Tropici CIAT899 Research Articles

Specific primers for the rapid detection and quantitation of Rhizobium elite strains of common beans in the plant and environment

Tools for adequately detecting and monitoring elite strains used for common bean inoculation are lacking. These tools help evaluate strain competitiveness and nodule occupancy rates and study their ecology in the environment. For this reason, we designed and tested specific primers for three Rhizobium elite strains widely used as common bean inoculants: Rhizobium tropici CIAT 899, R. tropici CPAC H12, and R. freirei PRF 81. Primer specificity was confirmed in PCR assays performed with the target strains and 45 strains of the Rhizobiaceae family. CIAT 899 and CPAC H12 showed an average nucleotide identity of 99.99 %; thus, all further evaluations were performed with CIAT 899 and PRF 81. Primer amplification efficiency, sensitivity, determination coefficients, and dissociation curves were evaluated. In addition, three pairs of primers for each target strain were evaluated for their capacities to quantify their targets in the nodules, rhizosphere, and roots. Eleven and seven strain-specific primer pairs were selected for CIAT 899 and PRF 81, respectively. They amplified DNA isolated from bacterial suspensions, nodules, and nodule extracts used directly in PCR with average amplification efficiencies between 91 % and 105 % and 96 % and 107 % for CIAT 899 and PRF 81 primers and detection limits of 32 and 34 copies of target DNA, respectively. Their use showed that populations increased in the roots and decreased in the rhizospheric soil over 15 days. Nodular occupancy rates were above 90 % when the strains were inoculated individually, and a high co-infection rate was observed upon co-inoculation, with CIAT 899 and PRF 81 detected in 100 % and 73 % of the nodules, respectively. We conclude that the primers and methodologies shown here are suitable for detecting the target strains with high specificity and sensitivity. Our approach also demonstrated that R. tropici CIAT 899 and R. freirei PRF 81 were competitive and effective in infecting common beans under the tested conditions.

Synergistic Interaction of Rhizobium tropici, Rhizophagus irregularis and Serendipita indica in Promoting Snap Bean Growth

The overuse of chemical pesticides and fertilizers in crop farming has led to a decrease in crop quality and negative impacts on soil and the environment. It is crucial to adopt alternative strategies to maintain soil and environmental quality while enhancing crop growth and yield. To explore this, a study was conducted under greenhouse conditions to investigate the effect of Rhizobium tropici CIAT 899 alone, as well as in association with mycorrhizae (Rhizophagus irregularis) and endophytic fungus (Serendipita indica), on the growth, yield, and nutrient status of snap bean plants. At harvest, the rhizobial strain CIAT 899 demonstrated the highest effectiveness. It significantly increased the number of nodules in both Contender and Garrafal Enana varieties by 6.97% and 14.81%, respectively, compared with the control without inoculation. Furthermore, the results indicated that co-inoculation of Rhizobium and symbiotic fungi had positive effects on nitrogen content, phosphorus availability, and overall plant growth. Regardless of the variety, plants inoculated with R. tropici CIAT 899 and Serendipita indica exhibited the highest values for plant growth parameters. This combination resulted in 168% and 135% increases in root dry biomass, as well as 140% and 225% increases in the number of pods for Contender and Garrafal Enana, respectively, compared with the control at harvest. Additionally, this study highlights the potential benefits of combining R. tropici with either Serendipita indica or Rhizophagus irregularis in terms of nitrogen and phosphorus uptake. These symbiotic microorganisms demonstrated synergistic interactions with snap bean plants, leading to improved mineral nutrition and enhanced growth. Overall, these findings suggest that utilizing these symbiotic microorganisms can effectively enhance the mineral nutrition and growth of snap bean plants.

