Accumulation of specific flavonoids in soybean (Glycine max (L.) Merr.) as a function of the early tripartite symbiosis with arbuscular mycorrhizal fungi and Bradyrhizobium japonicum (Kirchner) Jordan

This study is the first report assessing the effect of soil inoculation on the signalling interaction of Bradyrhizobium japonicum, arbuscular mycorrhizal fungi (AMF) and soybean plants throughout the early stages of colonisation that lead to the tripartite symbiosis. In a study using soil disturbance to produce contrasting indigenous AMF treatments, the flavonoids daidzein, genistein and coumestrol were identified as possible signals for regulating the establishment of the tripartite symbiosis. However, it was unclear whether soil disturbance induced changes in flavonoid root accumulation other than through changing the potential for AMF colonization. In this study, soil treatments comprising all possible combinations of AMF and B. japonicum were established to test whether (1) modifications in root flavonoid accumulation depend on the potential for AMF colonization, and (2) synthesis and accumulation of flavonoids in the roots change over time as a function of the early plant-microbial interactions that lead to the tripartite symbiosis. The study was comprised of two phases. First, maize was grown over 3-week periods to promote the development of the AM fungus Glomus clarum. Second, the interaction between soybean, G. clarum and B. japonicum was evaluated at 6, 10, 14 and 40 days after plant emergence. Root colonization by G. clarum had a positive effect on nodulation 14 days after emergence, producing, 30% more nodules which were 40% heavier than those on roots solely inoculated with B. japonicum. The tripartite symbiosis resulted in 23% more N₂ being fixed than did the simpler symbiosis between soybean and B. japonicum. The presence of both symbionts changed accumulation of flavonoids in roots. Daidzein and coumestrol increased with plant growth. However, development of the tripartite symbiosis caused a decrease in coumestrol; accumulation of daidzein, the most abundant flavonoid, was reduced in the presence of AMF.     

Volatile organic compounds in the roots and rhizosphere of Pinus spp.

Plant roots normally release a complex mixture of chemicals which have important effects in the rhizosphere. Among these different root-emitted compounds, volatile isoprenoids have received very little attention, yet they may play important and diverse roles in the rhizosphere, contributing to the regulation of microbial activity and nutrient availability. It is therefore important to estimate their abundance in the rhizosphere, but so far, there is no reliable sampling method that can be used to measure realistic rates of root emissions from plants growing in field conditions, or even in pots. Here, we measured root content of volatile isoprenoids (specifically monoterpenes) for Pinus pinea, and explored the feasibility of using a dynamic bag enclosure method to measure emissions from roots of intact pot-grown plants with different degrees of root cleaning. We also investigated a passive diffusion method for exploring monoterpenes in soil at incremental distances from mature Pinus sylvestris trees growing in field conditions. Total monoterpene content of P. pinea roots was 415±50 μg g⁻¹ fresh wt in an initial screening study, and between 688±103 and 1144±208 μg g⁻¹ dry wt in subsequent investigations. Emissions from shaken-clean roots of intact plants and roots of intact plants washed to remove remaining soil after shaken-clean experiments were 119±14 and 26±5 μg g⁻¹ dry wt h⁻¹, respectively. Emissions from intact roots in soil-balls were an order of magnitude lower than from shaken-clean roots, and probably reflected the amount of emitted compounds taken up by physical, chemical or biological processes in the soil matrix surrounding the roots. Although monoterpene content was not significantly different in droughted roots, emission rates from droughted roots were generally significantly lower than from well-watered roots. Finally, passive sampling of monoterpenes in the soil at different distances from mature P. sylvestris trees in field conditions showed significantly decreasing sampling rates with increasing distance from the trunk. We conclude that it is feasible to measure volatile isoprenoid emissions from roots but the method of root preparation affects magnitude of measured emissions and therefore must be decided according to the application. We also conclude that the rhizosphere of Pinus species is a strong and previously un-characterized source of volatile isoprenoid emissions and these are likely to impact significantly on rhizosphere function.     

