Plant growth-promoting rhizobacteria alter rooting patterns and arbuscular mycorrhizal fungi colonization of field-grown spring wheat

The impact of plant growth-promoting rhizobacteria (PGPR) inoculants on the growth, yield and interactions of spring wheat with arbuscular mycorrhizal fungi (AMF) was assessed in field studies. The pseudomonad inoculants P. cepacia R55, R85, P. aeruginosa R80, P. fluorescens R92 and P. putida R104, which enhance growth and yield of winter wheat, were applied at a rate of ca. 10⁷–10⁸ cfu seed^-1 and plots established on pea stubble or summer fallow at two different sites in Saskatchewan. Plant shoot and root biomass, yield and AMF colonization were determined at four intervals. Plant growth responses were variable and dependent on the inoculant strain, harvest date and growth parameter evaluated. Significant increases or decreases were measured at different intervals but these were usually transient and final seed yield was not significantly affected. Harvest index was consistently increased by all pseudomonad inoculants; responses to strain R55 and R104 were significant. Root biomass to 60 cm depth was not significantly affected by inoculants except strain R104, which significantly reduced root dry weight. However, root distribution, root length and AMF colonization of roots within the soil profile to 60 cm were significantly altered by inoculants. Most of these responses were reductions in the assessed parameter and occurred at depths below 15 cm; however, strains R85 and R92 significantly increased root dry weight in the 0- to 15-cm zone. These results indicate that some PGPR inoculants may adversely affect mutualistic associations between plants and indigenous soil microorganisms, and suggest a possible reason as to why spring wheat growth was not consistently enhanced by these pseudomonad PGPR.     

Bacteria associated with Glomus clarum spores influence mycorrhizal activity

The effects of bacteria associated with arbuscular mycorrhizal fungal (AMF) spores on spore germination, growth in vitro and on the pea-AMF symbiosis were evaluated. Bacterial colonies were recovered from untreated Glomus clarum NT4 spores and NT4 spores decontaminated with 5% chloramine-T for 30, 45 and 60 min on five different media. Both G+ and G− bacteria were recovered from untreated NT4 spores, whereas only G+ bacteria were isolated from decontaminated spores. An in vitro assessment of the effect of spore-associated bacteria on clean, decontaminated NT4 spores revealed that (i) most of the bacteria isolated from untreated spores generally did not significantly alter spore function, (ii) some bacteria isolated from clean, decontaminated spores inhibited or stimulated NT4 spore germination, (iii) stimulation of spore germination occurred only when bacteria were in contact with spores, and (iv) inhibition of spore germination was the result of volatile bacterial metabolites. A stimulatory bacterial isolate, Bacillus pabuli LA3, significantly (P<0.05) enhanced the shoot growth, AMF-colonization, shoot N content and P use efficiency of NT4-inoculated 6 week-old pea plants over that of plants co-inoculated with an inhibitory bacterial isolate, Bacillus chitinosporus LA6a and NT4.     

Selective interactions between arbuscular mycorrhizal fungi and<Emphasis Type="Italic"> Rhizobium leguminosarum</Emphasis> bv.<Emphasis Type="Italic"> viceae</Emphasis> enhance pea yield and nutrition

