Dynamic changes of bacterial community under the influence of bacterial-feeding nematodes grazing in prometryne contaminated soil

Microcosm experiments were carried out to study the effects of bacterial-feeding nematodes and prometryne on soil bacterial communities in contaminated soil. Prometryne (5 or 10 mg kg⁻¹ dry soil, that is, P5 or P10) and bacterial-feeding nematodes (5 or 10 individuals g⁻¹ dry soil, that is, N5 or N10), singly and in combination (P5N5, P5N10, P10N5, P10N10), were added to a nematode-free soil. An uncontaminated nematode-free soil was studied for comparison (Control). Bacterial-feeding nematode grazing boosted soil enzyme activities in contaminated soils, thus speeding up prometryne degradation. In the initial stage of the experiment, prometryne enhanced the soil enzyme activities too, but served the opposite purpose later. Denaturing gradient gel electrophoresis (DGGE) analysis indicated that prometryne contamination and nematode grazing over the incubation period exerted an obvious impact on Species richness (S), Shannon–Wiener index (H′) and Evenness (E_H) of soil bacteria, which increased initially, then decreased and increased again later. The cluster analysis of DGGE profiles showed that the similarity of soil bacterial communities in all treatments with indigenous microbes, P5, P5N5, P5N10, P10, P10N5, and P10N10 and the Control was 75%, 44%, 78% and 49% at Day 0, Day 8, Day 18 and Day 30, respectively. Compared to the Control, DGGE profiles displayed a varying characteristic bands pattern in all treatments over the incubation period with certain bands present in the treatments while not in the Control and vice versa, suggesting that bacterial-feeding nematode grazing and prometryne contamination affected soil bacterial communities evidently. Consequently, when added to contaminated soil, bacterial-feeding nematodes can contribute to restoration of contaminated sites by degrading toxic compounds like prometryne through enhanced microbial activity.     

Changes in soil organic carbon dynamics in an Eastern Chinese coastal wetland following invasion by a C4 plant Spartina alterniflora

The exotic C₄ grass Spartina alterniflora was intentionally introduced to tidal coastal wetlands in Jiangsu province of China in 1982. Since then it has rapidly replaced the native C₃ plant Suaeda salsa, becoming one of the dominant vegetation types in the coastal wetlands of China. Although plant invasion can change soil organic carbon (SOC) storage, little is known about how plant invasion influences C storage within soil fractions. We investigated how S. alterniflora invasion across an 8, 12 and 14-year chronosequence affected SOC and soil nitrogen (N), using soil fractionation and stable δ¹³C isotope analyses. SOC and N concentrations at 0-10 cm depth in S. alterniflora soil increased during the S. alterniflora invasion chronosequence, ranging from 3.67 to 4.90 g C kg⁻¹ soil, and from 0.307 to 0.391 g N kg⁻¹ soil. These were significantly higher than the values in the Suaeda salsa community, by 27.0-69.6% for SOC, and 21.8-55.2% for total N. The S. alterniflora-derived SOC varied from 0.40 to 0.92 g C kg⁻¹ according to mixing calculations, assuming the two possible SOC sources of S. alterniflora and S. salsa, and accounted for 10.8-18.7% of total SOC in the colonized soils. The estimated accumulative rate of SOC from C₄ (S. alterniflora) was 64.1 C kg⁻¹ soil year⁻¹ and from C₃ sources was 78.1 mg C kg⁻¹. The concentration of S. alterniflora-derived SOC significantly decreased from coarse fraction to fine fraction, and linearly increased as the period of S. alterniflora invasion increased. The highest accumulative rate of SOC from a C₄ source occurred in macroaggregates, while the highest rate from C₃ was in microaggregates. The storage of SOC derived from S. alterniflora in the macroaggregates was 0.27-0.44 g C kg⁻¹ soil, accounting for 43.1-49.1% of the total C₄derived SOC in the soil. Our results suggest that S. alterniflora invasion in coastal wetlands could facilitate SOC storage, because of the high potential for accumulation of the C which has been newly derived from S. alterniflora litter and roots.     

