Mesorhizobium Strains Research Articles

Mesorhizobia strain and chickpea variety drive phenotypic plasticity of plant growth and nodulation

Chickpea (Cicer arietinum) establishes symbiotic relationships with several Mesorhizobium species and the three-way interaction between chickpea variety, Mesorhizobium strain, and environment, drives plant growth and nitrogen fixation. Here we quantified the phenotypic plasticity for shoot dry weight, nodule dry weight, nodules per plant, nodule colour, symbiotic effectiveness, and nitrogen cost in a factorial experiment combining five chickpea varieties, seven Mesorhizobium strains and three photothermal regimes. Plant growth and nitrogen fixation traits varied with variety, Mesorhizobium strain, photothermal environment and their interaction. Phenotypic plasticity was larger for nodules per plant (7.3-fold) than for shoot dry weight (2.7-fold), verifying a hierarchy of plasticities between these traits. Strain-driven plasticity of plant growth and nitrogen fixation traits was larger than variety-driven plasticity for our combination of varieties, strains, and photothermal environments, with strain-driven phenotypic plasticity being 2.7-fold vs 1.4-fold for shoot dry matter, 2.5-fold vs 1.7-fold for nodule dry weight, 7.3-fold vs 2.1-fold for nodules per plant, 3.7-fold vs 1.7-fold for nodule color, 2.9-fold vs 1.6-fold for symbiotic effectiveness, and 2.3-fold vs 1.6-fold for nitrogen cost. Our study provides insights on the phenotypic plasticity of the legume-rhizobia interaction by considering the plants as part of the rhizobia environment and vice-versa.

Evolution of novel Mesorhizobium genospecies that competitively and effectively nodulate Cicer arietinum following inoculation with the Australian commercial inoculant strain M. ciceri CC1192

Abstract Background and aims Mesorhizobium ciceri CC1192 is the commercial inoculant strain for Cicer arietinum (chickpea) cultivation in Australia, including in the Ord River Irrigation Area (ORIA), where C. arietinum cropping began in 1985. Mesorhizobium strains are known to gain the capacity to nodulate legumes through acquisition of symbiosis Integrative and Conjugative Elements (ICEs), leading to the evolution of novel rhizobia. Here, we assess the impact of symbiosis ICE transfer and compare the genomic diversity and symbiotic effectiveness of C. arietinum nodulating rhizobia from the ORIA. Methods Nodule isolates collected from field cultivated C. arietinum were genotyped by RAPD-PCR, and representative strains from each genotype were whole genome sequenced and symbiotically phenotyped in glasshouse conditions to assess N2 fixation effectiveness against CC1192. Results A total of 68 nodule isolates, all harbouring the CC1192 symbiosis ICE (ICEMcSym1192), were analysed, with 12 identified as the inoculant strain, and 56 novel strains clustering into ten genotypes. These novel genotypes dominated as nodule occupants across the majority of sites sampled. Nine of the ten representative strains were as effective at N2 fixation as CC1192, with WSM4904 the only ineffective strain. Core genome phylogeny showed the ten strains represent four novel Mesorhizobium genospecies. Novel strains WSM4904 and WSM4906 shared 98.7% sequence identity, yet exhibited very different symbiotic phenotypes. Conclusions The CC1192 symbiosis ICE has transferred to a wide diversity of Mesorhizobium spp. in the ORIA. These evolved strains are competitive against CC1192 at nodulating C. arietinum, and the majority are effective symbiotic N2 fixers.

Chickpea (Cicer arietinum L.) growth, nodulation, and yield as affected by varieties, Mesorhizobium strains, and NPSB fertilizer in Southern Ethiopia.