This study evaluated the response of pea (Pisum sativum cv. Trapper) to arbuscular mycorrhizal fungi (AMF) and Rhizobium leguminosarum bv. viceae strains varying in their effectiveness on pea. Plants were inoculated with the AMF species Glomus clarum NT4 or G. mosseae NT6 and/or ten Rhizobium strains, and grown for 90 days in soil containing indigenous AMF and rhizobia. The effectiveness of the Rhizobium strains on the growth (P <0.046; r =0.64) and N nutrition (P <0.04; r =0.65) of 6-week-old pea grown under gnotobiotic conditions was correlated with the growth and N nutrition of 90-day-old pea grown in natural soil for all strains except LX48. The growth and yield response of pea to co-inoculation with AMF and Rhizobium strains depended on the particular AMF-Rhizobium strain combination. In some cases, the yield and N nutrition of pea inoculated with a superior Rhizobium strain was significantly (P <0.05) enhanced by an apparently compatible AMF species compared to the Rhizobium treatment. On the other hand, an apparently incompatible AMF species significantly (P <0.05) reduced the performance of an effective Rhizobium strain. In general, treatments with effective Rhizobium strains or co-inoculation treatments with effective Rhizobium strains and a compatible AMF species produced the best results. Changes in total shoot dry matter production was significantly (P <0.05) correlated with the total shoot N (P <0.0001; r =0.95) and P content (P <0.0001; r =0.87), indicating that this response was mediated by enhanced N and P nutrition. Growth, yield and nutrition of pea were not related to AMF colonization of roots. Our results suggest that careful co-selection of AMF species and Rhizobium strains can enhance pea yield and nutrition.     

Influence of Pseudomonas syringae R25 and P. putida R105 on the growth and N2 fixation (acetylene reduction activity) of pea (Pisum sativum L.) and field bean (Phaseolus vulgaris L.)

The effects of inoculating field peas (Pisum sativum L.) with Rhizobium leguminosarum and field beans (Phaseolus vulgaris L.) with R. phaseoli, alone or in combination with Pseudomonas syringae R25 and/or P. putida R105, were assessed under gnotobiotic conditions in growth pouches and in potted soil in a growth chamber. Inoculation of peas with P. syringae R25 or P. putida R105 alone had no effect on plant growth in pouches. In soil, however, the isolate R25 inhibited nitrogenase activity (as assessed by acetylene reduction assay) of nodules formed by indigenous rhizobia; strain R105 stimulated pea seedling emergence and nodulation. P. syringae R25 inhibited the growth of beans in either plant-growth system. P. putida R105, however, had no effects on beans in pouches, but reduced plant root biomass and nodulation by indigenous rhizobia in soil. Coinoculation of pea seeds with R. leguminosarum and either of the pseudomonads significantly (P<0.01) increased shoot, root, and total plant weight in growth pouches, but had no effect in soil. Co-inoculation of field beans with R. phaseoli and P. putida R105 had no effects on plant biomass in growth pouches or in soil, but the number of nodules and the acetylene reduction activity was significantly (P<0.01) increased in the soil. In contrast, co-inoculation of beans with rhizobia and P. syringae R25 had severe, deleterious effects on seedling mergence, plant biomass, and nodulation in soil and growth pouches. Isolate R25 was responsible for the deleterious effects observed. Although plant growth-promoting rhizobacteria may interact synergistically with root-nodulating rhizobia, the PGPR selected for one crop should be assessed for potential hazardous effects on other crops before being used as inoculants.     

A bradyrhizobial-<Emphasis Type="Italic">Penicillium</Emphasis> spp. biofilm with nitrogenase activity improves N<Subscript>2</Subscript> fixing symbiosis of soybean

A bradyrhizobial-fungal biofilm (i.e. Bradyrhizobium elkanii SEMIA 5019-Penicillium spp.) developed in vitro was assayed for its nitrogenase activity and was evaluated for N₂-fixing symbiosis with soybean under greenhouse conditions. The biofilm showed nitrogenase activity, but the bradyrhizobial strain alone did not. Shoot and root growth, nodulation and N accumulation of soybean increased significantly with an inoculum developed from the biofilm. This study concludes that such biofilmed inoculants can improve N₂-fixing symbiosis in legumes, and can also directly contribute to soil N fertility in the long term. Further studies should be conducted to investigate the performance of these inoculants under field conditions.     

