Comparative effectiveness of Pseudomonas and Serratia sp. containing ACC-deaminase for coinoculation with Rhizobium leguminosarum to improve growth, nodulation, and yield of lentil

Three plant growth-promoting rhizobacteria strains containing ACC-deaminase (Pseudomonas jessenii, Pseudomonas fragi, and Serratia fonticola) and Rhizobium leguminosarum were selected and characterized by conducting some experiments under axenic condition. The selected isolates had the potential to improve the growth of lentil seedlings under axenic conditions. Pot and field experiments were conducted to evaluate the potential of these selected strains for improving growth and yield of lentil under natural conditions. A classical triple response (reduction of stem elongation, swelling of hypocotyle, and change in the direction of growth) bioassay was also conducted to evaluate the effect of high ethylene concentration on the growth of etiolated lentil seedlings, and the performance of coinoculation was evaluated to reduce the classical triple response in comparison with cobalt (Co²⁺), a chemical inhibitor of ethylene. Results showed that coinoculation of Pseudomonas and Serratia sp. with R. leguminosarum significantly increased the growth and yield of lentil. However, synergistic/coinoculation effect of P. jessenii with R. leguminosarum was more pronounced compared to that with P. fragi and S. fonticola. It increased the number of pods per plant, number of nodules per plant, dry nodule weight, grain yield, and straw yield up to 76%, 196%, 109%, 150%, and 164% under pot and up to 98%, 98%, 100%, 82%, and 78%, respectively, under field conditions as compared to uninoculated control. Similarly, combined inoculation significantly increased N concentration of grains under both pot and field conditions. The results from classical triple response assay showed that the effects of classical triple response decreased due to coinoculation in etiolated lentil seedlings and due to a decrease in the ethylene concentration. It is suggested that the strategy adopted by Pseudomonas sp. containing ACC-deaminase with Rhizobium to promote nodulation and yield by adjusting ethylene levels could be exploited as an effective tool for improving growth, nodulation, and yield of lentil.     

Production of growth-promoting substances and high colonization ability of rhizobacteria enhance the nitrogen fixation of soybean when coinoculated with Bradyrhizobium japonicum

Understanding the interaction mechanisms between plant growth-promoting rhizobacteria (PGPR), leguminous crops, and rhizobia is necessary to effectively use PGPR in increasing the biological nitrogen fixation of legumes. We determined the coinoculation effects of Bradyrhizobium japonicum A1017 and a gusA-marked strain of Pseudomonas fluorescens 2137, P. fluorescens WCS365, Azomonas agilis 125, and Azospirillum lipoferum 137 on soybean [Glycine max (L.) Merr] cv. Enrei grown under axenic conditions. The gusA-marked rhizobacteria effectively colonized the root tips and surfaces near the roots tips with a colonization rate ranging from 7.50 to 8.62 log colony forming units (cfu) gfw^-1. P. fluorescens 2137 had the highest colonization activity on soybean roots whether inoculated alone or coinoculated with B. japonicum A1017. Coinoculation of P. fluorescens 2137 and B. japonicum A1017 increased the colonization of B. japonicum A1017 on soybean roots, nodule number, and acetylene reduction activity (ARA) at 10 and 20 days after inoculation. Moreover, the addition of sterile spent medium of P. fluorescens 2137 increased the growth of B. japonicum A1017 in yeast mannitol broth (YMB), indicating that P. fluorescens 2137 may have released substances that increased the rhizobial population. The results of this study suggest that the enhanced nodulation and ARA of soybean due to the high colonization of P. fluorescens on soybean roots could depend on the production of growth-promoting substances that stimulate the growth of B. japonicum. However, coinoculation with P. fluorescens WCS365 decreased the nodule number and ARA, despite its slight stimulation of the growth of B. japonicum on the roots, indicating that coinoculation effects are strain dependent.     