Decomposition of N-labelled maize leaves in soil affected by endogeic geophagous Aporrectodea caliginosa

A microcosm experiment was carried out for 56 days at 12 °C to evaluate the feeding effects of the endogeic geophagous earthworm species Aporrectodea caliginosa on the microbial use of ¹⁵N-labelled maize leaves (Zea mays) added as 5 mm particles equivalent to 1 mg C and 57 μg N g⁻¹ soil. The dry weight of A. caliginosa biomass decreased in the no-maize treatment by 10% during the incubation and increased in the maize leaf treatments by 18%. Roughly 5% and 10% of the added maize leaf-C and leaf-N, respectively, were incorporated into the biomass of A. caliginosa. About 29% and 33% of the added maize leaf-C were mineralised to CO₂ in the no-earthworm and earthworm treatments, respectively. The presence of A. caliginosa significantly increased soil-derived CO₂ production by 90 μg g⁻¹ soil in the no-maize and maize leaf treatments, but increased the maize-derived CO₂ production only by 40 μg g⁻¹ soil. About 10.5% of maize leaf-C and leaf-N was incorporated into the soil microbial biomass in the absence of earthworms, but only 6% of the maize leaf-C and 3% of the maize leaf-N in the presence of earthworms. A. caliginosa preferentially fed on N rich, maize leaf-colonizing microorganisms to meet its N demand. This led to a significantly increased C/N ratio of the unconsumed microbial biomass in soil. The ergosterol-to-microbial biomass C ratio was not significantly decreased by the presence of earthworms. A. caliginosa did not directly contribute to comminution of plant residues, as indicated by the absence of any effects on the contents of the different particulate organic matter fractions, but mainly to grazing of residue-colonizing microorganisms, increasing their turnover considerably.     

Influence of bacterial-feeding nematodes on nitrification and the ammonia-oxidizing bacteria (AOB) community composition

The effects of bacterial-feeding nematodes on nitrification and the ammonia-oxidizing bacteria (AOB) community composition were studied in soil microcosms. Sterilized soils were inoculated with mixed soil bacteria (obtained by filtering) or with bacteria and bacterial-feeding nematodes, after which the dynamic inorganic nitrogen concentration was measured weekly. After 28 days of incubation, denaturing gradient gel electrophoresis (DGGE) based on PCR amplification of the amoA gene was used to analyze the AOB community composition. In addition, a clone library from the amoA gene fragments was established using clones randomly selected and sequenced from the two treatments. The results showed that the presence of bacterial-feeding nematodes led to significantly greater NH₄⁺ and NO₃⁻ contents over the entire incubation period, indicating that bacterial-feeding nematodes promoted both N mineralization and nitrification. The results of DGGE showed that the AOB community composition was significantly changed in the presence of bacterial-feeding nematodes. Furthermore, the sequencing results suggested that Nitrosospira sp. was the dominant species in the treatment without nematodes, while Nitrosomonas sp. and Nitrosospira sp. were the dominant species in the treatment with nematodes. Such changes in the AOB community may be one of explanation of the important role that nematodes play in promoting nitrification.     

DIBOA: Fate in soil and effects on root-knot nematode egg numbers

The benzoxazinoid 2,4-dihydroxy-1,4-benzoxazin-3-one (DIBOA) is produced by rye (Secale cereale) and may contribute to plant-parasitic nematode suppression when rye plants are incorporated as a green manure. We investigated the fate of DIBOA in soil and DIBOA's effects on nematode reproduction. Soil in plastic bags was treated with DIBOA at concentrations ranging from 1.1 to 18 μg g⁻¹ dry soil, and with the root-knot nematode Meloidogyne incognita. Control soils were treated with water or with 0.31% methanol, with or without nematodes. DIBOA concentrations extracted from the soil were measured at selected times for 5 consecutive days. The soil from each bag was then placed into a pot in the greenhouse, and a cucumber seedling was transplanted into each pot. Five weeks later, only the highest DIBOA concentration, 18 μg g⁻¹ soil, reduced nematode egg numbers. At 0 h, DIBOA measured in soil ranged from 19.68 to 35.51% of the initial DIBOA concentration, and was dependent on the concentration added to the soil. DIBOA half-life was from 18 to 22 h, and very little DIBOA was present in soil after 120 h. Identified breakdown products accounted for only 4% at maximum of the initially added DIBOA. The results of our study demonstrate that high soil concentrations of DIBOA are necessary to suppress M. incognita; DIBOA may not be a major factor in nematode suppression by a rye cover crop.     

