Field performance of three Bradyrhizobium japonicum strains with two soybean (Glycine max. L.) cultivars

Summary A field experiment was condutced in a clay loam soil to study the performance of three Bradyrhizobium japonicum strains; USDA 110, USDA 138 and TAL 379, in relation to their N₂-fixing potential and competitiveness on two soybean cultivars (Clark and Calland). Inoculation of soybean cultivars with these strains, either singly or in combination, induced significant increases in plant dry weight, N₂ fixation and seed yields. However, no significant differences were found between the rhizobial strains and/or their mixtures in N₂ fixation and increased seed yield for both cultivars. The two soybean cultivars gave similar responses to inoculation. No significant differences in seed yield were observed between Clark and Calland cultivars. The interaction between inoculant strain and soybean cultivar was not significant. The competition between strains for nodulation was assessed. Strain USDA 110 was the most competitive, followed by USDA 138. Strain TAL 379 was always less competitive on both cultivars. The incidence of double-strain occupancy of nodules varied from 8% to 40%.     

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.     

Competitiveness of native Bradyrhizobium japonicum strains in two different soil types

Interstrain competitiveness is a key factor affecting the performance of rhizobium inoculant. In the present study five native strains of Bradyrhizobium japonicum, namely SSF 4, SSF 5, SSF 6, SSF 7 and SSF 8, were assessed for their competitiveness in nodulating soybean using serological methods. The strains were inoculated individually or with the type strain USDA 110 at a 1:1 ratio. Nodule occupancy determined by immunofluorescence and dot immunoblot assay revealed that under in vitro conditions SSF 8 is more competitive than USDA 110 whereas the others were less competitive. The competitive ability of these strains was also estimated in pot culture in the field. In red soil both SSF 8 and USDA 110 were equally competitive whereas in black soil SSF 8 competed better than USDA 110 and produced more nodules. In a black soil field trial using a randomized block design, USDA 110 or SSF 8, when inoculated alone, occupied the majority of the nodules and enhanced nodule dry weight and shoot biomass. SSF 8 was more competitive when the strains were co-inoculated. Received: 1 November 1996     

Yield and N accumulation responses of late‐season determinate soybean to irrigation and inoculation with various strains ofBradyrhizobium japonicum

Abstract

Cobb and Coker 488, late‐season (maturity group VIII) cultivars of soybean [Glycine max(L.) Merr], were grovn under irrigated and non‐irrigated conditions on a Norfolk loamy sand in a two‐year field experiment. Each cultivar was inoculated withBradyrhizobium japonicumstrains [USDA 3I1b110; Brazil 587; NifTAL 184 and 102 (NifTAL cultures of Brazil 587 and USDA 110, respectively); and North Carolina 1001, 1004, 1005, 1010, and 1029). Drought conditions were present both years, and irrigation significantly increased the overall yield (2.49 vs 1.92 Mg ha^‐1). Coker 488 was significantly higher in seed yield than Cobb (2.55 vs 2.02 Mg ha^‐1). Strain ofB.japonicumalso affected seed yields. NC1010‐inoculated soybean was significantly higher in seed yield rank than all other soybean at the P<0.01 level, when compared by single degree of freedom contrast (sdfc). The yield ranking of soybean inoculated with NC1001 was significantly lower than soybean inoculated with all other strains, when compared by sdfc (P<0.10). Other strains differed in responses which ranged from good to poor inoculants under specific water management conditions. For instance, under nonirrigated conditions, soybean inoculated with strains ofB.japonicumfrom North Carolina was significantly higher in seed yield than those inoculated with the cultures of USDA 110, B587, or the control, when compared by sdfc (P>0.03, 0.05, 0.06, respectively). Since soybean inoculated with either strain of USDA 110 was generally high in yield rank under irrigated conditions, their response to irrigation was large relative to soybean inoculated with the NC strain (P<0.04). Neither seed nitrogen nor xylem water potential was highly correlated to seed yield. Since seed yield and N content were not highly correlated, the amount of N accumulated in soybean dry mass and that removed in seed were not highly correlated. Thus, the amount of N returned to the soil would be affected by management combinations of late‐season determinate soybean cultivar,B.japonicumstrain, and irrigation     

