The Faculty of Agronomy, University of Montenegro, Cacak, Serbia
This paper presents a study on the effect of inoculation of maize seed (NSSC-640) by asymbiotic nitrogen-fixing bacteria (Azotobacter chroococcum strain 84) and increasing rates of mineral nitrogen on quantitative composition of microorganisms (total number of bacteria, number of actionomycetes and azotobacter) in smonitza under maize and on yield.
The following fertilization treatments were studied: NiPK (90:75:60 kg ha'1); N2PK (120:75:60 kg ha"1); N3PK (150:75:60 kg ha"1), as well as the treatment with presowing inoculation of maize seed.
Microbiological analysis included the assessment of total number of microorganisms, actinomycetes and azotobacters in the rhizosphere and edaphosphere during maize vegetation. Total numbers of microorganisms were determined by growth on the medium for the total number with an appropriate amount (0.5 mL) of 10"6 soil dilution, numbers of actinomycetes - by growing on synthetic agar according to Krasil'nikov with 10"4 of soil dilution and azotobacters - by growing on Fyodorov's agar with 10"2 of soil dilution.
The study results showed that the numbers of microorganism groups were affected by the type and rate of fertilizers applied, as well as by the time and sampling zone. The application of Azotobacter chroococcum, strain 84, resulted in an increase of the total numbers of microorganisms, actinomycetes and azotobacters, especially in the rhizosphere soil at the onset, and even in the middle of maize vegetation. Lower rates of nitrogen fertilizers (90 and 120 kg ha"1) led to a significant increase of total bacteria numbers, as well as an insignificant change in azotobacter numbers, whereas their high rate (150 kg ha"1) had a depressive effect on the mentioned microorganisms, particularly in the edaphosphere of maize. In contrast, actinomycete numbers were not reduced even with this nitrogen treatment.
Under the studied agroecological conditions, the highest maize yield, but not economically justified one, was obtained with the highest nitrogen rate. Seed inoculation with Azotobacter chroococcum, strain 84, caused an insignificant rise in maize yield, which can be associated with acid reaction of the soil studied and slower release of nitrogen accumulated in their cells.
EFFECT OF INCREASING PHOSPHORUS CONCENTRATIONS ON THE GROWTH AND PHYSIOLOGY OF RHIZOBIUM-INOCULATED AND NITROGEN-SUPPLIED COWPEA (VIGNA UNGUICULATA L. WALP) PLANTS
M.L. Izaguirre-Mayoral, 0. Carballo
Instituto Venezolano de Investigaciones Científicas, Centro de Microbiología y Biología Celular, Laboratorio de Biotecnología y Virología Vegetal, Apdo 21827, Caracas 1020-A, Venezuela
Cowpea is an important grain crop in Venezuela. However, local varieties do not respond to Rhizobium inoculation. Nodulated plants always produce a smaller biomass and lower yields. Thus, farmers do not rely upon inoculation for better yields, even though the price of fertilizers has increased over the years. Since cowpea crops are planted mainly in savanna soils, characterized by low phosphorus (P) content, it could be hypothesized that P is the limiting factor for the physiological performance of cowpea plants. The aim of this investigation was, therefore, to analyze the effect of increasing P concentrations on the growth of Rhizobium-inoculated and nitrogen-supplied cowpea plants. For this purpose, plants inoculated with Bradyrhizobium 1-125 (R+) or provided with 15 mM nitrogen (N+) were grown in nutrient solutions supplied with the following P concentrations (Pc): 0.05 mM (PI), 0.1 mM (P2), 0.25 mM (P3), 0.5 mM (P4), 1 mM (P5), 2 mM (P6), 3 mM (P7) and 4 mM (P8). At the flowering stage, plants were divided into roots, shoots, aerial and nodule biomass. Concentration of chlorophyll, N, P, total reducing sugars (TRS) and N-compounds were determined in the individual plant components following standard protocols. The experiment was conducted under controlled conditions and the pH of the solution was maintained at 6.2. Increasing Pc up to P6 enhanced the aerial mass of R+ and N+ plants but reduced their root mass regardless of the mode of N uptake. Concomitantly, increasing P up to P4 enhanced the nodule mass in R+ plants. The ureides in R+ plants as well as the nitrate concentration in N+ plants remained constant between PI and P6. Nevertheless, the aerial mass in R+ plants was always smaller than that of N+ plants in spite of the higher TRS, a-amino-N, chlorophyll and P concentrations in the leaf tissues. Metabolic constraints for ureide degradation in leaves seem to be the mechanism underlying the poor response of plants to rhizobial inoculation at high Pc. The P7 and P8 concentrations proved to be toxic for R+ and N+ plants. From present results we may conclude that P is not the key factor for the improvement of the symbiotic process and growth of cowpea plants under savanna conditions.
