Diversity and symbiotic effectiveness of Rhizobium leguminosarum bv. trifolii isolates from pasture soils in south-western Australia

Soil Research ◽  
2002 ◽  
Vol 40 (8) ◽  
pp. 1319 ◽  
Author(s):  
M. T. Collins ◽  
J. E. Thies ◽  
L. K. Abbott
Keyword(s):  
Western Australia ◽  
Rhizobium Leguminosarum ◽  
Subterranean Clover ◽  
Most Probable Number ◽  
Enzyme Linked Immunosorbent Assay ◽  
Symbiotic Effectiveness ◽  
Probable Number ◽  
Pcr Fingerprinting ◽  
Rapd Pcr ◽  
Inoculant Strain

The abundance of the Australian inoculant strain of Rhizobium leguminosarum bv. trifolii for subterraneum clover (WU95) and the diversity of naturalised rhizobia were assessed in 3 subterranean clover pastures in the Albany region of south-western Western Australia. Most probable number, enzyme linked immunosorbent assay (ELISA), and polymerase chain reaction (PCR) techniques were used. A putative strain similar to inoculant strain WU96 was uncommon at one site (South Stirling) and not isolated at 2 other sites. Randomly amplified polymorphic DNA (RAPD) PCR fingerprinting using the RPO1 primer identified 45 different profiles amongst the 208 isolates examined. RAPD-PCR fingerprinting using the primers RPO4 and RPO5 confirmed most groupings based on RPO1 fingerprint patterns and revealed further genetic diversity within some groups. Overall, 54 putative strains were defined by RAPD-PCR fingerprint profiles across the 3 sites. Subterranean clover rhizobia at the Manypeaks and Mount Shadforth sites were dominated by isolates with 1 or 2 RPO1 RAPD profiles at 2 sampling times, while the population at South Stirling was much more diverse. The symbiotic effectiveness of 11 rhizobial isolates, representing the major RPO1 RAPD profile groups within naturalised rhizobial populations, were compared in pot culture with those of the 2 commercial inoculant strains for subterranean clover, WU95 and TA1, on 3 cultivars. Differences in effectiveness among 3 of the 11 isolates were observed in comparison to both the commercial strains and other naturalised isolates. The nitrogen fixing effectiveness of 8 isolates representing different subgroups from one RP01 group was not the same. The use of all 3 primers increased the precision in defining putative strains of Rhizobium leguminosarum bv. trifolii, and although naturalised rhizobia from these pastures are saprophytically competent, their dominance in nodules does not appear to be linked to symbiotic effectiveness.

Symbiotic performance of Mediterranean Trifolium spp. with naturalised soil rhizobia

2011 ◽  
Vol 62 (10) ◽  
pp. 903 ◽  
Author(s):  
E. A. Drew ◽  
N. Charman ◽  
R. Dingemanse ◽  
E. Hall ◽  
R. A. Ballard
Keyword(s):  
Dry Matter ◽  
Rhizobium Leguminosarum ◽  
Subterranean Clover ◽  
Trifolium Subterraneum ◽  
Most Probable Number ◽  
Diverse Populations ◽  
Symbiotic Efficiency ◽  
Probable Number ◽  
Mediterranean Origin ◽  
Inoculant Strain

Naturalised soil rhizobia that nodulate clover occur in high number and are known to vary in their symbiotic performance (SP) with subterranean clover (Trifolium subterraneum L.). However, the extent of suboptimal fixation across a range of other clover species is not well understood. T. subterraneum and nine other annual clover species of Mediterranean origin were evaluated for their SP in combination with the naturalised clover rhizobia in 71 Australian soils and five strains of Rhizobium leguminosarum bv. trifolii that have been used in the inoculants produced for clovers. The most probable number method, using subterranean clover as the trap plant was used to estimate the number of clover rhizobia in the soils. Ninety-two percent of soils tested contained more than 1000 rhizobia/g. An extract of each soil, or strain of rhizobia was used to inoculate plants growing in N-deficient media in the greenhouse. Plants were grown for 4 weeks after inoculation and shoot dry matter determined and expressed as a percentage of the ‘best’ soil rhizobia treatment, to provide a proportional measure of SP for each clover species. SP (mean of clover species) ranged from 96% with the current inoculant strain for annual clovers (WSM1325) down to 48% with former inoculant strain WU95. When inoculated with soils predominantly from mainland Australia, SP (mean of soil treatments) of the different Trifolium spp. was 55% (resupinatum), 53–47% (subterraneum), 50% (nigrescens), 49% (michelianum), 48% (isthmocarpum), 38% (hirtum), 35% (purpureum), 32% (vesiculosum), 25% (spumosum) and 21% (glanduliferum). Within each of the clover species, SP resulting from individual soil treatments ranged from 100% (by definition for the best soil treatment) down to close to zero. Trifolium glanduliferum formed nodules readily with the inoculant strains but nodulation was erratic with the rhizobia in many soils. It is therefore proposed that the naturalised rhizobia in many soils are unlikely to be inoculant strains. This research demonstrates symbiotic efficiency across annual clover species is compromised where diverse populations of clover rhizobia have naturalised in soils.

