Population Densities of Rhizobium japonicum Strain 123 Estimated Directly in Soil and Rhizospheres

Rhizobium japonicum serotype 123 was enumerated in soil and rhizospheres by fluorescent antibody techniques. Counting efficiency was estimated to be about 30%. Indigenous populations of strain 123 ranged from a few hundred to a few thousand per gram of field soil before planting. Rhizosphere effects from field-grown soybean plants were modest, reaching a maximum of about 2 × 10⁴ cells of strain 123 per g of inner rhizosphere soil in young (16-day-old) plants. Comparably slight rhizosphere stimulation was observed with field corn. High populations of strain 123 (2 × 10⁶ to 3 × 10⁶ cells per g) were found only in the disintegrating taproot rhizospheres of mature soybeans at harvest, and these populations declined rapidly after harvest. Pot experiments with the same soil provided data similar to those derived from the field experiments. Populations of strain 123 reached a maximum of about 10⁵ cells per g of soybean rhizosphere soil, but most values were lower and were only slightly higher than values in wheat rhizosphere soil. Nitrogen treatments had little effect on strain 123 densities in legume and nonlegume rhizospheres or on the nodulation success of strain 123. No evidence was obtained for the widely accepted theory of specific stimulation, which has been proposed to account for the initiation of the Rhizobium-legume symbiosis.     

Early Infection and Competition for Nodulation of Soybean by Bradyrhizobium japonicum 123 and 138

Interactions of soybean with Bradyrhizobium japonicum 123 (serogroup 123) and 138 (serogroup c1) were used to examine the relationship between early infection rates, competition for nodulation, and patterns of nodule occupancy. Both strains formed more infections in autoclaved soil (sterile soil) than in untreated soil (unsterile soil). Inoculation did not increase numbers of infection threads in unsterile soil-grown plants, where infection of proximal portions of primary roots was complete by 5 days after planting. Both strains infected and nodulated at similar rates in sterile soil. Nodules were always clustered on the upper root system, regardless of inoculation and soil treatment. Sixty-seven percent of the nodules of uninoculated plants grown in unsterile soil were occupied by rhizobia belonging to serogroups other than 123 or c1. Inoculation with strain 123 or 138 increased occupancy by that strain at the expense of residency by other rhizobia. Eighty-three percent of all nodules on plants dually inoculated with both strains in sterile soil contained strain 138. The corresponding value for plants inoculated in unsterile soil was 31%. Neither inoculum strain dominated occupancy of first-formed nodules in unsterile soil. It appears that north central Missouri soil may not have populations of highly competitive serogroup 123 and that early infection and nodulation rates do not contribute to the competitive success of strain 138.     

The Soybean Rj4 Allele Restricts Nodulation by Bradyrhizobium japonicum Serogroup 123 Strains

Of nine Bradyrhizobium japonicum serogroup 123 strains examined, 44% were found to be restricted for nodulation by cultivar Hill. Nodulation studies with soybean isoline BARC-2 confirmed that the soybean Rj4 allele restricts nodulation by the same serogroup 123 isolates. Immunological analyses indicated that B. japonicum strains in serogroups 123 and 31 share at least one surface somatic antigen.     

Nodulation and Competition for Nodulation of Selected Soybean Genotypes among Bradyrhizobium japonicum Serogroup 123 Isolates

Twenty recently obtained field isolates of Bradyrhizobium japonicum serogroup 123 were tested for their nodule mass production on the standard commercial soybean (Glycine max (L.) Merr. cv. Williams) and on two soybean plant introduction (PI) genotypes previously determined to restrict nodulation by strain USDA 123. Four of the field isolates showed similar restricted nodulation on the two genotypes, while all 20 isolates produced a normal amount of nodules on G. max cv. Williams. Serological analyses with adsorbed fluorescent antibodies showed that members of the 123 serotype ranked low in nodulation of the two PIs, in contrast to members of serotypes 127 and 129. Competition studies on the PIs indicated that isolates which were restricted were not competitive for nodule occupancy against strain USDA 110. However, unrestricted isolates of serogroup 123 were very competitive against USDA 110. On G. max cv. Williams, all serogroup 123 isolates tested were very competitive against USDA 110.     

