Indigenous Soil Microflora Research Articles

Biological control of Striga hermonthica by Cubitermes termite mound powder amendment in sorghum culture

Striga hermonthica (Del.) Benth is an obligate root hemi-parasite of several cereals. Its effect on cereal crops is the main constraint for food production in sub-Saharan Africa. Various control methods have been already proposed, but the infestation by these parasitic plants persists. An appropriated method for Striga management adapted for the African farmer is very much needed. In this study, amendment of soil infested by this phytoparasite with Cubitermes mound powder is proposed as chemical amendment and natural microbial inoculum, to promote plant growth and reduce damage by S. hermonthica on sorghum ( Sorghum bicolor L.). The influence of Cubitermes mound powder on the development of several microbial groups (arbuscular mycorrhizal fungi, actinomycetes, saprophytic fungi) was investigated in a pot experiment with sorghum cultured in a sandy soil infested by S. hermonthica. In the amended soil, sorghum growth and mycorrhizal colonization of sorghum plants were significantly greater than in the control treatment. Mycorrhizal colonization was negatively correlated with the number of emerged Striga plants per pot and positively correlated with sorghum growth. The relationship with substrate-induced respiration (SIR) responses showed that amended soil was characterized by its response to hydroxybutyric acid (catabolic marker of mycorrhizal colonization) and non-amended soil by its response to phenylalanine. We noted that the number of emerged Striga plants in amended pots was significantly decreased. Since Cubitermes mound suspensions did not affect Striga seed germination under axenic conditions, it suggests that the amendment with Cubitermes powder reduces S. hermonthica infestation indirectly, i.e. via its effect on the indigenous soil microflora. Overall, it appears that management of Cubitermes mounds is a promising strategy to consider for effective protection of sorghum from Striga infestation.

Bioremediation <i>In Situ</i> of an Alkaline Soil Polluted with Heavy Metals

An experimental plot consisting of alkaline soil heavily polluted with heavy metals (copper, zinc, cadmium) was treated in situ by stimulating the activity of the indigenous soil microflora, which contained different metal-solubilizing microorganisms. This was achieved by adding to the soil solid biodegradable organic substrates (cow manure, plant compost, straw), zeolite saturated with ammonium phosphate and aqueous solution containing acetate, lactate and magnesium chloride. An efficient removal of the above-mentioned heavy metals was achieved via the drainage water and their residual contents in the soil decreased to below the relevant permissible levels within 18 months of treatment. The metals were solubilized mainly as complexes with the organic acids added by the irrigating water solutions or secreted by the indigenous heterotrophic microorganisms. The effluents containing dissolved metals were efficiently treated by means of a wetland constructed near to the experimental plot.

Rhamnolipid Produced from Agroindustrial Wastes Enhances Hydrocarbon Biodegradation in Contaminated Soil

A crude biosurfactant solution was produced by Pseudomonas aeruginosa growing on agroindustrial wastes as the substrate and used to study its effect on hydrocarbon biodegradation by the indigenous soil microflora under laboratory conditions. Two concentrations were studied at first and 1 mg of biosurfactant/g of soil showed to be the most efficient for the total petroleum hydrocarbon reduction, which reached 85% at the first 20 days in soil microcosms. Respirometric and microbial analyses showed that the biosurfactant added did not have toxic effects over the microbial population. The use of a biosurfactant for bioremediation has been limited because of its high cost production. Biosurfactants produced from cost-free by-products combines waste minimization with economic potential bioremediation process.

