Some factors affecting growth and survival ofRhizobium spp. in soil-peat cultures

Summary The growth and survival of rhizobium were studied in neutralized and sterilized soil-peat cultures containing alder bog peat, old moss peat, young reed peat, or young moss peat enriched with lucerne meal and sucrose. Although all these media proved to be excellent carriers for rhizobium, old moss peat from the 0–20 cm layer was less favourable than old moss peat from the 20–40 and 40–60 cm layer, while young moss peat proved to be the least satisfactory type of peat. A low storage temperature is always beneficial for the survival of rhizobia. Neutralization with CaCO₃ is to be preferred to that with CaCO₃+KH₂PO₄. Neutralization with NH₄OH exerted a detrimental effect. Much higher numbers of rhizobium were found in sterilized than in unsterilized soil-peat cultures. An antagonistic bacillus, isolated from peat, exerted a marked growth depression on rhizobium when both organisms were inoculated in sterilized soil-peat or in quartz sand media. Sterilization of the media permitted a rapid growth of the rhizobia and favoured their viability during storage, especially in autoclaved media containing nutrients. For the rhizobium ofLotonus bainesii sterilization of the peat proved essential for good growth. A harmful effect on the numbers of rhizobia was noted during the first week after the inoculation of the soil-peat mixtures when autoclaving had been carried out for 5 hours. This harmful effect proved, however, to be of a temporary nature.     

Removal of Hexane in Biofilters Packed with Perlite and a Peat–Perlite Mixture

Removal of hexane from air–hexane mixtures in biofilters packed with different solid media under nitrogen supplementation was performed for 70 days. Two columns containing Perlite or a mixture of peat and Perlite, were used. The solid media were supplemented with nitrogen source up to 1 kg/m³ per week for high nutrient supplementation and 0.2 kg/m³ per month for low nutrient supplementation. A high rate of hexane removal: 95 g/m³ h was achieved under high nutrient supplementation, high air flow rate and high hexane concentration. However, the percentage of hexane removal decreased with increasing air flow rate and hexane inlet concentration. For high nutrient supplementation the type of solid medium did not significantly affect the biodegradation capacity. With low nutrient supplementation, the highest removal rate was achieved in the column containing the peat–perlite mixture. The column containing perlite had a significantly lower pressure drop (20 Pa/m) than the 2400–2930 Pa/m observed for the column containing the mixture. Perlite offers an opportunity of running a biofiltration process at a lower and stable pressure drop if the nutrient supplementation is managed properly.     

Effects of some chemical factors on flagellation and swarming ofVibrio alginolyticus

Vibrio alginolyticus strains recently isolated from Dutch coastal seawater changed flagellar organization when cultivated in the presence of certain chemical agents. On agar media with more than 4.0% (w/v) NaCl the number of lateral flagella per cell decreased with increasing salt concentration. Both on agar media and in broth cultures with 6.0–9.0% (w/v) NaCl, cells with polar tufts of 2–4 sheathed or unsheathed flagella were frequently found. Cells grown on agar media with 7.3–9.8% (w/v) Na₂SO₄ had drastically reduced numbers of lateral flagella, but lacked polar tufts. EDTA suppressed growth, but did not affect flagellar arrangement. In the presence of 0.1–0.3% boric acid or 0.05–0.1% aluminium hydroxide, cells in liquid media tended to produce lateral, in addition to the polar flagella normally observed in broth cultures. Of a number of surface-active agents tested, Tween 80 and Na-taurocholate, even in high concentrations, did not affect flagellation. Bile salts (0.1%) and Na-deoxycholate (0.05%) strongly reduced the number of both polar and lateral flagella. In agar cultures, Na-lauryl sulphate (0.01–0.1%) inhibited the formation of lateral, but increased the incidence of polar flagella. Teepol (0.05–0.2%) had a similar effect and also it had a deteriorating effect on the sheaths of the polar flagella. Concomitant with the reduction in the number of lateral flagella, induced by these agents, swarming on agar media was inhibited.     