Long-Term Effects of Metal-Rich Sewage Sludge Application on Soil Populations of Bradyrhizobium japonicum

The application of sewage sludge to land may increase the concentration of heavy metals in soil. Of considerable concern is the effect of heavy metals on soil microorganisms, especially those involved in the biocycling of elements important to soil productivity. Bradyrhizobium japonicum is a soil bacterium involved in symbiotic nitrogen fixation with Glycine max, the common soybean. To examine the effect of metal-rich sludge application on B. japonicum, the MICs for Pb, Cu, Al, Fe, Ni, Zn, Cd, and Hg were determined in minimal media by using laboratory reference strains representing 11 common serogroups of B. japonicum. Marked differences were found among the B. japonicum strains for sensitivity to Cu, Cd, Zn, and Ni. Strain USDA 123 was most sensitive to these metals, whereas strain USDA 122 was most resistant. In field studies, a silt loam soil amended 11 years ago with 0, 56, or 112 Mg of digested sludge per ha was examined for total numbers of B. japonicum by using the most probable number method. Nodule isolates from soybean nodules grown on this soil were serologically typed, and their metal sensitivity was determined. The number of soybean rhizobia in the sludge-amended soils was found to increase with increasing rates of sludge. Soybean rhizobia strains from 11 serogroups were identified in the soils; however, no differences in serogroup distribution or proportion of resistant strains were found between the soils. Thus, the application of heavy metal-containing sewage sludge did not have a long-term detrimental effect on soil rhizobial numbers, nor did it result in a shift in nodule serogroup distribution.     

Exclusion of inefficient Bradyrhizobium japonicum serogroups by soybean genotypes

Soybean [Glycine max (L.) Merr.] forms a symbiosis with serogroups of Bradyrhizobium japonicum that differ in their dinitrogen fixing abilities. The objectives of this study were to identify soybean genotypes that would restrict nodulation by relatively inefficient serogroups indigenous to a large portion of the southeastern USA, and then characterize the nodulation responses of selected genotypes with specific bradyrhizobial strains under controlled conditions. From field screening trials followed by controlled single and competitive inoculations of serogroups USDA 31, 76 and 110, twelve soybean genotypes out of 382 tested were identified with varying levels of exclusion abilities. Soybean nodule occupancies and nodulation characteristics were influenced by plant genotype, environment (i.e. field or greenhouse), bradyrhizobial serogroup, and location of nodules (i.e. tap or lateral root). The cultivar Centennial sustains high seed yields even though it nodulates to a high degree with the inefficient serogroup USDA 31. In contrast, data from the released cultivars Braxton, Centennial and Coker 368 indicate that they may have been selected to exclude the inefficient serogroup USDA 76 from their tap root nodules, possibly contributing to high seed yield.     

Ecology of Indigenous Soil Rhizobia: Response of Bradyrhizobium japonicum to Readily Available Substrates

Populations of indigenous Bradyrhizobium japonicum serocluster 123 and serogroups 110 and 138 were studied after various sugars were added to their soil habitat. Loam soil with approximately 10⁴ cells of each group per g of soil were amended every 3 days with 0.1% glucose, sucrose, arabinose, xylose, or galactose. Enumerations of the populations were made every 12 days by immunofluorescence assay. Each B. japonicum population in the sugar-treated soils increased by about 1 log during the first 12 days, to a maximum of about 10⁶ cells by day 36 or 48, irrespective of the sugar added. Maximum growth rates were similar for each group and occurred during the 12-day incubation period. The most rapid growth was in response to arabinose, with a mean generation time of about 3.0 days. Other mean doubling times were 4.0 days with glucose and galactose treatments, 4.5 days with xylose treatment, and 5.4 days with sucrose amendment. These data provide the first direct evidence that indigenous soil rhizobia can compete successfully with other soil bacteria for readily available substrates in soil in the absence of host legume roots or other rhizospheres. The growth rates in soil of the specific B. japonicum populations studied were nearly the same with a given sugar treatment but varied considerably with different sugars. The mean generation times of 3 to 5 days are among the first reported growth rates for heterotrophic bacteria in natural soil.     

Rhizobium japonicum Serogroup and Hydrogenase Phenotype Distribution in 12 States

A survey was conducted in 1980 on 972 isolates of Rhizobium japonicum obtained from 65 soybean field locations in 12 states. Isolates were examined for the hydrogenase (Hup) phenotype and somatic serogroup identity. Only 20% of the isolates were Hup⁺, with a majority of Hup⁻ isolates occurring in 10 of the 12 states. The most predominant serogroup was 31 (21.5%), followed by 123 (13.6%). Although most serogroups contained a majority of Hup⁻ isolates, marked differences occurred. None of the isolates in serogroup 135 were Hup⁺, but 93% of the isolates in serogroup 122 were Hup⁺. The serogroups with relatively high frequencies of Hup⁺ isolates (122 and 110) constitute only a small part (<5% each) of the R. japonicum field population in the 12 states.     

