high concentrations of TCA cycle intermediates independent of the chemotaxis systems. An inorganic phosphate uptake system (PstSCAB) and a putative methyltransferase (encoded by RL3464) were found to be critical to the response, although their downstream effectors were not identified. Finally, we characterised the importance of the chemotaxis and flagella systems of Rlv3841 at sequential stages of the symbiosis. This involved marking bacteria with luminescence, fluorescence and enzymatic activity and assaying the bacteria by optical imaging, flow cytometry and staining. We found that there are several critical stages in which chemotaxis and motility are essential and that these occur after the plant root has been colonised by the rhizobia. In particular, we found that Che1 and flagella are important for movement around the root and to root hair tips and that Che2 is involved in infection thread formation and bacteroid differentiation. Overall, this work improves our understanding of the chemotaxis and motility systems of Rlv3841 and their importance across the whole Rhizobium-legume symbiosis cycle.

Authors: Aroney, S

• DOI: 10.5287/ora-w40qzxa7x
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: Rhizobium leguminosarum; rhizobia; chemotaxis; motility; Rhizobia-legume symbiosis
• Subjects: Microbiology