Effects of Plant-Growth-Promoting Rhizobacteria on Yield,Growth, and Some Physiological Characteristics of Wheat and Barley Plants

In 2009 a greenhouse experiment was conducted to determine the effects of boron (B) and plant growth-promoting rhizobacteria (PGPR) treatments, applied either alone or in combination, on yield, plant growth, leaf total chlorophyll content, stomatal conductance, membrane leakage, and leaf relative water content of wheat (Triticum aestivum L. cv. Bezostiya) and barley (Hordeum vulgare L. cv. Tokak) plants. Results showed that alone or combined B (0, 1, 3, 6, 9 kg ha^?1) and PGPR (Bacillus megaterium M3, Bacillus subtilis OSU142, Azospirillum brasilense Sp245, and Raoultella terrigena) treatments positively affected dry weight and physiological parameters searched in both species. Statistically significant differences were observed between bacterial inoculation and B fertilizer on root and shoot dry weight under non-cold-stress (NCS) and cold-stress (CS) conditions. Leaf total chlorophyll content (LTCC), stomatal conductance (SC), leaf relative water content (LRWC), and membrane leakage (ML) were negatively affected by CS conditions and decreased with reduced temperatures of media, but B and PGPR application alleviate the low-temperature deleterious effect in both species. The greatest SC and LRWC, and the lowest ML, were obtained by 6 kg B ha^?1 combined with R. terrigena treatment. The greatest LTCC in both NCS and CS conditions was observed with B. megaterium M3 application alone.     

Yield enhancement and phosphorus economy in lentil (Lens culinaris Medikus) with integrated use of phosphorus,Rhizobium and plant growth promoting rhizobacteria

The study evaluated the effects of phosphorus (0, 20, 30, and 40 kg P₂O₅ ha^?1) and biofertilizers [Rhizobium (Rhizobium leguminosarum bv viciae), plant growth promoting rhizobacteria (PGPR) (Pseudomonas fluorescens), Rhizobium + PGPR, and uninoculated control] in lentil. Application of 40 kg P₂O₅ ha^?1 resulted in the highest number of nodules, nodule dry weight, leghemoglobin content in nodules, chlorophyll content, yield attributes, and grain yield. Coinoculated treatment performed better than uninoculated control, and individual inoculations of Rhizobium and PGPR in terms of all above mentioned parameters. Application of 20 kg P₂O₅ ha^?1 + Rhizobium inoculation gave statistically similar and 20 kg P₂O₅ ha^?1 + Rhizobium + PGPR inoculation gave significantly higher grain yield than that by 40 kg P₂O₅ ha^?1 alone. The use of Rhizobium alone and Rhizobium + PGPR consortium can save not only 20 kg P₂O₅ ha^?1 but also increase the grain yield of lentil.     

Response of Soybean to dual Inoculation with Rhizobium japonicum and Phosphate dissolving Bacteria

A field experiment in silty clay soil was carried out to evaluate the effect of dual inoculation with Rhizobium japonicum and phosphate dissolving bacteria (PDB) on nitrogen and phosphorus uptakes as well as seed yield of soybean. Seed inoculation with PDB before planting did not induce marked enrichment of PDB counts in the rhizosphere. Inoculation with R. japonicum alone increased the dry weight, N-uptake and seed yield significantly. Further significant increases in P-uptake and seed yield were recorded by inoculation with rhizobia and fertilization with superphosphate. However, the inoculation with PDB in combination with R. japonicum in the presence of rock phosphate or superphosphate did not show significant increases in dry weight, N and P uptakes or seed yield as compared with the treatments inoculated with rhizobia alone.     

EFFECT OF PLANT GROWTH-PROMOTING RHIZOBACTERIA STRAIN ON FREEZING INJURY AND ANTIOXIDANT ENZYME ACTIVITY OF WHEAT AND BARLEY

In 2009 a greenhouse experiment was conducted to determine the effects of boron (B) and plant growth-promoting rhizobacteria (PGPR) on wheat (Triticum aestivum spp. vulgare cv ‘Bezostiya’) and barley (Hordeum vulgare cv ‘Tokak’) on plant growth, freezing injury, and antioxidant enzyme capacity. Results showed that boron (0, 1, 3, 6, 9 kg B ha^?1) and PGPR application (Bacillus megaterium M3, Bacillus subtilis OSU142, Azospirillum brasilense Sp245 and Raoultella terrigena) at which 50% of leaves were injured (LT₅₀) values and ice nucleation activities in both plants were found statistically significant. Boron application with all PGPR strains decreased LT₅₀ values in wheat and barley plants under noncold stress (NCS) and cold stress conditions (CS). There were statistically significant differences between bacterial inoculation and B fertilizer in terms of root and shoot dry weight under NCS and CS conditions. Reactive oxidative oxygen species (ROS) and antioxidant enzyme activities (SOD, POD, CAT) were negatively affected CS conditions and decreased with reduced temperatures of media, but B and PGPR applications alleviated the low-temperature deleterious effects in both plants species tested. The lowest ROS and antioxidant enzyme (SOD, POD, CAT) of wheat and barley were observed with 6 kg B ha^?1 with R. terrigena.     