Development of microbial degradation of cypermethrin and diazinon in organically and conventionally managed soils

The behaviour of pesticides in the soil is a complex issue and is controlled by the physical, chemical and biological properties of the soil. The ability of microorganisms to degrade pesticides is not only controlled by the bioavailability of a chemical but also by their capacity to develop the ability to utilise available chemicals. The development of catabolism in the indigenous soil microflora of four organically and one conventionally managed soils was investigated for two pesticides: cypermethrin and diazinon. Soils were amended with cypermethrin and diazinon and aged for 0, 2, 4, 6, 8, and 14 weeks and, at each time point, mineralisation of freshly added 14C-cypermethrin or 14C-diazinon was measured by trapping 14CO2. In general, contact times between the soil and the pesticide resulted in a reduction in the lag phase (the period of time before mineralisation exceeded 5% of the added activity), followed by increases in the extent of mineralisation. Cypermethrin was mineralised significantly in all soils; whereas, diazinon was only appreciably mineralised in two of the soils, most notably in the organic soil from Redesdale. Statistical analysis showed pH and organic matter content of the soil had a significant effect on the extent of mineralisation (P< or = 0.05) of the cypermethrin in the soils.

Enhancement of pyrene mineralization in soil by wood-decaying fungi

The mineralization of [14C]pyrene in sterilized and non-sterile soil was investigated using the wood-decaying fungi Kuehneromyces mutabilis and Agrocybe aegerita in a period of 63 days. In sterilized soil 5.1% and 1.5% of the pyrene was mineralized to 14CO2 by K. mutabilis and by A. aegerita, respectively. In non-sterile soil, 27.3% of pyrene was mineralized by indigenous soil microflora including a Mycobacterium gilvum strain. During soil inoculation with fungi the mineralization was higher (47.7% for K. mutabilis and 38.5% for A. aegerita). For a mass balance analysis the soil was extracted with toluene and methanolic KOH (humic acid extraction). Considering the sum of mineralization and formation of bound residues (non-extractable radioactivity), about 50% (sterilized soil) and 75% (non-sterile soil) of pyrene were eliminated by K. mutabilis. In comparison with indigenous soil microflora, K. mutabilis enhanced pyrene elimination up to 42%.

Linking Catabolism to Cyclodextrin Extractability: Determination of the Microbial Availability of PAHs in Soil

When assessing the potential of a contaminated site for bioremediation, it is desirable to know how much of the contaminant(s) is available for microbial degradation, thus allowing the likelihood of successful bioremediation to be predicted. The aims of this study were to investigate the degradation of PAHs in two soils by a catabolic inoculum and indigenous soil microflora and link this to the cyclodextrin extractable fraction in the presence of transformer oil (0.05, 0.01, or 0.005%). This study showed very little difference between indigenous and inoculum-derived degradation for phenanthrene in laboratory-aged soil, and strong relationships were also observed between both of the microbial degradative conditions and the amount of phenanthrene extracted by cyclodextrin. Furthermore, the indigenous degradation of PAHs in a field-contaminated soil showed significant linear correlations with the cyclodextrin extractable fraction, with gradients approximating to 1. There are several novel facets to this study. First, in aged, contaminated soils, indigenous microflora gave an equally sensitive determination of degradative availability as that measured by the catabolic inoculum. Second, this is the first time intrinsic biodegradation of PAHs has been predicted by the cyclodextrin extraction in laboratory-spiked and field-contaminated soils. The cyclodextrin extraction technique represents a powerful tool for predicting the extent of intrinsic and augmented microbial degradation and will be useful in the assessment of contaminated land prior to bioremediation.

Pilot Scale Bioremediation of Creosote-Contaminated Soil—Efficacy of Enhanced Natural Attenuation and Bioaugmentation Strategies

ABSTRACT In this study, the efficacy of bioremediation strategies (enhanced natural attenuation with nitrate and phosphate addition [ENA] and bioaugmentation) for the remediation of creosote-contaminated soil (7767 ± 1286 mg kg−1 of the 16 EPA priority PAHs) was investigated at pilot scale. Bioaugmentation of creosote-contaminated soil with freshly grown or freeze dried Mycobacterium sp. strain 1B (a PAH degrading microorganism) was applied following bench scale studies that indicated that the indigenous soil microflora had a limited PAH metabolic activity. After 182 days, the total PAH concentration in creosote-contaminated soil was reduced from 7767 ± 1286 mg kg−1 to 5579 ± 321 mg kg−1, 2250 ± 71 mg kg−1, 2050 ± 354 mg kg−1 and 1950 ± 70 mg kg−1 in natural attenuation (no additions) and ENA biopiles and biopiles augmented with freshly grown or freeze dried Mycobacterium sp. strain 1B respectively. In ENA and bioaugmentation biopiles, between 82% and 99% of three-ring compounds (acenaphthene, anthracene, fluorene, phenanthrene) were removed while four-ring PAH removal ranged from 33 to 81%. However, the extent of PAH degradation did not vary significantly between the ENA treatment and biopiles augmented with Mycobacterium sp. strain 1B. Four-ring PAH removal followed the order fluoranthene > pyrene > benz[a]anthracene > chrysene. The high residual concentration of some four-ring PAHs may be attributable to bioavailability issues rather than a lack of microbial catabolic activity. Comparable results between ENA and bioaugmentation at pilot scale were surprising given the limited degradative capacity of the microbial consortia enriched from the creosote-contaminated soil.