Interactions between residue placement and earthworm ecological strategy affect aggregate turnover and N2O dynamics in agricultural soil

Previous laboratory studies using epigeic and anecic earthworms have shown that earthworm activity can considerably increase nitrous oxide (N₂O) emissions from crop residues in soils. However, the universality of this effect across earthworm functional groups and its underlying mechanisms remain unclear. The aims of this study were (i) to determine whether earthworms with an endogeic strategy also affect N₂O emissions; (ii) to quantify possible interactions with epigeic earthworms; and (iii) to link these effects to earthworm-induced differences in selected soil properties. We initiated a 90-day ¹⁵N-tracer mesocosm study with the endogeic earthworm species Aporrectodea caliginosa (Savigny) and the epigeic species Lumbricus rubellus (Hoffmeister). ¹⁵N-labeled radish (Raphanus sativus cv. Adagio L.) residue was placed on top or incorporated into the loamy (Fluvaquent) soil. When residue was incorporated, only A. caliginosa significantly (p < 0.01) increased cumulative N₂O emissions from 1350 to 2223 μg N₂O-N kg⁻¹ soil, with a corresponding increase in the turnover rate of macroaggregates. When residue was applied on top, L. rubellus significantly (p < 0.001) increased emissions from 524 to 929 μg N₂O-N kg⁻¹, and a significant (p < 0.05) interaction between the two earthworm species increased emissions to 1397 μg N₂O-N kg⁻¹. These effects coincided with an 84% increase in incorporation of residue ¹⁵N into the microaggregate fraction by A. caliginosa (p = 0.003) and an 85% increase in incorporation into the macroaggregate fraction by L. rubellus (p = 0.018). Cumulative CO₂ fluxes were only significantly increased by earthworm activity (from 473.9 to 593.6 mg CO₂-C kg⁻¹ soil; p = 0.037) in the presence of L. rubellus when residue was applied on top. We conclude that earthworm-induced N₂O emissions reflect earthworm feeding strategies: epigeic earthworms can increase N₂O emissions when residue is applied on top; endogeic earthworms when residue is incorporated into the soil by humans (tillage) or by other earthworm species. The effects of residue placement and earthworm addition are accompanied by changes in aggregate and SOM turnover, possibly controlling carbon, nitrogen and oxygen availability and therefore denitrification. Our results contribute to understanding the important but intricate relations between (functional) soil biodiversity and the soil greenhouse gas balance. Further research should focus on elucidating the links between the observed changes in soil aggregation and controls on denitrification, including the microbial community.     

Impacts of different N management regimes on nitrifier and denitrifier communities and N cycling in soil microenvironments

Real-time quantitative PCR assays, targeting part of the ammonia monooxygenase (amoA), nitrous oxide reductase (nosZ), and 16S rRNA genes were coupled with ¹⁵N pool dilution techniques to investigate the effects of long-term agricultural management practices on potential gross N mineralization and nitrification rates, as well as ammonia-oxidizing bacteria (AOB), denitrifier, and total bacterial community sizes within different soil microenvironments. Three soil microenvironments [coarse particulate organic matter (cPOM; >250 μm), microaggregate (53-250 μm), and silt-and-clay fraction (<53 μm)] were physically isolated from soil samples collected across the cropping season from conventional, low-input, and organic maize-tomato systems (Zea mays L.-Lycopersicum esculentum L.). We hypothesized that (i) the higher N inputs and soil N content of the organic system foster larger AOB and denitrifier communities than in the conventional and low-input systems, (ii) differences in potential gross N mineralization and nitrification rates across the systems correspond with AOB and denitrifier abundances, and (iii) amoA, nosZ, and 16S rRNA gene abundances are higher in the microaggregates than in the cPOM and silt-and-clay microenvironments. Despite 13 years of different soil management and greater soil C and N content in the organic compared to the conventional and low-input systems, total bacterial communities within the whole soil were similar in size across the three systems (∼5.15 × 10⁸ copies g⁻¹ soil). However, amoA gene densities were ∼2 times higher in the organic (1.75 × 10⁸ copies g⁻¹ soil) than the other systems at the start of the season and nosZ gene abundances were ∼2 times greater in the conventional (7.65 × 10⁷ copies g⁻¹ soil) than in the other systems by the end of the season. Because organic management did not consistently lead to larger AOB and denitrifier communities than the other two systems, our first hypothesis was not corroborated. Our second hypothesis was also not corroborated because canonical correspondence analyses revealed that AOB and denitrifier abundances were decoupled from potential gross N mineralization and nitrification rates and from inorganic N concentrations. Our third hypothesis was supported by the overall larger nitrifier, denitrifier, and total bacterial communities measured in the soil microaggregates compared to the cPOM and silt-and-clay. These results suggest that the microaggregates are microenvironments that preferentially stabilize C, and concomitantly promote the growth of nitrifier and denitrifier communities, thereby serving as potential hotspots for N₂O losses.     