Behavior of Phytophthora clandestina propagules at a field site in Western Australia

Soil Research ◽  
1986 ◽  
Vol 24 (4) ◽  
pp. 485 ◽  
Author(s):  
DH Wong ◽  
MJ Barbetti ◽  
K Sivasithamparam
Keyword(s):  
Soil Moisture ◽  
Western Australia ◽  
Marked Decrease ◽  
Subterranean Clover ◽  
Sole Source ◽  
Disease Index ◽  
Most Probable Number ◽  
Field Soil ◽  
Field Site ◽  
Probable Number

The number of infective propagules of Phytophthora clandestina, an important pathogen of subterranean clover, was estimated by the most probable number (MPN) method in soil at a field site in Western Australia. The behaviour of the propagules of this fungus was studied in relation to root disease index (RDI), disease suppressiveness index (DSI), rainfall, soil moisture and soil temperature for a period of 12 months. There was an increase in the number of infective propagules in the field soil in January and no further increase was observed until May, then marked decrease occurred in June and July with the numbers remaining low through to December. There was no significant correlation between MPN and RDI, DSI, rainfall, soil moisture or temperature. The RDI was, however, negatively correlated (P < 0.05) with DSI linearly (r = 0.76) and quadratically (r = 0.81). Subterranean clover appears to be the sole source of P. clandestina inoculum in the pasture sward of mixed plant species.

Symbiotic effectiveness of Rhizobium leguminosarum bv. trifolii collected from pastures in south-western Victoria

1997 ◽  
Vol 37 (6) ◽  
pp. 623 ◽  
Author(s):  
P. E. Quigley ◽  
P. J. Cunningham ◽  
M. Hannah ◽  
G. N. Ward ◽  
T. Morgan
Keyword(s):  
Rhizobium Leguminosarum ◽  
Subterranean Clover ◽  
Annual Rainfall ◽  
Test Plant ◽  
Available Phosphorus ◽  
Symbiotic Effectiveness ◽  
Plant Weight ◽  
Commercial Inoculant ◽  
Shoot Weight ◽  
Western Victoria

Summary. The whole-soil inoculation method was used to assess the symbiotic effectiveness of rhizobia populations in soils collected from 18 randomly-selected pastures in south-western Victoria. This was part of a larger study which described the condition of pasture within this region. Based on the shoot weights of test subterranean clover plants, cv. Mount Barker, effectiveness varied from 36 to 94% depending on the site of rhizobia collection. This range was wider than that found in an earlier survey of rhizobia effectiveness conducted nearby. WU95, the commercial inoculant for subterranean clover, was significantly more effective than 9 of the rhizobia samples. Rhizobia from 2 sites were especially poor and their effectiveness (37%) was not significantly different from the nil inoculum control (28%). Symbiotic effectiveness was not related to soil pH, available sulfur, available phosphorus, total nitrogen or mean annual rainfall for each site where rhizobia were collected. After pooling data for all sites, the shoot weights were significantly related to the proportions of plants with nodules assigned high nodulation scores. In contrast, low scores, within 1 of 6 categories, did not significantly affect shoot weight. The technique of using mean nodulation score was less capable of discriminating differences in symbiotic effectiveness, compared with assessment based on test plant weight.

scholarly journals Seasonal Variation of Ralstonia solanacearum Biovar 2 Populations in a Spanish River: Recovery of Stressed Cells at Low Temperatures

2005 ◽  
Vol 71 (1) ◽  
pp. 140-148 ◽  
Author(s):  
Paola Caruso ◽  
Jose Luis Palomo ◽  
Edson Bertolini ◽  
Bel�n �lvarez ◽  
Mar�a M. L�pez ◽  
...  
Keyword(s):  
Ralstonia Solanacearum ◽  
Water Samples ◽  
Low Temperatures ◽  
Most Probable Number ◽  
Enzyme Linked Immunosorbent Assay ◽  
Probable Number ◽  
Solid Media ◽  
Nonculturable State ◽  
Enrichment Protocol ◽  
Stressed Cells

