Cloning and characterization of a NADP+-malic enzyme gene from Bradyrhizobium japonicum USDA110

Malic enzymes have been considered to play a key role in energy metabolism for nitrogenase reaction in bacteroids. To elucidate the physiological role of the malic enzymes in Bradyrhizobium japonicum bacteroids, a putative malic enzyme gene Bjtme1 was cloned by polymerase chain reaction (PCR) using degenerated primers from conserved regions of the protein sequences of bacterial malic enzymes and draft sequence data of the Bradyrhizobium japonicum USDA110 genome sequence project. To confirm the characteristics of the Bjtme1 gene, the protein encoded by this gene was over-expressed using a pET32a(+) system and it exhibited a NADP⁺-malic enzyme (EC 1.1.1.40) activity, indicating that Bjtme1 was the gene of the NADP⁺-malic enzyme. This is the first report on the cloning and characterization of the NADP⁺-malic enzyme gene from B. japonicum, and the gene structure was compared with that of NADP⁺-malic enzyme genes of other rhizobia.     

Genetic diversity of fast-and slow-growing soybean rhizobia determined by random amplified polymorphic DNA analysis

The genetic relationships among six strains of rhizobia, including three strains of Rhizobium fredii and three strains of Bradyrhizobium japonicum, was determined using random amplified polymorphic DNA (RAPD) technique. In this study, 46 arbitrary 10mer primers were employed for RAPD, generating a total of 251 informative fragments. A dendrogram of phylogenetic relationships among the six strains was constructed. The results indicated that geographical distribution may affect phylogeny, as there were closer relationships among the four Taiwanese strains, SB138, SB562, SB368 and SB651, than between these strains and USDA192, which originated from mainland China. The strain USDA110, obtained from the United States, was used in the parsimony analysis. The greatest similarity (55.6%), existed between two strains of B. japonicum, SB562 and SB138, which both, and the lowest R. fredii (44.4%) between two strains of R. fredii, SB368 and USDA192. We also found a RAPD marker specific to the four Taiwanese SB strains used in the study. The RAPD technique is a potential tool for the identification of the genetics and systematics of different populations. Received: 23 January 1997     

Genetic diversity and phylogeny of indigenous rhizobia from cowpea [<Emphasis Type="Italic">Vigna unguiculata</Emphasis> (L.) Walp.]

16S rRNA RFLP, 16S rRNA sequencing, 16S-23S rRNA Intergenetic Spacer (IGS) RFLP and G-C rich random amplified polymorphic DNA (RAPD) assays were conducted to genetically characterise indigenous cowpea [Vigna unguiculata (L.) Walp.] rhizobia from different geographic regions of China. Isolated cowpea rhizobia comprised six 16S rRNA genospecies. Genotype I was composed of 14 isolated strains and the reference strains of B. japonicum and B. liaoningense. This group was divided into two sub-groups respectively related to B. japonicum and B. liaoningense by 16S rRNA sequencing, IGS restriction fragment length polymorphism and RAPD assays. Genotype II composed of 27 isolates from a variety of geographic regions. Four different assays confirmed this group was genetically distinct from B. japonicum and B. liaoningense and probably represent an uncharacterised species. Strains isolated from Hongan, Central China and B. elkanii were grouped to genotype III. Strain DdE4 was solely clustered into genotype IV and related to Rhizobium leguminosarum. Genotypes V and VI consisted of six fast-growing isolates and clustered with reference strain of Sinorhizobium fredii. Comparing with the miscellaneous slow-growing isolates, fast-growing isolates mainly isolated from cowpea cultivar Egang I exhibited strict microbe–host specificity except SjzZ4. Nucleotide sequences reported were deposited in the GenBank with the accession numbers DQ786795–DQ786804.     

Genetic variability of soybean bradyrhizobia populations under different soil managements

Due to their ecologic and economic importance, bradyrhizobia have been extensively studied in recent years. Since 1992, Bradyrhizobium elkanii SEMIA 587 and SEMIA 5019 and Bradyrhizobium japonicum SEMIA 5079 and SEMIA 5080 have been widely used in most Brazilian soybean fields. The objective of this work was to estimate the genetic variability of bradyrhizobial isolates recovered from soils under rhizobial inoculation and different soil managements. Only 25% of the isolates demonstrated high similarities to the original strains, and a strong correlation was obtained between the bradyrhizobial genetic variability and soil management. A high level of genetic diversity was observed both within isolates (H = 5.46) as well as among the different soil practices. Soil under no-tillage presented a higher bradyrhizobia diversity compared with bradyrhizobia isolated from soil under conventional tillage. Serological characterization also indicated that B. elkanii strains SEMIA 587 and SEMIA 5019 were more competitive and presented a higher nodular occupancy capacity than strains belonging to B. japonicum species in Southern Brazilian soils.     