Influence of earthworm activity on aggregate-associated carbon and nitrogen dynamics differs with agroecosystem management

Earthworms are known to be important regulators of soil structure and soil organic matter (SOM) dynamics, however, quantifying their influence on carbon (C) and nitrogen (N) stabilization in agroecosystems remains a pertinent task. We manipulated population densities of the earthworm Aporrectodea rosea in three maize-tomato cropping systems [conventional (i.e., mineral fertilizer), organic (i.e., composted manure and legume cover crop), and an intermediate low-input system (i.e., alternating years of legume cover crop and mineral fertilizer)] to examine their influence on C and N incorporation into soil aggregates. Two treatments, no-earthworm versus the addition of five A. rosea adults, were established in paired microcosms using electro-shocking. A ¹³C and ¹⁵N labeled cover crop was incorporated into the soil of the organic and low-input systems, while ¹⁵N mineral fertilizer was applied in the conventional system. Soil samples were collected during the growing season and wet-sieved to obtain three aggregate size classes: macroaggregates (>250 μm), microaggregates (53-250 μm) and silt and clay fraction (<53 μm). Macroaggregates were further separated into coarse particulate organic matter (cPOM), microaggregates and the silt and clay fraction. Total C, ¹³C, total N and ¹⁵N were measured for all fractions and the bulk soil. Significant earthworm influences were restricted to the low-input and conventional systems on the final sampling date. In the low-input system, earthworms increased the incorporation of new C into microaggregates within macroaggregates by 35% (2.8 g m⁻² increase; P=0.03), compared to the no-earthworm treatment. Within this same cropping system, earthworms increased new N in the cPOM and the silt and clay fractions within macroaggregates, by 49% (0.21 g m⁻²; P<0.01) and 38% (0.19 g m⁻²; P=0.02), respectively. In the conventional system, earthworms appeared to decrease the incorporation of new N into free microaggregates and macroaggregates by 49% (1.38 g m⁻²; P=0.04) and 41% (0.51 g m⁻²; P=0.057), respectively. These results indicate that earthworms can play an important role in C and N dynamics and that agroecosystem management greatly influences the magnitude and direction of their effect.     

Natural N abundance of plant and soil inorganic-N as evidence for over-fertilization with compost

To test the hypothesis that N isotope composition can be used as evidence of excessive compost application, we measured variation in patterns of N concentrations and corresponding δ¹⁵N values of plants and soil after compost application. To do so, a pot experiment with Chinese cabbage (Brassica campestris L. cv. Maeryok) was conducted for 42 days. Compost was applied at rates of 0 (SC0), 500 (SC1), 1000 (SC2), and 1500 mg N kg⁻¹ soil (SC3). Plant-N uptake linearly increased with compost application (r² = 0.956, P < 0.05) with an uptake efficiency of 76 g N kg⁻¹ of compost-N at 42 days after application, while dry-mass accumulation did not show such linear increases. Net N mineralized from compost-N increased linearly (r² = 0.998, P < 0.01) with a slope of 122 g N kg⁻¹ of compost-N. Plant-δ¹⁵N increased curvilinearly with increasing compost application, but this increase was insignificant between SC2 and SC3 treatments. The δ¹⁵N of soil inorganic-N (particularly NO₃⁻-N) increased with compost application. We found that plant-δ¹⁵N reflected the N isotope signal of soil NO₃⁻-N at each measurement during plant growth, and that δ¹⁵N of inner leaves and soil NO₃⁻-N was similar when initial NO₃⁻ in the compost was abundant. Therefore, we concluded that δ¹⁵N of whole plant (more obviously in newer plant parts) and soil NO₃⁻-N could reveal whether compost application was excessive, suggesting a possible use of δ¹⁵N in plants and soil as evidence of excess compost application.     