Mycorrhizal fungi inoculation effect on plant growth and phosphorus metabolism of snap bean variety "Contender"

Biofertilizers management for sustainable agriculture is a challenge to improve farmers ‘food security today and correct nutrient deficiencies (potassium, phosphorus and nitrogen) in plant. As biofertilizers, mycorrhizal symbiosis plays an important role in nutrient uptake and the productivity of plants. The aim of this study was the Rhizobium and mycorrhizal inoculations effect on plant growth and mineral nutrition of snap bean variety "Contender" in greenhouse condition. The experiment was a randomized block design with four treatments: a control non-inoculated, dual inoculation with Serendipita indica (S.indica) and Rhizobium tropici Ciat 899 strain (Ciat 899), dual inoculation with Rhizophagus irregularis (R. irregularis) and Ciat 899, and combined inoculation with R. irregularis, S.indica and Ciat 899 ; using eight replicates per treatment. Ciat 899 associated with S.indica or R. irregularis significantly increased plant growth parameters, and shoots and root phosphorus concentrations. It shows higher biomass production, pod number, nodule number and phosphorus content than plants inoculated with triple strains and non-mycorrhizal plants. Thus, the colonization rate exceeds 80%. The results of this study show that dual inoculation with Rhizobium and mycorrhizal fungi enhance significantly productivity in common bean.

Rhizobacteria control damping-off and promote growth of lima bean with and without co-inoculation with Rhizobium tropici CIAT899.

Rhizoctonia solani compromises the production of lima bean, an alternative and low-input food source in many tropical regions. Inoculation of bacterial strains has been used, but research on their biocontrol and growth promotion potential on lima bean is scarce. The objective of this study was to evaluate the effects of inoculation with rhizobacterial strains of the genera Bacillus, Brevibacillus, Paenibacillus, Burkholderia, Pseudomonas, and Rhizobium in combination or not with N2-fixing Rhizobium tropici on the control of damping-off disease and growth promotion in lima bean plants. Greenhouse experiments were conducted to evaluate the inoculation with bacterial strains with biocontrol potential in combination or not with R. tropici in substrate infected with R. solani CML 1846. Growth promotion of these strains was also assessed. Strains of Brevibacillus (UFLA 02-286), Pseudomonas (UFLA 02-281 and UFLA 04-885), Rhizobium (UFLA 04-195), and Burkholderia (UFLA 04-227) co-inoculated with the strain CIAT 899 (Rhizobium tropici) were the most effective in controlling R. solani, reducing the disease incidence in 47-60% on lima bean. The promising strains used in the biocontrol assays were also responsive in promoting growth of lima bean under disease and sterile conditions. A positive synergistic effect of co-inoculation of different genera contributed to plant growth, and these outcomes are important first steps to improve lima bean production.

Rhizobium tropici and Riboflavin Amendment Condition Arbuscular Mycorrhiza Colonization in Phaseolus vulgaris L.

Phaseolus vulgaris L. (Fabaceae) forms symbioses with arbuscular mycorrhizal fungi (AMF) and nitrogen-fixing rhizobia (NFB). The tripartite relationship uses molecular singals to establish intracellular symbioses in roots. The goal of this study was to determine if Rhizobium tropici CIAT 899 and exogenous riboflavin (vitamin B2) have an effect on AMF species selection and root colonization of P. vulgaris. Using SSU rRNA fragment amplification of DNA extracted from P. vulgaris roots, we found that the presence of R. tropici altered the relative distribution of AMF species. Dominikia bernensis (Ohel) was the most abundant AMF species in P. vulgaris roots but when R. tropici was co-inoculated, Glomus species dominated. Rhizobacteria such as R. tropici, secrete riboflavin and could affect AMF symbiosis. Addition of 50 μM riboflavin to P. vulgaris, increased plant growth (28%), dry nodule weight (18%), AMF colonization (248%) and mycorrhizal vesicle frequency (56%) in bean roots. 3.12 and 12.5 µM riboflavin favored the presence of Glomus macrocarpum in P. vulgaris roots. This work provides the basis to further study of rhizobial and mycorrhizal co-inoculation of Phaseolus vulgaris bean.