Ability of free-living cells ofBradyrhizobium japonicum to denitrify in soils

Summary Experiments to assess the ability of free-living cells of six strains of soybean rhizobia (Bradyrhizobium japonicum USDA 76, 94, 110, 122, 123, and 135) to denitrify nitrate in five soils showed that although some strains ofB. japonicum have the capacity to rapidly denitrify nitrate in soils under anaerobic conditions, it is unlikely that the numbers of soybean rhizobia commonly found under field conditions are sufficient to significantly influence either the extent or the products of denitrification in soil. It is our general conclusion that the advantages, if any, that the ability to denitrify conveys to rhizobia or to the rhizobia-legume symbiosis are not offset by increased losses of plant-available N when denitrifying strains of rhizobia are present as free-living cells in soil.     

Intrinsic antibiotic resistance and competition in fast- and slow-growing soybean rhizobia on a hybrid of Asian and US cultivars

Summary Six fast-growing soybean rhizobia (Rhizobium fredii) and thirteen slow-growing soybean rhizobia (Bradyrhizobium japonicum) were examined for resistance to 10 antibiotics. Axenic studies were carried out to determine the competitiveness of dual-strain inocula consisting of fast- and slow-growing rhizobia isolated from subtropical-tropical soils for nodule occupancy on a hybrid of Asian and US soybean cultivars. Nodule occupancy was determined by intrinsic resistance to erythromycin and neomycin. The results showed wide variability in resistance to 10 antibiotics for fast- and slow-growing rhizobia. The intrinsic antibiotic resistance of fast- and slow-growing rhizobia was extremely high against nalidixic acid (400 g ml^–1) and penicillin (200 g ml^–1). The competitive ability of inoculant strains for nodule occupancy varied for different combination sets and with the plant growing media. Our results show that fast-growing rhizobia nodulate a hybrid of Asian and US soybean cultivars. Fast-growing soybean rhizobia did not completely exclude nodulation by the slow-growing strains, which formed 0–79% nodules, depending on the strain used in the inoculum.     

Polyphasic characterization of mung bean (Vigna radiata L.) rhizobia from different geographical regions of China

Polyphasic characterization of 54 indigenous mung bean (Vigna radiata L.) rhizobia from different geographic regions of China was determined by analyzing the variability of 16S rRNA gene RFLP, 16S–23S rRNA gene Intergenetic Spacer (IGS) RFLP, G-C rich RAPD and phenotype assays. Based on these characteristics, mung bean rhizobia were clustered into four groups. Group I comprised 16 slow-growing isolates from a variety of geographic regions. This group was genetically distinct from Bradyrhizobium japonicum and Bradyrhizobium liaoningense, and may represent a new species. Group II was composed of 18 isolates, which could be sub-divided into two sub-groups that were respectively related to B. japonicum and B. liaoningense. Group III comprised 12 isolates from South China and clustered together with Bradyrhizobium elkanii. Group IV formed a miscellany of 8 fast-growing isolates variously related to the genera Sinorhizobium, Rhizobium and Mesorhizobium.     

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.     

Nitrogen fixation capacity and nodule occupancy by Bradyrhizobium japonicum and B. elkanii strains

 In a previous study soybean Bradyrhizobium strains, used in Brazilian studies and inoculants over the last 30 years, and strains adapted to the Brazilian Cerrados, a region frequently submitted to environmental and nutritional stresses, were analyzed for 32 morphological and physiological parameters in vivo and in vitro. A cluster analysis allowed the subdivision of these strains into species Bradyrhizobium japonicum, Bradyrhizobium elkanii and a mixed genotype. In this study, the bacteria were analyzed for nodulation, N₂ fixation capacity, nodule occupancy and the ability to increase yield. The goal was to find a relationship between the strain groups and the symbiotic performance. Two strains of Brazilian B. japonicum showed higher rates of N₂ fixation and nodule efficiency (mg of N mg^–1 of nodules) under axenic conditions. These strains also showed greater yield increases in field experiments when compared to B. elkanii strains. However, no differences were detected between B. japonicum and B. elkanii strains when comparing nodule occupancy capacity. The adapted strains belonging to the serogroup B. elkanii SEMIA 566, most clustered in a mixed genotype, were more competitive than the parental strain, and some showed a higher capacity of N₂ fixation. Some of the adapted strains, such as S-370 and S-372, have shown similar N₂ fixation rates and nodulation competitiveness to two Brazilian strains of B. japonicum. This similarity demonstrates the possibility of enhancing N₂ fixing ability, after local adaptation, even within B. elkanii species. Differences in the DNA profiles were also detected between the parental SEMIA 566 and the adapted strains by analyses with the ERIC and REP-PCR techniques. Consequently, genetic, morphological and physiological changes can be a result of adaptation of rhizobia to the soil. This variability can be used to select strains capable of increasing the contribution of N₂ fixation to soybean nutrition. Received: 28 May 1997     