Survival of several <Emphasis Type="Italic">Rhizobium/Bradyrhizobium</Emphasis> strains on different inoculant formulations and inoculated seeds

The effect of a variety factors on the survival of several rhizobia strains on inoculants and inoculated seeds has been evaluated. Since the rhizobia strains showed different cell-density-evolution patterns on peat-based inoculants and on inoculated seeds, several inoculant formulations with highly effective Rhizobium/Bradyrhizobium strains (for Lupinus, Hedysarum, Phaseolus and Glycine max.) were monitored under the following storage conditions: (a) the inoculants were kept refrigerated (at 4 °C), or (b) at room temperature (25 °C). The effect of water content (30–50%, w/w) in the inoculants as well as that of several seed-coating adhesives were also investigated. Alternative carriers including perlite and vermiculite were tested. For all of the strains, survival on sterile peat-based inoculants was higher than on the corresponding unsterile peat formulation; for the latter, refrigerated storage conditions are recommended to ensure high bacterial densities. The water content of the inoculants had a differential effect on strain survival depending on the sterility of the peat, such that a high water content was more detrimental when unsterilized peat was employed. The best adherent for rhizobia survival was a gum arabic/water solution. Perlite was as effective as peat in maintaining a high population of rhizobia, at least for 6 months of storage. Electronic Publication     

A dry granular inoculant of Rhizobium for soil application

A dry granular inoculant of Rhizobium was prepared from sodium alginate and peralite. High numbers of two groundnut (Arachis hypogaea) Rhizobium strains, NC 92 and TAL 1000 used to prepare inoculants survived in dry granules beyond 180 days. The viable counts were 9.72 and 9.91 log₁₀ rhizobia g^-1 of dry granules for NC 92 and TAL 1000, respectively compared to 8.0 log₁₀ rhizobia g^-1 of peat inoculant for NC 92 at the end of six months storage. The granular inoculant was free from contaminants. In a pot culture experiment the granular inoculant applied to the soil gave similar results when seeds were dressed with a peat inoculant; nodulation and growth of groundnut were similar. The major advantage of this inoculant is that, it can be stored in a dry state without losing much viability.     

Morphological Changes of Rhizobia in Peat Cultures

Morphological changes that take place in peat cultures of several species of rhizobia were examined. These changes seemed to be associated with enhanced survival of cells in peat and after inoculation onto plastic beads, which were used as a model system for seeds. Cell wall changes, in which the periplasmic space appeared to be occluded with electron-dense material, were observed in Rhizobium sp. strain SU343 and Bradyrhizobium lupini WU425 cells after 7 and 14 days in peat, respectively. Nutrient limitation and low O₂ concentration in peat are suggested to be factors involved in the induction of the morphological changes. Polyhydroxybutyrate reserves, which were present in broth-cultured cells of both species of rhizobia, were mobilized after transfer into peat but did not appear to influence survival after inoculation onto beads. Enhanced expression of an iron-manganese superoxide dismutase was also observed after the cells were transferred into peat. We conclude that cell wall thickening in rhizobia after transfer from broth cultures into peat is an adaptive response for long-term survival under nutrient-limited conditions in peat. Cells with thickened walls may also be more resistant to other types of stress, such as that encountered on a seed surface.     

Short-term survival of rhizobia on arrowleaf clover seed sown at different depths

Seed of arrowleaf clover (Trifolium vesiculosum Savi) were inoculated with a streptomycin resistant mutant ofRhizobium leguminosarum biovartrifolii and planted on the surface of a Norwood fine sandy loam and at 10 and 25 mm depths. Populations of rhizobia declined from an excess of 10,000 seed⁻¹ immediately after inoculation to less than 100 within three to four days after sowing on the soil surface when water was the peat inoculant adhesive. Gum arabic as the adhesive promoted the survival of rhizobia. Populations of rhizobia on surface sown seed declined much more rapidly than on seed buried in soil. Although, the soil was nearly air dry, rhizobia on buried seed survived at populations exceeding 1,000 seed⁻¹. The maximum soil temperatures ranged between 21 and 36°C over the sampling time and did not seem to have a major influence on short term survival of rhizobia. Delayed germination of seed due to the higher temperature would indirectly influence the number of viable rhizobia present at germination.     