Growth of cassava cultivar TMS 30572 as affected by alley-cropping and mycorrhizal inoculation

The effect of inoculation with Glomus clarum, a vesicular-arbuscular mycorrhiza fungus, and alley-cropping on the growth of the cassava cultivar, TMS 30572, was investigated under field conditions in a low nutrient tropical soil. Cassava was grown either interplanted between two hedgerow tree species (alley-cropped) or sole-cropped. Sub-plots were either inoculated with G. clarum or were not inoculated. No effort was made to destroy the indigenous mycorrhizal fungi. Three months after planting, no significant influence of G. clarum inoculation was observed on the growth of roots, shoots or leaf area index (LAI). However, with time, inoculation and system of cropping enhanced these growth parameters. Nine months after planting, the total biomass of alley-cropped cassava was significantly higher than that of inoculated and non-inoculated sole-cropped cassava. Inoculation had led to an increase in the fresh tuber yield of both the alley- and sole-cropped cassava 12 months after planting. The LAI of both alley- and sole-cropped cassava inoculated with G. clarum increased. Received: 6 December 1996     

Enhanced Growth and nodule occupancy of red kidney bean and soybean inoculated with soil aggregate-based inoculant

Volcanic ash soil, which is widely distributed in Japan, contains a large amount of well-structured soil aggregates. By using these aggregates as carrier materials, we prepared (brady)rhizobial inoculants for red kidney bean (Phaseolus vulgaris) and soybean (Glycine max). Autoclaved soil aggregates were inoculated with Rhizobium tropici CIATS99R or Bradyrhizobium japonicum USDA110R, incubated for 15 or 21 d at 30°C, slowly air-dried at 20°C to prepare the aggregate-based inoculants, and stored at various temperatures. The populations of CIATS99R and USDA110R in the aggregate-based inoculants were maintained during several months of storage at 20°C. When the aggregate-based inoculants were mixed with soil, CIATS99R and USDA110R cells showed a remarkably improved survival in soils compared with those mixed with soil without carrier material. The effect of the aggregate-based inoculants on the growth of red kidney bean and soybean was examined in pot experiments. By placing a small amount of the inoculant just beneath the seeds at the time of sowing, plant growth was significantly enhanced compared with the use of traditional peat-based inoculant. In addition, nodule formation on the upper part of soybean roots and nodule occupancy by the inoculated strain were remarkably enhanced by the aggregate-based inoculant. It is suggested that soil aggregates might be suitable carrier materials for preparing cheap and effective (brady)rhizobial inoculants.     

Inoculation of rice plants with the endophytic diazotrophs Herbaspirillum seropedicae and Burkholderia spp.

 Four experiments were performed under gnotobiotic conditions to select strains of the endophytic diazotrophs Herbaspirillum seropedicae and Burkholderia spp. as inocula of rice plants. Eighty strains of H. seropedicae originally isolated from rice, sorghum and maize plants, were tested in test tube cultures with N-free agar as the substrate. Rice plants showed medium and high increases in their fresh weight in response to inoculation with nineteen strains. These strains were tested again, and six strains were then selected to evaluate their contribution to the N of the plant via biological N₂ fixation (BNF) using an agar growth medium containing 5 mg N l^–1of ¹⁵N-labelled (NH₄)₂SO₄. The contribution of the strains to plant N via BNF varied from 54% when rice plants were inoculated with strain ZAE94, to 31% when strain ZAE67 was used. These results were confirmed in the fourth gnotobiotic experiment, which also included strains of the new N-fixing bacteria belonging to the genus Burkholderia, isolated from rice, as well as a strain of Burkholderia vietnamiensis, isolated from rice rhizosphere. Burkholderia spp. strains showed similar effects to those observed for H. seropedicae strains, while B. vietnamiensis fixed only 19% of plant total N. The best four strains were tested in a pot experiment where pre-germinated, inoculated rice seedlings were grown in soil labelled with ¹⁵N. The results confirmed the gnotobiotic experiments, although the levels of N in the rice plants derived from BNF of the selected H. seropedicae and Burkholderia spp. strains were lower. Nevertheless, there was an increase in N content in grains of inoculated plants, and the results showed that the method used for strain selection is very useful and can be applied to other strains of N₂-fixing bacteria and plants. Received: 4 May 1999     