Response of soybean cultivars toward inoculation with three arbuscular mycorrhizal fungi and Bradyrhizobium japonicum in the alluvial soil

The aim of this study was to assess the comparative efficacy of three arbuscular mycorrhizal fungi (AMF) combined with cultivar specific Bradyrhizobium japonicum (CSBJ) in soybean under greenhouse conditions. Soybean seeds of four cultivars namely JS 335, JS 71-05, NRC 2 and NRC 7 were inoculated with three AM fungi (Glomus intraradices, Acaulospora tuberculata and Gigaspora gigantea) and CSBJ isolates, individually or in combination, and were grown in pots using autoclaved alluvial soil of a non-legume cultivated field of Ajmer (Rajasthan). Assessment of the data on nodulation, plant growth and seed yield revealed that amongst the single inoculations of three AMF, G. intraradices produced the largest increases in the parameters studied followed by A. tuberculata and G. gigantea indicating that plant acted selectively on AMF symbiosis. The dual inoculation with AMF + CSBJ further improved these parameters demonstrating synergism between the two microsymbionts. Among all the dual treatments, G. intraradices + B. japonicum brought about the largest increases in the studied characteristics particularly in seed weight per plant that increased up to 115.19%, which suggested that a strong selective synergistic relationship existed between AMF and B. japonicum. The cv. JS 335 exhibited maximum positive response towards inoculation. The variations in efficacy of different treatments with different soybean cultivars indicate the specificity of the inoculation response. These results provide a basis for selection of an appropriate combination of specific AMF and Bradyrhizobium which could further be utilized for verifying the symbiotic effectiveness and competitive ability of microsymbionts under field conditions of Ajmer region.     

Application of repetitive extragenic palindromic (REP)-PCR and enterobacterial repetitive intergenic consensus (ERIC)-PCR analysis to the identification and classification of Japan and Thai local isolates of Bradyrhizobium japonicum,Shinorhizobium meliloti,Rhizobium leguminosarum

Repetitive extragenic palindromic (REP)-PCR and enterobacterial repetitive intergenic consensus (ERIC)-PCR analysis was applied to the identification and classification of local isolates of 44 Bradyrhizobium japonicum, 7 Sinorhizobium meliloti, 10 Rhizobium leguminosarum strains from Japan and Thai. Using genomic DNA of the 61 strains, both REP and ERIC primers induced reproducible PCR band patterns, although REP-PCR generated more bands and appeared to be more useful for distinguishing the isolates from each other. Using mixed matrix data from both REP- and ERIC-PCR data, it become possible to distinguish all the isolates analyzed in this experiment from each other. When cluster analysis was applied to both PCR matrix data of 44 B. japonicum isolates, only the REP-PCR dendrogram showed a grouping profile corresponding to the exo-polysaccharide phenotype with a exceptions. When the matrix data of R. leguminosarum and S. meliloti were subjected to cluster analysis, S. meliloti appeared to form a different subgroup from R. leguminosarum in the dendrogram of REP-PCR data except for one strain. In the case of ERIC-PCR, isolates of R. leguminosarum from northern Thailand formed a separate subgroup from other R. leguminosarum and S. meliloti which were dispersed in the dendrogram. These data suggest that REP-PCR and ERIC-PCR were effective for the identification of individual isolates even though the isolates showed a wide genetic diversity and the same phenotype. When the data of the local isolates from Japan and Thailand were subjected to cluster analysis, REP- and ERIC-PCR analysis revealed different grouping characteristics.     