The effect of forestry drainage on vertical distributions of major plant nutrients in peat soils

Vertical distributions of total N, P, K, Ca and Mg in a 0–60 cm surface peat layer were studied at 80 pine mire sites in southern Finland. The sites fell into two categories according to the soil nutrient regime: Meso-oligotrophic and oligo-ombrotrophic, and formed a chronosequence from undrained sites to sites drained 55 years ago. A statistically significant drainage age effect on the gravimetric (mg g-1) concentration profile forms was detected for all nutrients except K. In oligo-ombrotrophic sites the concentration of N increased following drainage in the topmost layer (0–10 cm) and that of P in all layers. In meso-oligotrophic sites the changes in N and P profiles were obscure. The concentration profiles of K remained clearly surface-enriched in both site type groups, but there was a general drop in the concentration values immediately after drainage. Ca and Mg decreased, especially in the 10–20 and 25–35 cm layers in both site type groups. The volumetric (kg m-3) nutrient concentrations clearly reflected the increase in the bulk density of the surface peat occurring after drainage. The compaction of peat had compensated for the effect of the processes removing nutrients from the soil (increased tree stand uptake, leaching); for Ca and Mg to a lesser degree than for the other nutrients. It was concluded that the N, P and K profiles did not show changes that would be likely to affect site productivity, whereas the net loss of Ca and Mg may cause problems in the longer term. As the total K capital of the sites was in general rather small, a disturbance in the biological cycle, such as cutting of the tree stand, may be critical. This revised version was published online in August 2006 with corrections to the Cover Date.     

A simplein situ method for the characterization of porosity in growth media

Summary The method presented was based on a very simple approach of the forces operating in the pores of porous media. The standardized time (t_v) needed to infiltrate a given volume of a liquid into a porous medium in a defined state was throught to be an integrated measure of pore geometry and continuity. The state of the pore system was defined by external suction (S) and medium porosity characterized by the parameters k₁ and k₂ in the equation t_v=k₁+k₂ S^−0.5. The method theory was not rejected by experiments with glass beads and selected peat based growth media. The method ranked the media with respect to the probability for satisfactory gas exchange in the order of peat, peat and 26% perlite, peat and 34% perlite, and peat and 44% mineral wool. This ranking was achieved 95 days after the media were filled in containers and exposed to a daily watering procedure. Before this time, the ranking of the media was slightly different, if at all possible. Five days after the containers were filled, only peat and 44% mineral wood was significantly different from the other media. Judged by the standardized time method, the probability for satisfactory gas exchange decreased significantly during the 95 days experiment. This aggravation was supported by measurements of the volume fraction of pores filled with gas. The changes with time were least marked in the medium containing mineral wool.     

Phylogenetic composition of bacterial communities in small boreal lakes and ombrotrophic bogs of the upper Volga basin

Fluorescent in situ hybridization (FISH) with rRNA-specific oligonucleotide probes was used to assess the numbers and phylogenetic diversity of prokaryotic microorganisms in the water of small boreal lakes and peatland catchments of the swampy upper Volga basin. The abundance of bacterioplankton in lake water was found to vary from 1.6 to 8.7 × 10⁶ cells ml⁻¹, with the highest values detected in neutral eutrophic lakes. The total cell numbers in the peat of ombrotrophic bogs were 3.9–4.3 × 10⁸ cells g⁻¹ of wet peat. The proportion of bacteria identified by the group-specific probes decreased from 79–85% in neutral (pH 6.6–6.9) mesotrophic and eutrophic lakes to 65–69% in acidic (pH 4.4–5.5) dystrophic lakes and to 51–58% in the peat of acidic (pH 3.6–3.9) ombrotrophic bogs. The diversity of bacterial communities was highest in lakes with neutral water. These communities were dominated by members of the phylum Actinobacteria (31–44% of the total bacterial number), while the contribution of Alphaproteobacteria (16–19%), Bacteroidetes (6–16%), Betaproteobacteria (6–7%), Planctomycetes (2–8%), and Gammaproteobacteria (4–5%) was also significant. In acidic dystrophic lakes, Actinobacteria (25–35%) and Betaproteobacteria (25–34%) predominated, while peatland catchments were dominated by the Alphaproteobacteria (20–23%). The presence of acidobacteria and some planctomycetes common for bogs in the water of acidic dystrophic lakes, as well as the high proportion of bacteria (31–49%) that were not identified by the group-specific probes, suggest the impact of microbial processes in peatland catchments on the microbial composition of the receiving waters.     