A significant legume crop in Ethiopia, chickpeas (Cicer arietinum L.) have several advantages, including high nutritional value and the capacity to improve soils deficient in nitrogen through biological nitrogen fixation using several endosymbiotic Mesorhizobium spp. strains. However, the host variety, the soil's capacity to hold nutrients, and the endosymbiont's innate physiological traits all affect how efficient the strains are. The primary obstacles to its cultivation in the research area are inadequate agronomic methods and low soil fertility [low nitrogen (N), low soil organic matter (OM), low accessible phosphorous (P), sulfur (S), and boron (B)], which results in ineffective nodulation. To evaluate the effects of NPSB fertilization and inoculation, a field experiment was carried out in Buchach Kebele's Cheha area during the primary cropping season of 2021/22. The trial included two chickpea kinds (Local and Arerti), two NPSB levels (zero and 121 kg NPSB ha-1), and four levels of Mesorhizobium strains (CP-M41, CP-EAL 029, CP-M20b, and un-inoculated control). A randomized complete block design (RCBD) was used to organize the treatments in a factorial form with three replications. In comparison to the single application and the control, the interaction impact of strains, NPSB fertilizer, and variety greatly increased nodulation parameters, growth parameters, yield, and yield components. The Arerti variety combined with the CP-M41 Mesorhizobium strain and NPSB fertilizer had the maximum grain production (3177.16 kg ha-1). It yielded 15.96%, 24.06%, and 37.93% more than the Arerti with CP-M41 strain, Arerti with NPSB, and the control treatments, respectively. The partial budget analysis of the study treatments showed that the Arerti variety with the combined application of 121 kg NPSB ha-1 and Mesorhizobium strain CP-M41 inoculation produced the highest net return (102,092.6 ETB ha-1) with an acceptable marginal rate of return (618%). It has been found that the CP-M41 strain and the Arerti variety, when combined with 121 kg NPSB ha-1 application, is a suitable treatment combination to achieve increased chickpea crop yield and profit in the studied area. However, the results need further validation in the farmer's field before recommending to farmers.

Viruses of Nitrogen-Fixing Mesorhizobium Bacteria in Globally Distributed Chickpea Root Nodules

Legume nodules are specialized environments on plant roots that are induced and dominated by nitrogen-fixing bacteria. Bacteriophages (phages) in these nodules could potentially provide top-down controls on the population size and, therefore, the function of nitrogen-fixing symbionts. Here we sought to characterize the diversity and biogeographical patterns of phages that infect nitrogen-fixing Mesorhizobium symbionts isolated from root nodules, leveraging 266 genomes of Mesorhizobium isolated from nodules and 648 nodule metagenomes collected from three species of chickpea plants ( Cicer spp.) under different agricultural management practices, spanning eight countries on five continents. We identified 106 phage populations (viral operational taxonomic units [vOTUs]) in Mesorhizobium draft genomes, 37% of which were confirmed as likely prophages. These vOTUs were detected in 64% of the Mesorhizobium-dominated nodule metagenomes and 58% of the Mesorhizobium isolates. Per metagenome, 1 to 16 putative Mesorhizobium vOTUs were detected, with more than half of the nodules containing only one such vOTU. The majority of vOTUs were detected exclusively in Ethiopia, followed by India and Morocco, with the lowest richness of putative Mesorhizobium phages in countries that applied industrial Mesorhizobium inoculants to crops. Two vOTUs were identified in five or more countries and in nodules dominated by different strains of Mesorhizobium, suggesting infection of diverse Mesorhizobium hosts and long-term interactions. Beta-diversity of these Mesorhizobium phage assemblages was significantly correlated with the dominant Mesorhizobium strain, but not with measured environmental parameters. Our findings indicate that nitrogen-fixing nodules in chickpea plants can contain distinct viral assemblages, with potential impacts on the nodule microbiome that bear further exploration.

Mesorhizobium liriopis sp. nov., isolated from the fermented fruit of Liriope platyphylla a medicinal plant.

A facultative anaerobic and Gram-negative strain, designated RP14T, was isolated from the fruit of Liriope platyphylla fermented for 60 days at 25°C. Strain RP14T showed 98.0 % 16S rRNA similarity to Mesorhizobium huakuii IFO 15243T, but in the phylogenetic tree, Mesorhizobium terrae NIBRBAC000500504T was its closest neighbour. The average nucleotide identity and digital DNA-DNA hybridization values between strain RP14T and 15 genomes of type strains of Mesorhizobium, were 73.8-74.4% and 16.4-20.2 %, respectively, which were lower than the recommended thresholds for species delineation. The strain grew at 25-32°C (optimum, 28°C), at pH 7.0-12.0 (optimum, pH 9.0) and with 0-2% NaCl (optimum, 0 %; w/v). Cells of strain RP14T were catalase-positive, oxidase-negative, rod-shaped and formed yellow-coloured colonies. The major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol and phosphatidylglycerol. The major fatty acid were C16 : 0, C19 : 0 cyclo ω8c and summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c). The DNA G+C content was 62.8 mol%. Based on polyphasic evidence, we propose Mesorhizobium liriopis sp. nov as a novel species within the genus Mesorhizobium. The type strain is RP14T (=KACC 22720T=TBRC 16341T).