Molecular characterization and PCR detection of a nitrogen-fixing Pseudomonas strain promoting rice growth

Nitrogen-fixing plant growth-promoting rhizobacteria (PGPR) from the genus Pseudomonas have received little attention so far. In the present study, a nitrogen-fixing phytohormone-producing bacterial isolate from kallar grass (strain K1) was identified as Pseudomonas sp. by rrs (16S ribosomal RNA gene) sequence analysis. rrs identity level was high with an uncharacterized marine bacterium (99%), Pseudomonas sp. PCP2 (98%), uncultured bacteria (98%), and Pseudomonas alcaligenes (97%). Partial nifH gene amplified from strain K1 showed 93% and 91% sequence similarities to those of Azotobacter chroococcum and Pseudomonas stutzeri, respectively. The effect of Pseudomonas strain K1 on rice varieties Super Basmati and Basmati 385 was compared with those of three non-Pseudomonas nitrogen-fixing PGPR (Azospirillum brasilense strain Wb3, Azospirillum lipoferum strain N4 and Zoogloea strain Ky1) used as single-strain inoculants. Pseudomonas sp. K1 was detected in the rhizosphere of inoculated plants by enrichment culture in nitrogen-free growth medium, which was followed by observation under the microscope as well as by PCR using a rrs-specific primer. For both rice varieties, an increase in shoot biomass and/or grain yield over that of noninoculated control plants was recorded in each inoculated treatment. The effect of Pseudomonas strain K1 on grain yield was comparable to those of A. brasilense Wb3 and Zoogloea sp. Ky1 for both rice varieties. These results show that nitrogen-fixing pseudomonads deserve attention as potential PGPR inoculants for rice.     

Phosphate-solubilizing rhizobacteria enhance the growth and yield but not phosphorus uptake of canola (Brassica napus L.)

The ability of phosphate-solubilizing rhizobacteria to enhance the growth and phosphorus uptake of canola (Brassica napus L., cv. Legend) was studied in potted soil experiments in the growth chamber. One hundred and eleven bacteria isolated from the rhizosphere of field-grown plants, and a collection of nine bacteria known to be effective plant growth-promoting rhizobacteria (PGPR), were screened for P-solubilization in vitro. All rhizobacteria were identified using whole-cell fatty acids methyl ester (FAME) profiles. The best P-solubilizing isolates were two Bacillus brevis strains, B. megaterium, B. polymyxa, B. sphaericus, B. thuringiensis, and Xanthomonas maltophilia (PGPR strain R85). The P-solubilizers were tested for their effects on growth and P-uptake of canola plants in a P-deficient soil amended with rock phosphate. Although some of the P-solubilizing rhizobacteria significantly increased plant height or pod yield, none increased P-uptake. The most effective inoculant was a B. thuringiensis isolate which significantly increased the number and weight of pods and seed yield without rock phosphate. Xanthomonas maltophilia increased plant height, whereas the other bacilli increased the number on weight of pods. These results demonstrate the potential use of these P-solubilizing rhizobacteria as inoculants for canola, but indicate that P-solubilization was not the main mechanism responsible for positive growth response. Received: 8 February 1996     

Synergistic effects of vesicular-arbuscular mycorrhizal fungi and diazotrophic bacteria on nutrition and growth of sweet potato (Ipomoea batatas)