Competition in the Phaseolus vulgaris-Rhizobium symbiosis and the role of resident soil rhizobia in determining the outcomes of inoculation

Background and AimsInoculation of legumes with effective N2-fixing rhizobia is a common practice to improve farming profitability and sustainability. To succeed, inoculant rhizobia must overcome competition for nodulation by resident soil rhizobia that fix N2 ineffectively. In Kenya, where Phaseolus vulgaris (common bean) is inoculated with highly effective Rhizobium tropici CIAT899 from Colombia, response to inoculation is low, possibly due to competition from ineffective resident soil rhizobia. Here, we evaluate the competitiveness of CIAT899 against diverse rhizobia isolated from cultivated Kenyan P. vulgaris.MethodsThe ability of 28 Kenyan P. vulgaris strains to nodulate this host when co-inoculated with CIAT899 was assessed. Rhizosphere competence of a subset of strains and the ability of seed inoculated CIAT899 to nodulate P. vulgaris when sown into soil with pre-existing populations of rhizobia was analyzed.ResultsCompetitiveness varied widely, with only 27% of the test strains more competitive than CIAT899 at nodulating P. vulgaris. While competitiveness did not correlate with symbiotic effectiveness, five strains were competitive against CIAT899 and symbiotically effective. In contrast, rhizosphere competence strongly correlated with competitiveness. Soil rhizobia had a position-dependent numerical advantage, outcompeting seed-inoculated CIAT899 for nodulation of P. vulgaris, unless the resident strain was poorly competitive.ConclusionSuboptimally effective rhizobia can outcompete CIAT899 for nodulation of P. vulgaris. If these strains are widespread in Kenyan soils, they may largely explain the poor response to inoculation. The five competitive and effective strains characterized here are candidates for inoculant development and may prove better adapted to Kenyan conditions than CIAT899.

Hydroponic Common-Bean Performance under Reduced N-Supply Level and Rhizobia Application.

This study aims to explore the possibility of a reduced application of inorganic nitrogen (N) fertiliser on the yield, yield qualities, and biological nitrogen fixation (BNF) of the hydroponic common bean (Phaseolus vulgaris L.), without compromising plant performance, by utilizing the inherent ability of this plant to symbiotically fix N2. Until the flowering stage, plants were supplied with a nutrient solution containing N-concentrations of either a, 100%, conventional standard-practice, 13.8 mM; b, 75% of the standard, 10.35 mM; or c, 50% of the standard, 6.9 mM. During the subsequent reproductive stage, inorganic-N treatments b and c were decreased to 25% of the standard, and the standard (100% level) N-application was not altered. The three different inorganic-N supply treatments were combined with two different rhizobia strains, and a control (no-inoculation) treatment, in a two-factorial experiment. The rhizobia strains applied were either the indigenous strain Rhizobium sophoriradicis PVTN21 or the commercially supplied Rhizobium tropici CIAT 899. Results showed that the 50-25% mineral-N application regime led to significant increases in nodulation, BNF, and fresh-pod yield, compared to the other treatment, with a reduced inorganic-N supply. On the other hand, the 75-25% mineral-N regime applied during the vegetative stage restricted nodulation and BNF, thus incurring significant yield losses. Both rhizobia strains stimulated nodulation and BNF. However, the BNF capacity they facilitated was suppressed as the inorganic-N input increased. In addition, strain PVTN21 was superior to CIAT 899-as 50-25% N-treated plants inoculated with the former showed a yield loss of 11%, compared to the 100%-N-treated plants. In conclusion, N-use efficiency optimises BNF, reduces mineral-N-input dependency, and therefore may reduce any consequential negative environmental consequences of mineral-N over-application.

Non-Ionic Osmotic Stress Induces the Biosynthesis of Nodulation Factors and Affects Other Symbiotic Traits in Sinorhizobium fredii HH103.