A Mesorhizobium japonicum quorum sensing circuit that involves three linked genes and an unusual acyl-homoserine lactone signal.

Members of the genus Mesorhizobium, which are core components of the rhizosphere and specific symbionts of legume plants, possess genes for acyl-homoserine lactone (AHL) quorum sensing (QS). Here we show Mesorhizobium japonicum MAFF 303099 (formerly M. loti) synthesizes and responds to N-[(2E, 4E)-2,4-dodecadienoyl] homoserine lactone (2E, 4E-C12:2-HSL). We show that the 2E, 4E-C12:2-HSL QS circuit involves one of four luxR-luxI-type genes found in the sequenced genome of MAFF 303099. We refer to this circuit, which appears to be conserved among Mesorhizobium species, as R1-I1. We show that two other Mesorhizobium strains also produce 2E, 4E-C12:2-HSL. The 2E, 4E-C12:2-HSL is unique among known AHLs in its arrangement of two trans double bonds. The R1 response to 2E, 4E-C12:2-HSL is extremely selective in comparison with other LuxR homologs, and the trans double bonds appear critical for R1 signal recognition. Most well-studied LuxI-like proteins use S-adenosylmethionine and an acyl-acyl carrier protein as substrates for synthesis of AHLs. Others that form a subgroup of LuxI-type proteins use acyl-coenzyme A substrates rather than acyl-acyl carrier proteins. I1 clusters with the acyl-coenzyme A-type AHL synthases. We show that a gene linked to the I1 AHL synthase is involved in the production of the QS signal. The discovery of the unique I1 product enforces the view that further study of acyl-coenzyme A-dependent LuxI homologs will expand our knowledge of AHL diversity. The involvement of an additional enzyme in AHL generation leads us to consider this system a three-component QS circuit. IMPORTANCE We report a Mesorhizobium japonicum quorum sensing (QS) system involving a novel acyl-homoserine lactone (AHL) signal. This system is known to be involved in root nodule symbiosis with host plants. The chemistry of the newly described QS signal indicated that there may be a dedicated cellular enzyme involved in its synthesis in addition to the types known for production of other AHLs. Indeed, we report that an additional gene is required for synthesis of the unique signal, and we propose that this is a three-component QS circuit as opposed to the canonical two-component AHL QS circuits. The signaling system is exquisitely selective. The selectivity may be important when this species resides in the complex microbial communities around host plants and may make this system useful in various synthetic biology applications of QS circuits.

Mesorhizobium improves chickpea growth under chromium stress and alleviates chromium contamination of soil

Environmental pollution has become a transnational issue that impacts ecosystems, soil, water, and air and is directly related to human health and well-being. Chromium pollution decreases the development of plant and microbial populations. It warrants the need to remediate chromium-contaminated soil. Decontaminating chromium-stressed soils via phytoremediation is a cost-effective and environmentally benign method. Using multifunctional plant growth-promoting rhizobacteria (PGPR) lower chromium levels and facilitates chromium removal. PGPR work by altering root architecture, secreting chemicals that bind metals in the rhizosphere, and reducing phytotoxicity brought on by chromium. The present study aimed to investigate the chromium bioremediation capacity of metal-tolerant PGPR isolate while promoting the growth of chickpeas in the presence of varying levels of chromium (15.13, 30.26, and 60.52 mg/kg of chromium). The isolate, Mesorhizobium strain RC3, substantially reduced chromium content (60.52 mg/kg) in the soil. It enhanced the root length by 10.87%, the shoot length by 12.38%, the number of nodules by 6.64%, and nodule dry weight by 13.77% at 90 days. After 135 days of sowing, more improvement in the root length (18.05), shoot length (21.60%)the chlorophyll content (6.83%), leghaemoglobin content (9.47%), and the highest growth in the crop seed yield (27.45%) and crop protein content (16.83%)The isolate reduced chromium accumulation in roots, shoots, and grains chickpea. Due to chromium bioremediation and its plant growth-promoting and chromium-attenuating qualities, Mesorhizobium strain RC3 could be used as a green bioinoculant for plant growth promotion under chromium stress.

Nodulation and Growth Promotion of Chickpea by Mesorhizobium Isolates from Diverse Sources.