Summary Sweet potatoes were micropropagated and then transplanted from axnic conditions to fumigated soil in pots in the greenhouse. Spores of Glomus clarum were obtained from Brachiaria decumbens or from sweet potatoes grown in soil infected with this fungus and with an enrichment culture of Acetobacter diazotrophicus. Three experiments were carried out to measure the beneficial effects of vesicular-arbuscular mycorrhizal (VAM) fungi-diazotroph interactions on growth, nutrition, and infection of sweet potato by A. diazotrophicus and other diazotrophs obtained from sweet potato roots. In two of these experiments the soils had been mixed with ¹⁵N-containing organic matter. The greatest effects of mycorrhizal inoculation were observed with co-inoculation of A. diazotrophicus and/or mixed cultures of diazotrophs containing A. diazotrophicus and Klebsiella sp. The tuber production was dependent on mycorrhization, and total N and P accumulation were increased when diazotrophs and G. clarum were applied together with VAM fungal spores. A. diazotrophicus infected aerial plant parts only when inoculated together with VAM fungi or when present within G. clarum spores. More pronounced effects on root colonization and intraradical sporulation of G. clarum were observed when A. diazotrophicus was co-inoculated. In non-fumigated soil, dual inoculation effects, however, were of lower magnitude. ¹⁵N analysis of the aerial parts and roots and tubers at the early growth stage (70 days) showed no statistical differences between treatments except for the VAM+Klebsiella sp. treatment. This indicates that the effects of A. diazotrophicus and other diazotrophs on sweet potato growth were caused by enhanced mycorrhization and, consequently, a more efficient assimilation of nutrients from the soil than by N₂ fixation. The possible interactions between these effects are discussed.     

Interaction among free-living N-fixing bacteria isolated from Drosera villosa var. villosa and AM fungi (Glomus clarum) in rice (Oryza sativa)

Rice is usually grown in N-deficient soils, demanding that the element be supplied to the field by commercially available N fertilizers. Unfortunately, a substantial amount of the urea-N or NO₃-N applied as fertilizers is lost through different mechanisms, causing environmental pollution problems. Utilization of biological N₂ fixation (BNF) technology can decrease the application of N fertilizers, reducing environmental risks. This study evaluated the effects of four free-living N-fixing bacterial species, isolated from oligotrophic soil conditions, as single inoculants or combined with arbuscular mycorrhizal fungi (Glomus clarum), on the development of rice plants grown as flooded or upland rice, in the greenhouse. Upland rice roots were inoculated with Methylobacterium sp., Burkholderia sp. and Sphingomonas sp., whereas the species Burkholderia sp., Pseudomonas sp. and Sphingomonas sp., were inoculated on flooded rice. Inoculants consisted of individual bacterial species or their mixtures, with or without G. clarum. Controls included non-bacteria/non-AM fungi, and AM fungi alone. Experiments were carried out in five replicates. The presence of G. clarum decreased or did not significantly affect plant growth under the different culture conditions. The presence of AM fungi stimulated the N-fixing bacterial population of upland rice. Bacterial species had different effects, under both culture conditions, and some genera of N-fixing bacteria increased root and shoot growth at different plant growth stages. The level of mycorrhiza colonization had no influence on plant growth     

Biostimulant effects of Bacillus strains on wheat from in vitro towards field conditions are modulated by nitrogen supply

Bacillus velezensis strains, belonging to plant growth‐promoting rhizobacteria (PGPR), are increasingly used as microbial biostimulant. However, their field application to winter wheat under temperate climate remains poorly documented. Therefore, three B. velezensis strains IT45, FZB24 and FZB42 were tested for their efficacy under these conditions. Two biological interaction systems were firstly developed under gnotobiotic and greenhouse conditions combined with sterile or non‐sterile soil, respectively, and finally assayed in the field during two years coupled with different N fertilization rates. Under gnotobiotic conditions, all three strains significantly increased root growth of 14 d‐old spring and winter wheat seedlings. In the greenhouse using non‐sterile soil, only FZB24 significantly increased root biomass of spring wheat (+31%). The three strains were able to improve nutrient uptake of the spring wheat grown in the greenhouse, particularly for the micronutrients Fe, Mn, Zn, and Cu, but the observed increases in nutrient uptake were dependent on the organs and the elements. The root biomass increases in inoculated plants coincided with lowered nutrient concentrations of P and K. In 2014, under field conditions and absence of any N fertilizer supply, FZB24 significantly increased grain yields by 983 kg ha^?1, or 14.9%, in relation to non‐inoculated controls. The three strains in the 2015 field trial failed to confirm the previous positive results, likely due to the low temperatures occurring during and after inoculations. The Zeleny sedimentation value, indicative of flour quality, was unaffected by the inoculants. The results are discussed in the perspective of bacterial application to wheat under temperate agricultural practices.     

