| Summary: | <p>Some soil-dwelling bacteria in the genus Rhizobium can form symbioses with leguminous plants, including important crop species such as peas and beans. During symbiosis, rhizobial bacteroids inhabiting root nodules fix atmospheric dinitrogen into ammonium, which is exchanged with the host plant for fixed carbon from photosynthesis. Inoculation of leguminous crops with highly effective rhizobia can significantly increase crop yields and reduce inorganic fertiliser usage. However, potential yield benefits are reduced or lost if inoculant strains are unable to compete with indigenous rhizobia. </p>
<p>High-throughput transposon insertion sequencing (INSeq) techniques were used to assess genes for importance in different stages of symbiosis, including poorly understood genetic factors influencing competitive nodulation ability and development of nitrogen-fixing nodules. INSeq classifications were used as starting points for experimental investigation of mutations found to affect abundance of rhizobia in nodules, including mutations which have both positive and negative effects on fitness. Previous results have revealed that timing of infective and developmental events is likely crucial to nodulation ability, hence a novel mutant screening strategy, aiming to identify mutants delayed in these events, was developed using Lux- and GFP-based biosensors then tested in a combined growth and non-invasive imaging system. </p>
<p>Rhizobial mutant strains were generated for 47 genes of interest using established and novel mutagenesis systems. Phenotypes were found for 17 of 35 mutants examined, including seven mutants with significantly impaired competitive nodulation ability, two mutants (ΔRL0339 & ΔRL3565) with enhanced competitive nodulation ability and one mutant (ΔRL3624) showing enhanced growth promotion under nitrogen-limited conditions. The results in this thesis provided the basis for further investigation of nodulation kinetics and characterisation of genes which may be utilised for future development of more effective inoculant strains.</p>
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