Single and Multistrain Rhizobial Inocula for Pinto and Black Bean Cultivars

ABSTRACT

Common bean (Phaseolus vulgaris L.) is relatively poor in dinitrogen (N₂) fixation, so selecting compatible host cultivar and Rhizobium strain combinations may offer an improvement. The effectiveness of six rhizobial strains was evaluated using five bean cultivars of bean (three pinto and two black bean) in a growth-room experiment. We subsequently selected the three best strains to assess whether multi-strain inoculation had advantages over single-strain inoculation in growth-room and field experiments. In the first-growth-room experiment, Rhizobium strains UMR 1899, RCR 3618, and USDA 2676 were selected for high nodulation, plant dry weight, shoot nitrogen (N), and N₂ fixation. In a second growth-room experiment, the individual strains and a mixture of the three strains generally did not differ in the parameters evaluated. Total shoot N accumulated ranged from 172.9 to 162.8 mg plant^?1, of which 32.1% to 33.6% (equivalent to 54.0 to 59.2 mg plant^{? 1}) was fixed. In field experiments, plant biomass and seed N₂ fixed did not differ among the inoculants at any site. These results suggest that the three strains were equally effective and that the multi-strain inoculant offered no consistent advantage over the single-strain inoculants.     

Alternatives to peat as a carrier for rhizobia inoculants: Solid and liquid formulations

Many of the microbial inoculants all over the world are based on solid peat formulations. This has been mostly true for well developed legume inoculants based on selected rhizobial strains, due to peat bacterial protection properties. Six carriers (bagasse, cork compost, attapulgite, sepiolite, perlite and amorphous silica) were evaluated as alternatives to peat. Compost from the cork industry and perlite were superior to peat in maintaining survival of different rhizospheric bacteria. Other tested materials were discarded as potential carriers for soybean rhizobia. Also, different liquid culture media have been assayed employing mannitol or glycerol as C sources. Some media maintained more than 10⁹ cfu ml^?1 of Sinorhizobium (Ensifer) fredii SMH12 or Bradyrhizobium japonicum USDA110 after 3 months of storage. Rhizobial survival on pre-inoculated seeds with both solid and liquid formulations previously cured for 15 days led to a higher bacterial numbers in comparison with recently made inoculants. An additional curing time of solid inoculants up to 120 days had a beneficial effect on rhizobial survival on seeds. The performance of different formulations of two highly effective soybean rhizobia strains was assayed under field conditions. Soybean inoculated with cork compost, perlite and liquid formulations produced seed yields that were not significantly different to those produced by peat-based inoculants.     

Multiphase characterization of wild Vigna associated root nodule bacteria from Japanese subtropical islands unveiled novel high temperature resistant Bradyrhizobium strains having high symbiotic compatibility with soybean and mungbean

ABSTRACT

Vigna riukiuensis plant – a rare type of vigna, found only in Taiwan and the islands of Okinawa prefecture, Japan – possesses intrinsic property of high level of salt and heat tolerance. To understand the diversity and identify suitable rhizobia, multiphase characterization of root nodule bacteria associated with V. riukiuensis grown in Ishigaki and Iriomote Islands of Okinawa prefecture was performed. Multigene phylogenetic analysis of housekeeping genes based on 16S rRNA gene sequences, 16S-23S rRNA gene internal transcribed spacer (ITS) and 23S rRNA gene sequences identified three main groups closely similar to Bradyrhizobium japonicum, B. elkanii and B. jicamae family. However, analysis of symbiotic nifH and nodD1 genes and their phylogenetic trees showed similar topology, having only few discrepancies in comparison to the housekeeping gene phylogeny. Interestingly, for some of the isolates having similarity with B. elkanii, growth was observed at 40°C, which exceed the highest record for B. elkanii to the best of our knowledge. All the isolates were observed to have the capability of forming root nodules and fix nitrogen in their original host plant V. riukiuensis and two other crops: soybean and mungbean. Most of the isolates showed similar or higher nitrogen-fixing capability in comparison with B. diazoefficiens USDA110 in V. riukiuensis and V. radiata (mungbean), and Iri 5/6 in V. riukiuensis, Iri 5/12 in soybean and Ishi 7/2 in mungbean showed highest acetylene reduction assay (in µmol/h/gm nodule dry weight) activity, which was significantly higher than B. diazoefficiens USDA110. In addition, six isolates attained higher soybean biomass production compared with B. diazoefficiens USDA110, suggesting high symbiotic compatibility with soybean. Among them, Iri 5/7 of B. elkanii group contributed 29% higher soybean biomass production than B. diazoefficiens USDA110 and could grow at 40°C, hence it could be a promising soybean inoculant in the tropics.     