Prediction of polycyclic aromatic hydrocarbon biodegradation in contaminated soils using an aqueous hydroxypropyl‐β‐cyclodextrin extraction technique

This study investigated the use of an aqueous hydroxypropyl-beta-cyclodextrin (HPCD) shake extraction to predict the degree of microbial degradation of polycyclic aromatic hydrocarbons (PAHs) in soils. Three different aged PAH-contaminated soils were studied: A soil from a former coke works (CW) and two artificially contaminated soils (AC1 and AC2). First, the catabolic activity of the indigenous soil microflora was assessed with 14C-respirometry, using a range of 14C-labeled aromatic compounds. Extensive mineralization of several compounds occurred in the CW and the AC2 soils, suggesting that both soils contained catabolically active microorganisms. No significant mineralization occurred in the AC1 soil, implying that either it did not contain an indigenous PAH-degrading microbial population or that degradation, but not mineralization, occurred. The soils then were subjected to three sets of analyses: dichloromethane (DCM) soxhlet extraction, six-week biodegradation assay followed by DCM extraction, and extraction with HPCD followed by DCM extraction. A general decrease in PAHs present in the soils occurred after the biodegradation assay. In the CW and the AC1 soils, strong correlations were observed between the amount of PAHs biodegraded and the fraction of PAHs removed from the soils using the HPCD extraction. However, the AC2 soil showed a more modest correlation between the biodegradable fraction and the HPCD extractable fraction, with the HPCD extraction slightly underestimating the extent of PAH biodegradation. The results of this study indicated that an aqueous HPCD extraction may be a useful tool in assessing the microbial availability of aged contaminant mixtures in soils, although further validation is required.

Leaf litter quality and decomposition rates of yellow birch and sugar maple seedlings grown in mono-culture and mixed-culture pots at three soil fertility levels

Seedlings of yellow birch (Betula alleghaniensis Britton) and sugar maple (Acer saccharum Marsh.) were grown for 2 years in mono-culture and mixed-culture and at three fertility levels. Following the second growing season, senescent leaves were analysed for N concentration, acid hydrolysable substances (AHS), and nonhydrolysable remains (NHR). A litter sub-sample was then inoculated with indigenous soil microflora, incubated 14 weeks, and mass loss was measured. Litter-N was significantly higher at medium than at poor fertility, as well as in yellow birch than in sugar maple litter. The species effect on litter-N increased with increasing fertility. At medium fertility, litter-N of sugar maple litter was lower in mixed-culture than in mono-culture. AHS, NHR as well the NHR/N ratio were significantly higher in yellow birch than in sugar maple litter. At medium fertility, the NHR/N ratio of sugar maple litter was significantly lower in mono-culture than in mixed-culture. Mass loss was significantly greater at medium and rich fertility than at poor fertility, and in yellow birch than in sugar maple litter. At poor fertility, mixed-litter decomposed at a rate comparable to yellow birch, whereas at medium and rich fertility, mixed-litter decomposed at a rate comparable to sugar maple. There was a significant positive relationship between litter-N and mass loss. A similar positive relationship between NHR and mass loss was presumed to be a “species” effect on decomposition. Results support the hypothesis that species × fertility and species × mixture interactions can be important determinants of litter quality and, by implication, of site nutrient cycling.