(1) Background: Some rhizobia, such as Rhizobium tropici CIAT 899, activate nodulation genes when grown under osmotic stress. This work aims to determine whether this phenomenon also takes place in Sinorhizobium fredii HH103. (2) Methods: HH103 was grown with and without 400 mM mannitol. β-galactosidase assays, nodulation factor extraction, purification and identification by mass spectrometry, transcriptomics by RNA sequencing, motility assays, analysis of acyl-homoserine lactones, and indole acetic acid quantification were performed. (3) Results: Non-ionic osmotic stress induced the production of nodulation factors. Forty-two different factors were detected, compared to 14 found in the absence of mannitol. Transcriptomics indicated that hundreds of genes were either activated or repressed upon non-ionic osmotic stress. The presence of 400 mM mannitol induced the production of indole acetic acid and acyl homoserine lactones, abolished swimming, and promoted surface motility. (4) Conclusions: In this work, we show that non-ionic stress in S. fredii HH103, caused by growth in the presence of 400 mM mannitol, provokes notable changes not only in gene expression but also in various bacterial traits, including the production of nodulation factors and other symbiotic signals.

The Rhizobium tropici CIAT 899 NodD2 protein promotes symbiosis and extends rhizobial nodulation range by constitutive nodulation factor synthesis.

In the symbiotic associations between rhizobia and legumes, the NodD regulators orchestrate the transcription of the specific nodulation genes. This set of genes is involved in the synthesis of nodulation factors, which are responsible for initiating the nodulation process. Rhizobium tropici CIAT 899 is the most successful symbiont of Phaseolus vulgaris and can nodulate a variety of legumes. Among the five NodD regulators present in this rhizobium, only NodD1 and NodD2 seem to have a role in the symbiotic process. However, the individual role of each NodD in the absence of the other proteins has remained elusive. In this work, we show that the CIAT 899 NodD2 does not require activation by inducers to promote the synthesis of nodulation factors. A CIAT 899 strain overexpressing nodD2, but lacking all additional nodD genes, can nodulate three different legumes as efficiently as the wild type. Interestingly, CIAT 899 NodD2-mediated gain of nodulation can be extended to another rhizobial species, since its overproduction in Sinorhizobium fredii HH103 not only increases the number of nitrogen-fixing nodules in two host legumes but also results in nodule development in incompatible legumes. These findings potentially open exciting opportunities to develop rhizobial inoculants and increase legume crop production.

Priming of defense-related genes in Brassica oleracea var. capitata using concentrated metabolites produced by Rhizobium tropici CIAT 899.

To verify the potential of metabolites extracted from Rhizobium tropici to trigger the priming of defense responses in cruciferous plants, we analyzed the expression of defense-related genes by qRT-PCR. Brassica oleracea var. capitata, susceptible to Xanthomonas campestris pv. campestris, were grown in greenhouse conditions. At 18days after sowing, plants were inoculated with 1mL of 1% concentrated metabolites produced by R. tropici (CM-RT) in the root. In a second experiment, leaves were sprayed with 1mL of a solution containing 1% CM-RT. Aerial and root tissue were collected separately at 0 (non-treated control condition), 24, and 48h after application, submitted to RNA extraction and gene expression analysis by qRT-PCR. The results showed that, after root treatment with CM-RT, most evaluated genes were upregulated at 24h after application and downregulated at 48h after application in roots, while in leaves, genes were downregulated both at 24 and 48h after application. On the other hand, leaf treatment with CM-RT showed that most evaluated genes in leaves and roots were upregulated at 24 and 48h after application. These results indicate that the effect of CM-RT applied in roots seems restricted to the applied region and is not sustained, while the application in leaves results in a more systemic response and maintenance of the effect of CM-RT for a longer period. The results obtained in this study emphasize the biotechnological potential of using metabolites of R. tropici as an elicitor of active defense responses in plants.