Effect of Acidity on the Composition of an Indigenous Soil Population of Rhizobium trifolii Found in Nodules of Trifolium subterraneum L.

Acidity affected which members of an indigenous soil population of Rhizobium trifolii nodulated Trifolium subterraneum L. cv. Mt. Barker. In three experiments involving plants grown either in mineral salts agar adjusted to pH 4.8 or 6.8 and inoculated with a soil suspension or grown directly in samples of unamended soil (pH 4.8) or soil amended with CaCO₃ (pH 6.4), 121 of 151 isolates of R. trifolii were placed into four serogroups. Seventy-nine of these isolates were placed into two serogroups (6 and 36) whose nodulating ability was affected by the pH of the plant root environment. Representatives of serogroup 6 occupied the greatest percentage of the nodules at the low pH in both mineral salts agar (77%) and in unlimed soil (47 and 57%). The same serogroup was a minor nodule occupant at the higher pH in mineral salts agar (0%) and in limed soil (0 and 10%). In contrast, serogroup 36 was virtually absent in nodules formed at the low pH, whereas it was the dominant serogroup at the higher pH in both mineral salts agar (32%) and in limed soil (35 and 49%). Despite the isolates from within each serogroup being antigenically identical, separation of cellular proteins by sodium dodecyl sulfate-polyacrylamide gradient gel electrophoresis revealed four and six different gel types within serogroups 6 and 36, respectively. Isolates represented by one or two gel types dominated the contribution of each serogroup to the nodule population. Further evidence for differences between isolates within each gel type were revealed from measurements of symbiotic effectiveness.     

Population Changes and Persistence of Rhizobium phaseoli in Soil and Rhizospheres

The impact of legume cultivation on the establishment and persistence of an inoculant strain of Rhizobium phaseoli and its ability to compete with a resident population of R. phaseoli for nodule occupancy was examined utilizing strain-specific fluorescent antibodies. The soil (Hubbard loamy sand) was inoculated homogeneously with 5 × 10⁵ cells per g of soil and confined in plastic cylinders kept in field plots. Inoculated and uninoculated cylinders were either left fallow or planted to two seeds of legumes. Two hosts, navy bean (Phaseolus vulgaris L.) cv. Seafarer and snap bean cv. Picker, as well as a nonhost, soybean (Glycine max (L.) Merr.) cv. Wilkin, were used. Inoculant Viking 1 was highly stimulated in all three rhizospheres sampled at 6 (flowering), 10 (podfill), and 17 (decay) weeks and in the following spring, whereas counts in fallow soil decreased rapidly. Although the overwintering population remained highest in the vicinity of decaying host roots, Viking 1 persisted, even in fallow soil, to produce abundant nodulation of host plants the following spring. Viking 1 was an excellent competitor for nodulation sites on the roots of the hosts; it thoroughly outcompeted the resident population of R. phaseoli, occupying virtually 100% of the nodules under inoculated conditions in all experiments.     

Influence of Environmental Factors on Interstrain Competition in Rhizobium japonicum

The effect of several biotic and abiotic factors on the pattern of competition between two strains of Rhizobium japonicum was examined. In two Minnesota soils, Waseca and Waukegan, strain USDA 123 occupied 69% (Waseca) and 24% (Waukegan) of the root nodules on Glycine max L. Merrill cv. Chippewa. USDA 110 occupied 2% of the root nodules in the Waseca soil and 12% of the nodules in the Waukegan soil. Under a variety of other growth conditions—vermiculite, vermiculite amended with Waseca soil, and two Hawaiian soils devoid of naturalized Rhizobium japonicum strains—USDA 110 was more competitive than USDA 123. The addition of nitrate to or the presence of antibiotic-producing actinomycetes in the rhizosphere of soybeans did not affect the pattern of competition between the two strains. However, preexposure of young seedings to USDA 110 or USDA 123 before transplantation into soil altered the pattern of competition between the two strains significantly. In the Waseca soil, preexposure of cv. Chippewa to USDA 110 for 72 h increased the percentage of nodules occupied by USDA 110 from 2 to 55%. Similarly, in the Hawaiian soil Waimea, nodule occupancy by USDA 123 increased from 7 to 33% after a 72-h preexposure.     