A novel non-chemical nematicide for the control of root-knot nematodes

The efficacy of a novel bio-nematicide based on a strain of Bacillus firmus was investigated in a series of field and laboratory studies. The efficacy of this bio-nematicide was compared with standard fumigant nematicides and the bio-control agent Pasteuria penetrans. The impact of the application of this bio-nematicide on the size and activity of the soil microflora was also tested by measuring ninhydrin reactive N and FDA hydrolytic activity, respectively. The bio-nematicide was applied as powder at three dose levels 50, 70 and 90 g walking m −1 and incorporated into the top 0–20 cm of soil, which was naturally infected with Meloidogyne spp. In field studies, 1,3-dichloropropene and dazomet + sodium tetrathiocarbonate applications were generally superior to the bio-nematicide treatment. However, the recommended dose of the bio-nematicide (70 g walking m −1) significantly suppressed the numbers of second stage juveniles at the end of the cropping season in comparison with 1,3-dichloropropene. The broadcast application of the bio-nematicide was more efficient than the banded application, regardless of the dose levels, and as efficient as the oxamyl + sodium tetrathiocarbonate combined application for the control of root-knot nematodes. In pot-experiments, the bio-nematicide was generally more efficient in controlling root-knot nematodes than the bio-control agent P. penetrans. Application of the bio-nematicide increased both the size and the activity of the soil microbial biomass, especially at the higher dose rates of 70 and 90 g walking m −1. This was probably due mainly to the stimulation of the indigenous soil microflora caused by the addition of animal and plant extracts contained in the bio-nematicide formulation and partly to the added spores of B. firmus. Overall, the results indicate that the novel bio-nematicide offers a satisfactory and environmentally friendly solution for the control of root-knot nematodes.

Indigenous soil bacteria and low moisture may limit but allow faecal bacteria to multiply and become a minor population in tropical soils

The soil environment in Hawaii is generally characterised as sub-optimal but permissive to support the in situ growth of E. coli and enterococci. However, soil desiccation and competition for nutrients by major indigenous soil microflora have been identified as potential factors that could limit a rapid and continual growth of faecal indicator bacteria in this soil environment. Despite these limitations, the genetic capacities of E. coli and enterococci are robust enough to enable these bacteria to become established as minor populations of Hawaii's soil microflora. Although the concentrations of E. coli and enterococci may have represented a fraction of the total soil microbiota, their presence in this habitat was very significant, for two important reasons: (a) soil was a major environmental source of E. coli and enterococci, and (b) the elevated counts of these bacteria in streams that routinely exceeded the EPA standards were due to run-off from soil. As a result, E. coli and enterococci were inadequate indicators to measure the degree of faecal contamination and potential presence of sewage-borne pathogens in Hawaiian streams.

Bioremediation of Crude Oil Contaminated Soil by Bioaugmentation of Pseudomonas fluorescens NS1

A feasibility study was conducted to evaluate the efficiency of Pseudomonas fluorescens NS1 bioaugmented to stimulate in situ bioremediation of crude oil-contaminated soil with different amendments in treatment units. Pure culture of P. fluorescens NS1 was isolated from a petroleum-contaminated soil. The rate of degradation of petroleum hydrocarbons by the indigenous soil microflora and in the presence of P. fluorescens NS1 was assessed with the addition of nutrients and bulking agents for a period of about 35 days. The study showed that addition of wheat bran as bulking agent rapidly enhanced bioremediation of crude oil-contaminated soil compared to amendments in other treatment units.