ABSTRACT The presence of Ralstonia solanacearum biovar 2 in the watercourses of European countries is increasing, but little is known about its ecology in aquatic habitats. The detection of this pathogen in 2000 in one Spanish river led us to study its population density at different locations on the river over a period of 3 years. During 2000 and 2001, the pathogen was recovered at low densities (10 to 80 CFU/ml) by direct plating on modified SMSA agar from water samples at 14�C or higher, but its isolation was usually unsuccessful at temperatures below 9�C. To monitor the pathogen's abundance in winter, we used two liquid selective media for enrichment (at 29 and 35�C) and compared them by using spiked river water samples: modified Wilbrink broth (MWB) was more efficient than modified SMSA broth for double-antibody-sandwich indirect enzyme-linked immunosorbent assay (DASI-ELISA) detection of R. solanacearum. Enrichment in MWB at both temperatures allowed us to recover R. solanacearum cells that were nonculturable on solid media up to 25 days after their entry into the viable but nonculturable state. When we applied this technique to water samples during the cold months of 2001 and 2002, we obtained the best detection results by the most-probable-number method after enrichment at 35�C with MWB. The enrichment protocol was combined with DASI-ELISA and validated by Co-PCR to detect both naturally and artificially starved and cold-stressed cells in water, which were still infective. Overall, the data from this study demonstrate the effects of temperature variation on the population and culturability of R. solanacearum cells on solid media and their survival at low temperatures.

Influence of chemical and biological properties of field grown pea and lentil rhizospheres on nodulation status and resident rhizobial population in acidic soils of Assam, India

2015 ◽  
Vol 38 (6) ◽  
Author(s):  
D. J. Nath ◽  
D. Gogoi ◽  
A. Gayan ◽  
A. Chelleng
Keyword(s):  
Significant Variation ◽  
Microbial Biomass Carbon ◽  
Biological Properties ◽  
Most Probable Number ◽  
Symbiotic Effectiveness ◽  
Number Count ◽  
Probable Number ◽  
Nodule Number ◽  
Nodulation Status ◽  
Rhizobial Population

The influence of rhizospheric properties on nodulation status and resident rhizobial population of pea and lentil grown at twenty three sites of Assam were assessed. Besides the plant infectivity of rhizobia isolated from pea and lentil nodules were tested for symbiotic effectiveness. The survey established significant variation of nodule number ((35.-223.plant)), nodule dry weight (6.6 to 271. mgplant) and most probable number count ( to 147g soil) across the twenty three diverse sites. Rhizospheric properties <italic>viz</italic> pH, organic carbon, available P<sub>2</sub>O<sub>5</sub> and K<sub>2</sub>O could establish significant correlation with nodule number (r=0.892*, 0.771*, 0.753* and 0.669*) and most probable number(r=0.784*, 0.580*, 0.615* and 0.649*) counts, correspondingly. Microbial biomass carbon showed significant correlation with nodule number(r=0.672*). Dehydrogenase and phosphomonoesterase activities too showed significant correlation with nodule number(r=0.695* and 0.634*) and most probable number (r=.575* and 0.534*) count, respectively. The symbiotic effectiveness of isolated rhizobia demonstrated significant variation of nodule score (1.33-3.67). Only 6 isolates could attain nodule score of ≥3.0, irrespective of resident rhizobial population.

Nitrogen fixation characteristics of Rhizobium surviving in soils 'equilibrated' with sewage biosolids

2001 ◽  
Vol 52 (10) ◽  
pp. 963 ◽  
Author(s):  
Kellie J. Munn ◽  
Jeffrey Evans ◽  
Phillip M. Chalk
Keyword(s):  
N2 Fixation ◽  
Soil Type ◽  
Rhizobium Leguminosarum ◽  
Sewage Treatment Plant ◽  
Treatment Plant ◽  
Subterranean Clover ◽  
Isotope Dilution Method ◽  
Dilution Method ◽  
Symbiotic Effectiveness ◽  
Amended Soils