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     

Evaluation of effective Myanmar Bradyrhizobium strains isolated from Myanmar soybean and effects of coinoculation with Streptomyces griseoflavus P4 for nitrogen fixation

In order to substitute the use of chemical fertilizers in legume production, there is a need for the production of rhizobial inoculants which are capable of being used as biofertilizers. To achieve this, an effective symbiotic nitrogen (N) fixation between legumes and root nodule bacteria will be essential. Evaluation of effective Myanmar Bradyrhizobium (Jordan 1982) strains isolated from Myanmar soybean (Glycine max L. Merr.) and effects of coinoculation with Streptomyces griseoflavus Krainsky 1914 P4 for N fixation were studied in pot experiments using sterilized vermiculite and Hoagland solution in the Phytotron (25°C and 70% relative humidity) with completely randomized design and three replicates. N fixation ability of soybean was evaluated by acetylene reduction activity (ARA) by gas chromatography. It was found that MAS23 showed a relatively high degree of stability and a high level of ARA per plant on both Yezin-3 and Yezin-6 soybean varieties. In the symbiotic relationship between Bradyrhizobium strains and P4 experiments, the treatments consisted of six Bradyrhizobium strains (MAS23, MAS33, MAS34, MAS43, MAS48 and USDA110) and Streptomyces griseoflavus P4 were evaluated with four Myanmar soybean varieties (Yezin-3, Yezin-6, Hinthada and Shan Sein). In the Yezin-3 soybean variety, the best treatment for ARA per plant was found in the dual inoculation of P4 and MAS34. In the Yezin-6 soybean variety, the highest nodule dry weight was found in dual inoculation of P4 with MAS34 but the highest ARA per plant was observed in the dual inoculation of P4 and MAS23. On the other hand, single inoculation of MAS43 and coinoculation of P4 with MAS48 were significantly higher in N fixation of Hinthada, and coinoculation of P4 with MAS33 was significant improvement of ARA per plant (P < 0.05) in Shan Sein soybean.     

Response of field-grown bean (Phaseolus vulgaris L.) to Rhizobium inoculation and nitrogen fertilization in two Cerrados soils

 Most soils sown with field beans (Phaseolus vulgaris L.) contain indigenous rhizobia which might interfere with the establishment of inoculated strains. As a consequence, the benefits of bean inoculation are usually questioned, and the use of N fertilizer is gradually becoming a common practice. The present study had the objective of evaluating the effectiveness of inoculation and N fertilization in field soil with (site 1) and without (site 2) a previous bean-cropping history. At site 1, which had a rhizobial population of 7×10² cells g^–1 soil, inoculation had no effect on nodulation or yield, whereas at site 2 (<10 cells g^–1 soil) inoculation increased nodulation, nodule occupancy by the inoculated strain and grain yield. N fertilizer decreased nodulation at both sites, but increased grain yield at site 1 but not at site 2, indicating that the response to inoculation and N fertilization depends on the cropping history. When bean was cultivated for the first time, indigenous populations of rhizobia were low and high yields were accomplished solely with seed inoculation, with no further response to N fertilizer. In contrast, previous cultivation of bean increases soil rhizobia, preventing nodule formation by inoculated strains, and N fertilizer may be necessary for maximum yields. A significant interaction effect between N fertilizer and inoculation was detected for serogroup distribution only at site 2, with N fertilizer decreasing nodule occupancy by the inoculated strain and increasing the occurrence of indigenous strains. Consequently, although no benefits were obtained by the combination of inoculation and N fertilizer, this practice may be feasible with the selection of appropriate N-tolerant strains from the indigenous rhizobial population. Received: 26 May 1999     

