Competition Among Rhizobium spp. for Nodulation of Leucaena leucocephala in Two Tropical Soils

The successful nodulation of legumes by a Rhizobium strain is determined by the competitive ability of that strain against the mixture of other native and inoculant rhizobia. Competition among six Leucaena rhizobial strains in single and multistrain inoculants were studied. Field inoculation trials were conducted in an oxisol and a mollisol soil, both of which contained indigenous Leucaena-nodulating rhizobia. Strain-specific fluorescent antibodies were used for the identification of the strains in Leucaena nodules. Mixtures of three recommended inoculum strains for Leucaena spp. (TAL82, TAL582, and TAL1145) were used in peat-based inocula either alone or with one of the three other strains isolated from the sites, B213, B214, and B215. Each of these latter three strains was also used as single-strain inocula to study their competition with the native rhizobia in the two soil systems. In the oxisol soil, strains B213 and B215, when used as single-strain inocula, outcompeted the native rhizobia and formed 92 and 62% of the nodules, respectively. Strain B214 was the least competitive in oxisol soil, where it formed 30% of the nodules, and the best in mollisol soil, where it formed 70% of the nodules. The most successful competitor for nodulation in multistrain inocula was strain TAL1145, which outcompeted native and other inoculum Leucaena rhizobia in both soils. None of the strains in single or multistrain inoculants was capable of completely overcoming the resident rhizobia, which formed 4 to 70% of the total nodules in oxisol soil and 12 to 72% in mollisol soil. No strong relationship was detected between the size of the rhizosphere population of a strain and its successful occupation of nodules.     

Methods To Alter the Recovery and Nodule Location of Bradyrhizobium japonicum Inoculant Strains on Field-Grown Soybeans

Three strains of Bradyrhizobium japonicum, I17, 110, and 61A76, were evaluated for their ability to form nodules on field-grown soybeans in soil with a highly competitive indigenous B. japonicum population. The predominant indigenous strain, 0336, in the field site used was unlike the more common isolates from Midwestern soils which belong to the 123 or 138 serogroups. This strain persisted in the soil for at least 30 years without any soybean crops. The three inoculant strains differed in their ability to compete with indigenous strains for nodule formation. Four different inoculation treatments were tested in three adjacent fields. When the amount of inoculum was increased, a higher proportion of nodules contained the inoculant strain. The most competitive inoculant strain was I17, a recent field isolate. Strain 61A76 was better than 110. There was no difference in recovery of the inoculant strains on the Hodgson or Corsoy soybean cultivars, nor was there a difference in recovery of the inoculant strains during the growing season. The vertical distribution of nodules containing the inoculant strains was affected by the method of adding the inoculant to the soil. Inoculant added to the seed furrow produced nodules mainly in the top region of the soybean root. Inoculant tilled into the soil produced nodules primarily in the bottom part of the root. The nodules that were produced in the bottom part of the root are younger and may contribute significant amounts of fixed nitrogen to the soybean during seed formation.     

Soybeans inoculated with root zone soils of Canadian native legumes harbour diverse and novel Bradyrhizobium spp. that possess agricultural potential

An assessment was made of the evolutionary relationships of soybean nodulating bacteria associated with legumes native to eastern Canada to identify potential new sources of soybean inoculant strains.Short season soybeans were used to selectively trap bacteria from root zone soils of four native legume species. Screening of more than 800 bacterial isolates from soybean root nodules by analysis of recA gene sequences followed by analyses of selected genotypes using six core and two symbiosis (nodC and nifH) gene sequences permitted identification of diverse taxa that included eight novel and four named Bradyrhizobium species as well as lineages attributed to the genera Afipia and Tardiphaga.Plant tests showed that symbionts related to four named species as well as a novel Bradyrhizobium lineage were highly efficient with regard to nitrogen fixation on soybeans relative to an inoculant strain.A new symbiovar (sv. septentrionalis) is proposed based on a group of four novel Bradyrhizobium spp. that possess distinctive nodC and nifH gene sequences and symbiotic characteristics.Evidence is provided for horizontal transfer of sv. septentrionalis symbiosis genes between novel Bradyrhizobium spp., a process that rendered recipient bacteria ineffective on soybeans.Diverse lineages of non-symbiotic and symbiotic Bradyrhizobium spp. co-occured within monophyletic clusters in a phylogenetic tree of concatenated core genes, suggesting that loss and/or gain of symbiosis genes has occurred in the evolutionary history of the bacterial genus.Our data suggest that symbiont populations associated with legumes native to eastern Canada harbour elite strains of Bradyrhizobium for soybean inoculation.     