Compatibility of Rhizobial Genotypes within Natural Populations of Rhizobium leguminosarum Biovar viciae for Nodulation of Host Legumes

Populations of Rhizobium leguminosarum biovar viciae were sampled from two bulk soils, rhizosphere, and nodules of host legumes, fava bean (Vicia faba) and pea (Pisum sativum) grown in the same soils. Additional populations nodulating peas, fava beans, and vetches (Vicia sativa) grown in other soils and fava bean-nodulating strains from various geographic sites were also analyzed. The rhizobia were characterized by repetitive extragenomic palindromic-PCR fingerprinting and/or PCR-restriction fragment length polymorphism (RFLP) of 16S-23S ribosomal DNA intergenic spacers as markers of the genomic background and PCR-RFLP of a nodulation gene region, nodD, as a marker of the symbiotic component of the genome. Pairwise comparisons showed differences among the genetic structures of the bulk soil, rhizosphere, and nodule populations and in the degree of host specificity within the Vicieae cross-inoculation group. With fava bean, the symbiotic genotype appeared to be the preponderant determinant of the success in nodule occupancy of rhizobial genotypes independently of the associated genomic background, the plant genotype, and the soil sampled. The interaction between one particular rhizobial symbiotic genotype and fava bean seems to be highly specific for nodulation and linked to the efficiency of nitrogen fixation. By contrast with bulk soil and fava bean-nodulating populations, the analysis of pea-nodulating populations showed preferential associations between genomic backgrounds and symbiotic genotypes. Both components of the rhizobial genome may influence competitiveness for nodulation of pea, and rhizosphere colonization may be a decisive step in competition for nodule occupancy.     

Influence of Glycine spp. on Competitiveness of Bradyrhizobium japonicum and Rhizobium fredii

The displacement of indigenous Bradyrhizobium japonicum in soybean nodules with more effective strains offers the possibility of enhanced N₂ fixation in soybean (Glycine max (L.) Merr.). Our objective was to determine whether the wild soybean (G. soja Sieb. & Zucc.) genotype PI 468397 would cause reduced competitiveness of important indigenous B. japonicum strains USDA 31, 76, and 123 and thereby permit nodulation by Rhizobium fredii, the fast-growing microsymbiont of soybean. In an initial experiment, PI 468397 nodulated and fixed moderate amounts of N₂ with USDA 31 and 76 but, despite the formation of nodules, fixed essentially no N₂ with USDA 123. In contrast, PI 468397 formed a highly effective symbiosis with R. fredii strain USDA 193. In two subsequent experiments, Williams soybean and PI 468397 were grown in a pasteurized soil mixture or in soybean rhizobium-free soil and inoculated with both USDA 123 and USDA 193. In each experiment, more than 90% of the nodules of Williams contained USDA 123, while only a maximum of 2% were occupied with USDA 193. In contrast, in the two experiments, 16 and 11%, respectively, of the nodules produced on PI 468397 were occupied by USDA 123, while in both experiments 87% contained USDA 193. Thus, in relation to the cultivar Williams, which is commonly grown and used as a parent in soybean breeding programs in the United States, PI 468397 substantially reduced the competitive ability of B. japonicum strain USDA 123 in relation to R. fredii strain USDA 193.     

DNA Hybridization Probe for Use in Determining Restricted Nodulation among Bradyrhizobium japonicum Serocluster 123 Field Isolates

Several soybean plant introduction (PI) genotypes have recently been described which restrict nodulation of Bradyrhizobium japonicum serocluster 123 in an apparently serogroup-specific manner. While PI 371607 restricts nodulation of strains in serogroup 123 and some in serogroup 127, those in serogroup 129 are not restricted. When DNA regions within and around the B. japonicum I-110 common nodulation genes were used as probes to genomic DNA from the serogroup strains USDA 123, USDA 127, and USDA 129, several of the probes differentially hybridized to the nodulation-restricted and -unrestricted strains. One of the gene regions, cloned in plasmid pMJS12, was subsequently shown to hybridize to 4.6-kilobase EcoRI fragments from DNAs from nodulation-restricted strains and to larger fragments in nodulation-unrestricted strains. To determine if the different hybridization patterns could be used to predict nodulation restriction, we hybridized pMJS12 to EcoRI-digested genomic DNAs from uncharacterized serocluster 123 field isolates. Of the 36 strains examined, 15 were found to have single, major, 4.6-kilobase hybridizing EcoRI fragments. When tested for nodulation, 80% (12 of 15) of the strains were correctly predicted to be restricted for nodulation of the PI genotypes. In addition, hybridization patterns obtained with pMJS12 and nodulation phenotypes on PI 371607 indicated that there are at least three types of serogroup 127 strains. Our results suggest that the pMJS12 gene probe may be useful in selecting compatible host-strain combinations and in determining the suitability of field sites for the placement of soybean genotypes containing restrictive nodulation alleles.     