The cultivation of chickpea (Cicer arietinum L.) in South Africa is dependent on the application of suitable Mesorhizobium inoculants. Therefore, we evaluated the symbiotic effectiveness of several Mesorhizobium strains with different chickpea genotypes under controlled conditions. The tested parameters included shoot dry weight (SDW), nodule fresh weight (NFW), plant height, relative symbiotic effectiveness (RSE) on the plant as well as indole acetic acid (IAA) production and phosphate solubilization on the rhizobia. Twenty-one Mesorhizobium strains and six desi chickpea genotypes were laid out in a completely randomized design (CRD) with three replicates in a glasshouse pot experiment. The factors, chickpea genotype and Mesorhizobium strain, had significant effects on the measured parameters (p < 0.001) but lacked significant interactions based on the analysis of variance (ANOVA). The light variety desi genotype outperformed the other chickpea genotypes on all tested parameters. In general, inoculation with strains LMG15046, CC1192, XAP4, XAP10, and LMG14989 performed best for all the tested parameters. All the strains were able to produce IAA and solubilize phosphate except the South African field isolates, which could not solubilize phosphate. Taken together, inoculation with compatible Mesorhizobium promoted chickpea growth. This is the first study to report on chickpea-compatible Mesorhizobium strains isolated from uninoculated South African soils with no history of chickpea production; although, their plant growth promotion ability was poorer compared to some of the globally sourced strains. Since this study was conducted under controlled conditions, we recommend field studies to assess the performance of the five highlighted strains under environmental conditions in South Africa.

​Plant Growth Promoting Mesorhizobia as a Potential Inoculant for Chickpea (Cicer arietinum L.): A Review

Chickpea establish symbiotic relationship with Mesorhizobium ciceri and fixes atmospheric nitrogen, which depends on the efficacy of the Mesorhizobium strains, crop management, soil and environmental factors. A multi-trait Mesorhizobium isolate having more than one Plant growth promoting (PGP) traits often stimulates plant growth directly by IAA and GA production, enhances nutrient uptake by solubilizing phosphorus and siderophore production while indirect growth promotion is by prevention of some pathogenic microorganisms by producing HCN. Several studies revealed that plant growth attributes and yield was increased with inoculation of multi trait Rhizobium isolates. The occurrence of different PGP traits of Mesorhizobium and their effects on plant growth will be discussed in this review paper.

Evolution of novel strains of Ensifer nodulating the invasive legume Leucaena leucocephala (Lam.) de Wit in different climatic regions of India through lateral gene transfer.

More than 200 root-nodule bacterial strains were isolated from Leucaena leucocephala growing at 42 sampling sites across 12 states and three union territories of India. Genetic diversity was observed among 114 strains from various climatic zones; based on recA, these were identified as strains of Ensifer, Mesorhizobium, Rhizobium, and Bradyrhizobium. In multilocus sequence analysis (MLSA) strains clustered into several novel clades and lineages. Ensifer were predominant nodulating genotype isolated from majority of alkaline soils, while Mesorhizobium and Rhizobium strains were isolated from a limited sampling in North-Eastern states with acidic soils. Positive nodulation assays of selected Ensifer representing different genetic combinations of housekeeping and sym genes suggested their broad host range within the closely related mimosoid genera Vachellia, Senegalia, Mimosa, and Prosopis. Leucaena selected diverse strains of Ensifer and Mesorhizobium as symbionts depending on available soil pH, climatic, and other edaphic conditions in India. Lateral gene transfer seems to play a major role in genetic diversification of Ensifer exhibited in terms of Old World vs. Neotropical genetic make-up and mixed populations at several sites. Although Neotropical Ensifer strains were most symbiotically effective on Leucaena, the native Ensifer are promiscuous and particularly well-adapted to a wide range of sampling sites with varied climates and edaphic factors.

Mesorhizobium ventifaucium sp. nov. and Mesorhizobium escarrei sp. nov., two novel root-nodulating species isolated from Anthyllis vulneraria