The nodD1 Gene of Sinorhizobium fredii HH103 Restores Nodulation Capacity on Bean in a Rhizobium tropici CIAT 899 nodD1/nodD2 Mutant, but the Secondary Symbiotic Regulators nolR, nodD2 or syrM Prevent HH103 to Nodulate with This Legume.

Rhizobial NodD proteins and appropriate flavonoids induce rhizobial nodulation gene expression. In this study, we show that the nodD1 gene of Sinorhizobium fredii HH103, but not the nodD2 gene, can restore the nodulation capacity of a double nodD1/nodD2 mutant of Rhizobium tropici CIAT 899 in bean plants (Phaseolus vulgaris). S. fredii HH103 only induces pseudonodules in beans. We have also studied whether the mutation of different symbiotic regulatory genes may affect the symbiotic interaction of HH103 with beans: ttsI (the positive regulator of the symbiotic type 3 protein secretion system), and nodD2, nolR and syrM (all of them controlling the level of Nod factor production). Inactivation of either nodD2, nolR or syrM, but not that of ttsI, affected positively the symbiotic behavior of HH103 with beans, leading to the formation of colonized nodules. Acetylene reduction assays showed certain levels of nitrogenase activity that were higher in the case of the nodD2 and nolR mutants. Similar results have been previously obtained by our group with the model legume Lotus japonicus. Hence, the results obtained in the present work confirm that repression of Nod factor production, provided by either NodD2, NolR or SyrM, prevents HH103 to effectively nodulate several putative host plants.

Co-inoculation of Rhizobium and Bradyrhizobium promotes growth and yield of common beans

Some strains of Bradyrhizobium benefit the biological N2 fixation (BNF) and growth of common beans (Phaseolus vulgaris L.) when co-inoculated with Rhizobium tropici under greenhouse conditions. Here, we evaluated the benefit of Bradyrhizobium diazoefficiens strain CPAC 7 co-inoculated with Rhizobium tropici strain CIAT 899 on common bean in the field. Three Bradyrhizobium strains were co-inoculated with R. tropici CIAT 899 in one experiment in the greenhouse and two in the field. The greenhouse experiment combined each of three Bradyrhizobium strains (29w, USDA 110, and CPAC 7) in co-inoculation with CIAT 899 in Leonard jars. Then, two of these strains, 29w and CPAC 7, were co-inoculated with Rhizobium on-seeds (1.2 × 106 CFU seed−1) or both on-seed and in-furrow (1.08 × 108 CFU m−1) in a field experiment. Finally, a second field experiment evaluated strain CPAC 7 at doses ranging from 1.5 × 105 to 1.2 × 106 CFU seed−1. Co-inoculation with strain CPAC 7 improved shoot biomass accumulation in Leonard jars. In the field, co-inoculation of CPAC 7 with CIAT 899 on-seed enhanced grain yield by 20% compared with inoculation of CIAT 899 only (p < 0.10). The percentage of N derived from the air (%Ndfa) was between 50 and 60%, with only a tendency towards co-inoculation benefit. Given that the %Ndfa of the inoculated plants was not statistically different between treatments, the increased grain yield was not entirely due to BNF but also due to an increase of soil N uptake. In the second field trial, larger doses of inoculum increased nodulation and root biomass, but significant differences in grain yield were not observed. Our results show that the Rhizobium-Bradyrhizobium co-inoculation can enhance growth and grain yield of common beans and indicate that a plant growth-promoting mechanism besides BNF may be involved in the beneficial co-inoculation effect. However, further adjustments are needed to ensure a better response under field conditions.