Antibiotic-resistant mutants identified from nodules of uninoculated soybeans grown in a strongly acidic soil

Two hypotheses that antibiotic-resistant nodule isolates from uninoculated soybeans grown in a strongly acidic soil were naturally occurring rhizobia which had acquired resistance to spectinomycin and streptomycin or were contaminants from adjacent, inoculated treatments, were tested in laboratory experiments. Soybean nodule isolates (166) as well as 48 cowpea and 89 Rhizobium japonicum strains were used in tests of resistance to six concentrations (0–500 μg ml^?1) of kanamycin, spectinomycin and streptomycin, tolerance of stresses of pH 4.6, with or without 50 μm Al, and serological cross-reactivity.More strains from the strongly acidic soil were resistant to the antibiotics than from slightly acidic soils, but resistance to antibiotics did not necessarily entail resistance to pH 4.6 or to 50 μm Al. Twenty-three nodule isolates which were resistant to spectinomycin or streptomycin cross-reacted with antisera of the inoculum strains, indicating that they were contaminants. None of 59 antibiotic-sensitive nodule isolates from uninoculated plants and none of 31 from inoculated plants cross-reacted with the antisera. All 53 antibiotic-resistant isolates from nodules of plants in inoculated plots cross-reacted with test antisera, indicating stability of the antibiotic markers.Cowpea rhizobia were generally more resistant to the antibiotics and more tolerant of pH 4.6 and 50 μm Al than were R. japonicum. Among strains of R. japonicum, slow growers were more resistant to antibiotics than moderately fast growers.     

Effects of some Nigerian Bradyrhizobium inoculants on the performance of three cowpea cultivars (Vigna unguiculata) in two field plots

Summary We assessed the effectiveness of three locally made lignite, subbituminous coal and cowmanure-based cowpea Bradyrhizobium inoculants in comparison with a peat-based imported Bradyrhizobium incoculant in a two-field plot investigation. The local inoculants were prepared by incorporating three rhizobia strains (Ife CR9, Ife CR15 and Bradyrhizobium japonicum) into each of the above carrier materials and were used to inoculate three cowpea seed varieties: TVU 1190, IT 82E-60 and Ife brown. With lignite-based He CR9 inoculated into TVU 1190 seeds, total N content of the plants was 178.6 mg/plant compared with only 64.3 mg/plant for the uninoculated nitrate-free control plants. With Nigerian lignite, sub-bituminous coal and cow manure as carriers for cowpea rhizobia, the cowpea yield of the inoculated plants increased by 72%, 54% and 10%, respectively, compared with uninoculated plants, while the peat-based inoculant gave a 25% increase in cowpea yield. With lignite-based Ife CR9 inoculated into Ife brown seeds, total N content of the plants was 149.1 mg/plant, but with inoculation by lignite-based B. japonicum, total N content of the treated Ife brown plant was 132.4 mg/plant. Thus, the native Ife CR9 strain seems to be slightly better adapted to tropical conditions than the imported B. japonicum.     

Rhizobiophage effects on nodulation,nitrogen fixation,and yield of field-grown soybeans (Glycine max L. Merr.)

Summary Previous laboratory and greenhouse studies have shown that phages significantly reduce soil populations of homologous rhizobia. Reductions in nodulation and N₂ fixation have also been observed. The purpose of the current study was to examine the effect of a phage specific ofBradyrhizobium japonicum USDA 117 on nodulation, nodule occupancy, N₂ fixation and soybean growth and yield under field conditions. The phage was inoculated in combination withB. japonicum USDA 117 and/orB. japonicum USDA 110 (resistant strain) into a rhizobia-free sandy loam soil and planted toGlycine max (L.) Merr. Williams. When the phage was applied to soil inoculated withB. japonicum USDA 117 alone, significant reductions in nodule weight and number, shoot weight, foliar N, nitrogenase activity, and seed index were observed. When, however, the soil also contained the non-homologous strain,B. japonicum USDA 110, no significant effects on any of these parameters were found. Nodule occupancy by competing strains ofB. japonicum USDA 110 and USDA 117 was also affected by the phage. In soil which did not contain the phage, 46% and 44% of the identified nodules were occupied by USDA 110 and 117, respectively. When the phage was present in the soil, nodule occupancy byB. japonicum USDA 117 was reduced to 23%, while occupancy byB. japonicum USDA 110 was increased to 71%. These results suggest that nodulation by selected strains of rhizobia can be restricted and nodulation by more effective, inoculated strains can be increased through the introduction of a homologous phage to soils.     