Distribution of monoterpenes between organic resources in upper soil horizons under monocultures of Picea abies, Picea sitchensis and Pinus sylvestris

The aim of this study was to compare the monoterpene content and distribution in litters and roots of three conifer species: Picea abies (L.) Karst, Picea sitchensis (Bong.) Carr. and Pinus sylvestris (L.). We analysed the monoterpene content of green needles, needle litter, F (fermentation) layer material and roots collected from monoculture plots. The rate of loss of monoterpenes from freshly fallen litter in the field was also studied at two monthly intervals over 10 months, to assess the length of time that monoterpenes entering the litter layer remain. Monoterpene analysis was carried out by extracting homogenised samples in hexane and identifying and quantifying the resulting monoterpenes using gas chromatography with flame ionisation detection (GC-FID) and gas chromatography-mass spectrometry (GC-MS). Mean total monoterpene concentrations varied significantly between the three species examined (e.g. in freshly fallen litter 1531 ± 96, 100 ± 5 and 1175 ± 122 μg g⁻¹ d. wt for P. abies, P. sitchensis and P. sylvestris); each species had distinctive and consistent monoterpene profiles associated with each type of tissue, and total monoterpene concentrations in green needles varied between individual trees of the same species, particularly for P. sitchensis. A substantial proportion of the monoterpene content of green needles remained in the needles after litter fall for P. abies (42%), P. sitchensis (11%) and P. sylvestris (30%). Although rates of monoterpene loss from needle litters varied initially (P. sylvestris > P. abies > P. sitchensis), the majority of the monoterpene content was lost after 4-6 months. Maximum monoterpene emission rates from decaying litter were calculated of 39 (P. abies), 1.7 (P. sitchensis) and 39 μg m⁻² h⁻¹ (P. sylvestris). Monoterpene concentrations in F layer material were very low (<10 μg g⁻¹ d. wt). Roots, particularly in P. sylvestris, represented a significant pool of monoterpenes (185 ± 16, P. abies; 258 ± 54, P. sitchensis; 2133 ± 200 μg g⁻¹ d. wt, P. sylvestris). The monoterpene profile was similar between roots and litter of P. sylvestris (α-pinene most abundant), and for P. sitchensis, (limonene and α-pinene most abundant), although a different pattern was observed between needle litter (most abundant β-pinene) and roots (most abundant myrcene) of P. abies. The relatively high concentrations and different profiles of monoterpenes characterised in upper organic soil horizons here emphasise the need for their influence on soil ecological processes to be assessed.     

Lead tolerance in Aporrectodea rosea earthworms from a clay pigeon shooting site

Lead tolerance in individuals of the earthworm species Aporrectodea rosea collected from a clay pigeon shooting site was investigated. Lead concentrations in the shooting site soil and the un-shot control site were 6410±2250 and 296±98 mg_Pb kg⁻¹ dry weight, respectively. Of these concentrations 1050±240 and 12±9 mg_Pb kg⁻¹ dry weight were suggested to be available, using ammonium acetate (1 M), respectively. With respect to earthworm body burdens of lead the shooting site earthworms had a body burden of 6.1±1.2 mg_Pb g⁻¹ dry weight while the uncontaminated site earthworms had almost a 1000-times lower body burden of 7.1±9.0 μg_Pb g⁻¹ dry weight. Lead tolerance was assessed in uncontaminated soil that had been augmented with lead, using lead nitrate solutions, to obtain lead concentrations in soil of 0.5, 5 and 50 mg_Pb kg⁻¹ dry weight. Earthworms were exposed for 28 days during which time a semi-qualitative assessment was made of their condition. Results showed no decrease in condition in the shooting site earthworms with increasing exposure time or concentration. In contrast, earthworms collected from an uncontaminated site showed a significant (p<0.05) decrease in condition when exposed to lead concentrations above, and including, a concentration of 5 mg kg⁻¹ dry weight soil. These results suggested lead tolerance in the shooting site earthworms.     