Effect of compost,nitrogen salts,and NPK fertilizers on methane oxidation potential at different temperatures

The effects of compost, nitrogen salts, and nitrogen–phosphorous–potassium (NPK) fertilizers on the methane oxidation potential (MOP) of landfill cover soil at various temperatures were assessed. For this, we used batch assays conducted at 5°C, 15°C, and 25°C with microcosms containing landfill cover soil slurries amended with these elements. Results indicated variable impacts dependent on the type of amendment and the incubation temperature. For a given incubation temperature, MOP varied from one compost to another and with the amount of compost added, except for the shrimp/peat compost. With this latter compost, independent of the amount, MOP values remained similar and were significantly higher than those obtained with other composts. Amendment with most of the tested nitrogen salts led to similar improvements in methanotrophic activity, except for urea. MOP with NPK fertilizer addition was amongst the highest in this study; the minimum value obtained with NPK (20–0–20) suggested the importance of P for methanotrophs. MOP generally increased with temperature, and nutrient limitation became less important at higher temperatures. Overall, at each of the three temperatures tested, MOP with NPK fertilizer amendments provided the best results and was comparable to those observed with the addition of the shrimp/peat compost. The results of this study provide the first evidence of the following: (1) compost addition to improve methanotrophic activity in a landfill cover soil should consider the amount and type of compost used and (2) the importance of using NPK fertilizers rather than nitrogen salts, in enhancing this activity, primarily at low temperatures. One can also consider the potential beneficial impact of adding these elements to enhance plant growth, which is an advantage for MOP.     

Influence of Rhizobial inoculation on yield of wheat (Triticum aestivum L.)

Summary Soil + charcoal (1∶3) carrier based and liquid cultures of Rhizobia were used to inoculate wheat seed cv. HD2329. The plants received 100 kg N in two equal splits and 60 kg P₂O₅ and 40 kg K₂₀ ha⁻¹. Inoculation with rhizobia had little effect on grain yield of wheat. Significant increase in straw yield and N-uptake occurred due to inoculation. A comparison of results of a similar experiment conducted during 1983–84, showed that inoculation with the same strains of rhizobia and application 50 kg N ha⁻¹ as basal dressing, was more effective in increasing yield and N-uptake in wheat cv. HD2329. It appears reasonable to assume occurrence of nitrogen fixation by root nodule bacteria in rhizosphere of wheat.     

Nodulation of lucerne (Medicago sativa L.) in an acid soil: Effects of inoculum size and lime-pelleting

The effects of inoculum level and lime-pelleting were studied in an acid soil with respect to the nodulation and growth of lucerne (Medicago sativa cv Resis) and the population dynamics of Rhizobium meliloti. In small root-boxes (rhizotrons), the in-situ survival of inoculated rhizobia was studied in the micro-environment around the seed for a period of 12 days after sowing. During the initial 24 hours, a strong increase in rhizobial numbers was measured, concomitantly with the development of roots. As a result of lime-pelleting, rhizobial numbers were higher only at 3 days after sowing (P<0.05). Later, this difference diminished steadily. Addition of lime did not increase the adhesion of the rhizobia to the seedling tap root. Plant responses to inoculation were studied in pots. To obtain optimal nodulation, the soil had to be neutralized around the seed with lime and at least 10⁵ cells of R. meliloti were required. With more than 10⁵ rhizobia per seed, lime-pelleting increased the number of crown-nodulated seedlings from 24% to 77%. Higher numbers of rhizobia could not compensate the effect of lime. A strong correlation was found between crown nodulation, nitrogen content and dry weight of the shoots.     

Sodium Chloride as a Cause of Low Numbers of Rhizobium in Legume Inoculants

S ummary . A rapid reduction, after manufacture, in numbers of Rhizobium trifolii in commercial legume inoculants was observed in 1971 and explained by the presence of high concentrations of sodium chloride in the peat used as carrier material. A strain of R. trifolii growing in broth proved to be more sensitive to sodium ions than did 2 strains of R. meliloti but was more tolerant of up to 0.3% of chloride. The effect of salt differed somewhat according to the source of peat used as carrier, but peat containing °0.2% of chloride (expressed as a % of dry peat) may cause considerable loss of viability of rhizobia.     