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     

Effects of symbiosis with Rhizobium fredii on transport of fixed nitrogen in the xylem of soybean plants

An experiment was conducted to identify the main nitrogenous compound transported in the xylem sap of soybean plants nodulated with Rhizobium fredii. Soybean (Glycine max L. Merr.) cultivars, wild type Bragg (nod⁺, fix⁺) and its nitrate tolerant, hypernodulating mutant ntsll16 (nod⁺⁺, fix⁺) were used for this experiment. These soybean plants were inoculated with a slowgrowing rhizobium, Bradyrhizobium japonicum USDAllO or fast-growing rhizobia consisting of a mixture of R. fredii USDA191, USDA193, and USDA-194 and grown in a phytotron under natural light and controlled temperature conditions. Xylem sap was collected from Bragg and ntsll16 plants at the flowering and pod elongation stages. Acetylene reduction activity per plant or per nodule weight was not different between soybean lines and inoculums. The composition of the nitrogenous compounds in the xylem sap was compared between the symbionts, with B. japonicum and R. fredii. At the flowering stage, ureide-N and amide-N accounted for 53 to 70% and 20 to 27% respectively of the total N in the sap collected from the plants inoculated either with B. japonicum or R. fredii. At the pod elongation stage, ureide-N and amide-N accounted for 74 to 85%, and 7 to 19% of total sap N. With the growth of the soybean plants, the ratio of ureide-N in the xylem sap increased. These results suggest that in the case of wild soybean and the hypernodulating mutant line nodulated by R. fredii, ureide is transported as the main nitrogenous compound of fixed nitrogen in the xylem sap in the same way as in plants nodulated with B. japonicum.     

Preheating Method for the Determination of Total Phosphorus Content in Andosols

Abstract

The identification of rhizobial strains is a major problem in studies for the evaluation of the symbiotic effectiveness of specific strains in soils containing native rhizobia. Bradyrhizobium japonicum which includes most strains of soybean-nodulating bacteria is known to display a wide range of genetic diversity (Miyashita 1987). It is, therefore, necessary to develop a reliable taxonomic system based on the genetic traits, which would enable to differentiate and identify of the strains.     

Field response of legumes to inoculation with plant growth-promoting rhizobacteria

Carrier-based (soil/FYM, 1:1) plant growth-promoting rhizobacteria (PGPR) isolates (Bacillus subtilis, Klebsiella planticola and Proteus vulgaris) were tested individually and in combination with Bradyrhizobium japonicum and Rhizobium leguminosarum biovar viciae under field conditions on soybean and lentil crops, respectively, under field conditions. Inoculation of soybean (Glycine max) cv. Pusa 22 with B. subtilis produced maximum nodule number, mass and nitrogenase activity (acetylene reduction activity, ARA) followed by B. japonicum (SB 271). Maximum soybean yield was registered with the coinoculation of B. japonicum and B. subtilis over an uninoculated control. Maximum nodulation in the lentil (Lens culinaris) cv. L 4147 was obtained with a combination of R. leguminosarum (L-12-87) and P. vulgaris inoculation followed by a single inoculation with Rhizobium and B. subtilis. None of the PGPR isolates either singly or in coinoculation with R. leguminosarum could significantly influence the yield of the lentil crop.     

Inoculation with the native Rhizobium gallicum 8a3 improves osmotic stress tolerance in common bean drought-sensitive cultivar

Abstract

Symbiotic nitrogen fixation potential in common bean is considered to be low in comparison with other grain legumes. However, it may be possible to improve the nitrogen fixation potential of common bean using efficient rhizobia. In order to improve osmotic stress tolerance of a drought-sensitive common bean cultivar (COCOT) consumed in Tunisia, plants were inoculated either by the reference strain Rhizobium tropici CIAT 899 or by inoculation with rhizobia isolated from native soils Rhizobium gallicum 8a3. Fifteen days after sowing, osmotic stress was applied by means of 25 mM mannitol (low stress level) or by 75 mM mannitol (high stress level). Fifteen days after treatment plants were harvested and different physiological and biochemical parameters were analysed. Results showed no significant differences between the studied symbioses under control conditions. However after exposure to osmotic stress our results showed better tolerance of COCOT to osmotic stress when inoculated with the native R. gallicum 8a3. This can be partially explained by better water-use efficiency in both leaves and nodules, better relative water content in nodules and better efficiency in utilization of rhizobial symbiosis as compared with COCOT-CIAT 899 symbiosis. Hence, the present study suggested the better use of native soil isolated strains for the inoculation of common bean in order to improve its performance and nitrogen fixation potential under stressful conditions.     