Effect of some fungicide seed treatments on the survival of Bradyrhizobium japonicum and on the nodulation and yield of soybean [Glycine max. (L) Merr.]

Several commercial fungicide seed treatments were evaluated for their possible effect on the survival of Bradyrhizobium japonicum on seeds and on the nodulation and yield of soybeans in a greenhouse and a field experiment. quinolate Pro (carbendazim and oxine copper), Vitavax 200FF (carboxin and thiram), and Monceren (pencycuron) had a small effect or no effect on the survival of B. japonicum and on the nodulation and yield of soybeans. They can thus be considered compatible with soybean seed inoculation. Germipro UFB (carbendazim and iprodione), Apron 35J (metalaxyl), and Tachigaren (hymexazol) decreased B. japonicum survival and the nodulation and yield of soybeans and thus cannot be considered compatible with soybean seed inoculation.     

Selection of type A and type B strains for improving symbiotic effectiveness on non-Rj and Rj4 genotype soybean varieties

The selection of effective rhizobia for higher efficiency nitrogen fixation is one of the most important steps for inoculant production. Therefore, this experiment was conducted to select the most effective type A and type B strains for specific Rj-gene harboring soybean varieties and to test the symbiotic effectiveness of selected strains on different Rj-gene harboring soybean varieties. Screening experiments using the specific soybean varieties were done with a completely randomized design and three replications in this study. Evaluation of the effective Myanmar Bradyrhizobium strains for plant growth, nodulation and N₂ fixation were studied in pot experiments using sterilized vermiculite in the Phytotron (controlled-environmental condition). Then, a pot experiment was conducted using Futsukaichi soil in the screen house (natural environmental condition). The N₂ fixation ability of soybean was evaluated by acetylene reduction activity (ARA) and the relative ureide index method. In the first screening experiment, type A and type B strains with higher nitrogen fixation and proper nodulation on their respective soybean cultivars were selected for the next screening. In the second screening, Bradyrhizobium elkanii AHY3-1 (type A), Bradyrhizobium japonicum SAY3-7 (type A), B. elkanii BLY3-8 (type B) and B. japonicum SAY3-10 (type B) isolates, which showed higher nitrogen fixation and nodulation in Yezin-3 (Rj₄) and Yezin-6 (non-Rj), were selected for the next experiment. In the third screening experiment, SAY3-7 and BLY3-8, which had higher nitrogen fixing potential and proper nodulation, were selected as effective isolates. These two isolates were compatible with non-Rj and Rj₄ soybean varieties for nodulation and nitrogen fixation. Based on the results of the screening experiment, these two strains were tested for their symbiotic efficacy in Futsukaichi soil. This study shows that inoculation treatment of SAY3-7 and BLY3-8 significantly increased plant growth, nodulation, and N₂ fixation at the V6, R3.5 and R8 stages in Yezin-3 (Rj₄) and/or Yezin-6 (non-Rj), and the seed yield at R8 stage, in Yezin-3 (Rj₄) and Yezin-6 (non-Rj) soybean varieties compared with the control treatment. It can be concluded that SAY3-7 and BLY3-8 are suitable for inoculant production because of their higher nitrogen fixation ability, proper nodulation and better productivity of Myanmar soybean cultivars.     

Selection of Rhizobium isolates able to improve productivity of lentil (Lens culinaris Medik)

Abstract

Nitrogen-fixing efficiency is influenced by genetic combination of rhizobia strain/legume genotype. In this research, 48 rhizobia isolates from lentil (Lens culinaris Medik) were characterized for DNA polymorphism and assayed for their ability to improve, in open field, grain yield of lentil. By DNA polymorphism analysis, only four haplotypes were observed for Rhizobium isolates from lentil seeds, whereas a higher genetic variability was observed for Rhizobium isolates obtained from root-tubercles. The performance of the 11 rhizobia haplotypes was analysed in the field after inoculation on two Italian lentil ecotypes. Two isolates of rhizobia induced a significant improvement of grain yield (plus from 37% to 40% with respect to non-inoculated plants) on the Colfiorito ecotype, whereas no significant improvement of yield was obtained on Colliano.     