Food choice and reproductive success of Folsomia candida feeding on copper-contaminated mycelium of the soil fungus Alternaria alternata

We examined collembolan food preference for fungal mycelium grown on copper-contaminated medium, and the relationship between copper content, food selectivity and collembolan fitness when fed contaminated mycelium.To clarify whether collembolan food selectivity is related to fitness parameters, Folsomia candida were fed mycelium of the dark-pigmented fungus Alternaria alternata grown on medium with different copper concentrations. Copper-contaminated food (fungus grown on 50, 125, 250 and 500 μg Cu g⁻¹ medium, fresh wt.) was offered together with untreated food for 4 weeks. F. candida fed selectively on the provided mycelium and discriminated clearly between mycelium grown on high and low levels of contamination, distinctly preferring fungus grown on medium with a total copper concentration of 50 and 125 μg g⁻¹. In contrast, fungus grown on highly contaminated medium (250 and 500 μg g⁻¹) was avoided. Collembolan food preference generally matched fitness parameters. Reproduction was significantly affected by the total copper concentration of the fungal growth medium. When fed their preferred mycelium, collembolan reproduction was enhanced, whereas a diet of highly contaminated mycelium (250 or 500 μg g⁻¹) resulted in a strong decrease in reproduction. Adult survival was affected only marginally. Even though heavy metal contamination is a potential stress factor for many soil microarthropods, F. candida is able to discriminate between high and low quality food sources, and even benefits from moderately elevated copper concentrations.     

Adsorption and desorption of monomeric Bt (Bacillus thuringiensis) Cry1Aa toxin on montmorillonite and kaolinite

Genetically modified crops, which produce pesticidal proteins from Bacillus thuringiensis, release the toxins into soils through root exudates and upon decomposition of crop residues. Although the phenomena of gene transfer and emergence of resistance have been well documented, the fate of these toxins in soil has not yet been clearly elucidated. The aim of this study was to elucidate the adsorption and the desorbability of the Cry1Aa Bt insecticidal protein in contact with two sodium-saturated clays: montmorillonite and kaolinite. Because the toxin is released into soil in small quantities, it was assumed that it will be in a monomeric state in solution until it oligomerized on cell membranes. The originality of this study was to focus on the monomeric form of the protein. Specific sample conditions were required to avoid polymerisation. A pH above 6.5 and an ionic strength of at least 150 mM (NaCl) were necessary to keep the protein in solution and in a monomeric state. The adsorption isotherms obtained were of the L-type (low affinity) for both clays and fitted the Langmuir equation. The adsorption maximum of the toxin, calculated by the Langmuir nonlinear regression, decreased with increasing pH from 6.5, which was close to the isoelectric point, to 9. At pH 6.5, the calculated adsorption was 1.7 g g⁻¹ on montmorillonite and 0.04 g g⁻¹ on kaolinite. Desorbability measurements showed that a small fraction of toxin could be desorbed by water (up to 14%) and more by alkaline pH buffers (36 ± 7%), indicating that it was not tightly bound. Numerous surfactants were evaluated and the toxin was found to be easily desorbed from both clays when using zwitterionic and nonionic surfactants such as CHAPS, Triton-X-100, and Tween 20. This finding has important implications for the optimization of detection methods for Bt toxin in soil.     

Mycorrhizae activity and diversity in conventional and organic apple orchards from Brazil

