Competition, Nodule Occupancy, and Persistence of Inoculant Strains: Key Factors in the Rhizobium-Legume Symbioses

Biological nitrogen fixation by Rhizobium-legume symbioses represents an environmentally friendly and inexpensive alternative to the use of chemical nitrogen fertilizers in legume crops. Rhizobial inoculants, applied frequently as biofertilizers, play an important role in sustainable agriculture. However, inoculants often fail to compete for nodule occupancy against native rhizobia with inferior nitrogen-fixing abilities, resulting in low yields. Strains with excellent performance under controlled conditions are typically selected as inoculants, but the rates of nodule occupancy compared to native strains are rarely investigated. Lack of persistence in the field after agricultural cycles, usually due to the transfer of symbiotic genes from the inoculant strain to naturalized populations, also limits the suitability of commercial inoculants. When rhizobial inoculants are based on native strains with a high nitrogen fixation ability, they often have superior performance in the field due to their genetic adaptations to the local environment. Therefore, knowledge from laboratory studies assessing competition and understanding how diverse strains of rhizobia behave, together with assays done under field conditions, may allow us to exploit the effectiveness of native populations selected as elite strains and to breed specific host cultivar-rhizobial strain combinations. Here, we review current knowledge at the molecular level on competition for nodulation and the advances in molecular tools for assessing competitiveness. We then describe ongoing approaches for inoculant development based on native strains and emphasize future perspectives and applications using a multidisciplinary approach to ensure optimal performance of both symbiotic partners.

Stimulation of Soybean (Glycine max) growth and yield using Bradyrhizobium inoculants in the semi-arid environment of Botswana

Background: Crop yields in the semi-arid regions are low due to climatic and soil related constraints.Soybean as one of the most important legume crops grown worldwide, has a role to contribute nitrogen to improve nutrient poor soils in Africa. A study was conducted to examine the effects of Bradyrhizobium spp inoculations on the growth and yield of soybean varieties in a glasshouse.Method: The study was arranged in a randomized complete block factorial design, with factor A being two soybean varieties (Bimha and Status) while factor B was inoculation using four Bradyrhizobium strains and the uninoculated control. Results: Bradyrhizobium inoculation significantly (P less than 0.001)affected days to 50% flowering, days to emergence, nodule number, root dry weight and grain yield and yield traits. Parameters that were affected by both inoculant strain and variety included days to 50% flowering, days to emergence, number of pods per plant, pod weight and number of seeds per pod. The interaction effect of variety and Bradyrhizobium inoculant strain was observed only on number of pods per plants. Our study shows that soybean grows well when inoculated with Bradyrhizobium inoculants, in semi-arid conditions of Botswana.

Evolution of diverse effective N2-fixing microsymbionts of Cicer arietinum following horizontal transfer of the Mesorhizobium ciceri CC1192 symbiosis integrative and conjugative element