Rapid quantification of Bacillus subtilis antibiotics in the rhizosphere

Biological control agents like Bacillus subtilis offer an alternative and supplement to synthetic pesticides. Antibiotic production by biocontrol strains of B. subtilis can play a major role in plant disease suppression. Our current understanding of B. subtilis antibiosis comes from culture media measurements of antibiotic production and in vitro suppression of pathogens. Quantifying the antibiotic metabolite chemistry of B. subtilis biofilms growing on root surfaces provides a more accurate understanding of in vivo antibiotic production. An analytical method based on solid-phase extraction (SPE) with high-performance liquid chromatography (HPLC) and mass spectroscopy (MS) has been developed to quantify antibiotics produced by B. subtilis growing on plant roots. Cucumber (Cucumis sativus) was grown in composted soil and potting media inoculated with B. subtilis strain QST 713 (AgraQuest, USA). Two important B. subtilis antibiotics, surfactin and iturin A, were extracted from root and rhizosphere soil using acidified organic solvents followed by cleaning and concentration using SPE. HPLC and HPLC-MS were used to measure surfactin and iturin A. Rhizosphere concentrations of both antibiotics increased with plant age. For plants grown in peat-based potting media, surfactin concentrations increased from 9 μg g⁻¹ root fresh weight (RFW) at 15 d to 30 μg g⁻¹ RFW at 43 d. Iturin concentrations were 7 μg g⁻¹ RFW at 15 d and 180 μg g⁻¹ RFW at 43 d. In an initial field trial in a composted fine sandy loam, we demonstrated rhizosphere production of surfactin and iturin under competition and predation by the myriad macro- and microfauna existing in a fertile high-organic soil, with mature B. subtilis-inoculated cucumber roots yielding 33 μg g⁻¹ RFW surfactin and 630 μg g⁻¹ RFW iturin at 78 d.     

Evidence for acclimation of N cycling to episodic N inputs in anthropogenically-affected intertidal salt marsh sediments

Nitrate removal was compared in anthropogenically-impacted and unimpacted salt marsh sediments in microcosms using the acetylene block technique and ¹⁵N natural abundance measurements. Potential denitrification rates were greater at the impacted site than the unimpacted site at all added NO₃⁻ concentrations (233, 467, or 700 μg N g dw⁻¹). Although the change in concentration of NH₄⁺ over time was small (69-104 μg N g dw⁻¹), the δ¹⁵N of accumulated NH₄⁺ increased significantly (0.26-13.22‰), and was more enriched for all NO₃⁻ treatments in the impacted sediments than the unimpacted site. The impacted site may be acclimated to episodic N inputs, and based on concentrations and ¹⁵N natural abundance had greater denitrification and N cycling than the unimpacted site.     

Evidence for gluconic acid production by Enterobacter intermedium as an efficient strategy to avoid protozoan grazing

The effect of pure gluconic acid and of gluconic-acid-producing bacteria on the activity of three protozoan species, Colpoda steinii (a ciliate), Vahlkampfia sp. (an amoeba) and Neobodo designis (a flagellate), was determined in vitro and in soil microcosms. Pure gluconic acid was shown to mediate disappearance of active cells, due to encystment and/or death of protozoa, at 0.15 mM in saline medium. Similarly, the presence of gluconic acid inhibited excystment of the three protozoa tested. Enterobacter intermedium 60-2G (Wt), a gluconic acid-producing rhizobacterium, elicited the same effects on protozoa when co-cultured in the presence of 5 g L⁻¹ glucose. However, the effect was not observed when glucose was omitted from the medium. Similarly, a pqqA isogenic mutant strain, unable to produce gluconic acid from glucose, exhibited a reduced effect on protozoan activity. Rhizosphere-microcosm studies performed with wheat (Triticum aestivum L.) confirmed the reduced ability of the pqqA mutant to limit protozoa reproduction compared to the Wt strain. Since the sodium salt of gluconic acid did not cause any significant stress to protozoa and considering that addition of 50 mM Tris-Cl (pH 7.2) abolished the deleterious effect of gluconic acid, acidification of the medium appeared as the key factor that induced encystment/death of protozoa. We propose that production and excretion of gluconic acid should be considered an efficient mechanism evolved by bacteria to escape, tolerate or defend themselves against protozoan grazing in rhizosphere environments.     

A mass-balance approach to measuring microbial uptake and pools of phosphorus in nutrient-amended soils

Soil microbial biomass P is usually determined through fumigation-extraction (FE), in which partially extractable P from lysed biomass is converted to biomass P using a conversion factor (K_p). Estimation of K_p has been usually based on cultured microorganisms, which may not adequately represent the soil microbial community in either nutrient-poor or in altered carbon and nutrient conditions following fertilisation. We report an alternative approach in which changes in microbial P storage are determined as the residual in a mass balance of extractable P before and after incubation. This approach was applied in three low-fertility sandy soils of southwestern Australia, to determine microbial P immobilisation during 5-day incubations in response to the amendment by 2.323 mg C g⁻¹, 100 μg N g⁻¹ and 20 μg P g⁻¹. The net P immobilisation during the amended incubations determined to be 18.1, 14.1 and 16.3 μg P g⁻¹ in the three soils, accounting for 70.6-90.5% of P added through amendment. Such estimates do not rely on fumigation and K_p values, but for comparison with the FE method we estimated ‘nominal’ K_p values to be 0.20-0.31 for the soils under the amended conditions. Our results showed that microbial P immobilisation was a dominant process regulating P concentration in soil water following the CNP amendment. The mass-balance approach provides information not only about changes in the microbial P compartment, but also about other major P-pools and their fluxes in regulating soil-water P concentrations under substrate- and nutrient-amended conditions.     