The Effect of Interaction Between White-rot Fungi and Indigenous Microorganisms on Degradation of Polycyclic Aromatic Hydrocarbons in Soil

White-rot fungi applied for soil bioremediation have to compete with indigenous soil microorganisms. The effect of competition on both indigenous soil microflora and white-rot fungi was evaluated with regard to degradation of polycyclic aromatic hydrocarbons (PAH) with different persistence in soil. Sterile and non-sterile soil was artificially contaminated with 14C-labeled PAH consisting of three (anthracene), four (pyrene, benz[a]anthracene) and five fused aromatic rings (benzo[a]pyrene, dibenz[a,h]anthracene). The two fungi tested,Dichomitus squalens and Pleurotus ostreatus, produced similar amounts of ligninolytic enzymes in soil, but PAH mineralization by P. ostreatus was significantly higher. Compared to the indigenous soil microflora, P.ostreatus mineralized 5-ring PAH to a larger extent, while the indigenous microflora was superior in mineralizing 3-ring and 4-ring PAH. In coculture the special capabilities of both soil microflora and P. ostreatus were partly restricted due to antagonistic interactions, but essentially preserved. Thus, soil inoculation with P. ostreatus significantly increased the mineralization of high-molecular-weight PAH, and at the same time reduced the mineralization of anthracene and pyrene. Regarding the mineralization of low-molecular-weight PAH, the stimulation of indigenous soil microorganisms by straw amendment was more efficient than application of white-rot fungi.

Long-Term Maintenance of Rapid Atrazine Degradation in Soils Inoculated with Atrazine Degraders

Two different microbial communities able to degrade atrazine (atz) were inoculated in four different soils. The most critical factor affecting the success of inoculation was the soil pH and its organic matter (OM) content. In two alkaline soils (pH > 7), some inoculations led immediately to a strong increase of the biodegradation rate. In a third slightly acidic soil (pH = 6.1), only one inoculum could enhance atz degradation. In a soil amended with organic matter and straw (pH = 5.7, OM = 16.5%), inoculation had only little effect on atz dissipation on the short as well as on the long-term. Nine months after the microflora inoculations, atz was added again and rapid biodegradation in all alkaline inoculated soils was recorded, indicating the long-term efficiency of inoculation. In these soils, the number of atz degraders was estimated at between 6.5 × 103 and 1.5 × 106 (g of soil)-1, using the most probable number (MPN) method. Furthermore, the presence of the atz degraders was confirmed by the detection of the gene atzA in these soils. Denaturing gradient gel electrophoresis (DGGE) analysis of the 16S rDNA genes indicated that the inoculated bacterial communities had little effect on the patterns of the indigenous soil microflora.

Growth in coculture stimulates metabolism of the phenylurea herbicide isoproturon by Sphingomonas sp. strain SRS2.

Metabolism of the phenylurea herbicide isoproturon by Sphingomonas sp. strain SRS2 was significantly enhanced when the strain was grown in coculture with a soil bacterium (designated strain SRS1). Both members of this consortium were isolated from a highly enriched isoproturon-degrading culture derived from an agricultural soil previously treated regularly with the herbicide. Based on analysis of the 16S rRNA gene, strain SRS1 was assigned to the beta-subdivision of the proteobacteria and probably represents a new genus. Strain SRS1 was unable to degrade either isoproturon or its known metabolites 3-(4-isopropylphenyl)-1-methylurea, 3-(4-isopropylphenyl)-urea, or 4-isopropyl-aniline. Pure culture studies indicate that Sphingomonas sp. SRS2 is auxotrophic and requires components supplied by association with other soil bacteria. A specific mixture of amino acids appeared to meet these requirements, and it was shown that methionine was essential for Sphingomonas sp. SRS2. This suggests that strain SRS1 supplies amino acids to Sphingomonas sp. SRS2, thereby leading to rapid metabolism of (14)C-labeled isoproturon to (14)CO(2) and corresponding growth of strain SRS2. Proliferation of strain SRS1 suggests that isoproturon metabolism by Sphingomonas sp. SRS2 provides unknown metabolites or cell debris that supports growth of strain SRS1. The role of strain SRS1 in the consortium was not ubiquitous among soil bacteria; however, the indigenous soil microflora and some strains from culture collections also stimulate isoproturon metabolism by Sphingomonas sp. strain SRS2 to a similar extent.

Effect of biocontrol strains of Trichoderma on plant growth, Pythium ultimum polulations, soil microbial communities and soil enzyme activities.