Rhizobia are soil bacteria capable of forming N2-fixing symbioses with legumes, with highly effective strains often selected in agriculture as inoculants to maximize symbiotic N2 fixation. When rhizobia in the genus Mesorhizobium have been introduced with exotic legumes into farming systems, horizontal transfer of symbiosis Integrative and Conjugative Elements (ICEs) from the inoculant strain to soil bacteria has resulted in the evolution of ineffective N2-fixing rhizobia that are competitive for nodulation with the target legume. In Australia, Cicer arietinum (chickpea) has been inoculated since the 1970’s with Mesorhizobium ciceri sv. ciceri CC1192, a highly effective strain from Israel. Although the full genome sequence of this organism is available, little is known about the mobility of its symbiosis genes and the diversity of cultivated C. arietinum-nodulating organisms. Here, we show the CC1192 genome harbors a 419-kb symbiosis ICE (ICEMcSym1192) and a 648-kb repABC-type plasmid pMC1192 carrying putative fix genes. We sequenced the genomes of 11 C. arietinum nodule isolates from a field site exclusively inoculated with CC1192 and showed they were diverse unrelated Mesorhizobium carrying ICEMcSym1192, indicating they had acquired the ICE by environmental transfer. No exconjugants harboured pMc1192 and the plasmid was not essential for N2 fixation in CC1192. Laboratory conjugation experiments confirmed ICEMcSym1192 is mobile, integrating site-specifically within the 3’ end of one of the four ser-tRNA genes in the R7ANS recipient genome. Strikingly, all ICEMcSym1192 exconjugants were as efficient at fixing N2 with C. arietinum as CC1192, demonstrating ICE transfer does not necessarily yield ineffective microsymbionts as previously observed.Importance Symbiotic N2 fixation is a key component of sustainable agriculture and in many parts of the world legumes are inoculated with highly efficient strains of rhizobia to maximise fixed N2 inputs into farming systems. Symbiosis genes for Mesorhizobium spp. are often encoded chromosomally within mobile gene clusters called Integrative and Conjugative Elements or ICEs. In Australia, where all agricultural legumes and their rhizobia are exotic, horizontal transfer of ICEs from inoculant Mesorhizobium strains to native rhizobia has led to the evolution of inefficient strains that outcompete the original inoculant, with the potential to render it ineffective. However, the commercial inoculant strain for Cicer arietinum (chickpea), M. ciceri CC1192, has a mobile symbiosis ICE (ICEMcSym1192) which can support high rates of N2 fixation following either environmental or laboratory transfer into diverse Mesorhizobium backgrounds, demonstrating ICE transfer does not necessarily yield ineffective microsymbionts as previously observed.

Draft Genome Sequence of Sinorhizobium meliloti AK555

The inoculation of legume seeds with Sinorhizobium bacteria significantly improves pasture production. Here, we report the draft genome sequence of symbiotically efficient and salt-tolerant Sinorhizobium meliloti inoculant strain AK555, which substantially increases biomass yield of a number of Medicago sativa subsp.

Increased lucerne nodulation in acid soils with Sinorhizobium meliloti and lucerne tolerant to low pH and high aluminium

Acidic conditions with damaging levels of available aluminium (Al3+) currently limit lucerne (Medicago sativa) production on soils in the New Zealand high country and in large areas of Australia. Increased lucerne nodulation could be achieved by using an Al3+-tolerant strain of Sinorhizobium meliloti to inoculate an Al3+-tolerant lucerne line. The Al3+ tolerance of the current commercial Australasian inoculant strain for lucerne, S. meliloti RRI128, was compared with strain SRDI736, selected in Australia for tolerance to low pH. Four Al3+ levels (0, 2, 4 and 8 µm) were created at pH 5.1 in a hydroponic system. The rhizobial strains were evaluated on SARDI AT7, a lucerne line selected for improved growth and nodulation in acidic solution culture with Al3+, and on Stamina 5, a commercial cultivar commonly grown in Australasia. SARDI AT7 when inoculated with strain SRDI736 produced more nodules per plant (3.6 vs 2.4) and had higher nodulation percentage (>80% vs <50%) at all Al3+ levels than when inoculated with RRI128. The percentage of nodulated Stamina 5 plants after inoculation with the commercial strain was lower than after inoculation with strain SRDI736 (10–16% vs 25–70%) at all Al3+ levels. The potential of S. meliloti strains SRDI736, SRDI672 and RRI128 and rates of lime to increase lucerne nodulation and dry matter production in soils of low pH (<5.5, in water) and high Al3+ (>3 mg kg–1 soil) was also investigated in a pot trial. Lime had a more consistent effect than inoculant strain for increasing nodulation. At 0.5 and 2 t lime ha–1, plants inoculated with strains SRDI672 and SRDI736 had more nodules per plant than plants inoculated with the commercial strain. At 4 t lime ha–1, the number of nodules per plant was highest for all three inoculants, and there were no differences among them. This confirms the importance of lime to increase lucerne nodulation in low-pH, high-Al3+ soils. However, where liming is uneconomic or impractical, the results show that it was possible to select rhizobial strains to increase lucerne nodulation in acidic soils with damaging levels of available Al3+.