Ten mesorhizobial strains isolated from root-nodules of Anthyllis vulneraria by trapping using soils from southern France were studied to resolve their taxonomy. Their 16S rDNA sequences were identical and indicated that they are affiliated to the genus Mesorhizobium within the group M. prunaredense/M. delmotii/M. temperatum/M. mediterraneum/M. wenxiniae and M. robiniae as the closest defined species. Their evolutionary relationships with validated species were further characterized by multilocus sequence analysis (MLSA) using 4 protein-coding housekeeping genes (recA, atpD, glnII and dnaK), that divides the strains in two groups, and suggest that they belong to two distinct species. These results were well-supported by MALDI-TOF mass spectrometry analyses, wet-lab DNA-DNA hybridization (≤58%), and genome-based species delineation methods (ANI < 96%, in silico DDH < 70%), confirming their affiliation to two novel species. Based on these differences, Mesorhizobium ventifaucium (STM4922T = LMG 29643T = CFBP 8438T) and Mesorhizobium escarrei (type strain STM5069T = LMG 29642T = CFBP 8439T) are proposed as names for these two novel species. The phylogeny of nodulation genes nodC and nodA allocated the type strains into symbiovar anthyllidis as well as those of M. metallidurans STM2683T, M. delmotii STM4623T and M. prunaredense STM4891T, all recovered from the same legume species.

Rhizobial diversity is associated with inoculation history at a two-continent scale.

A total of 120 Mesorhizobium strains collected from the central dry zone of Myanmar were analyzed in a pot experiment to evaluate nodulation and symbiotic effectiveness (SE%) in chickpea plants. Phylogenetic analyses revealed all strains belonged to the genus Mesorhizobium according to 16–23S rDNA IGS and the majority of chickpea nodulating rhizobia in Myanmar soils were most closely related to M. gobiense, M. muleiense, M. silamurunense, M. tamadayense and M. temperatum. Around two-thirds of the Myanmar strains (68%) were most closely related to Indian strain IC-2058 (CA-181), which is also most closely related to M. gobiense. There were no strains that were closely related to the cognate rhizobial species to nodulate chickpea: M. ciceri and M. mediterraneum. Strains with diverse 16S–23S rDNA IGS shared similar nodC and nifH gene sequences with chickpea symbionts. Detailed sequence analysis of nodC and nifH found that the strains in Myanmar were somewhat divergent from the group including M. ciceri and were more closely related to M. muleiense and IC-2058. A cross-continent analysis between strains isolated in Australia compared with Myanmar found that there was little overlap in species, where Australian soils were dominated with M. ciceri, M. temperatum and M. huakuii. The only co-occurring species found in both Myanmar and Australia were M. tamadayense and M. silumurunense. Continued inoculation with CC1192 may have reduced diversity of chickpea strains in Australian soils. Isolated strains in Australian and Myanmar had similar adaptive traits, which in some cases were also phylogenetically related. The genetic discrepancy between chickpea nodulating strains in Australia and Myanmar is not only due to inoculation history but to adaptation to soil conditions and crop management over a long period, and there has been virtually no loss of symbiotic efficiency over this time in strains isolated from soils in Myanmar.

Simple experiment on legume-rhizobium symbiosis aimed at students without laboratory experience

ABSTRACT The curriculum for undergraduate Agricultural Engineering does not usually include much laboratory practice. In order to address this, here we propose the design of a practical class on legume-rhizobia symbiosis, using an interdisciplinary approach that takes elements from botany, agriculture, and microbiology. The students perform an assay to stimulate nodulation in chickpea seedlings through inoculation with Mesorhizobium strains recommended as commercial inoculants, and evaluate improvement in plant growth as a result of nitrogen fixation. The experiment could serve as a springboard to discuss the importance of plant-bacteria interactions in the cultivation of a regionally relevant crop. It may also encourage the use of abilities of different kinds, including the appropriate manipulation of lab materials, the active engagement of scientific thought, the application of knowledge to specific real world scenarios and collaborative teamwork, all of which will be of paramount importance throughout the students’ careers. The class can be easily adapted for other degrees connected to environmental microbiology, as well as for other symbiotic pairs that may include significant crops from other regions around the world.

Native Mesorhizobium strains improve yield and nutrient composition of the common bird's-foot trefoil grown in an acid soil