Secondary Metabolites of Rhizobium tropici CIAT 899 Added to Bradyrhizobium spp. Inoculant Promote Soybean Growth and Increase Yield

Secondary Metabolites of Rhizobium tropici CIAT 899 Added to Bradyrhizobium spp. Inoculant Promote Soybean Growth and Increase Yield

Inoculation with plant growth-promoting bacteria and reduction of nitrogen fertilizer in herbage accumulation and nutritional value of Mavuno grass

Strategies that improve the use of nitrogen (N), by tropical forage grasses, can bring environmental and social gains. Thus, this study aimed to evaluate the forage productivity and the nutritional value (NV) of the hybrid Urochloa spp. cv. ´Mavuno`, under inoculation with plant growth-promoting bacteria (PGPB) and doses of N. The experiment was carried out in the field, for a period of 14 months, in a randomized block design, with nine treatments and four replications in plots of 9 m². Seed inoculations were tested with: Azospirillum brasilense Ab-V5 and Ab-V6 strains; Pseudomonas fluorescens CCTB 03 and co-inoculation with Rhizobium tropici CIAT 899 and A. brasilense Ab-V6, combined with 50 and 100 kg ha-1 of added mineral N; fertilization with N-mineral with these doses, without inoculation and the control treatment, without N and without inoculation. The bacteria were inoculated to the grass seeds. The forage was evaluated by determining the shoot dry weight yield (SDWY) and the levels of crude protein (CP), neutral detergent insoluble fiber (NDF), acid detergent insoluble fiber (ADF) and in vitro dry matter digestibility (IVDMD). The results were subjected to analysis of variance by the F test (p ≤ 0.05) and compared to each other by the Scott-Knott test at 5% probability. Inoculation with PGPB resulted in greater SDWY when associated with a dose of 50 kg ha-1 of N. In plants inoculated with A. brasilense Ab-V5 + Ab-V6 and P. fluorescens CCTB 03, associated with a dose of 50 kg ha-1 of N-mineral, SDWY was similar to that obtained with double the dose of N in the absence of inoculation. The inoculation with PGPB resulted in an increase in the NV of Mavuno grass, with emphasis on A. brasilense Ab-V5 + Ab-V6 and P. fluorescens CCTB 03, associated with the dose of 100 kg ha-1 of N-mineral that improved the nutritional value in all evaluated items. The inoculation with A. brasilense Ab-V5 + Ab-V6 was the most promising, followed by P. fluorescens CCTB 03, mainly in association with 50 kg ha-1 of N.

Effect of a mix of oligogalacturonides on symbiotic nitrogen fixation in common bean

The objective of this study was to evaluate the effect of a mix of pectic oligosaccharides in the common bean-rhizobia symbiosis. Cuba Cueto-25-9-N bean seeds were inoculated with Rhizobium tropici CIAT 899 and treated with a mix of oligogalacturonides, either by application to the seed (10 mg L-1, 1 ml per seed) at sowing and inoculation or by foliar spray (5 or 100 mg L-1, 1.5 ml per plant) to plants with two trefoils. Plant growth, nodulation, nitrogenase activity, and the gene expression of glutamine synthetase and glutamate synthase in nodules were evaluated in inoculated plants and oligogalacturonide-treated inoculated plants at 18 d post-inoculation. The oligogalacturonide-treated plants showed increases in shoot and root growth and number of nodules. Regarding nodule function, the oligogalacturonides increased the nitrogenase activity and the expression level of the gene coding for NADH-dependent glutamate synthase. The positive effects of oligogalacturonides resulted in higher effectiveness of the symbiotic nitrogen fixation. The application of oligogalacturonides can offer an alternative to increasing the symbiotic nitrogen fixation in common bean plants.