Interactions of Glomus clarum with Acetobacter diazotrophicus in infection of sweet potato (Ipomoea batatas), sugarcane (Saccharum spp.), and sweet sorghum (Sorghum vulgare)

Summary Spores of the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus clarum obtained from sweet potatoes grown in soil inoculated with this fungus and with an enrichment culture of Acetobacter diazotrophicus contained A. diazotrophicus and several other bacteria, including a diazotrophic Klebsiella sp. Inoculation of micropropagated sweet potatoes with G. clarum and A. diazotrophicus enhanced spore formation in soil compared to VAM inoculation alone. Plants inoculated with VAM spores containing the bacteria showed additional increases in the number of spores formed within roots. A. diazotrophicus infected aerial plant parts only when inoculated together with VAM or when present within VAM spores. Micropropagated sugarcane seedlings inoculated with the same VAM spores containing the diazotrophs also contained much higher numbers of A. diazotrophicus in aerial parts than seedlings inoculated in vitro with the bacteria alone. When grown in non-sterile soil, the sugarcane seedlings again showed the greatest infection of aerial parts after inoculation with VAM spores containing the diazotrophs. This treatment also increased VAM colonization and the numbers of spores formed within roots. Similar effects were observed in sweet sorghum except that the aerial plant parts were not infected by A. diazotrophicus.     

Nodulation competitiveness of Rhizobium leguminosarum bv. phaseoli and Rhizobium tropici strains measured by glucuronidase (gus) gene fusion

Summary The nodulation competitiveness of 17 Rhizobium leguminosarum bv. phaseoli and 3 R. tropici strains was analysed in growth pouches, at pH 5.2 and 6.4. All 20 strains were coinoculated with a gus ⁺ strain of R. leguminosarum bv. phaseoli strain KIM5s. The gus⁺ phenotype, carrying the glucuronidase gene, was used to type nodules directly in the growth pouches. Nodule occupancy ranged from 4% for the least competitive to 96% for the most competitive R. leguminosarum bv. phaseoli strain. The R. tropici strains showed low rates of nodule occupancy at pH 6.4 but their competitiveness improved significantly under acid conditions. CIAT 895 was the only R. leguminosarum bv. phaseoli strain that was less competitive (P<0.05) at the lower pH. The competitiveness of all the other R. leguminosarum bv. phaseoli strains was unaffected by pH. Various physiological and genetic properties of the strains were analysed in search of correlations with nodulation competitiveness. Hybridisation patterns with three different DNA probes (nif KDH, common nod genes, and hup genes) and the metabolism of 53 different C sources were compared. No general correlations were found between hybridisation or growth pattern and competitiveness. The less competitive R. tropici strains had a unique DNA hybridisation pattern and were not able to use shikimate, ferulate, coumarate, or asparagine as C sources. Most of the less competitive R. leguminosarum bv. phaseoli strains could not metabolize either ferulate or coumarate. This might indicate a relationship between nodulation competitiveness and the ability to degrade aromatic compounds.     