Performance of phaseolus bean rhizobia in soils from the major production sites in the Nile Delta

The symbiotic and competitive performances of two highly effective rhizobia nodulating French bean P. vulgaris were studied in silty loam and clayey soils. The experiments were carried out to address the performance of two rhizobia strains (CE3 and Ph. 163] and the mixture thereof with the two major cultivated bean cultivars in two soil types from major growing French bean areas in Egypt. Clay and silty loam soils from Menoufia and Ismailia respectively were planted with Bronco and Giza 6 phaseolus bean cultivars. The data obtained from this study indicated that rhizobial inoculation of Giza 6 cultivar in clayey soil showed a positive response to inoculation in terms of nodule numbers and dry weight. This response was also positive in dry matter and biomass accumulation by the plants. The inoculant of strain CE3 enhanced plant growth and N-uptake relative to Ph. 163. However, the mixed inoculant strains were not always as good as single strain inoculants. The competition for nodulation was assessed using two techniques namely fluorescent antibody testing (FA) and REP-PCR fingerprinting. The nodule occupancy by inoculant strain Ph. 163 in both soils occupied 30-40% and 38-50 of nodules of cultivar Bronco. The mixed inocula resulted in higher proportions of nodules containing CE3 in silty loam soil and Ph. 163 in clayey soil. The native rhizobia occupied at least 50% of the nodules on the Bronco cultivar. For cultivar Giza 6, the native rhizobia were more competitive with the inoculant strains. Therefore, we suggest using the studied strains as commercial inocula for phaseolus bean.     

Identification of Bradyrhizobium japonicum Nodule Isolates from Wisconsin Soybean Farms

One-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis was a more discriminating method than serotyping for identifying strains of Bradyrhizobium japonicum. Analysis of 543 nodule isolates from southeastern Wisconsin soybean farms revealed that none of the isolates were formed by any of the inoculant strains supplied by either of two inoculant companies. Twenty-nine indigenous strains and six inoculant strains were identified. Strain 61A76, the most competitive indigenous strain, formed 21% of the nodules. Indigenous strains 3030, 3058, 0336, and 3052 formed 15, 11, 9, and 9% of the nodules, respectively. These predominant strains were not associated with a particular soybean cultivar, soil type, or farm location.     

Evaluation of strain competitiveness in Rhizobium leguminosarum bv phaseoli using a nod + fix− natural mutant

Competition from native soil rhizobia is likely to be an important factor limiting Phaseolus vulgaris L. inoculant response in Latin America. We used UMR 1116, a nod ⁺ fix^– natural mutant of Rhizobium leguminosarum bv phaseoli strain CC511, as a reference strain to study competition for nodulation sites in this species. When P. vulgaris cv Carioca was planted in soils containing different proportions of UMR 1116 and the effective and competitive strain UMR 1899, UMR 1116 occupied more than 50% of the nodules at all inoculant ratios tested, though increasing the proportion of UMR 1899 in the inoculant did enhance the number and percentage of effective nodules and plant dry weight. Sixty two strains of bean rhizobia were tested in competition with UMR 1116. An inoculant ratio of 1:1 was used, with all strains applied to the soil rather than to seeds. Strains varied in the number and percentage of effective nodules produced in competition with UMR 1116, and in plant dry weight, and there was a strong correlation between variation in each of these traits and plant N accumulation. Seven of the strains (UMR 1073, 1084, 1102, 1125, 1165, 1378 and 1384) were identified as both superior in competitive ability and active in N₂ fixation. Site of placement of the inoculant and ambient temperature influenced strain response.Journal paper 16736, Agricultural Experiment Station, University of Minnesota, St. Paul, MN 55108, USA     

Inoculation Response of Legumes in Relation to the Number and Effectiveness of Indigenous Rhizobium Populations