Wood, nematodes, and the nematode-trapping fungus Arthrobotrys oligospora

Researchers have proposed that Arthrobotrys oligospora and related fungi trap soil nematodes to obtain nitrogen and thereby compete saprophytically for carbon and energy in nitrogen-poor environments, including litter and wood. The current study tested two hypotheses concerning this model. The first was that wood decomposition would be enhanced if both large numbers of nematodes (a potential nitrogen supply) and A. oligospora (a cellulolytic organism that can use that N supply) were present. The second was that A. oligospora trapping activity would increase if large numbers of nematodes were added to soil containing abundant carbon (a wood dowel or chip). Although the first hypothesis was supported by an in vitro experiment on agar (A. oligospora degraded much more wood when nematodes were present), neither hypothesis was supported by an experiment in vials containing field soil. In soil, wood decomposition was unaffected by the addition of A. oligospora or large numbers of nematodes. Whereas A. oligospora trapped virtually all nematodes added to agar cultures, it trapped few or no nematodes added to soil. Given that the fungal isolate was obtained from the same soil and that the fungus increased to large numbers (>1×10³ propagules g⁻¹ soil), the failure of A. oligospora to trap nematodes in soil is difficult to explain. Soil nitrate levels, however, were high (71 mg kg⁻¹), and it is possible that with lower nitrate levels, trapping in soil might be stimulated by wood and nematodes.     

Rhizosphere isoflavones (daidzein and genistein) levels and their relation to the microbial community structure of mono-cropped soybean soil in field and controlled conditions

Despite an increase in the understanding of the soybean isoflavones involved in root-colonizing symbioses, relatively little is known about their levels in the rhizosphere and their interactions with the soil microbial community. Based on a 13-year experiment of continuous soybean monocultures, in the present study we quantified isoflavones in the soybean rhizosphere and analyzed the soil microbial community structure by examining its phospholipid fatty acid (PLFA) profile. Two isoflavones, daidzein (7, 4′-dihydroxyisoflavone) and genistein (5,7,4′- trihydroxyisoflavone), were detected in the rhizosphere soil of soybean plants, with the concentrations in the field varying with duration of mono-cropping. Genistein concentrations ranged from 0.4 to 1.2 μg g⁻¹ dry soil over different years, while daidzein concentrations rarely exceeded 0.6 μg g⁻¹ dry soil. PLFA profiling showed that the signature lipid biomarkers of bacteria and fungi varied throughout the years of the study, particularly in mono-cropping year 2, and mono-cropping years 6-8. Principal component analysis clearly identified differences in the composition of PLFA during different years under mono-cropping. There was a positive correlation between the daidzein concentrations and soil fungi, whereas the genistein concentration showed a correlation with the total PLFA, fungi, bacteria, Gram (+) bacteria and aerobic bacteria in the soil microbial community. Both isoflavones were easily degraded in soil, resulting in short half-lives. Concentrations as small as 1 μg g⁻¹ dry soil were sufficient to elicit changes in microbial community structure. A discriminant analysis of PLFA patterns showed that changes in microbial community structures were induced by both the addition of daidzein or genistein and incubation time. We conclude that daidzein and genistein released into the soybean rhizosphere may act as allelochemicals in the interactions between root and soil microbial community in a long-term mono-cropped soybean field.     

Elevated CO2, but not defoliation, enhances N cycling and increases short-term soil N immobilization regardless of N addition in a semiarid grassland