Inoculation response in kidney beans (Phaseolus vulgaris,L.) to vesicular-arbuscular mycorrhizal fungi and rhizobia in non-sterilized soil

The effect of dual inoculation on three local cultivars (Miss Kelly, Portland Red, Round Red) of red kidney beans (Phaseolus vulgaris, L.) with four strains of Rhizobium leguminosarum bv. phaseoli and three species of vesicular-arbuscular mycorrhizal (VAM) fungi was examined in a clay loam soil. Rhizobial strains B 17 and B 36, each paired with Glomus pallidum or G. aggregatum, were the most effective pairings for cv. Miss Kelly. Inoculation of Miss Kelly with any of these pairings significantly (P=0.05) increased growth, number of nodules, nodule dry weight, mycorrhizal colonization, and shoot N and P content than other pairings. The growth response by cv. Portland Red was significantly improved by pairings of B 36 or B 17 with any of the three VAM fungi. For both cultivars (Miss Kelly and Portland Red), CIAT 652 or T 2 paired with VAM fungi did not give a positive growth response. In contrast, for cv Round Red the T 2 rhizobial strain in combination with any of the three VAM fungi showed a significant (P=0.05) growth improvement in all parameters. Our results suggest that while dual inoculation of VAM fungi and rhizobia significantly improves the growth response by red kidney beans, the best pairings of VAM fungus and rhizobia for each cultivar need to be carefully selected.     

Renodulation and characterization of Rhizobium isolates from cicer milkvetch (Astragalus cicer L.)

In 1993 and 1994, 12 bacterial isolates were isolated from root nodules of cicer milkvetch (Astragalus cicer). In the tests for nodulation of A. cicer by these bacterial isolates, five were found to form hypertrophic structures, while only two formed true nodules. These true nodules were formed in a sterilized soil system. This system might be able to act as a DNA donor to provide residual DNA to other microbes in the soil. The rhizobial isolates were thought to have lost genetic material crucial to nodulation during the isolation process. This hypothesis was supported by an experiment in which isolate B2 was able to nodulate A. cicer in vermiculite culture after being mixed with heat-killed rhizobia, Rhizobium leguminosarum bv. trifolii and R. loti. The nodulation would not occur in vermiculite culture system without the heat-killed rhizobia. Based on the biochemical data, the B2 and 9462L, which formed true nodules with A. cicer, were closely related. The rhizobia type cultures that nodulate A. cicer include Bradyrhizobium japonicum, Rhizobium leguminosarum bv. trifolii, R. leguminosarum bv. viceae, and R. loti. All of these rhizobia were from different cross-inoculation groups. The B2 and 9462L isolates could only nodulate Medicago sativa, Phaseolus vulgaris, and Melilotus officinalis, but not these species within the genus from which they were isolated: Astragalus. The traditional cross-inoculation group concept obviously does not fit well in the classification of rhizobia associated with Astragalus. The rhizobia isolated from A. cicer can be quite different, and the rhizobia able to renodulate A. cicer also quite diverse. Received: 27 June 1996     

Combined effects of plant growth‐promoting rhizobacteria and genistein on nitrogen fixation in soybean at suboptimal root zone temperatures

Application of plant growth‐promoting rhizobacteria (PGPR) or the plant to bacteria signal molecule genistein has been shown to increase nodulation and nitrogen (N) fixation by soybean [Glycine max (L.) Merr.] over a range of root zone temperatures (RZTs) and, specifically, off‐sets at least some of the ill‐effects of low RZTs. Two sets of controlled‐environment experiments, one on a growth bench and the other in a greenhouse, were conducted to examine the combined ability of both PGPR and genistein to reduce the negative effects of low RZT on soybean nodulation and N fixation. Each of two the PGPR strains, Serratia proteamaculans 1–102 and Serratia liquefaciens 2–68 were co‐inoculated with Bradyrhizobium japonicum USDA 110 preincubated with 17.5 (somewhat inhibitory), and 15°C (very inhibitory). At RZTs of 25 and 17.5°C PGPR strains and genistein in combination increased the number of nodules and the amount of Nn fixed. The most stimulatory effect was observed at 17.5°C for the combination: S. proteamaculans 1–102 plus B. japonicum USDA 110 pre‐incubated in 15 μM genistein under greenhouse conditions. For most treatment combinations the stimulatory effects of PGPR and genistein were additive at RZTs of 17.5 and 25°C. Surprisingly, the combination of these two factors resulted in antagonism at the very inhibitory RZT of 15°C. The results suggest that the negative effects of certain low RZTs could be more effectively off‐set by combined treatments of PGPR plus geneistin pre‐incubation of rhizobial cultures than by their individual treatment.     

