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Characterisation of rhizobia for the new annual pasture legume Scorpiurus muricatus targeted for medium-to-low rainfall areas of southern Australia

Burns, Kit Alexander (2019) Characterisation of rhizobia for the new annual pasture legume Scorpiurus muricatus targeted for medium-to-low rainfall areas of southern Australia. Honours thesis, Murdoch University.

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Legumes play an integral role in increasing agricultural productivity, particularly in low input agricultural systems in Australia, due to their ability to form symbiotic interactions with a group of soil bacteria called rhizobia. However, in medium-to-low rainfall areas of southern Australia, there is a lack of suitable annual pasture legumes, which is limiting agricultural productivity and profitability in these farming systems. Scorpiurus muricatus is an annual legume from the Mediterranean which possesses high nutritive value and palatability for livestock, is high yielding, capable of self-seeding and is well-adapted to hot and dry summers. As such, S. muricatus is currently being evaluated as a new pasture legume for southern Australia. Crucial to the success of introducing this legume will be the availability of a highly effective rhizobial inoculant strain. This thesis therefore sought to characterise the phylogeny, free-living and symbiotic phenotype of a range of bacteria isolated from Scorpiurus spp.

A total of 19 strains were investigated, with 16s rRNA sequencing demonstrating that 18 of these strains belonged to the genus Mesorhizobium, with the remaining strain (WSM1184) most closely related to Agrobacterium tumefaciens. Analysis of nifH and nodC symbiosis genes further showed that the characterised Mesorhizobium strains generally shared highly similar sequences for these loci, indicating a comparatively high degree of genetic similarity. In particular, WSM1343 (isolated from Scorpiurus sulcatus growing in Morocco) and WSM1386 (isolated from S. sulcatus in Manjimup, Western Australia) were shown to share highly similar symbiosis genes, but divergent 16S rRNA genes, suggesting the possibility that these strains may contain symbiosis genes on mobile Integrative and Conjugative Elements (ICEs).

While the temperature tolerance and apparent optimum growth temperature of the test strains of 28°C was consistent with that commonly reported for Mesorhizobium spp., their growth rate was atypical for this genus, with 15 of the 18 strains having a growth rate on YMA at 28°C slower than that generally described for Mesorhizobium. This slower growth rate may be a common feature of rhizobia from S. muricatus nodules and therefore should be considered when isolating organisms from this legume.

Symbiotic effectiveness experiments showed all Mesorhizobium strains nodulated S. muricatus and fixed N2 on this host, with the most effective strain producing 67.5% of the mean shoot dry weight of the N-fed control plants. Host range experiments demonstrated a subset of the Mesorhizobium strains nodulate existing Australian commercial pasture legumes Biserrula pelecinus and Lotus corniculatus, with the effectiveness data suggesting these strains fix N2 poorly on both hosts. In contrast, none of the strains tested were able to nodulate the grain legume Cicer arietinum.

While this thesis has characterised the phylogeny, free-living and symbiotic phenotype of a range of S. muricatus microsymbionts, further work is required before a suitable commercial inoculant strain can be recommended for this pasture legume. First, all the strains tested in this thesis were isolated from S. sulcatus plants or soils with Scorpiurus spp. present, rather than S. muricatus and it is not known whether strains from either species are cross-compatible for effective N2 fixation. Future studies may therefore locate more effective N2-fixing rhizobia for S. muricatus by isolating microsymbionts from this host in the field. Second, experiments testing the ability of commercial inoculants for already-established pasture legumes B. pelecinus (WSM1497), Lotus sp. (SU343, CC829) and the grain legume C. arietinum (CC1192) to nodulate and fix N2 on S. muricatus need to be conducted to determine whether these inoculants will interact with this legume. Finally, the data strongly suggest that S. muricatus-nodulating Mesorhizobium spp. may contain symbiosis genes on mobile symbiosis ICEs. Given that the phenomenon of ICE transfer has led to the evolution of poorly effective microsymbionts for B. pelecinus, it is imperative that these S. muricatus strains be interrogated for the presence and transfer of symbiosis ICEs, in order to manage this mobility in any future commercial inoculant strain that is released for this pasture legume species.

Item Type: Thesis (Honours)
Murdoch Affiliation(s): School of Veterinary and Life Sciences
United Nations SDGs: Goal 12: Responsible Consumption and Production
Supervisor(s): Terpolilli, Jason, O'Hara, Graham and Yates, Ron
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