A multi-site field evaluation of granular inoculants for legume nodulation

The most common method of inoculating legume crops in Australia is the application of peat slurry inoculant to seed. The recent introduction of granular (solid) formulations of inoculants into the Australian market has provided the potential to apply rhizobia with greater ease, but their efficacy has not been independently evaluated. Here, we compare the efficacy of a range of experimental and commercially-available granular inoculants on chickpea, faba bean, lentil, lupin and pea crops in comparison with un-inoculated treatments, and with conventional seed-applied peat slurry inoculants. Thirty-seven field experiments were established in Victoria, South Australia and southern New South Wales over five years. Peat slurry inoculants provided effective nodulation of all legumes. Granular inoculants varied markedly in their ability to improve grain legume nodulation. The size of response depended inversely on background nodulation from soil rhizobial populations. At sites with median background nodulation, peat granules and attapulgite clay granules placed with seed resulted in nodulation similar to peat-slurry-based inoculation, but treatments with bentonite clay granules did not increase nodule numbers much above those in un-inoculated treatments. The generally lower numbers of rhizobia g⁻¹ in the bentonite granules, translated to lower rhizobia application rate to the soil. However, differences in number of rhizobia g⁻¹ granule did not fully explain the nodulation differences between granules. Granule moisture content and granule particle size differed markedly between granule types but their influence on nodulation was not tested. Grain yields did not differ between attapulgite granules placed with seed, peat granules and peat slurry inoculants (all well-nodulated treatments), but were lower with bentonite granule inoculants. Yield differences within sites were related to nodulation and the differences between treatments attenuated as background nodulation increased. Overall, these studies demonstrate that certain granule types have the potential to be used in Australia with grain legumes, particularly in circumstances when seed-applied inoculants are problematic, such as where seed fungicides or insecticides need to be applied. However, granular inoculant formulations differ substantially in their potential to produce nodules on a range of grain legumes.     

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.     

Competitive ability of selected Cyclopia Vent. rhizobia under glasshouse and field conditions

This study tested the competitive ability of three locally isolated Cyclopia rhizobia and strain PPRICI3, the strain currently recommended for the cultivation of Cyclopia, a tea-producing legume. Under sterile glasshouse conditions, the three locally isolated strains were equally competitive with strain PPRICI3. In field soils, the inoculant strains were largely outcompeted by native rhizobia present in the soil, although nodule occupancy was higher in nodules growing close to the root crown (the original inoculation area). In glasshouse experiments using field soil, the test strains again performed poorly, gaining less than 6% nodule occupancy in the one soil type. The presence of Cyclopia-compatible rhizobia in field soils, together with the poor competitive ability of inoculant strains, resulted in inoculation having no effect on Cyclopia yield, nodule number or nodule mass. The native rhizobial population did not only effectively nodulate uninoculated control plants, they also out-competed introduced strains for nodule occupancy in inoculated plants. Nonetheless, the Cyclopia produced high crop yields, possibly due to an adequate supply of soil N.     

Benefits of inoculation of the common bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis>) crop with efficient and competitive<Emphasis Type="Italic"> Rhizobium tropici</Emphasis> strains

Cropping in low fertility soils, especially those poor in N, contributes greatly to the low common bean (Phaseolus vulgaris L.) yield, and therefore the benefits of biological nitrogen fixation must be intensively explored to increase yields at a low cost. Six field experiments were performed in oxisols of Paraná State, southern Brazil, with a high population of indigenous common bean rhizobia, estimated at a minimum of 10³ cells g^–1 soil. Despite the high population, inoculation allowed an increase in rhizobial population and in nodule occupancy, and further increases were obtained with reinoculation in the following seasons. Thus, considering the treatments inoculated with the most effective strains (H 12, H 20, PRF 81 and CIAT 899), nodule occupancy increased from an average of 28% in the first experiment to 56% after four inoculation procedures. The establishment of the selected strains increased nodulation, N₂ fixation rates (evaluated by total N and N-ureide) and on average for the six experiments the strains H 12 and H 20 showed increases of 437 and 465 kg ha^–1, respectively,in relation to the indigenous rhizobial population. A synergistic effect between low levels of N fertilizer and inoculation with superior strains was also observed, resulting in yield increases in two other experiments. The soil rhizobial population decreased 1 year after the last cropping, but remained high in the plots that had been inoculated. DGGE analysis of soil extracts showed that the massive inoculation apparently did not affect the composition of the bacterial community.     