Acid soils occupy more than 3.95 billion ha of the world soils, and finding an adequate solution for the limitation of crop production on these soils is indispensable. Using highly effective rhizobia tolerant to low pH enables successful nodulation and quality crop production of legumes in acid soils. In this study, isolation and characterization of native rhizobia associated with root nodules of bird's-foot trefoil (Lotus corniculatus L.) from Serbia were conducted. Their effects on the plant yield and nutrient composition of bird's-foot trefoil grown in an acid soil (pH 5.4), in a pot experiment were evaluated. Out of 72 strains isolated, 40 could nodulate bird's-foot trefoil when reinoculated in the test tubes under gnotobiotic conditions, and 23 isolates showed high nitrogen-fixing efficiency. Overall, all isolates could grow well in medium with a pH between 4.5 and 8. Indole-3-acetic acid (IAA) production was detected in all nodulating isolates and 24 could solubilize inorganic phosphates. The identification of selected isolates showed that all belong to Mesorhizobium genus (M. tianshanense, M. erdmanii, M. cantuariense, M. loti, M. jarvisii and M. caraganae). Four acid-tolerant isolates (1M12, 631oz, U1C, and 754) with high nitrogen-fixing efficiency in vitro and particular PGP traits were selected for the pot experiment with acid soil. All applied bacterial treatments (except 1M12) increased the shoot dry weight of bird's-foot trefoil plants (up to 50%), compared to the control. In addition, N uptake and N% were increased up to 20% by inoculation. All applied treatments influenced the concentrations and improved uptake of macro (P, K, Ca, and Mg) and micronutrients (Cu, Fe, Mn, Ni, Zn, and B) in the plant material. The obtained results indicated that satisfactory yield and mineral composition of L. corniculatus in acid soils could be achieved by inoculation with selected Mesorhizobium strains.

Ecological performance of multifunctional pesticide tolerant strains of Mesorhizobium sp. in chickpea with recommended pendimethalin, ready-mix of pendimethalin and imazethpyr, carbendazim and chlorpyrifos application.

The present study was designed to screen the Mesorhizobium strains (50) for tolerance with four recommended pesticides in chickpea. In-vitro, robust pesticide tolerant strains were developed in pesticides amended media over several generations. Further, verification of the multifunctional traits of pesticide tolerant mesorhizobia under pesticide stress was conducted in-vitro. Among different pesticides, significantly high tolerance in Mesorhizobium strains was observed with recommended doses of pendimethalin (37%) and ready-mix (36%) followed by chlorpyrifos (31%) and carbendazim (30%), on an overall basis. Based on multifunctional traits, Mesorhizobium strains viz. MR2, MR17 and recommended MR33 were the most promising. Ecological performance of the potential Mesorhizobium strains alone and in dual-inoculation with recommended PGP rhizobacterium strain RB-1 (Pseudomonas argenttinensis JX239745.1) was subsequently analyzed in field following standard pesticide application in PBG-7 and GPF-2 chickpea varieties for two consecutive rabi seasons (2015 and 2016). Dual-inoculant treatments; recommended RB-1 + MR33 (4.1%) and RB-1 + MR2 (3.8%) significantly increased the grain yield over Mesorhizobium alone treatments viz MR33 and MR2, respectively. Grain yield in PBG7 variety was significantly affected (7.3%) by the microbial inoculant treatments over GPF2 variety. Therefore, the potential pesticide tolerant strains MR2 and MR33 can be further explored as compatible dual-inoculants with recommended RB-1 for chickpea under environmentally stressed conditions (pesticide application) at multiple locations. Our approach using robust multifunctional pesticide tolerant Mesorhizobium for bio-augmentation of chickpea might be helpful in the formulation of effective bio-inoculants consortia in establishing successful chickpea-Mesorhizobium symbiosis.

Definition of the novel symbiovar canariense within Mesorhizobium neociceri sp. nov., a new species of genus Mesorhizobium nodulating Cicer canariense in the “Caldera de Taburiente” National Park (La Palma, Canary Islands)

Cicer canariense is a highly promiscuous wild chickpea nodulated by Mesorhizobium strains in La Palma Island located at Canary archipelago. Four of these strains, CCANP34, CCANP35T, CCANP38 and CCANP95 belong to a group phylogenetically close to Mesorhizobium caraganae with 100% similarity values in the 16S rRNA gene. However, the genomes of the strains CCANP35T and M. caraganae LMG 24397T obtained in this work showed ANIb and dDDH values of 90.02% and 44.1%, respectively. These values are lower than those currently accepted for different bacterial species showing that the Canarian strains do not belong to the species M. caraganae. The Canarian strains also differ from M. caraganae in the amounts of several fatty acids and in several phenotypic traits. Based on the obtained results the Canarian strains belong to a novel species for which we propose the name Mesorhizobium neociceri sp. nov. and whose type strain is CCANP35T. The results of the phylogenetic analyses of nodC and nifH symbiotic genes showed that the Canarian strains represent a novel symbiovar within genus Mesorhizobium phylogenetically divergent to that encompassing M. caraganae. We propose the names canariense and caraganae for the symbiovars encompassing the strains of M. neociceri and M. caraganae, respectively.