To determine the effects of urban sewage biosolids on the symbiotic effectiveness of Rhizobium leguminosarum bv. trifolii and N2 fixation, glasshouse and laboratory studies were carried out with several soils, biosolids, and biosolid application levels. Symbiotic effectiveness of R. l. trifolii was estimated as the dry weight or N content of seedlings of subterranean clover grown with only N2 fixation and seed N as the available nitrogen sources. The N fixed by legumes in unamended and biosolid-amended soils was determined using the 15N isotope dilution method. Six soils were represented in the experiments. Each of these was equilibrated over a period of 12 months with dried, finely ground biosolids (DWS) from the Malabar sewage treatment plant, at biosolids levels ranging from the equivalent of 60 to 240 t DWS/ha. One of the soils was also equilibrated with each of 4 other biosolids. The maximal concentration of heavy metals in soil amended with biosolids was 1026 mg/kg. The effect of biosolids on symbiotic effectiveness depended on the soil type and biosolid applications level. Thus, biosolids reduced the symbiotic effectiveness of R. l. trifolii in 2 of the 6 soils, although at different levels of biosolid. In most soil treatments N2 fixation was detected in subterranean clover, confirming the persistence of symbiotically effective rhizobia in most biosolids-amended soils. In addition, in strongly acidic soils plant N and N2 fixation increased significantly with biosolids addition, probably in response to higher soil pH, exchangeable Ca, and available P. In the treatments in which the symbiotic effectiveness of R. l. trifolii was reduced by biosolids, this was reflected in poor N2 fixation. However, symbiotic effectiveness did not correlate well with N2 fixation, probably because increases in soil nitrate at higher biosolids levels inhibited N2 fixation. Nevertheless, there were instances at 240 t DWS/ha where this was unlikely to explain the decrease in N2 fixation. It was concluded that adverse effects of biosolids on symbiotic effectiveness depend first on soil type, and then on biosolid type and application level; and the response in symbiotic effectiveness to adding biosolids to soil needs to be determined for each distinctively different site of biosolids reuse.

Agronomic and environmental drivers of population size and symbiotic performance of Rhizobium leguminosarum bv. viciae in Mediterranean-type environments

2012 ◽  
Vol 63 (5) ◽  
pp. 467 ◽  
Author(s):  
E. A. Drew ◽  
M. D. Denton ◽  
V. O. Sadras ◽  
R. A. Ballard
Keyword(s):  
Population Size ◽  
Dry Matter ◽  
Rhizobium Leguminosarum ◽  
Field Pea ◽  
Environmental Drivers ◽  
Annual Rainfall ◽  
Maximum Temperature ◽  
Most Probable Number ◽  
Probable Number ◽  
Commercial Strain

The population size and symbiotic performance (ability to fix N2) of rhizobia (Rhizobium leguminosarum bv. viciae) capable of nodulating field pea (Pisum sativum) were assessed in 114 soils from Mediterranean-type environments of southern Australia. All soils were collected in autumn, before the growing season, and had a history of crop legumes including field pea, faba bean, lentil, or vetch. The most probable number (MPN) technique, with vetch as a trap plant, was used to estimate the numbers of pea rhizobia in soils. Of the soils tested, 29% had low numbers of pea rhizobia (<100 rhizobia/g), 38% had moderate numbers (100–1000/g), and the remaining 33% had >1000/g. Soil pH, the frequency of a host crop in the rotation, and the number of summer days with a maximum temperature >35°C were strongly correlated with the pea rhizobia population size. Symbiotic performance (SP) of pea rhizobia in soils was assessed for soils with a MPN >100 rhizobia/g. An extract of the soils was used to inoculate two field pea cultivars growing in a nitrogen-deficient potting media in the greenhouse. Plants were grown for 5 weeks after inoculation and shoot dry matter was expressed as a percentage of the dry matter of plants grown with a commercial strain R. leguminosarum bv. viciae, SU303. Symbiotic performance ranged from 25 to 125%. One-quarter of the soils assessed had suboptimal SP (i.e. <70%). Soil and climatic variables were weakly associated with SP, with pH and average annual rainfall accounting for 17% of the variance. This research highlights the complexity of factors influencing population size and symbiotic performance of pea rhizobia in soils. Options for the improved management of populations of pea rhizobia in Mediterranean environments are discussed. Specifically, our data indicate that inoculation of pea crops is likely to be beneficial where pH(H2O) <6.6, particularly when summers have been hot and dry and when a host has been absent for ≥5 years, as numbers of rhizobia are likely to be below the thresholds needed to optimise nodulation and crop growth. New inoculation technologies and plant breeding will be required to overcome large populations of pea rhizobia with suboptimal SP.