Symbiotic effectiveness of Bradyrhizobium japonicum in acid soils can be predicted from their sensitivity to acid soil stress factors in acidic agar media

In acid soil, low pH, reduced availability of nutrients, and toxicity of Al and Mn limit plant growth and the survival and effectiveness of rhizobia. The symbiosis between legumes and rhizobia is particularly sensitive to acid soil stress. A pot experiment evaluated whether Bradyrhizobium japonicum strain growth on acidic agar media would predict ability to colonize the rhizosphere and form effective nodules in acidic soils. Three Indonesian strains of B. japonicum with similar effectiveness at neutral pH in sand culture but with different tolerance of acid soil stress factors in agar media, and an acid-tolerant commercial strain (CB1809) of comparable effectiveness, were tested in three acid soils using the Al tolerant soybean (Glycine max cv PI 416937). At 7 days after inoculation all strains had achieved large rhizosphere populations, but by day 14 the rhizosphere population of the acid-sensitive strain had decreased, while the more acid-tolerant strains increased. The acid-tolerant strains had significantly greater nodulation and symbiotic effectiveness than plants inoculated with the acid-sensitive strain. Laboratory prescreening of B. japonicum for acid, Al and Mn tolerance in acid media successfully identified strains which were symbiotically competent in low pH soils.     

Differences in numbers of nodules and lateral roots between soybean (Glycine max L. Merr.) cultivars,Kitamusume and Toyosuzu

Two cultivars of soybean (Glycine max L. Merr .), Kitamusume and Toyosuzu were grown with commercially-purchased granulated soil in a greenhouse. Kitamusume formed a larger number of nodules per g shoot dry weight and its nodules showed a smaller average diameter and average dry weight per nodule than Toyosuzu regardless of plant age or rhizobial strains (Bradyrhizobium japonicum (AI017, J5033, 646, J1B140), B. elkanii (USDA94), and Rhizobium fredii (MAFF210054)) inoculated at the rate of 10⁸-10⁹ cells per 3 L pot. These differences were observed in the nodules formed on both primary and lateral roots. With a lower inoculation dose of rhizobia (10² and 10⁵ cells per pot), Kitamusume formed a smaller number of larger nodules than Toyosuzu inoculated with 10⁸-10⁹ cells per pot. This result indicated that the number of nodules was the factor directly controlled by host, because the size of the nodules was not determined by the genetic background, but changed depending on the number of nodules formed. The number of the first order lateral roots of 21 d old Kitamusume was 1.49 times larger than that of Toyosuzu. The relationship between the number of nodules and the number of first order lateral roots of 14 cultivars showed a significant positive correlation. These results indicated that the formation of nodules and lateral roots may be similarly controlled by a certain factor in different cultivars.     

Nodulation competition among Bradyrhizobium japonicum strains as influenced by rhizosphere bacteria and iron availability

Summary Bacteria isolated from the root zones of field-grown soybean plants [Glycine max (L.) Merr.] were examined in a series of glasshouse experiments for an ability to affect nodulation competition among three strains of Bradyrhizobium japonicum (USDA 31, USDA 110, and USDA 123). Inocula applied at planting contained competing strains of B. japonicum with or without one of eleven isolates of rhizosphere bacteria. Tap-root nodules were harvested 28 days after planting, and nodule occupancies were determined for the bradyrhizobia strains originally applied. Under conditions of low iron availability, five isolates (four Pseudomonas spp. plus one Serratia sp.) caused significant changes in nodule occupancy relative to the corresponding control which was not inoculated with rhizosphere bacteria. During subsequent glasshouse experiments designed to verify and further characterize these effects, three fluorescent Pseudomonas spp. consistently altered nodulation competition among certain combinations of bradyrhizobia strains when the rooting medium did not contain added iron. This alteration typically reflected enhanced nodulation by USDA 110. Two of these isolates produced similar, although less pronounced, effects when ferric hydroxide was added to the rooting medium. The results suggest that certain rhizosphere bacteria, particularly fluorescent Pseudomonas spp., can affect nodulation competition among strains of R. japonicum. An additional implication is that iron availability may be an important factor modifying interactions involving the soybean plant, B. japonicum, and associated microorganisms in the host rhizosphere.Paper No. 10648 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh, NC 27695-7601, USA