Synergistic effects of plant-growth promoting rhizobacteria and Rhizobium on nodulation and nitrogen fixation by pigeonpea (Cajanus cajan)

Plant‐growth promoting rhizobacteria (PGPR), in conjuction with efficient Rhizobium, can affect the growth and nitrogen fixation in pigeonpea by inducing the occupancy of introduced Rhizobium in the nodules of the legume. This study assessed the effect of different plant‐growth promoting rhizobacteria (Azotobacter chroococcum , Azospirillum brasilense, Pseudomonas fluorescens, Pseudomonas putida and Bacillus cereus) on pigeonpea (Cajanus cajan (L) Milsp.) cv. P‐921 inoculated with Rhizobium sp. (AR‐2–2 k). A glasshouse experiment was carried out with a sandy‐loam soil in which the seeds were treated with Rhizobium alone or in combination with several PGPR isolates. It was monitored on the basis of nodulation, N₂ fixation, shoot biomass, total N content in shoot and legume grain yield. The competitive ability of the introduced Rhizobium strain was assessed by calculating nodule occupancy. The PGPR isolates used did not antagonize the introduced Rhizobium strain and the dual inoculation with either Pseudomonas putida, P. fluorescens or Bacillus cereus resulted in a significant increase in plant growth, nodulation and enzyme activity over Rhizobium‐inoculated and uninoculated control plants. The nodule occupancy of the introduced Rhizobium strain increased from 50% (with Rhizobium alone) to 85% in the presence of Pseudomonas putida. This study enabled us to select an ideal combination of efficient Rhizobium strain and PGPR for pigeonpea grown in the semiarid tropics.     

Characterization of local rhizobia in Thailand and distribution of malic enzymes

TWenty-six isolates were obtained from nodules of various legume plants (Glycine max, Vigna sinensis, Arachis hypogaea, Desmanthus virgatus, Acacia mangium, Centrosema pascuorum, Pterocarpus indicus, Xylia xylocarpa, and Sesbania rostrata) in Thailand. After confirming their nodulation and nitrogen-fixing abilities, they were identified by 16S rRNA gene analysis as Bradyrhizobium japonicum, Bradyrhizobium elkanii, Rhizobium leguminosarum, Rhizobium gallicum, and Rhizobium galegae. Using these local isolates, the distribution of the activities of both NAD⁺-dependent (DME: EC 1.1.1.39) and NADP⁺-dependent (TME: EC 1.1.1.40) malic enzymes was surveyed. The malic enzyme activities were present in all the isolated rhizobia and in other 17 local Bradyrhizobium strains in Thailand. In almost all the rhizobia, the DME activity predominated whereas the TME activity predominated only in the Rhizobium gallicum strains that were major symbionts of Sesbania rostrata. Southern hybridization analysis was performed to survey the distribution of the malic enzyme genes among the local rhizobia, which are similar to those of B. japonicum. DNA probes (ME1 for DME and ME2 for TME) were prepared by polymerase chain reaction (PCR) using degenerated primers from conserved regions of the protein sequences of bacterial malic enzymes. Southern blot analysis with ME1 as a probe showed a single band in about half of the isolates, especially in B. japonicum and R. leguminosarum strains, suggesting the wide distribution of such DME genes among local rhizobia. In contrast, Southern blot analysis with ME2 as a probe detected a single band only in five B. japonicum strains, suggesting that the TME genes, which are similar to those of B. japonicum, would be unique in a group of B. japonicum.     