The aim of this study was to characterize the impacts of a conventional, an organic apple orchard and a native grassland on the activity and diversity of arbuscular mycorrhizal fungi (AMF) located at the south of Brazil during winter and summer. AMF activity was measured by the mycorrhizal inoculum potential (MIP), mycorrhizal fungal hyphal length (HL), easily extractable and total Bradford-reactive soil protein (BRSP). AMF diversity was represented by richness, Shannon diversity index and number of spores. Orchards reduced the MIP of the soil and increased the HL when compared to the grassland site. The amount of easily extractable BRSP was not different among orchards and between seasons evaluated, with overall mean value of 1.23 mg g⁻¹. However, the amount of total-BRSP was smaller in the conventional orchard (4.55 mg g⁻¹) than in the organic orchard (4.91 mg g⁻¹) and in the native grassland (5.12 mg g⁻¹). T-BRSP and total organic carbon were strongly correlated in the grassland during the winter, suggesting the contribution of this protein for carbon stocks in the native soil of this region. The organic orchard presented the highest AMF richness, but sporulation and Shannon diversity index were larger in the conventional orchard. Our data suggest that the conventional orchard promoted higher impacts on the natural condition of AMF activity, being considered an unadvisable practice to soil conservation.     

Facilitation of pentachlorophenol degradation in the rhizosphere of ryegrass (Lolium perenne L.)

The phytoremediation of xenobiotics depends upon plant-microbe interactions in the rhizosphere, but the extent and intensity of these effects are currently unknown. To investigate rhizosphere effects on the biodegradation of xenobiotics, a glasshouse experiment was conducted using a specially designed rhizobox where ryegrass seedlings were grown for 53 days in a soil spiked with pentachlorophenol (PCP) at concentrations of 8.7±0.5 and 18±0.5 mg kg⁻¹ soil. The soil in the rhizobox was divided into six separate compartments at various distances from the root surface. Changes in PCP concentrations with increasing distance from the root compartment of the rhizobox were then assessed. The largest and most rapid loss of PCP in planted soil was at 3 mm from the root zone where total PCP decreased to 0.20 and 0.65 mg kg⁻¹, respectively with the two PCP treatments. The degradation gradient followed the order: near-rhizosphere>root compartment>far-rhizosphere soil zones for both concentrations where ryegrass was grown. In contrast, there was no difference in PCP concentration with distance in the unplanted soil. The increases in both soil microbial biomass carbon and the activities of soil urease and phosphatase were accompanied by the enhanced degradation of PCP, which was higher in the near-rhizosphere than far-rhizosphere soil. The results suggest that the effect of root proximity is important in the degradation of xenobiotics such as PCP in soil.     

Effect of indigenous mycorrhizal colonization on phosphorus‐acquisition efficiency in soybean and sunflower

Despite a general consent about the beneficial contribution of arbuscular mycorrhizal fungi (AMF) on natural ecosystems, there is an intense debate about their role in agricultural systems. In this work, soybean (Glycine max L.) and sunflower (Helianthus annuus L.) field plots with different P availabilities were sampled across the Pampean Region of Argentina (> 150 samples from Mollisols) to characterize the relationship between available soil P and indigenous mycorrhizal colonization. A subsequent pot experiment with soybean and sunflower was carried out to evaluate the effect of P supply (0, 12, and 52 mg P kg^–1) and AMF inoculation on AMF colonization and crop responsiveness to P in a Mollisol. Both crops showed high AMF colonization in the field (average: 55% for soybean and 44% for sunflower). While mycorrhizal colonization in soybean was significantly and negatively related to available soil P, no such trends were apparent in sunflower. Also, total biomass was 3.5 and 2.0 times higher in mycorrhizal than in nonmycorrhizal pot‐grown soybean under low‐ and medium‐P conditions, respectively. Sunflower, on the other hand, did not benefit from AMF symbiosis under medium and high P supply. While mycorrhization stimulated P‐uptake efficiency in soybean, the generally high P efficiency in sunflower was not associated with AMF symbiosis.     