The heat increment of feeding in house wren chicks: magnitude, duration, and substitution for thermostatic costs

The heat increment of feeding (HIF), a transient postprandial increase in metabolic rate, is the energy cost of processing a meal. We measured HIF in house wren chicks (Troglodytes aedon) ranging in mass from 1.6 to 10.3 g. This mass range (age 2–10 days) spanned a transition from blind, naked, ectothermic chicks through alert, endothermic birds with nearly complete feathering. We fed chicks crickets (2.7–10% of chick body mass) and determined HIF from continuous measurements of oxygen consumption rate (O₂) before and after meals. At warm ambient temperatures (T _a) of 33–36 °C, the magnitude of HIF (in ml O₂ or joules) was linearly related to meal mass and was not affected by chick mass. HIF accounted for 6.3% of ingested energy, which is within the range of results for other carnivorous vertebrates. The duration of HIF was inversely related to chick mass; 10-g chicks processed a standard meal approximately twice as fast as 2-g chicks. HIF duration increased with increasing meal mass. The peak O₂ during HIF, expressed as the factorial increase above resting metabolism, was independent of body mass and meal mass. In large, endothermic chicks ( > 8 g), HIF substituted for thermoregulatory heat production at low T _a. Accepted: 11 December 1996     

Improvement of English walnut somatic embryo germination and conversion by desiccation treatments and plantlet development by lower medium salts

Summary Well-developed somatic embryos were selected from a repetivively somatic embryo line derived from embryonic axes of immature zygotic embryos of English walnut ‘No. 120’ (Juglans regia L.) for germination and conversion studies. In germinating dishes, somatic embryos germinated into only shoots, only roots, or both shoots and roots. Without any pretreatment, 28% somatic embryos germinated, while those treated with 2.5–5.0 mg 1⁻¹ (7.2–14.4 μmol) gibberellic acid (GA₃) germinated at 25–28% and those receiving a cold treatment of 2–3 mo. at 3–4°C germinated at 30–43%. However, only 4–19% of the germinating embryos showed both shoots and roots. Treated with desiccation, either with CaCl₂·6H₂O or Ca(NO₃)₂·4H₂O at 20°C in the dark for 3 d, somatic embryos germinated at 85–91%, 57–69% of which had both shoots and roots. Treatment with 2 mo. cold storage in combination with desiccation using Ca(NO₃)₂·4H₂O resulted in 92% of somatic embryos germinating, 70% of which showed both shoots and roots. No significant differences were observed between solid and liquid germination media. After transferring the germinating embryos to plantlet development media, 52–63% of those with both shoots and roots developed into plantlets while 11% with only shoots or 9% with only roots converted into plantlets. Plantlet development was improved by using lower medium salts and sucrose concentrations. The addition of activated charcoal enhanced root development, particularly root branching. Of 131 plants transplanted, 91 plants were acclimatized to a greenhouse.     

Spent mushroom substrates as component of growing media for germination and growth of horticultural plants

This research work was conducted in order to investigate the possibility of using spent mushroom substrate (SMS) in the production of horticultural seedlings replacing part of the peat in the growing media. Three vegetable species with different salt sensitivities, the less sensitive being tomato (Lycopersicon esculentum var. Muchamiel), the moderately salt-sensitive being courgette (Cucurbita pepo L. var. Afrodite F1) and the most salt-sensitive being pepper (Capsicum annum L. var. Lamuyo F1) were grown in 12 media containing SMS of two types of mushroom (Agaricus bisporus (SMS-AB) and Pleurotus ostreatus (SMS-PO)) or a mixture of both 50% (v/v) (SMS-50), as well as peat in various ratios. The proportions of each residue in the mixtures elaborated with peat were 25%, 50%, 75% and 100% v/v residue. A substrate of 100% peat was used as control. The experiment was arranged in a completely-randomised design with two replicates per treatment under greenhouse conditions. Prior to sowing, some physical, physico-chemical and chemical properties of the growing media were determined and seed germination and fresh weight of seedling were also measured. In most of the cases, the addition of SMS to the growing media produced an increase in the pH values, salt contents, macro and micronutrient concentrations and a decrease in the water holding capacity contents in comparison to peat, whereas great differences were found in the air capacity values between SMS-based substrates and peat. Up to 75% SMS can be used in mixtures with peat for seed germination of the plant species studied. Regarding the most suitable SMS-based substrates for plant growth, any substrate could be used for tomato seedling production. However, all SMS-AB-based substrates and the media containing low dose of SMS-PO and SMS-50 were adequate for growth of courgette and pepper.     