Elevated CO₂ and defoliation effects on nitrogen (N) cycling in rangeland soils remain poorly understood. Here we tested whether effects of elevated CO₂ (720 μl L⁻¹) and defoliation (clipping to 2.5 cm height) on N cycling depended on soil N availability (addition of 1 vs. 11 g N m⁻²) in intact mesocosms extracted from a semiarid grassland. Mesocosms were kept inside growth chambers for one growing season, and the experiment was repeated the next year. We added ¹⁵N (1 g m⁻²) to all mesocosms at the start of the growing season. We measured total N and ¹⁵N in plant, soil inorganic, microbial and soil organic pools at different times of the growing season. We combined the plant, soil inorganic, and microbial N pools into one pool (PIM-N pool) to separate biotic + inorganic from abiotic N residing in soil organic matter (SOM). With the ¹⁵N measurements we were then able to calculate transfer rates of N from the active PIM-N pool into SOM (soil N immobilization) and vice versa (soil N mobilization) throughout the growing season. We observed significant interactive effects of elevated CO₂ with N addition and defoliation with N addition on soil N mobilization and immobilization. However, no interactive effects were observed for net transfer rates. Net N transfer from the PIM-N pool into SOM increased under elevated CO₂, but was unaffected by defoliation. Elevated CO₂ and defoliation effects on the net transfer of N into SOM may not depend on soil N availability in semiarid grasslands, but may depend on the balance of root litter production affecting soil N immobilization and root exudation affecting soil N mobilization. We observed no interactive effects of elevated CO₂ with defoliation. We conclude that elevated CO₂, but not defoliation, may limit plant productivity in the long-term through increased soil N immobilization.     

Growth promoting effects of corn (Zea mays) bacterial isolates under greenhouse and field conditions

Fertilizer costs are a major component of corn production. The use of biofertilizers may be one way of reducing production costs. In this study we present isolation and identification of three plant growth promoting bacteria that were identified as Enterobacter cloacae (CR1), Pseudomonas putida (CR7) and Stenotrophomonas maltophilia (CR3). All bacterial strains produced IAA in the presence of 100 mg l⁻¹ of tryptophan and antifungal metabolites to several soilborne pathogens. S. maltophilia and E. cloacae had broad spectrum activity against most Fusarium species. The only strain that was positive for nitrogen fixation was E. cloacae and it, and P. putida, were also positive for phosphate solubilization. These bacteria and the corn isolate Sphingobacterium canadense CR11, and known plant growth promoting bacterium Burkholderia phytofirmans E24 were used to inoculate corn seed to examine growth promotion of two lines of corn, varieties 39D82 and 39M27 under greenhouse conditions. When grown in sterilized sand varieties 39M27 and 39D82 showed significant increases in total dry weights of root and shoot of 10-20% and 13-28% and 17-32% and 21-31% respectively. Plants of the two varieties grown in soil collected from a corn field had respective increases in dry weights of root and shoot of 10-30% and 12-35% and 11-19% and 10-18%. In sand, a bacterial mixture was highly effective whereas in soil individual bacteria namely P. putida CR7 and E. cloacae CR1 gave the best results with 39M27 and 39D82 respectively. These isolates and another corn isolate, Azospirillum zeae N7, were tested in a sandy soil with a 55 and 110 kg ha⁻¹ of nitrogen fertility at the Delhi research Station of Agriculture and Agri-Food Canada over two years. Although out of seven bacterial treatments, no treatment provided a statistically significant yield increase over control plots but S. canadense CR11 and A. zeae N7 provided statistically significant yield increase as compared to other bacteria. The 110 kg rate of nitrogen provided significant yield increase compared to the 55 kg rate in both years.     

Nematode suppression with brassicaceous amendments: application based upon glucosinolate profiles

Glucosinolate profiles differ among plant species and their isothiocyanate (ITC) derivatives differ in toxicity to nematodes. Successful management of plant-parasitic nematodes by ITCs requires the incorporation of appropriate amounts of glucosinolate-containing biomass. Plant materials, containing glucosinolate-precursors of the ITCs most toxic to nematodes, were selected and applied to soil based upon ITC lethal concentration (LC) values. This provided a reliable and repeatable basis for application rates for suppression of Meloidogyne javanica and Tylenchulus semipenetrans by Brassica hirta and M. javanica by B. juncea. Sufficient biomass of B. hirta to potentially yield 0.03-0.12 μmol ml⁻¹ of glucotropeolin reduced nematode survival compared to similar amounts of broccoli (Brassica oleraceae var. botrytis). At biomass levels providing >0.37 μmol ml⁻¹ of glucotropeolin, mortality of M. javanica was 100% with B. hirta. Biomass of B. juncea potentially yielding 2.82 μmol ml⁻¹ of sinigrin reduced M. javanica survival 65% below that obtained by a similar amount of broccoli. Rates of B. juncea to yield lethal levels of allyl ITC to reduce T. semipenetrans survival underestimated the glucosinolate application rates for this amendment. Application of plant biomass to soil >2.9% w/w reduced M. javanica survival regardless of the glucosinolate concentration of the amendment material. Application of brassicaceous amendments to soil initiates complex and dynamic biological and chemical processes. Despite the inherent complexity, we find that brassicaceous amendments can be applied to achieve consistent and repeatable nematode suppression when based upon the chemistry of the incorporated material.     