Influence of Lime and Phosphate on Nodulation of Soil-Grown Trifolium subterraneum L. by Indigenous Rhizobium trifolii

Previous research had identified four serogroups of Rhizobium trifolii indigenous to the acidic Abiqua soil (fine, mixed, mesic Cumulic Ultic Haploxeroll). Nodulation of subterranean clover (Trifolium subterraneum L.) by two of the serogroups, 6 and 36, was differentially influenced by an application of CaCO₃ which raised the pH of the soil from 5.0 to 6.5. These studies were designed to characterize this phenomenon more comprehensively. Liming the soil with either CaCO₃, Ca(OH)₂, MgO, or K₂CO₃ significantly (P = 0.05) increased the percent nodule occupancy by serogroup 36, whereas the percent nodule occupancy by serogroup 6 was decreased, but the decrease was significant (P = 0.05) only after application of either CaCO₃ or Ca(OH)₂. Application of KH₂PO₄ (25 mg of P kg of soil⁻¹), which did not change soil pH, also significantly (P = 0.05) increased the percent nodule occupancy by serogroup 36. Application of KH₂PO₄ in combination with Ca(OH)₂ produced the same increase in nodule occupancy by serogroup 36 as did individual application of the two materials. Soil populations of serogroup 36 consistently, and in the majority of cases significantly (P = 0.05), outnumbered those of serogroup 6 before planting and after harvest regardless of soil treatment or the outcome of nodulation. Soil chemical and plant analyses provided no evidence that liming was simulating phosphate addition by increasing the availability and subsequent uptake of soil P_i by the subclover plants. Liming did, however, result in a significant transformation (30 to 50 mg of P kg of soil⁻¹) of P_i from the residual soil P_i fraction into an NaOH-extractable organic P fraction during the preplant equilibration period.     

Competitiveness of indigenous populations of Bradyrhizobium japonicum serocluster 123 as determined using a root-tip marking procedure in growth pouches

Soil Bradyrhizobium populations limit nodule occupancy of soybean by symbiotically-superior inoculant strains throughout much of the American midwest. In this study, the competitiveness of indigenous populations of B. japonicum serocluster 123 from Waukegan and Webster soils was evaluated in growth pouches using a root-tip marking procedure. The native rhizobia were from soils incubated 0–8 h in soybean root exudate (SRE) or plant nutrient solution (PNS) prior to inoculation. Populations of serocluster 123 strains in soil and nodule occupancy by these strains were assessed using fluorescent antibodies prepared against B. japonicum USDA 123. There were no significant differences in populations that came from SRE or PNS incubated soils: both populations increased in number over the incubation period. Nodule occupancy by both populations in growth pouches was similar to that previously encountered in field studies with these two soils. With the Waukegan soil, the serocluster 123 population dominated nodulation forming 69 and 62% of taproot nodules above and below the root tip mark, respectively. However, for the more alkaline Webster soil, serocluster 123 strains were much less competitive, producing only 9 and 13%, respectively, of the nodules formed above and below the root tip mark. In growth pouches, soil populations of bradyrhizobia from the Webster soil produced significantly more nodules than those from the Waukegan soil, but both strains and a pure culture of USDA 110 had a similar distribution of nodules.     

Bradyrhizobium japonicum Serocluster 123 and Diversity among Member Isolates

Diversity was examined within a group of 79 isolates of Bradyrhizobium japonicum reactive to fluorescent antibodies (FAs) prepared against B. japonicum USDA 123. Analyses were by means of cross-adsorbed FAs, bacteriophage typing, and endonuclease restriction digest patterns. Serogroups 127 and 129 shared antigenic determinants with serogroup 123 but not with each other. Bacteriophage and DNA digest patterns reflected more common features between serogroups 123 and 127 than between 123 and 129. Serogroups 129 and 122 showed FA cross-reactivity. The term serocluster was proposed to reflect interrelationships observed among the serogroups.     