Five strains of Trichoderma with known biocontrol activities were assessed for their effect upon pea growth and their antagonistic activity against large Pythium ultimum inocula. The effect of Trichoderma inocula upon the indigenous soil microflora and soil enzyme activities in the presence and absence of Pythium is assessed. In the absence of Pythium, Trichoderma strain N47 significantly increased the wet shoot weight by 15% but did not significantly affect the dry weight, whilst strains T4 and N47 significantly increased the root weights by 22% and 80%) respectively. Strains TH1 and N47 resulted in significantly greater root lengths. Pythium inoculation significantly reduced the root length and the number of lateral roots and nodules, and significantly increased the root and rhizosphere soil fungal populations. Pythium inoculation significantly reduced the plant wet and dry shoot weights and significantly increased the wet and the dry shoot/root ratio. All the Trichoderma strains reduced the number of lesions caused by Pythium and increased the number of lateral roots. The effect of the Pythium on emergence and shoot growth was significantly reduced by all the Trichoderma strains except strain To10. Inoculation with Trichoderma strains TH1 and T4 resulted in significantly greater wet root weights (62% and 57%, respectively) in the presence of Pythium compared to the Pythium control. Strain N47 significantly increased the shoot/root ratio compared to the Pythium control. Inoculation with Trichoderma strains T4, T12 and N47 significantly reduced Pythium populations. Pythium increased the activity of C, N and P cycle enzymes, whilst four Trichoderma strains reduced this effect, indicating reduced plant damage and C leakage. Overall, strains T4 and N47 had the greatest beneficial characteristics, as both these strains improved plant growth in the absence of Pythium and reduced plant damage in the presence of Pythium. The dual properties of these strains improve the commercial application, giving them an advantage over single action inocula, especially in the absence of plant pathogens.

Effects of Elevated Carbon Dioxide and Nitrogen Fertilization on Mycorrhizal Fine Roots and the Soil Microbial Community in Beech-Spruce Ecosystems on Siliceous and Calcareous Soil.

To study the responses of forests to global change, model ecosystems consisting of beech and spruce trees were maintained in open top chambers for 4 years under four conditions, namely with normal and elevated CO2 and with low and high nitrogen input, each replicated four times. Each open top chamber (height 3 m, diameter 3 m, soil depth 1.5 m) contained two separate soil compartments containing nutrient-poor siliceous and nutrient-rich calcareous soil. Here, we focus on the fine roots and the soil microbial community in these model ecosystems. At the time of planting, the fine roots were cut back according to forestry practice, and the newly formed roots were colonized by the indigenous soil microflora. After 4 years, the total biomass of fine roots, when averaged over all treatments, was almost the same in each of the two soil types; it was highest in the top 100 mm of soil (60%) and decreased sharply in deeper soil layers. Fungal biomass associated with the fine roots, consisting mainly of ectomycorrhizal fungi, was estimated using the ergosterol content as a marker. It was much higher in fine roots in the siliceous than in the calcareous soil, indicating considerably enhanced ectomycorrhiza formation in the nutrient-poor siliceous soil. Elevated atmospheric CO2 stimulated fine root production by ca. 85% and 43% in the top 100 mm of calcareous and siliceous soils respectively. Increased nitrogen input caused a slightly reduced production of fine root biomass in the calcareous soil but increased it by 33% in the siliceous soil. Marker substances for microorganisms were analyzed in the root-free soil. The amounts of carbon released by fumigation/extraction (a general marker for microbial biomass) and chitin (a marker for fungal biomass) were significantly higher in the top layer of the siliceous than of the calcareous soil, but they did not respond significantly to the treatments with elevated CO2 or the nitrogen fertilizer. The total concentration of the phospholipid fatty acids (PLFAs) and the number of the PLFAs did not differ between the two soil types. However, four of the eight most abundant PLFAs differed significantly between the two soil types. Principal component analysis revealed clearly separated clusters for the two soil types. Although analysis did not reveal any significant changes in response to the treatments, the concentration of the PLFA typical for ectomycorrhizal fungi was significantly higher under conditions of elevated CO2 in the nutrient-rich calcareous soil.