A study of the impacts of Zn and Cu on two rhizobial species in soils of a long-term field experiment

Two agriculturally important species of rhizobia, Rhizobium leguminosarum biovar viciae (pea rhizobia) and R. leguminosarum bv. trifolii (white clover rhizobia), were enumerated in soils of a long-term field experiment to which sewage sludges contaminated predominantly with Zn or Cu, or Zn plus Cu, were added in the past. In addition to total soil Zn and Cu concentrations, soil pore water soluble Zn and free Zn²⁺, and soluble Cu concentrations are reported. Pea and white clover rhizobia were greatly reduced in soils containing ≥200 mg Zn kg^-1, and soil pore water soluble Zn and free Zn²⁺ concentrations ≥7 and ≥3 mg l^-1, respectively, in soils of pH 5.9–6. Copper also reduced rhizobial numbers, but only at high total soil concentrations (>250 mg kg^-1) and not to the same extent as Zn. Yields of field grown peas decreased significantly as total soil Zn, soil pore water soluble Zn and free Zn⁺² increased (R² = 0.79, 0.75 and 0.75, respectively; P < 0.001). A 50% reduction in seed yield occurred at a total soil Zn concentration of about 290 mg kg^-1, in soils of pH 5.9–6. The corresponding soil pore water soluble Zn and free Zn²⁺ concentrations were about 9 and 4 mg l^-1, respectively. Pea seed yields were not significantly correlated with total soil Cu (R² = 0.33) or soil pore water soluble Cu (R² = 0.39). Yield reductions were due to a combination of greatly reduced numbers of free-living rhizobia in the soil due to Zn toxicity, thus indirectly affecting N₂-fixation, and Zn phytotoxicity. These effects were exacerbated in slightly acidic soils due to increased solubility of Zn, and to some extent Cu, and an increase in the free Zn²⁺ fraction in soil pore water. The current United Kingdom, German and United States limits for Zn and Cu in soils are discussed in view of the current study. None of these limits are based on toxicity thresholds in soil pore water, which may have wider validity for different soil types and at different pH values than total soil concentrations. This revised version was published online in June 2006 with corrections to the Cover Date.     

Submerged culture of a mycelial formulation of a bioherbicidal strain of <Emphasis Type="Italic">Myrothecium </Emphasis><Emphasis Type="Italic">verrucaria</Emphasis> with mitigated mycotoxin production

A mycelial formulation of the fungus Myrothecium verrucaria (IMI 361690) containing 0.20% Silwet L-77 surfactant was found to be highly efficacious in controlling the exotic invasive weed kudzu. The mycelium can be rapidly (48–72 h) produced in several media, including an inexpensive soy flour–corn meal medium. Mycelial yields were 2, 10, and 25 g dry weight l⁻¹ in Czapek-Dox, Richard’s V-8, and soy flour–corn meal media, respectively. Scale-up production in soy flour–corn meal medium using laboratory fermenters (10–25 l), resulted in a mycelial formulation that caused 90% mortality of naturally-occurring mature (0.9–1.0 m in height) kudzu within 48 h after application in field experiments. HPLC analyses revealed that the mycelium produced in this liquid culture contained no detectable amounts of the trichothecene mycotoxins roridin A and verrucarin A (limit of detection 2 μg ml⁻¹). This has resulted in a safer, yet effective bioherbicidal product. We anticipate that these findings should improve the probability of EPA registration and subsequent commercial development of this bioherbicide.     

Dilution of Liquid Rhizobium Cultures To Increase Production Capacity of Inoculant Plants

Experiments were undertaken to test whether peat-based legume seed inoculants, which are prepared with liquid cultures that have been deliberately diluted, can attain and sustain acceptable numbers of viable rhizobia. Liquid cultures of Rhizobium japonicum and Rhizobium phaseoli were diluted to give 10⁸, 10⁷, or 10⁶ cells per ml, using either deionized water, quarter-strength yeast-mannitol broth, yeast-sucrose broth, or yeast-water. The variously diluted cultures were incorporated into gamma-irradiated peat, and the numbers of viable rhizobia were determined at intervals. In all of the inoculant formulations, the numbers of rhizobia reached similarly high ceiling values by 1 week after incorporation, irrespective not only of the number of cells added initially but also of the nature of the diluent. During week 1 of growth, similar multiplication patterns of the diluted liquid cultures were observed in two different peats. Numbers of rhizobia surviving in the various inoculant formulations were not markedly different after 6 months of storage at 28°C. The method of inoculant preparation did not affect the nitrogen fixation effectiveness of the Rhizobium strains.