Nodulating Competitiveness of a Nonmotile Tn7 Mutant of Bradyrhizobium japonicum in Nonsterile Soil

A nonmotile mutant of Bradyrhizobium japonicum serogroup 127 was generated by Tn7 mutagenesis and matched with the wild type against a common competitor in studies of soybean nodulation in nonsterile soil. The Tn7 mutant was very similar to the wild type in growth rate in culture, soybean lectin-binding ability, flagellar morphology, and nodulating capability, but it had a longer lag phase. Competing strains were distributed uniformly in soil in various ratios and at different population densities prior to planting. Mutant and wild type were equally prevalent in the seedling rhizosphere at about the time of nodule initiation, suggesting that motility conferred no advantage in rhizosphere colonization. Nodulation success of the Tn7 mutant was lower than that of the wild type under all test conditions. Differences were greatest at low soil populations of competitors and much less pronounced at initial populations of 10⁷ g⁻¹. The longer lag phase of the Tn7 mutant may have contributed to its decreased competitiveness, especially at the higher inoculation levels. The antibiotic and motility markers were stable, and the rifampin resistance derived from the parent did not affect adversely the competitiveness of the Tn7 mutant. We found motility to be of limited importance to the competitiveness of a strain in normal nonsterile soil, where the significance, if any, of this ability may be in migration at the immediate root surface in soils sparsely populated with rhizobial symbionts.     

Formation of Novel Polysaccharides by Bradyrhizobium japonicum Bacteroids in Soybean Nodules

Certain strains of Bradyrhizobium japonicum form a previously unknown polysaccharide in the root nodules of soybean plants (Glycine max (L.) Merr.). The polysaccharide accumulates inside of the symbiosome membrane—the plant-derived membrane enclosing the bacteroids. In older nodules (60 days after planting), the polysaccharide occupies most of the symbiosome volume and symbiosomes become enlarged so that there is little host cytoplasm in infected cells. The two different groups of B. japonicum which produce different types of polysaccharide in culture produce polysaccharides of similar composition in nodules. Polysaccharide formed by group I strains (e.g., USDA 5 and USDA 123) is composed of rhamnose, galactose, and 2-O-methylglucuronic acid, while polysaccharide formed by group II strains (e.g., USDA 31 and USDA 39) is composed of rhamnose and 4-O-methylglucuronic acid. That the polysaccharide is a bacterial product is indicated by its composition plus the fact that polysaccharide formation is independent of host genotype but is dependent on the bacterial genotype. Polysaccharide formation in nodules is common among strains in serogroups 123, 127, 129, and 31, with 27 of 39 strains (69%) testing positive. Polysaccharide formation in nodules is uncommon among other B. japonicum serogroups, with only 1 strain in 18 (6%) testing positive.     

Some USDA studies on the soybean-Rhizobium symbiosis

Summary The ecology, strain evaluation, genetics of host strain interactions and physiology of nitrogen fixation ofRhizobium japonicum in association with the soybean,Glycine max, were studied. Results of inoculation experiments with selected strains ofRhizobium japonicum indicated that indigenous strains occupied most of the nodules of soybeans grown in highRhizobium japonicum populated soils. Nodule sampling indicated that inoculation did not result in quicker nodulation or a higher incidence of root nodules (primary or secondary) than uninoculated checks. Rhizosphere studies indicated that colonization by introduced strains did occur but did not compete successfully with field strains for nodule sites. Recovery of specific serological types from nodules was influenced by planting intervals. The distribution of the serotypes varied with the time of planting and the age of the plant. Temperature studies indicated that the distribution of serotypes recovered from the nodules was influenced by temperature. Field studies showed the selectivity of soybean genotypes on strains ofRhizobium japonicum. Some strains were more common in the nodules of some varieties than in others. Closely related varieties had similar populations in their nodules. Three genes which control nodule response in soybeans are reported. Nitrogen fixation profiles were determined for some variety-strain interactions. Combinations previously classified as inefficient showed some nitrogenase activity as measured by the acetylene reduction technique. Research Microbiologist; Research Agronomist; Research Plant Physiologist, Soybean Investigations, Crops Research Division, Beltsville, Md. (USDA, ARS); and Plant Pathologist currently located at Michigan State University, East Lansing, Michigan.