Evaluation of the intrinsic competitiveness and saprophytic competence of Rhizobium tropici IIB strains

Inoculation of beans (Phaseolus vulgaris L.) with strains of R. tropici IIB and R. etli resulted in the disappearance of the R. tropici IIB stains from the nodule population and their replacement by other (non R. tropici IIB) bean symbionts (Vlassak et al. 1996). Coinoculation studies in monoxenic conditions and in soil core microcosms with plants harvested at two different growth stages indicated that the inoculated R. tropici IIB strains CIAT899 and F98.5 possess a good intrinsic competitiveness which declines, however, at a later plant growth stage and in soil conditions. The poor saprophytic competence of R. tropici IIB strain CIAT899 was further demonstrated by its poor survival in soil core microcosms after bean harvest. Strains were isolated from the field plots with a 3-year bean-planting history, characterized and evaluated for their competitiveness against R. tropici IIB strain CIAT899. Isolates from field plots, which had been repeatedly inoculated with R. tropici IIB strain CIAT899, showed a higher nodule occupancy compared to R. tropici IIB strain CIAT899, and this higher competitiveness exhibited by the field isolates might be an additional reason for the poor performance of R. tropici IIB strain CIAT899 in the field study. Plots with and without a history of bean production revealed after 3-year bean cultivation an almost totally different population that also significantly differed in competitiveness. Received: 12 February 1996     

Competition and persistence of Rhizobium tropici and Rhizobium etli in tropical soil during successive bean (Phaseolus vulgaris L.) cultures

Strains of Rhizobium tropici IIB, CIAT899 and F98.5, both showing good N₂ fixation, and a R. etli strain W16.3SB were introduced into a field which had no history of bean culture. Plant dilution estimates showed that in the presence of its host (Phaseolus vulgaris cv. Carioca) during the cropping seasons and the subsequent fallow summer periods, the bean rhizobial populations increased from less than 30 to 10³ g^–1 dry soil after 1 year and to 10⁴ g^–1 dry soil after 2 years. In the 1st year crop, the inoculated strains occupied most of the nodules, which resulted in a higher nodulation and C₂H₂ reduction activity. Without reinoculation for the second and third crops, however, little R. tropici IIB was recovered from the nodules and the bean population consisted mainly of R. etli, R. leguminosarum bv. phaseoli, and R. tropici IIA. Reinoculation with our superior R. tropici IIB strains before the second crop resulted in R. tropici IIB occupying the main part of the nodules and a positive effect on nodulation and C₂H₂ reduction activity, but reintroduction of the inoculant strain in the third season did not have any effect.     

Rhizobial inoculation improves drought tolerance,biomass and grain yields of common bean (Phaseolus vulgaris L.) and soybean (Glycine max L.) at Halaba and Boricha in Southern Ethiopia

ABSTRACT

While pulses are staple food-legumes in Ethiopia, their productivity is low due to low soil fertility. Elite rhizobial strains that significantly increased shoot dry weight and nitrogen (N) contents of common beans and soybeans in greenhouse were selected for two-year field trials to evaluate their effect on yields of the pulses in the field. Each pulse had six treatments, namely four rhizobial inoculants, uninoculated control, and synthetic N fertilizer. In the drought-affected year 2015, inoculated pulses tolerated moisture stress better than non-inoculated controls. Inoculation was conducive to higher or equivalent yields compared to synthetic N fertilizer. At Halaba, bean inoculated with strain HAMBI3562 gave the highest grain yield (1500 ± 81 kg ha^?1; mean±SE) while the control yielded only 653 ± 22 kg ha^?1. At Boricha, HAMBI3570 gave a grain yield (640 ± 35 kg ha^?1) comparable to synthetic N. When rainfall was optimal in 2016, inoculation with HAMBI3562 and HAMBI3570 gave grain yields (around 4300 kg ha^?1) equivalent to synthetic N. With soybean, strain HAMBI3513 produced consistently higher or comparable biomass and grain yields compared to synthetic N. In conclusion, HAMBI3562 and HAMBI3570 for beans and HAMBI3513 for soybeans can serve as inoculants for areas having similar conditions as the test areas.     