PLANT GROWTH PROMOTING RHIZOBACTERIA INCREASE GROWTH,YIELD AND NITROGEN FIXATION IN PHASEOLUS VULGARIS

Nitrogen (N) fixation by legume-Rhizobium symbiosis is important to agricultural productivity and is therefore of great economic interest. Growing evidence indicates that soil beneficial bacteria can positively affect symbiotic performance of rhizobia. The effect of co-inoculation with plant growth-promoting rhizobacteria (PGPR) and Rhizobium, on nodulation, nitrogen fixation, and yield of common bean (Phaseolus vulgaris L.) cultivars was investigated in two consecutive years under field conditions. The PGPR strains Pseudomonas fluorescens P-93 and Azospirillum lipoferum S-21 as well as two highly effective Rhizobium strains were used in this study. Common bean seeds of three cultivars were inoculated with Rhizobium singly or in a combination with PGPR to evaluate their effect on nodulation and nitrogen fixation. A significant variation of plant growth in response to inoculation with Rhizobium strains was observed. Treatment with PGPR significantly increased nodule number and dry weight, shoot dry weight, amount of nitrogen fixed as well as seed yield and protein content. Co-inoculation with Rhizobium and PGPR demonstrated a significant increase in the proportion of nitrogen derived from atmosphere. These results indicate that PGPR strains have potential to enhance the symbiotic potential of rhizobia.     

Bacillus subtilis NRRL B‐30408 inoculation enhances the symbiotic efficiency of Lens esculenta Moench at a Himalayan location

Field‐based experiments were conducted to evaluate the promotion abilities of Bacillus subtilis NRRL B‐30408 for growth of lentil (Lens esculenta Moench) at a mountain location of Indian Himalaya in two consecutive years. Observations were recorded for plant growth, yield, nodulation, root colonization by arbuscular mycorrhizal and endophytic fungi, and other related parameters. A positive influence of bacterial inoculation on plant biomass and yield‐related parameters was recorded in both years. The significant increase in growth and nodule numbers as well as leghaemoglobin and protein concentrations of nodules indicated an enhancement in efficiency of the Rhizobium–legume symbiosis due to bacterial inoculation. An increase in protein concentration was also recorded for shoots, leaves, and seeds. Due to bacterial inoculation, there was an increase in colonization by endophytic fungi and a simultaneous decrease in colonization by arbuscular mycorrhizal fungi in roots. Based on the results of this field study, inoculation with suitable plant growth–promoting rhizobacteria instead of dual inoculation is suggested as a better option for improving the yield and related attributes of a primary dietary legume such as lentil.     

Impact of single and co-inoculations with Rhizobial and PGPR isolates on chickpea (Cicer arietinum) in cereal-growing zone soil

Strains isolated from chickpea (Cicer arietinum L.) rhizospheric soil from selected sites in Algeria were screened for their plant-growth-promoting potential, for indole acetic acid production and P solubilization ability. Then, we selected native rhizobial strains with high nitrogen-fixing potential. On the basis of their efficiency under controlled conditions, two plant-growth-promoting rhizobacteria (PGPR) isolates and three nodulating bacteria were selected. Then, the effect of single PGPR isolates inoculation was compared to their combination with rhizobial inoculants on plant growth, on native cereal-growing soils under greenhouse conditions. No effects were observed on chickpea yield by using rhizobial inoculation alone, nor by PGPR-rhizobial co-inoculation on two soils presenting weak and no nodulation pattern in natural conditions. Only PGPR inoculation improved growth of plants on soil with no nodulation pattern. These findings emphasized inoculation on native soils at a little scale before large assays on field because no one could predict inocula behavior with native soil microflora.     

Effectiveness and competitiveness of spontaneous antibiotic-resistant mutants of Rhizobium leguminosarum and Rhizobium japonicum

Thirteen doubly-marked antibiotic-resistant isolates of Rhizobium leguminosarum and three resistant isolates of R. japonicum were developed and tested for effectiveness of N₂-fixation and competitiveness. Ninety-three percent of the mutants isolated were less competitive in nodule-forming ability, than their parents. However, only 38% of the R. leguminosarum isolates were reduced in N₂-fixation effectiveness when compared to their parents. Doubling times of the R. leguminosarum mutants increased when compared to their parents. No correlation was found between nodulation competitiveness and doubling time of the mutants. R. leguminosarum mutants in the nodules were evaluated using a new procedure, which facilitates the handling of large numbers of nodules.