Survival of the biocontrol agents Coniothyrium minitans and Bacillus subtilis MBI 600 introduced into pasteurised, sterilised and non-sterile soils

The biocontrol agents Coniothyrium minitans and Bacillus subtilis MBI 600 were added separately to three soil types that had been either sterilised, pasteurised or left non-sterile. Applied as a conidial suspension of 1×10⁶ cfu g⁻¹ soil, C. minitans showed good survival in all sterilised, pasteurised and non-sterile soils, remaining at the numerical level at which it was applied for the duration of the 30 d experiment. Applied at a lower rate of 1×10³ cfu g⁻¹ soil, C. minitans proliferated in sterilised soil to numbers slightly over 1×10⁶ cfu g⁻¹ soil, whereas no increase was seen in pasteurised or non-sterile soils from this lower application rate. However, although C. minitans was not easily recovered on plates from non-sterile soil, it did survive at the lower numerical level in pasteurised soil, and was recoverable throughout the experiment at the rate at which it was applied. B. subtilis MBI 600 survived well following introduction as a cell suspension into sterilised soil at a rate of 1×10⁶ cfu g⁻¹ soil. Spores were formed rapidly and, after 14 d, the introduced microorganism survived in this form rather than as vegetative cells. However, in non-sterile soil, the introduced microorganism did not compete well and decreased in number, with spores being formed in low numbers. Survival of B. subtilis MBI 600 in pasteurised soil was variable, but resembled the survival seen in non-sterile soil more than that seen in sterilised soil. More B. subtilis MBI 600 spores were formed in pasteurised soil than in non-sterile soil, however, and may have been important for survival in pasteurised soil. In conclusion, this work has shown that the biocontrol agent C. minitans can survive well in soil irrespective of whether the soil has been pasteurised or not and shows good promise as a soil inoculant for control of Sclerotinia sclerotiorum. Although soil pasteurisation does improve establishment of B. subtilis MBI 600 compared to non-sterile soil, survival is relatively poor when applied as cells. The best survival of B. subtilis MBI 600 occurred as spores in sterilised soil, and spore applications to pasteurised soil in an integrated control strategy may allow sufficient establishment of the biocontrol agent to target pathogens causing damping-off.     

Toxicological effects of TiO2 and ZnO nanoparticles in soil on earthworm Eisenia fetida

Nanoparticles (NPs) of TiO₂ and ZnO are receiving increasing attention due to their widespread applications. To evaluate their toxicities to the earthworm Eisenia fetida (Savigny, 1826) in soil, artificial soil systems containing distilled water, 0.1, 0.5, 1.0 or 5.0 g kg⁻¹ of NPs were prepared and earthworms were exposed for 7 days. Contents of Zn and Ti in earthworm, activities of antioxidant enzymes, DNA damage to earthworm, activity of cellulase and damage to mitochondria of gut cells were investigated after acute toxicity test. The results from response of the antioxidant system combined with DNA damage endpoint (comet assay) indicated that TiO₂ and ZnO NPs could induce significant damage to earthworms when doses were greater than 1.0 g kg⁻¹. We found that Ti and Zn, especially Zn, were bioaccumulated, and that mitochondria were damaged at the highest dose in soil (5.0 g kg⁻¹). The activity of cellulase was significantly inhibited when organisms were exposed to 5.0 g kg⁻¹ of ZnO NPs. Our study demonstrates that both TiO₂ and ZnO NPs exert harmful effects to E. fetida when their levels are higher than 1.0 g kg⁻¹ in soil and that toxicity of ZnO NPs was higher than TiO₂.     

Dynamics of microbial biomass and nitrogen supply during primary succession on blastfurnace slag dumps in dry tropics