Proteome responses of Rhizobium tropici CIAT 899 upon apigenin and salt stress induction

Rhizobium tropici establishes symbiosis with several leguminous species, inducing nitrogen fixing nodules. Communication between rhizobia and legumes starts when the flavonoids secreted by plant roots interact with the R. tropici NodD protein to recognize plant signaling molecules present in the rhizosphere and begin the nodulation process. Some studies in R. tropici CIAT 899 have demonstrated that the induction of nod genes by saline stress influences the formation of nodules in plant roots. The aim of this study was to analyze the proteome profile of R. tropici CIAT 899 to investigate the molecular response of the bacterium under apigenin (flavonoid) and salt stress induction. The bacterium was grown in TY medium, considered as the control condition (CC), and in the presence of apigenin (AC) and NaCl (SSC). Total proteins were extracted and analyzed by two-dimensional electrophoresis (2-DE) and the differential spots were identified by MALDI-TOF MS/MS. A total of 105 differentially abundant protein spots were observed, including 2 present only in CC, 2 exclusive to AC and 6 to SSC. A total of 73% of these protein spots were identified by mass spectrometry. Proteins involved in transport and cellular processes were mostly increased in AC:CC and AC:SSC comparison, while in SSC:CC, these proteins were mostly decreased. Under salt stress, the bacterium reduced the uptake of molecules and energy consumption, reducing the abundance of ABC transporter proteins. These proteins are crucial for bacterial adaptation in environmental conditions and can be important for the nodulation process.

OnfD, an AraC-Type Transcriptional Regulator Encoded by Rhizobium tropici CIAT 899 and Involved in Nod Factor Synthesis and Symbiosis.

Rhizobium tropici CIAT 899 is a broad-host-range rhizobial strain that establishes symbiotic interactions with legumes and tolerates different environmental stresses such as heat, acidity, or salinity. This rhizobial strain produces a wide variety of symbiotically active nodulation factors (NF) induced not only by the presence of plant-released flavonoids but also under osmotic stress conditions through the LysR-type transcriptional regulators NodD1 (flavonoids) and NodD2 (osmotic stress). However, the activation of NodD2 under high-osmotic-stress conditions remains elusive. Here, we have studied the role of a new AraC-type regulator (named as OnfD) in the symbiotic interaction of R. tropici CIAT 899 with Phaseolus vulgaris and Lotus plants. We determined that OnfD is required under salt stress conditions for the transcriptional activation of the nodulation genes and therefore the synthesis and export of NF, which are required for a successful symbiosis with P. vulgaris Moreover, using bacterial two-hybrid analysis, we demonstrated that the OnfD and NodD2 proteins form homodimers and OnfD/NodD2 form heterodimers, which could be involved in the production of NF in the presence of osmotic stress conditions since both regulators are required for NF synthesis in the presence of salt. A structural model of OnfD is presented and discussed.IMPORTANCE The synthesis and export of rhizobial NF are mediated by a conserved group of LysR-type regulators, the NodD proteins. Here, we have demonstrated that a non-LysR-type regulator, an AraC-type protein, is required for the transcriptional activation of symbiotic genes and for the synthesis of symbiotically active NF under salt stress conditions.

Comparative Assessment of Different Crop Rotation Schemes for Organic Common Bean Production

The aim of the current study was to contribute to the establishment of sustainable organic crop rotation schemes for common bean under mild-winter climatic conditions. Common bean was cultivated according to organic or conventional farming practices during spring-summer in two successive years with crop and treatment during the preceding winter as either: (a) organic broccoli, (b) conventional broccoli, (c) organic faba bean used as green manure, or (d) fallow. Common bean was either inoculated with Rhizobium tropici CIAT 899 or non-inoculated, while faba bean was inoculated or non-inoculated with Rhizobium laguerreae VFLE1. Inoculating faba bean with rhizobia enhanced dry biomass production and biological N-fixing ability in both experimental years. Furthermore, organic farming did not restrict the yield of broccoli compared to conventional practices during the first year, while the reverse was the case in the second year, due to reduced soil N availability. Furthermore, green manure enhanced the fresh pod yield in the following organic crop of common bean in both years. The lowest yield was recorded in organically grown common bean when the preceding winter crop was organically grown broccoli in both years. Rhizobia inoculation of the common bean during the first year slightly increased atmospheric N fixation by common bean.