POTENTIAL OF NITROGEN-FIXING SYMBIOSIS SYSTEMS FOR RE YE GETATION STRATEGIES IN MEDITERRANEAN ENVIRONMENTAL CONDITIONS
J. Cleyet-Marel, O. Domergue, L. Maure
Laboratoire des Symbioses Tropicales et Méditerranéennes IN R A/IRD/CIR A D/A GR O - M,
Anthropogenic degradation activities (overgrazing, non-regulated cultivation techniques, deforestation, quarry exploitation, etc.), together with a long dry and hot summer, with scarce, erratic, but torrential rainfalls, is a major threat to the sustainability of Mediterranean ecosystems. The removal of native trees and soil disturbance result in a lack of sufficient resident soil microflora and in many instances a number of nutrients, including N, P and K are not present in sufficient quantities to promote an acceptable plant growth.
Revegetation of degraded ecosystems requires selection of suitable plant species and adaptation of the plants to the unusual soil conditions. Therefore, nitrogen-fixing legumes are key components of the natural succession because their associated rhizobial symbioses constitute a source of N input to the ecosystem. Several Mediterranean native legumes have been selected from several areas in the South of France, some of them like Colutea, Coronilla, Genista, Medicago and Spartium belong to the shrub community and other like Astragalus, Dorycnium, Hippocrepis, Lotus, Medicago, Onobrychis and Ononis are classical herbaceous legumes.
Seeds of these plant species were collected and the rhizobial partners were isolated, characterized and the most efficient strains were selected for inoculum preparation. Legume plant seeds were germinated before seedlings were transferred in plastic containers used in forestry practice. All seedlings were inoculated with a previously selected rhizobial culture and allowed to grow for 6-9 months in an experimental nursery.
Plants were transferred in the chosen degraded ecosystem and experimental variables including survival rates, plant growth, seed production were tested. The most interesting legume species for reclamation and rehabilitation programs were identified to restore sustainable ecosystems.
DIVERSITY OF POPULATIONS OF RHIZOBIA THAT NODULATE PHASEOLUS VULGARIS IN FRANCE
N. Amarger, M. Bours, D. Depoil, K. Groud, F. Revoy
Laboratoire de Microbiologie des Sols, INRA, BP86510, 21065, Dijon, France
Phaseolus vulgaris is native to the Americas and was imported to Western Europe in the XVI century. The crop, being considered weak in nitrogen fixation and with a highly variable response to inoculation, usually receives N-fertilizer. Phaseolus rhizobia belong to several species and several biovars which possess different symbiotic properties. The strains that belong to the biovar (bv.) phaseoli of R. leguminosarum, R. etli, R. gallicum, R. giardinii, have a narrow host-range, they nodulate only Phaseolus spp. Strains of R. gallicum bv. gallicum, R. giardinii bv. giardinii. R. tropici have wider host ranges, not fully described yet, but that include at least Leucaena spp. in addition to Phaseolus spp. R. giardinii isolates, whether they belong to bv. phaseoli or bv. giardinii, induce ineffective nodules. The objectives of the study were to investigate the diversity of rhizobia at the origin of nodules in field grown Phaseolus, and to see whether this diversity fluctuates with geographical situation and plant genotype and could have agricultural implications.
Populations of 40 to 105 rhizobia were isolated from nodules of field grown common beans collected from seven locations in bean production areas in France. There were one or two years of sampling and one or several bean varieties per location. Nodule isolates were characterized by RFLP analysis of PCR amplified DNA fragments: 16S rDNA (Laguerre et al. 1994), 16S-23S ITS (Laguerre et al. 1996) and nif or nod gene (Laguerre et al. 2001) to determine the species, the intraspecies polymorphism, and the biovar, respectively.
The populations were found to be diverse at the species and/or at the intraspecies (biovar and 16S-23S ITS type) levels. Although some populations were composed of a single species, R. leguminosarum bv. phaseoli, two different species, R. leguminosarum bv. phaseoli and R. giardinii, R. etli or R. tropici were detected in the other populations but one. This latter population was dominated by R. giardinii bv. giardinii but a few isolates of R. etli, R. gallicum bv. gallicum, R. gallicum bv. phaseoli and R. giardinii bv. phaseoli were also identified. The composition varied among locations. Within locations, it could vary between years and between bean varieties. The proportion of R. giardinii, which has a Fix" phenotype and was present in half of the locations, could be rather high, which should have a negative effect on the amount of nitrogen fixed by the crop. The variability observed in the composition of the populations between years and plant genoypes shows that only a part of the rhizobial diversity present in a soil is revealed by sampling a given crop at a given time. This variability could be at the origin, of at least a part, of the variability observed in the crop response to inoculation.
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