Rhizobial isolates in active layer samples of permafrost soil of Spitsbergen, Arctic

Twenty-nine strains were isolated from two samples of the permafrost active layer of the Spitsbergen archipelago. The estimated number of bacteria ranged from 4.0⋅104 to 1.7⋅107 CFU∙g-1. As a result of sequencing of the 16S rRNA (rrs) genes, the isolates were assigned to 13 genera belonging to the phyla Actinobacteria, Proteobacteria (classes α, β, and γ), Bacteroidetes, and Firmicutes. Six isolates belonged to the rhizobial genus Mesorhizobium (order Rhizobiales). A plant nodulation assay with seedlings of legume plants Astragalus norvegicus, A. frigidus, A. subpolaris and Oxytropis sordida, originated from Khibiny (Murmansk region, Russia) and inoculated with Mesorhizobium isolates, showed the inability of these strains to form nodules on plant roots. Symbiotic (sym) genes nodC and nodD were not detected in Mesorhizobium strains either.

The leguminous trees Vachellia seyal (Del.) and Prosopis juliflora (Swartz) DC and their association with rhizobial strains from the root-influence zone of the grass Sporobolus robustus Kunth

This study focused on the genetic, symbiotic and phenotypic diversity of a collection of 12 Vachellia seyal and Prosopis juliflora rhizobial strains that were isolated from soils influenced by the roots of the grass Sporobolus robustus. Phylogenetic trees were constructed based on data from the 16S rRNA gene, from a multi-locus sequence analysis (MLSA) of the housekeeping genes recA, atpD, glnA and gyrB, as well as from the symbiotic genes nodA, nodC and nifH. Most of the mesorhizobial strains were clustered with the M. plurifarium type strain ORS 1032T and the 16S rRNA gene was as discriminatory as the other housekeeping genes. The Ensifer collection was closely related to E. alkalisoli YIC 4027T and E. fredii LMG 6217T using 16S rRNA and MLSA. Amplification of the Rhizobium collection was only observed with the 16S rRNA gene, where the strains were clustered with Rhizobium sp. 1B, Rhizobium sp. SEMIA 6411, Rhizobium sp. AS1-101a and Rhizobium sp. ORS 3441. The phylogeny of the nifH gene from Mesorhizobium and Ensifer strains were similar to those of nodulation genes (nodA and nodC). The nodA-nodC and nifH gene phylogenies were not consistent with those of 16S rRNA and the housekeeping genes. Ensifer genospecies tolerated high salinity better than the other genotypes after testing on a salt content gradient up to 49‰. We conclude that the rhizosphere of Sporobolus tussocks harbors a rhizobial microbiota, which could be capable of helping V. seyal and P. juliflora to grow better in saline soils.

Biserrula pelecinus L. is a promising forage legume for the central Ethiopian highlands

AbstractThe availability of effective inoculant rhizobia is often critical to the successful development of productive forage legumes. Biserrula pelecinus L. is a legume with potential as forage in Ethiopia to improve livestock feed quality and soil fertility. B. pelecinus can form N2‐fixing symbiosis with rhizobia in the genus Mesorhizobium. This study investigated the N2 fixation effectiveness of 15 B. pelecinus‐nodulating Mesorhizobium strains on two subspecies of B. pelecinus (B. pelecinus ssp. leiocarpa, native to Ethiopia, and the introduced B. pelecinus ssp. pelecinus). The most effective strain (WSM3873) on both subspecies was assessed at two sites; one with pre‐existing populations of B. pelecinus‐nodulating rhizobia (Modjo), and one without (Holeta). No inoculation response was observed at Modjo when B. pelecinus ssp. pelecinus was inoculated with WSM3873 alone, however, biomass yield was greatest (11.5 tonne DM/ha) following inoculation along with co‐application of phosphorus and nitrogen. At Holeta, a strong inoculation response was achieved with WSM3873 alone on B. pelecinus ssp. pelecinus. In contrast, B. pelecinus ssp. leiocarpa did not show any response at Modjo and failed to emerge after sowing at Holeta. While the native legume B. pelecinus ssp. leiocarpa appears poorly suited to development as a forage, B. pelecinus ssp. pelecinus and WSM3873 represents a promising legume‐rhizobia symbiosis that could benefit farming systems of the central Ethiopian highlands.

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.