Rapid in situ evolution of nodulating strains for Biserrula pelecinus L. through lateral transfer of a symbiosis island from the original mesorhizobial inoculant

Diverse rhizobia able to nodulate Biserrula pelecinus evolved following in situ transfer of nodA and nifH from an inoculant to soil bacteria. Transfer of these chromosomal genes and the presence of an identical integrase gene adjacent to a Phe tRNA gene in both the inoculant and recipients indicate that there was lateral transfer of a symbiosis island

Mining the sequence data of Rhizobium Leguminosarum BV Trifolii WSM1325 and WM2304

Most clover rhizobial inoculants form effective nitrogen-fixing symbioses with either annual or perennial species (and very few with both). This constraint provides a considerable barrier to agricultural productivity since background populations of R. trifolii may nodulate with an incompatible host but ineffectively fix nitrogen (Yates et al 2008)

Complete genome sequence of the Medicago microsymbiont Ensifer (Sinorhizobium) medicae strain WSM419

Ensifer (Sinorhizobium) medicae is an effective nitrogen fixing microsymbiont of a diverse range of annual Medicago (medic) species. Strain WSM419 is an aerobic, motile, non-spore forming, Gram-negative rod isolated from a M. murex root nodule collected in Sardinia, Italy in 1981. WSM419 was manufactured commercially in Australia as an inoculant for annual medics during 1985 to 1993 due to its nitrogen fixation, saprophytic competence and acid tolerance properties. Here we describe the basic features of this organism, together with the complete genome sequence, and annotation. This is the first report of a complete genome se-quence for a microsymbiont of the group of annual medic species adapted to acid soils. We reveal that its genome size is 6,817,576 bp encoding 6,518 protein-coding genes and 81 RNA only encoding genes. The genome contains a chromosome of size 3,781,904 bp and 3 plasmids of size 1,570,951 bp, 1,245,408 bp and 219,313 bp. The smallest plasmid is a fea-ture unique to this medic microsymbiont

Discerning the roles and relationships of EPS and the stringent response

Exopolysaccharides (EPS) represent an energy expensive composition of high molecular weight sugar polymers. Many physiological roles have been associated with EPS including pathogenesis, biofilm maturation, stress tolerance and symbiotic performance. Coincidentally, many of these functions are paralleled by the stringent response, a phase defined by a metabolic downshift in response to nutrient-limiting conditions. In many rhizobia, the synthesis of EPS and specific stationary phase regulators are required for the successful symbiosis with their legume host. Considerable research has examined the genetic and physical basis by which EPS is produced to initiate plant root infection, however, the physiological conditions that affect and coordinate these genes is not dearly understood. This work aimed to define the roles of EPS and the relationship to stationary phase in Sinorhizobiurn medicae whilst determining specific nutritional elements that affect the level of EPS synthesis

Mesorhizobium australicum sp. nov. and Mesorhizobium opportunistum sp. nov., isolated from Biserrula pelecinus L. in Australia

Biserrula pelecinus L. is a pasture legume that was introduced to Australia from the Mediterranean basin in 1993. Although the native rhizobial population could not nodulate B. pelecinus at the time of its introduction, recent research has shown the emergence of a diversity of strains (novel isolates) that are able to do so. Three novel isolates, WSM2073T, WSM2074 and WSM2076, had nearly identical 16S rRNA gene sequences, and clustered separately with all recognized species of the genus Mesorhizobium. Conversely, the novel isolate WSM2075T had >23 nt mismatches with the above three isolates. All four novel isolates shared 97-99% 16S rRNA gene sequence similarity with the type strains of all recognized Mesorhizobium species. However, strains WSM2073T, WSM2074 and WSM2076 showed <95.2% dnaK gene sequence similarity to the type strains of recognized Mesorhizobium species, and <92.9% to WSM2075T (which also shared <95.5% dnaK gene sequence similarity to the type strains of recognized Mesorhizobium species). Results for GSII gene sequencing were consistent with those for the dnaK gene. The fatty acid profiles of the novel isolates were diagnostic of root-nodule bacteria, but did not match those of recognized bacterial species. Strain WSM2075T had a significantly different fatty acid profile from the other three isolates. The above results indicated that strains WSM2073T, WSM2074 and WSM2076 represent the same species. Strain WSM2073T showed <45% DNA-DNA relatedness and WSM2075T <50% DNA-DNA relatedness with the type strains of recognized Mesorhizobium species; these two novel isolates shared 59% DNA-DNA relatedness. Collectively, these data indicate that strains WSM2073T, WSM2074 and WSM2076, and strain WSM2075T belong to two novel species of the genus Mesorhizobium, for which the names Mesorhizobium australicum sp. nov. and Mesorhizobium opportunistum sp. nov. are proposed, respectively. The type strain of Mesorhizobium australicum sp. nov. is WSM2073T (=LMG 24608T=HAMBI 3006T) and the type strain of Mesorhizobium opportunistum sp. nov. is WSM2075T (=LMG 24607T=HAMBI 3007T)