Occurrence of alfalfa mosaic and subterranean clover red leaf viruses in legume pastures in Western Australia

1995 ◽  
Vol 46 (4) ◽  
pp. 763 ◽  
Author(s):  
SJ McKirdy ◽  
RAC Jones
Keyword(s):  
Western Australia ◽  
Alfalfa Mosaic Virus ◽  
Subterranean Clover ◽  
Trifolium Subterraneum ◽  
Seed Transmission ◽  
Enzyme Linked Immunosorbent Assay ◽  
The South ◽  
Potential Threat ◽  
South West ◽  
Burr Medic

When leaf samples were collected from 94 Trifolium subterraneum (subterranean clover) pastures from six districts in spring 1993 in the south-west of Western Australia and tested by enzyme-linked immunosorbent assay, no alfalfa mosaic virus (AMV) or subterranean clover red leaf virus (SCRLV) was detected. In contrast, when 21 irrigated T. repens (white clover) pastures from one district (Bunbury) were sampled and tested in January (summer) 1994, AMV was detected in 16, with eight having infection levels >86%, while SCRLV was found in seven at infection levels of <12%. When a further five T. repens pastures were tested for AMV in October (spring) 1994, the virus was found in all with incidences up to 100%. None of the T. repens pastures with high levels of AMV infection had been resown with T. repens within the last 20 years, whereas those resown within the last five years had little or no infection. AMV was detected in 9/91 annual medic (Medicago spp.) pastures from seven wheatbelt districts sampled in spring 1991 or 1993; a single pasture of M. polymorpha (burr medic) cv. Serena was 21% infected, but the other eight infected ones had <3%. AMV seed transmission was detected in 1/19 commercial seed stocks of M. polymorpha harvested in 1991-93. AMV infection was followed over a 12-year period in M. murex (murex medic) cv. Zodiac seed stocks. It persisted readily through successive seed harvests during this period. It is concluded that infection with AMV and SCRLV is currently not a threat to T. subterraneum pastures in the south-west of Western Australia and that AMV seems not to be one in wheatbelt annual medic pastures provided these are sown with healthy medic seed. In contrast, AMV poses a potential threat to the productivity of irrigated T. repens pastures. SCRLV is also sometimes present in T. repens pastures, but was not found at serious levels.

Populations of Rhizobium leguminosarum biovars phaseoli and viceae in fields after bean or pea in rotation with nonlegumes

1989 ◽  
Vol 35 (6) ◽  
pp. 661-667 ◽  
Author(s):  
R. M. N. Kucey ◽  
M. F. Hynes
Keyword(s):  
Rhizobium Leguminosarum ◽  
Most Probable Number ◽  
Inoculum Potential ◽  
Phaseolus Vulgaris L ◽  
Probable Number ◽  
Pisum Sativum L ◽  
Plasmid Profiles ◽  
Southern Alberta ◽  
Rhizobium Competition ◽  
Legume Inoculants

Populations of Rhizobium leguminosarum bv. phaesoli and bv. viceae in southern Alberta soils were measured over a period of 4 years using a most probable number method. Five fields cropped to bean (Phaseolus vulgaris L.), five fields cropped to pea (Pisum sativum L.), and two fields cropped to wheat were used as test sites. Legume crops had received appropriate legume inoculants. Fields were sampled in the fall of the crop year and in the spring of the following 3 years during which fields were cropped to nonlegumes or left fallow. Numbers of R. leguminosarum bv. phaseoli were 100 to 1000 times higher in fields that had been planted to bean than in fields that had been planted to pea or wheat. Fields that had been planted to pea maintained populations of R. leguminosarum bv. viceae 10 to 100 times higher than fields that had been planted to bean or wheat. Wheat fields, which had never had legumes grown in them, contained between 1 and 100 rhizobia per gram of soil of both biovars of R. leguminosarum, indicating that both biovars are native to southern Alberta soils. The numbers of rhizobia did not decrease in proportion to the population of other bacteria in the soil over the duration of the experiment. Plasmid profiles of soil Rhizobium isolates obtained in the last year of the experiment showed that none of the isolates had plasmid profiles similar to those of strains added as inoculants in the 1st year of the experiment. These results show that fields cropped to legumes and receiving rhizobial inoculants in this study maintained high populations of rhizobia for several years after harvest of the legume crop.Key words: Rhizobium leguminosarum bv. phaseoli, Rhizobium leguminosarum bv. viceae, nodule, plasmid profiles, inoculum potential, rhizobium competition.

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