The response of legumes to inoculation with rhizobia can be affected by many factors. Little work has been undertaken to examine how indigenous populations or rhizobia affect this response. We conducted a series of inoculation trials in four Hawaiian soils with six legume species (Glycine max, Vigna unguiculata, Phaseolus lunatus, Leucaena leucocephala, Arachis hypogaea, and Phaseolus vulgaris) and characterized the native rhizobial populations for each species in terms of the number and effectiveness of the population for a particular host. Inoculated plants had, on average, 76% of the nodules formed by the inoculum strain, which effectively eliminated competition from native strains as a variable between soils. Rhizobia populations ranged from less than 6 × 10⁰/g of soil to 1 × 10⁴/g of soil. The concentration of nitrogen in shoots of inoculated plants was not higher than that in uninoculated controls when the most probable number MPN counts of rhizobia were at or above 2 × 10¹/g of soil unless the native population was completely ineffective. Tests of random isolates from nodules of uninoculated plants revealed that within most soil populations there was a wide range of effectiveness for N₂ fixation. All populations had isolates that were ineffective in fixing N₂. The inoculum strains generally did not fix more N₂ than the average isolate from the soil population in single-isolate tests. Even when the inoculum strain proved to be a better symbiont than the soil rhizobia, there was no response to inoculation. Enhanced N₂ fixation after inoculation was related to increased nodule dry weights. Although inoculation generally increased nodule number when there were less than 1 × 10² rhizobia per g of soil, there was no corresponding increase in nodule dry weight when native populations were effective. Most species compensated for reduced nodulation in soils with few rhizobia by increasing the size of nodules and therefore maintaining a nodule dry weight similar to that of inoculated plants with more nodules. Even when competition by native soil strains was overcome with a selected inoculum strain, it was not always possible to enhance N₂ fixation when soil populations were above a threshold number and had some effective strains.     

Interactive effects of synthetic nitrogen fertilizer and composted manure on ammonia volatilization from soils

The mutualism between legumes and nitrogen-fixing soil bacteria (rhizobia) is a key feature of many ecological and agricultural systems, yet little is known about how this relationship affects aboveground interactions between plants and herbivores. We investigated the effects of the rhizobia mutualism on the abundance of a specialized legume herbivore on soybean plants. In a field experiment, soybean aphid (Aphis glycines) abundances were measured on plants (Glycine max) that were either (1) treated with a commercial rhizobial inoculant, (2) associating solely with naturally occurring rhizobia, or (3) given nitrogen fertilizer. Plants associating with naturally occurring rhizobia strains exhibited lower aphid population densities compared to those inoculated with a commercial rhizobial preparation or given nitrogen fertilizer. Genetic analyses of rhizobia isolates cultured from field plants revealed that the commercial rhizobia strains were phylogenetically distinct from naturally occurring strains. Plant size, leaf nitrogen concentration, and nodulation density were similar among rhizobia-associated treatments and did not explain the observed differences in aphid abundance. Our results demonstrate that plant–rhizobia interactions influence plant resistance to insect herbivores and that some rhizobia strains confer greater resistance to their mutualist partners than do others.     

Phylogenetic relationship of Lotus uliginosus symbionts with bradyrhizobia nodulating genistoid legumes

Lotus species are legumes with potential for pastures in soils with low-fertility and environmental constraints. The aim of this work was to characterize bacteria that establish efficient nitrogen-fixing symbiosis with the forage species Lotus uliginosus. A total of 39 isolates were obtained from nodules of L. uliginosus naturally growing in two different locations of Portugal. Molecular identification of the isolates plus the commercial inoculant strain NZP2039 was performed by REP-PCR, 16S rRNA RFLP, and 16S rRNA, glnII and recA sequence analyses. Limited genetic diversity was found among the L. uliginosus symbionts, which showed a close phylogenetic relationship with the species Bradyrhizobium japonicum. The symbiotic nifH, nodA and nodC gene sequences were closely related with the corresponding genes of various Bradyrhizobium strains isolated from Lupinus and other genistoid legumes and therefore were phylogenetically separated from other Lotus spp. rhizobia. The L. uliginosus bradyrhizobia were able to nodulate and fix nitrogen in association with L. uliginosus, could nodulate Lotus corniculatus with generally poor nitrogen-fixing efficiency, formed nonfixing nodules in Lotus tenuis and Lupinus luteus roots and were unable to nodulate Glycine soja or Glycine max. Thus, L. uliginosus rhizobia seem closely related to B. japonicum biovar genistearum strains.     