Biostimulant effects of rhizobacteria on wheat growth and nutrient uptake depend on nitrogen application and plant development

The capacity of plant growth-promoting rhizobacteria (PGPR) – Bacillus amyloliquefaciens GB03 (BamGB03), B. megaterium SNji (BmeSNji), and Azospirillum brasilense 65B (Abr65B) – to enhance growth and nutrient uptake in wheat was evaluated under different mineral N fertilizer rates, in sterile and non-sterile soils, and at different developmental stages. In gnotobiotic conditions, the three strains significantly increased plant biomass irrespective of the N rates. Under greenhouse conditions using non-sterile soil, growth promotion was generally highest at a moderate N rate, followed by a full N dose, while no significant effect was observed for the inoculants in the absence of N fertilizer. At 50N, plant biomass was most significantly increased in roots (up to +45% with Abr65B) at stem-elongation stage and in the ears (+19–23% according to the strains) at flowering stages. For some nutrients (N, P, Mn, and Cu), the biomass increases in roots and ears were paralleled with reduced nutrient concentrations in the same organs. Nevertheless, growth stimulation resulted in a higher total nutrient uptake and higher nutrient uptake efficiency. Furthermore, Abr65B and BmeSNji counteracted the reduction of root development caused by a high N supply. Therefore, combining PGPR with a proper cultivated system, N rate, and plant stage could enhance their biostimulant effects.     

INOCULATION EFFECTS OF PANTOEA AGGLOMERANS STRAINS ON GROWTH AND CHEMICAL COMPOSITION OF PLUM

In this study, five non-pathogenic Pantoea agglomerans strains were utilized. The objective of this study was to test for biochemical characteristics of these strains, and to evaluate their inoculation effects on fruit set rate, fruit pomological traits, fruit chemical compositions and some vegetative parameters of plum cultivar ‘Stanley’. The results showed that some of the tested strains had beneficial effects on the fruit set rate, fruit pomological traits, fruit chemical composition and/or some vegetative growth parameters of plum in comparison to the control. In conclusion, tested Pantoea agglomerans strains (especially RK-85) are the suitable inoculants for plum cultivation in cold areas such as Erzurum, and these strains may be considered as biofertilizer and protector sensitive plants against frost damage by applying in suitable timing and dose.     

Establishment and persistence of Rhizobium trifolii in western australian soils

Large scale experiments with inoculated and drill sown Trifolium subterraneum, T. hirtum, and T. cherleri showed that recent isolates of Rhizobium trifolii from healthy plants in problem pastures were superior to the strains used in commercially available inoculants. The new rhizobia are also shown to persist in the soil longer than the commercial strains. Evidence was obtained of different levels of performance by R. trifolii strains on different soils. Following the inclusion of one of the superior isolates in commercial peat inoculants, a number of farmer-sown pastures were examined for strain persistence. The new isolates showed much improved persistence over the older inoculant strains.     

Characterization and evaluation of North American Trifolium ambiguum -nodulating rhizobia

Kura clover ( Trifolium ambiguum M. Bieb.) is a promising persistent forage legume, whose use is currently limited by establishment difficulties in part attributable to nodulation problems. In this study, we phenotypically characterized 18 Kura clover rhizobia including 12 newly isolated North American strains using 111 growth tests. The symbiotic performances of these 12 strains when used to inoculate hexaploid Kura clover plants were then evaluated in a growth chamber and compared to five strains commonly used in commercial Kura clover inoculants. Non-inoculated plants with or without N fertilization were used as controls. Field evaluations were also conducted in 2001 and 2002 in Montreal, Quebec and Becker, Minnesota. Hexaploid Kura clover was seeded with one of three North American strains or three commercial strains and compared to non-inoculated controls with or without N fertilizer. Phenotypic diversity observed among the 12 North American strains was limited. The North American strains were often more efficient than strains commonly used in commercial rhizobial inoculants in both growth chamber and field trials. In the growth chamber study, 60 days after seeding, shoot dry matter accumulation was overall 205% greater with North American strains than with commercial strains and 45% greater than with the best commercial strain tested. Some of the strains currently used in commercial inoculants failed to nodulate Kura clover plants effectively. North American strains tested in field trials (i.e., CT1-1, CT1-2, and WI4-4) overall increased total and shoot dry matter accumulation, 100 days after seeding, in three of four environments by 27% and 31% respectively, when compared to commercial inoculant strains. While these strains appear to have potential they still produced less dry matter than non-inoculated N-fertilized controls.