INTERACTIONS BETWEEN COMMON BEAN GENOTYPES AND RHIZOBIA STRAINS ISOLATED FROM MOROCCAN SOILS FOR GROWTH,PHOSPHATASE AND PHYTASE ACTIVITIES UNDER PHOSPHORUS DEFICIENCY CONDITIONS

The improvement of common bean production requires the selection of effective rhizobia strains and Phaseolus vulgaris genotypes adapted to available soil phosphorus limitations. The interactions between bean genotypes and rhizobia were studied in hydroponic culture using six genotypes and four strains, CIAT899 as reference and three strains isolated from nodule of farmer's fields in the Marrakech region. The phosphorus (P) sub-deficiency caused a significant reduction on shoot biomass in some bean genotype-rhizobia combinations. Nodule biomass is significantly more reduced under P limitation for several combinations tested. Bean plants inoculated with these local rhizobial strains showed higher nodulation and an increase of nodules phytase and phosphatase activities under phosphorus sub-deficiency especially for RhM11 strain. It was concluded that the studied bean-rhizobia symbiosis differ in their adaptation to phosphorus sub-deficiency and the nodule phosphatases and phytases activities may constitute a strategy of nodulated bean plants to adapt their nitrogen fixation to P deficiency.     

Effect of CO2-free air treatment on nitrogen fixation of soybeans inoculated with Bradyrhizobium japonicum and Rhizobium fredii

Soybean (Glycine max L. Merr.) cultivars Akisirome and Peking were inoculated with Bradyrhizobium japonicum Is-21 and Rhizobium fredii USDA 194, respectively, and were grown in cylindrical pots containing sterilized vermiculite which were aerated with CO₂-free air or ambient air to study the effects of CO₂ deficiency in the rhizosphere on plant growth, nodulation, and nitrogen fixation. The repressive effects of CO₂-free air treatment were more conspicuous in Peking than Akisirome, and nodule number, nodule mass, amount of biologically fixed N and plant growth of Peking were reduced remarkably by the CO₂-free air treatment.

Acetylene reduction activity (ARA) of Peking inoculated with USDA 194 and Akisirome inoculated Is-21 was assayed in the absence and presence of CO₂, ARA of Akisirome was not affected by the absence of CO₂, while that of Peking decreased drastically.

Based on these results, it was concluded that nitrogen fixation by Akisirome inoculated with B. japonicum Is-21 was not appreciably influenced by CO₂ deficiency in the rhizosphere, while that of Peking inoculated with R. fredii USDA 194 was severely repressed, and the decrease was estimated to be due to both the reduction of the nodule mass and specific nitrogen-fixing activity.     

Sampling effects on the assessment of genetic diversity of rhizobia associated with soybean and common bean

Biological nitrogen fixation plays a key role in agriculture sustainability, and assessment of rhizobial diversity contributes to worldwide knowledge of biodiversity of soil microorganisms, to the usefulness of rhizobial collections and to the establishment of long-term strategies aimed at increasing contributions of legume-fixed N to agriculture. Although in recent decades the use of molecular techniques has contributed greatly to enhancing knowledge of rhizobial diversity, concerns remain over simple issues such as the effects of sampling on estimates of diversity. In this study, rhizobia were isolated from nodules of plants grown under field conditions, in pots containing soil, or in Leonard jars receiving a 10⁻² or a 10⁻⁴ serially-diluted soil inoculum, using one exotic (soybean, Glycine max) and one indigenous (common bean, Phaseolus vulgaris) legume species. The experiments were performed using an oxisol with a high population (10⁵ cells g⁻¹ soil) of both soybean rhizobia, composed of naturalized strains introduced in inoculants and of indigenous common-bean rhizobia. BOX-PCR was used to evaluate strain diversity, while RFLP-PCR of the ITS (internally transcribed spacer) region with five restriction enzymes aimed at discriminating rhizobial species. In both analyses the genetic diversity of common-bean rhizobia was greater than that of soybean. For the common bean, diversity was greatly enhanced at the 10⁻⁴ dilution, while for the soybean dilution decreased diversity. Qualitative differences were also observed, as the DNA profiles differed for each treatment in both host plants. Differences obtained can be attributed to dissimilarity in the history of the introduction of both the host plant and the rhizobia (exotic vs. indigenous), to host-plant specificity, rhizobial competitiveness, and population structure, including ease with which some types are released from microcolonies in soil. Therefore, sampling method should be considered both in the interpretation and comparison of the results obtained in different studies, and in the setting of the goals of any study, e.g. selection of competitive strains, or collection of a larger spectrum of rhizobia. Furthermore, effects of sampling should be investigated for each symbiosis.