Competitive Abilities of Rhizobium meliloti Strains Considered to Have Potential as Inoculants

Twenty four strains of Rhizobium meliloti considered to have potential for inoculant production were grouped in pairs and tested for their ability to compete for nodulation on Medicago sativa, Medicago truncatula, and Medicago littoralis. At the outset, each pair of strains, which consisted of a wild type and a selected streptomycin-resistant mutant of another strain, was tested in an autoclaved soil. Six strain pairs, each consisting of a good and a poor competitor, reacted consistently when tested in each of five other autoclaved soils; eight pairs consisting of strains with comparable competitive abilities varied in their reactions in some of the soils, or even in the same soil when retested. An effect of soil pH on competitive ability was observed with some of these strains. Not all of the strains identified as good competitors on one or more of the Medicago spp. in the autoclaved soils were able to nodulate these plants satisfactorily in a field soil containing an established population of R. meliloti. Strain RF24, which seemed to be the best competitor on each of the three Medicago spp., grouped among the less effective strains on two of the legumes. Two strains of R. meliloti frequently used for inoculant production differed markedly with regard to competitive ability; this places some doubt on the relevancy of singling out competitive ability for special attention when selecting a strain for inoculant production.     

Diversity of a Soybean Bradyrhizobial Population Adapted to an Indian Soil

Soybean (Glycine max) is an introduced crop in India. Over the years it has been regularly inoculated with indigenous rhizobia. In this study genetic diversity has been studied at a site where soybean has been regularly grown with inoculation. Rhizobia were plant trapped using soybean varieties as host, and fingerprinted using BOX-PCR. BOX-PCR genomic fingerprints of 69 isolates from the nodules of 4 soybean varieties Pusa22, Bragg, PK1041 and PK1142 showed a high level of genetic diversity. The population profiles of the 69 isolates clustered them into 10 groups. Root nodule isolates from the four varieties were Bradyrhizobium japonicum types, growing in 4–7 days with typical colonies which were found to be genetically distinct from the USDA and SEMIA strains of B. japonicum and B. elkanii. Also the genotype of the host plant seemed to be one of the factors determining the diversity. The high diversity could be attributed both to lateral transfer of genetic material between inoculant and indigenous strains and to genomic rearrangements during the adaptation to the Indian soils.     

Diversity of a soybean rhizobial population adapted to a Cerrados soil

One hundred isolates were trapped by soybean (Glycine max) plants inoculated with a soil from the Cerrados, the main producing area in Brazil. The soil was originally void of rhizobia able to nodulate soybean, and 15 years before received inoculant containing Bradyrhizobium elkanii strains SEMIA 587 and SEMIA 5019; the area has been annually cropped with soybean since then, but with no further inoculation for the past 7 years. Enormous diversity was observed among the isolates, with thirteen serologically distinct groups, twelve protein and seven lipopolysaccharide profiles; no more than five isolates shared similar characteristics. An unexpected feature was that 48% of the isolates showed multiple reactions with the antisera to the serogroups established in the soils. Also 40% of the isolates reacted with the antiserum to B. japonicum strain SEMIA 566, that has never been introduced into the soil, probably due to dispersion from other cropping areas, associated with its high saprophytic competence; 13% of the isolates did not react with any of the antisera. Nodulation and N₂ fixation capacity also varied considerably among the isolates. Although one third of the isolates were fast growers with an acid reaction in vitro, and many formed pseudo-nodules on common bean (Phaseolus vulgaris), they shared several properties with the Bradyrhizobium inoculant strains. A high level of genetic diversity was confirmed when the DNAs were amplified with BOX and RPO1 primers, and several isolates were positioned in far different clusters in the analysis of interspersed repetitive or nif-directed sequences. Moreover, serological properties showed higher correlation with BOX than with RPO1 products. The high diversity could be attributed both to lateral transfer of genetic material between inoculant and indigenous strains and to genomic rearrangements during the adaptation to the Cerrados, and may play an important role as a biological buffer, avoiding the dominance of a particular strain.     

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.     