Biocontrol of Pythium in the pea rhizosphere by antifungal metabolite producing and non-producing Pseudomonas strains.

Four well-described strains of Pseudomonas fluorescens were assessed for their effect on pea growth and their antagonistic activity against large Pythium ultimum inocula. The effect of Pseudomonas strains on the indigenous soil microflora, soil enzyme activities and plant growth in the presence and absence of Pythium was assessed. Pythium inoculation reduced the shoot and root weights, root length, and the number of lateral roots. The effect of Pythium was reduced by the Pseudomonas strains. Strains F113, SBW25 and CHAO increased shoot weights (by 20%, 22% and 35%, respectively); strains Q2-87, SBW25 and CHAO increased root weights (14%, 14% and 52%). Strains SBW25 and CHAO increased root lengths (19% and 69%) and increased the number of lateral roots (14% and 29%). All the Pseudomonas strains reduced the number of lesions and the root and soil Pythium populations, while SBW25 and CHAO increased the number of lateral roots. Pythium inoculation increased root and soil microbial populations but the magnitude of this effect was Pseudomonas strain-specific. Pythium increased the activity of C, N and P cycle enzymes, while the Pseudomonas strains reduced this effect, indicating reduced plant damage. Strains SBW25 and CHAO had the greatest beneficial characteristics, as these strains produced the greatest reductions in the side effects of Pythium infection (microbial populations and enzyme activities) and resulted in significantly improved plant growth. Strain SBW25 does not produce antifungal metabolites, and its biocontrol activity was related to a greater colonization ability in the rhizosphere. This is the first critical comparison of such important strains of Ps. fluorescens showing disease biocontrol potential.

In situ bioremediation of soils contaminated with radioactive elements and toxic heavy metals

Two experimental plots of an agricultural land contaminated with radioactive elements (uranium, radium thorium) and toxic heavy metals (copper, zinc, cadmium) were treated by two different biotechnological in situ methods. The soil in this land was characterized by a negative net neutralization potential, and the soil pH was in a slightly acidic pH range (from 4 to 5). The contaminants were located mainly in the upper soil layers (mainly in the horizon A). Both methods were connected with the initial solubilization of the contaminants. Water acidified with sulphuric acid was used as a leach solution. The solubilization was mainly a result of the activity of the indigenous soil microflora. This activity was enhanced by suitable changes in the levels of some essential environmental factors, such as water, oxygen and nutrient content of the soil. The first method was then connected with the removal of the dissolved contaminants from the soil through the soil effluents. The second method was based on the transfer of the contaminants into the deeply located soil horizon B 2, where they were immobilized mainly as a result of the activity of the indigenous sulphate-reducing bacteria. Their activity was enhanced by injecting water solutions of organic compounds into the horizon B 2 through boreholes located in the relevant experimental plot.

Bioremediation of a soil contaminated with radioactive elements

Some agricultural lands located in the Vromos Bay area, near the Black Sea coast, Southeastern Bulgaria, have been contaminated with radioactive elements (uranium, radium and thorium) and toxic heavy metals (copper, cadmium and lead) as a result of mining and mineral processing of polymetallic ores. Laboratory experiments carried out with soil samples from these lands revealed that an efficient remediation of the soils was achieved by an in situ treatment method based on the activity of the indigenous soil microflora. The treatment was connected with the dissolution of the contaminants in the upper soil horizons and their transfer into the deeply located soil horizons (mainly to the horizon B 2) where they were immobilized as different insoluble compounds. The dissolution of contaminants was connected with the activity of both heterotrophic and chemolithotrophic aerobic microorganisms and the immobilization was due mainly to the anaerobic sulphate-reducing bacteria. The activity of these microorganisms was enhanced by suitable changes in the levels of some essential environmental factors such as water, oxygen and nutrient contents in the soil. On the basis of the above-mentioned laboratory results, the method was then applied under real field conditions in a heavily contaminated experimental plot of land located in the Vromos Bay area. Within 8 months of treatment, the contents of radioactive elements and toxic heavy metals in the soil were decreased below the relevant permissible levels.