The model legume Medicago truncatula A17 is poorly matched for N2fixation with the sequenced microsymbiont Sinorhizobium meliloti 1021

Medicago truncatula (barrel medic) A17 is currently being sequenced as a model legume, complementing the sequenced root nodule bacterial strain Sinorhizobium meliloti 1021 (Sm1021). In this study, the effectiveness of the Sm1021-M. truncatula symbiosis at fixing N2 was evaluated. • N2 fixation effectiveness was examined with eight Medicago species and three accessions of M. truncatula with Sm1021 and two other Sinorhizobium strains. Plant shoot dry weights, plant nitrogen content and nodule distribution, morphology and number were analysed. • Compared with nitrogen-fed controls, Sm1021 was ineffective or partially effective on all hosts tested (excluding M. sativa), as measured by reduced dry weights and shoot N content. Against an effective strain, Sm1021 on M. truncatula accessions produced more nodules, which were small, pale, more widely distributed on the root system and with fewer infected cells. • The Sm1021-M. truncatula symbiosis is poorly matched for N2 fixation and the strain could possess broader N2 fixation deficiencies. A possible origin for this reduction in effectiveness is discussed. An alternative sequenced strain, effective at N2 fixation on M. truncatula A17, is Sinorhizobium medicae WSM419

Characterisation of root-nodule bacteria isolated from perennial Southern African species of Lotononis

Lotononis is a genus of approximately 150 shrubs, herbaceous perennials and annuals belonging to the subfamily Fabaceae (Van Wyk, 1991). They are distributed mainly in southern Africa, with some species extending throughout Africa, southern Spain, Turkey, south-eastern Bulgaria and part of the Arabian Peninsula to the north-west of the Indian sub-continent (Van Wyk, 1991). Lotononis species have shown potential as perennial pasture legumes that can be used to help reduce the risk of dryland salinity in southern Australian agricultural systems. Species in the section Listia in particular may be useful as pasture legumes as they are perennial, stoloniferous, and lack the poisonous cyanogenic or alkaloid compounds found in some species of Lotononis. L. bainesii, from the Listia section, has been shown to grow well in southern Australia (Roberts & Carbon, 1969) and will grow on acid, sandy soils (R. Yates, pers. comm.). L. bainesii is nodulated by pink-pigmented root-nodule bacteria. Jaftha et al. (2002) characterised nine L. bainesii isolates and found them to be related to Methylobacterium. The genus Methylobacterium, often referred to as pink-pigmented facultative methylotrophs (PPFMs), are capable of growth on C1 compounds such as formate and methanol as sole carbon sources. PPFMs are ubiquitous in the plant phyllosphere and rhizosphere, where they utilize methanol and other C1 compounds that are the products of plant metabolism (Trotsenko et al, 2001). They can promote the germination or the growth of plants, probably because of their ability to synthesise auxins, cytokinins and other plant growth promoting substances (Holland & Polacco, 1994; Ivanova et al., 2000; Trotsenko et al., 2001). However, until the paper by Sy et al. (2001), describing Methylobacterium nodulans, which was isolated from nodules of Crotalaria species found in Senegal, no Methylobacterium species had been known to nodulate legumes, or indeed to fix nitrogen. The objectives in this study were to characterise root-nodule bacteria isolated from four species from the Listia section of Lotononis (L. angolensis, L. bainesii, L. listii and L. solitudinis) using a range of phenotypic and genetic techniques