Isolation and Characterization of Alfalfa-Nodulating Rhizobia Present in Acidic Soils of Central Argentina and Uruguay

We describe the isolation and characterization of alfalfa-nodulating rhizobia from acid soils of different locations in Central Argentina and Uruguay. A collection of 465 isolates was assembled, and the rhizobia were characterized for acid tolerance. Growth tests revealed the existence of 15 acid-tolerant (AT) isolates which were able to grow at pH 5.0 and formed nodules in alfalfa with a low rate of nitrogen fixation. Analysis of those isolates, including partial sequencing of the genes encoding 16S rRNA and genomic PCR-fingerprinting with MBOREP1 and BOXC1 primers, demonstrated that the new isolates share a genetic background closely related to that of the previously reported Rhizobium sp. Or191 recovered from an acid soil in Oregon (B. D. Eardly, J. P. Young, and R. K. Selander, Appl. Environ. Microbiol. 58:1809–1815, 1992). Growth curves, melanin production, temperature tolerance, and megaplasmid profiles of the AT isolates were all coincident with these characteristics in strain Or191. In addition to the ability of all of these strains to nodulate alfalfa (Medicago sativa) inefficiently, the AT isolates also nodulated the common bean and Leucaena leucocephala, showing an extended host range for nodulation of legumes. In alfalfa, the time course of nodule formation by the AT isolate LPU 83 showed a continued nodulation restricted to the emerging secondary roots, which was probably related to the low rate of nitrogen fixation by the largely ineffective nodules. Results demonstrate the complexity of the rhizobial populations present in the acidic soils represented by a main group of N₂-fixing rhizobia and a second group of ineffective and less-predominant isolates related to the AT strain Or191.     

Nodulation Efficiency of Legume Inoculation as Determined by Intrinsic Antibiotic Resistance

Patterns of intrinsic resistance and susceptibility to different levels of antibiotics were determined for strains of both fast- and slow-growing rhizobia. These patterns were stable to plant passage when they were used to identify Rhizobium strains in nodule suspensions or nodule isolates. The method of identification by intrinsic resistance and susceptibility patterns was reliable for identifying strains in field nodules when strains were first isolated from the nodules to provide a standard inoculum size and then typed on antibiotic-containing media. Other patterns of resistance were encountered during identification of field isolates; these patterns may have resulted from acquired resistance to certain antibiotics or from mixed infections of the nodules. The occurrence of resistance patterns identical to those of inoculant strains among native strains was directly related to the size of the soil population. High strain recovery was associated directly with high rates of inoculation.     

Variation in Preference for Rhizobium meliloti Within and Between Medicago sativa Cultivars Grown in Soil

Variation in nodulation preferences for Rhizobium strains within and between Medicago sativa cultivars was assessed in the greenhouse with plants grown in Leonard jars and two soils of diverse origin (Lanark and Ottawa), using inocula consisting of effective individual or paired strains of R. meliloti which could be recognized by high-concentration antibiotic resistance. The results indicated considerable variability in host preferences for R. meliloti among plants within cultivars but not between cultivars. The implications of this variation are discussed from the point of view of possible improvement of symbiotic nitrogen fixation. With one exception, the differences in nodulation success between inoculant R. meliloti strains were consistent in Leonard jars and both soils. All introduced strains formed significantly more nodules in Renfrew soil containing few native rhizobia than in Ottawa soil with a large resident R. meliloti population. Plants grown in Lanark soil without inoculation were ineffectively nodulated by native rhizobia and yielded significantly less growth than those receiving inoculation. In contrast, the yield of inoculated plants in Ottawa soil did not significantly differ from those without inoculation due to effective nodulation by native R. meliloti. The data indicated synergistic effects on yield by certain paired strain inocula relative to the same strains inoculated individually in Lanark but not in Ottawa soil or Leonard jars.     

Endophytic occupation of peanut root nodules by opportunistic Gammaproteobacteria

Several bacterial isolates were recovered from surface-sterilized root nodules of Arachis hypogaea L. (peanut) plants growing in soils from Córdoba, Argentina. The 16S rDNA sequences of seven fast-growing strains were obtained and the phylogenetic analysis showed that these isolates belonged to the Phylum Proteobacteria, Class Gammaproteobacteria, and included Pseudomonas spp., Enterobacter spp., and Klebsiella spp. After storage, these strains became unable to induce nodule formation in Arachis hypogaea L. plants, but they enhanced plant yield. When the isolates were co-inoculated with an infective Bradyrhizobium strain, they were even found colonizing pre-formed nodules. Analysis of symbiotic genes showed that the nifH gene was only detected for the Klebsiella-like isolates and the nodC gene could not be amplified by PCR or be detected by Southern blotting in any of the isolates. The results obtained support the idea that these isolates are opportunistic bacteria able to colonize nodules induced by rhizobia.     