Effect of nitrogen and phosphate fertilisers on microbial and nematode diversity in pasture soils

Soil microbial and nematode populations, soil microbial community structure, and microbial and nematode functional diversity were studied in two fertiliser trials on perennial pasture at three sampling times. The N trial involved the application of 0, 200 and 400 kg N ha⁻¹ y⁻¹ in the form of urea. The P trial involved the application of 0, 30, 50 and 100 kg P ha⁻¹ y⁻¹ as superphosphate. The purpose of this study was to determine biological characteristics that may be used as indicators of soil quality as affected by fertiliser inputs.The N or P treatments had no effect on total bacteria, cellulolytic microbes, or the fluorescein diacetate hydrolysis. The fungus Fusarium culmorum was found only in the 200 kg N treatment (P<0.01). Gliocladium roseum declined in isolation frequency with increasing N (P<0.05,) while other Gliocladium spp. increased (P<0.01).The microbial community structure, ecophysiological index (EP), and colony-development index (CD) were determined using: colony development rates in 1/10 tryptose soy agar (TSA), a Pseudomonas medium, and a nutrient poor medium. These parameters were not affected by the addition of the N or P fertilisers. In the N trial, the functional diversity of soil microbes, as determined by Shannon Diversity Index (H) and average well colour development (AWCD) (using Biolog gram negative microplates) was higher in the unfertilised than fertilised treatments. The values for H and AWCD were 4.2 and 0.78 in the unfertilised compared to 4.0 and 0.53 in fertilised treatment (P<0.01, 48 h, mean for both N treatments), respectively. There were no significant differences in these values in the P trial.Populations of the plant feeding nematodes Pratylenchus and Paratylenchus were greater (P<0.05) whereas those of Meloidogyne were lower (P<0.001), in soils fertilsed with N than in unfertilised soils. The genera Aporcelaimus, Dorylaimellus, and Tylencholaimellus were found only in control plots and their loss paralleled faunal changes resulting from pasture improvement reported elsewhere. Nematode Maturity Index (MI) values were 1.78, 1.85, and 1.53 for the N fertiliser treatments (P<0.05) suggesting a reduction at 400 kg N. The MI was not affected by the application of P (mean, 2.01), however, but all values in the P trial were greater than in the N trial. In the N and P trials an average of 29 and 35 nematode taxa were discriminated. The ratio of bacterial-feeding nematodes to bacterial-feeding plus fungal-feeding nematodes was similar across all treatments of the N (0.90–0.92) and P (0.84–0.90) trials, suggesting no relative change in the importance of bacterial- and fungal-mediated decomposition pathways in these soils as a result of fertiliser application.The finding that most microbiological characteristics did not respond to many years of fertiliser treatments suggests that the microbial community in the soils are similar and fertiliser amendments are insufficient to induce changes (either direct or indirect due to plant effects) in these communities. However, the consistent decrease in functional diversity of soil microflora and nematode populations with the application N, but not P, indicates that the N application can impact on community structure.     

Identification and localization of food-source microbial nucleic acids inside soil nematodes

Microorganisms (e.g., prokaryotes, fungi) are food sources for soil nematodes, but they can also be potential mutualists or pathogens. Understanding the linkages between microorganism and invertebrate diversity in soils requires the ability to distinguish between these microbial roles. We tested the potential of a taxon-specific fluorescent in situ hybridization (FISH) procedure for identifying and localizing microbial rRNA within the bodies of soil nematodes. Our objective was to determine whether the rate of digestion permitted detection and identification of food-source nucleic acids within the nematode digestive system (i.e., pharynges, intestines) before their breakdown. First, using laboratory cultures of Caenorhabditis elegans maintained on Escherichia coli, we were able to localize bacterial rRNA throughout the nematode pharynx with the universal bacterial-probe EUB338, although never in the intestines. Second, we applied the fungal rRNA probe FR1 to Aphelenchus avenae cultured on the fungus Rhizoctonia solani. We were unable to detect fungal rRNA within these nematodes, and it appears that this material may be digested rapidly. Next, we applied our technique to nematodes extracted directly from soils. We were able to localize bacterial rRNA within the pharynges of bacterial-feeding species of nematodes from desert soils. We also localized archaeal rRNA using the probe ARC344. Finally, application of EUB338 to desert soil nematodes revealed the presence of bacteria in the intestines of some nematodes and within the ovary of a single nematode. This technique has great potential for use in understanding the feeding behavior of bacterial-feeding soil nematodes and in studies of nematode:bacterial relationships.