Rhizobium dioscoreae sp. nov., a plant growth-promoting bacterium isolated from yam (Dioscorea species).

This study investigated endophytic nitrogen-fixing bacteria isolated from two species of yam (water yam, Dioscorea alata L.; lesser yam, Dioscorea esculenta L.) grown in nutrient-poor alkaline soil conditions on Miyako Island, Okinawa, Japan. Two bacterial strains of the genus Rhizobium , S-93T and S-62, were isolated. The phylogenetic tree, based on the almost-complete 16S rRNA gene sequences (1476 bp for each strain), placed them in a distinct clade, with Rhizobium miluonense CCBAU 41251T, Rhizobium hainanense I66T, Rhizobium multihospitium HAMBI 2975T, Rhizobium freirei PRF 81T and Rhizobium tropici CIAT 899T being their closest species. Their bacterial fatty acid profile, with major components of C19 : 0 cyclo ω8c and summed feature 8, as well as other phenotypic characteristics and DNA G+C content (59.65 mol%) indicated that the novel strains belong to the genus Rhizobium . Pairwise average nucleotide identity analyses separated the novel strains from their most closely related species with similarity values of 90.5, 88.9, 88.5, 84.5 and 84.4 % for R. multihospitium HAMBI 2975T, R. tropici CIAT 899T, R. hainanense CCBAU 57015T, R. miluonense HAMBI 2971T and R. freirei PRF 81T, respectively; digital DNA–DNA hybridization values were in the range of 26–42 %. Considering the phenotypic characteristics as well as the genomic data, it is suggested that strains S-93T and S-62 represent a new species, for which the name Rhizobium dioscoreae is proposed. The type strain is S-93T (=NRIC 0988T=NBRC 114257T=DSM 110498T).

In-Vitro Plant Growth Promotion of Rhizobium Strains Isolated from Lentil Root Nodules under Abiotic Stresses

Plant growth-promoting rhizobia are known to improve crop performance by multiple mechanisms. However, the interaction between host plants and Rhizobium strains is highly influenced by growing conditions, e.g., heat, cold, drought, soil salinity, nutrient scarcity, etc. The present study was undertaken to assess the use of Rhizobium as plant growth promoters under abiotic stress conditions. Fifteen Rhizobium strains isolated from lentil root nodules were tested for phosphate solubilization activity (PSA) and phytohormones production under salt and drought conditions. The results showed that 15 Rhizobium strains were significant phosphate solubilizers, and indole acedic acid (IAA) and gibberellic acid (GA3) producers based on least significant difference (LSD) analysis (p ≤ 0.05). The highest rate of PSA was attributed to three strains namely, 1145N5, 1159N11, and 1159N32 with a range of 144.6 to 205.6 P2O5 (µg/mL). The highest IAA production was recorded in the strain 686N5 with 57.68 ± 4.25 µg/mL as compared to 50.8667 ± 1.41 µg/mL and 37.32 ± 12.59 µg/mL for Rhizobium tropici CIAT 899 and Azospirillum brasilense DSM-1690, respectively. Strain 318N2111 produced 329.24 ± 7.84 µg/mL of GA3 as against 259.84 ± 25.55 µg/mL for A. brasilense DSM-1690. R. tropici CIAT 899 showed tolerance to salt (5% NaCl) and drought (ψ = −2.6 MPa) stress, whereas strain 686N5 showed an extremely high level of salt-tolerance (5% NaCl) and moderate level of drought tolerance (ψ = −0.75 MPa). These results indicate different pathways for drought and salt tolerance mechanisms. The assessment of plant growth promoting (PGP) activities of Rhizobium showed differences between bacterial viability and bacterial PGP activity in terms of abiotic stress tolerance where bacterial PGP activity is interrupted before reaching the bacterial tolerance threshold. These results integrate a new concept of PGPR screening based on PGP activity under abiotic stress.