Overcoming non-selective nodulation of Lessertia by soil-borne rhizobium in the presence of inoculant mesorhizobium

Background and aims

Legumes of the South African genus Lessertia, along with their microsymbionts, were introduced into the Western Australia wheatbelt. They achieved poor establishment followed by weak summer survival. This was caused in part by low levels of nodulation with the inoculant strains, and by ineffective nodulation with naturalized strains –an example of non-selective nodulation. The aims of this work were to assess Lessertia spp. symbiotic promiscuity, to study the effect of increased doses of an effective inoculant strain (WSM3565) with L. herbacea, and to study the competitive ability and symbiotic performance of different Mesorhizobium strains nodulating L. diffusa.

Methods

A glasshouse experiment was set up to evaluate the ability of L. diffusa, L. capitata, L. herbacea and L. excisa to nodulate with inoculants under current use in Western Australia. To assess competitive ability two field experiments were set up at Karridale, Western Australia. L. herbacea was inoculated with the strain WSM3565 at different doses and L. diffusa was inoculated with ten different Mesorhizobium strains. Rhizobia were re-isolated from nodules and their identity confirmed through PCR fingerprinting and sequencing of their partial dnaK.

Results

There were differences in promiscuity between different Lessertia spp., where L. herbacea proved to be highly promiscuous under controlled conditions. Increasing the inoculation dose of L. herbacea with WSM3565 did not improve establishment and survival of the legume in the field. Although WSM3565 nodule occupancy improved from 28 to 54 % with higher doses of inoculation, none of the treatments increased L. herbacea yield over the inoculated control. The inoculation of L. diffusa with the strains WSM3598, 3636, 3626 and 3565 resulted in greater biomass production than the uninoculated control. These strains were able to outcompete resident rhizobia and to occupy a high (>60 %) proportion of lateral root nodules. The naturalised strains that achieved nodulation were identified as R. leguminosarum.

Conclusion

The high numbers of resident rhizobia and their ability to rapidly nodulate Lessertia spp. are likely to be the main reasons for the low nodule occupancy achieved by some effective inoculant strains with L. diffusa and L. herbacea. Strains WSM 3636 and 3598 were very competitive on nodule occupancy and together with WSM 3565, WSM 3612 and WSM3626, effective on nodule formation and plant growth of L. diffusa despite the high numbers of resident soil rhizobia. These strains and L. diffusa have potential to be introduced as exotic legumes species and rhizobia strains to Western Australia.     

Survival and Competitiveness of Bradyrhizobium japonicum Strains 20 Years after Introduction into Field Locations in Poland

It was previously demonstrated that there are no indigenous strains of Bradyrhizobium japonicum forming nitrogen-fixing root nodule symbioses with soybean plants in arable field soils in Poland. However, bacteria currently classified within this species are present (together with Bradyrhizobium canariense) as indigenous populations of strains specific for nodulation of legumes in the Genisteae tribe. These rhizobia, infecting legumes such as lupins, are well established in Polish soils. The studies described here were based on soybean nodulation field experiments, established at the Poznań University of Life Sciences Experiment Station in Gorzyń, Poland, and initiated in the spring of 1994. Long-term research was then conducted in order to study the relation between B. japonicum USDA 110 and USDA 123, introduced together into the same location, where no soybean rhizobia were earlier detected, and nodulation and competitive success were followed over time. Here we report the extra-long-term saprophytic survival of B. japonicum strains nodulating soybeans that were introduced as inoculants 20 years earlier and where soybeans were not grown for the next 17 years. The strains remained viable and symbiotically competent, and molecular and immunochemical methods showed that the strains were undistinguishable from the original inoculum strains USDA 110 and USDA 123. We also show that the strains had balanced numbers and their mobility in soil was low. To our knowledge, this is the first report showing the extra-long-term persistence of soybean-nodulating strains introduced into Polish soils and the first analyzing the long-term competitive relations of USDA 110 and USDA 123 after the two strains, neither of which was native, were introduced into the environment almost 2 decades ago.     

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.