This paper reports the role of microbial biomass in the establishment of N pools in the substratum during primary succession (till 40-year age) in Blastfurnace Slag Dumps, an anthropogenically created land form in the tropics. Initially in the depressions in the slag dumps fine soil particles (silt+clay) accumulate, retaining moisture therein, and providing microsites for the accumulation of microbial biomass. In all sites microbial biomass showed distinct seasonality, with summer-peak and rainy season-low standing crops. During the summer season microbial biomass C ranged from 18.6 μg g⁻¹ in the 1-year old site to ca. 235 μg g⁻¹ in the 40-year old site; correspondingly, microbial biomass N ranged from 1.22 to 40 μg g⁻¹. On sites 2.5-years of age and younger, the microbial biomass N content accounted for more than 50% of the organic N in the soil, whereas the proportion of microbial biomass N was ca. 7% of organic N in 40-year old site. The strong correlation between microbial biomass and total N in soil indicated a significant role of microbes in the build-up of nitrogen during the initial stages of succession in the slag dumps. Though the organic N pool in the soil was low (594 mg kg⁻¹) even after 40 years of succession, the available N (NH₄-N and NO₃-N) contents in the soil were generally high through the entire age series (ca. 16-32 μg g⁻¹) during the rainy season (which supports active growth of the herbaceous community). The high mineral-N status on the slag dump was related with high N-mineralization rates, particularly in the young sites (20.6 and 13.9 μg g⁻¹ month⁻¹ at 1 and 2.5-year age). We suggest that along with the abiotic factors having strong effect on ecosystem functioning, the microbial biomass, an important biotic factor, shows considerable influence on soil nutrient build-up during early stages of primary succession on the slag dumps. The microbial biomass dynamics initiates biotic control in developing slag dumps ecosystem through its effect on nitrogen pools and availability.     

CO2 emission in an intensively cultivated loam as affected by long-term application of organic manure and nitrogen fertilizer

A long-term field experiment was conducted to examine the influence of mineral fertilizer and organic manure on the equilibrium dynamics of soil organic C in an intensively cultivated fluvo-aquic soil in the Fengqiu State Key Agro-Ecological Experimental Station (Fengqiu county, Henan province, China) since September 1989. Soil CO₂ flux was measured during the maize and wheat growing seasons in 2002-2003 and 2004 to evaluate the response of soil respiration to additions and/or alterations in mineral fertilizer, organic manure and various environmental factors. The study included seven treatments: organic manure (OM), half-organic manure plus half-fertilizer N (NOM), fertilizer NPK (NPK), fertilizer NP (NP), fertilizer NK (NK), fertilizer PK (PK) and control (CK). Organic C in soil and the soil heavy fraction (organo-mineral complex) was increased from 4.47 to 8.61 mg C g⁻¹ and from 3.32 to 5.68 mg C g⁻¹, respectively, after the 13 yr application of organic manure. In contrast, organic C and the soil heavy fraction increased in NPK soil to only 5.41 and 4.38 mg C g⁻¹, respectively. In the CK treatment, these parameters actually decreased from the initial C concentrations (4.47 and 3.32 mg C g⁻¹) to 3.77 and 3.11 mg C g⁻¹, respectively. Therefore, organic manure efficiently elevated soil organic C. However, only 66% of the increased soil organic C was combined with clay minerals in the OM treatment. Cumulative soil CO₂ emissions from inter-row soil in the OM and NPK treatments were 228 and 188 g C m⁻² during the 2002 maize growing season, 132 and 123 g C m⁻² during the 2002/2003 wheat growing season, and 401 and 346 g C m⁻² yr⁻¹ in 2002-2003, respectively. However, during the 2004 maize growing season, cumulative soil CO₂ emissions were as high as 617 and 556 g C m⁻², respectively, due to the contribution of rhizosphere respiration. The addition of organic manure contributed to a 16% increase in soil CO₂ emission in 2002-2003 (compared to NPK), where only 27%, 36% and 24% of applied organic C was released as CO₂ during the 2002 and 2004 maize growing seasons and in 2002-2003, respectively. During the 2002/2003 wheat growing season, soil CO₂ flux was significantly affected by soil temperature below 20 °C, but by soil moisture (WFPS) during the 2004 maize growing season at soil temperatures above 18 °C. Optimum soil WFPS for soil CO₂ flux was approximately 70%. When WFPS was below 50%, it no longer had a significant impact on soil CO₂ flux during the 2002 maize growing season. This study indicates the application of organic manure composted with wheat straw may be a preferred strategy for increasing soil organic C and sequestering C in soil.     

Diversity patterns of indigenous arbuscular mycorrhizal fungi associated with rhizosphere of cowpea (Vigna unguiculata (L.) Walp.) in Benin,West Africa

Assessment of diversity and understanding factors underlying species distribution are fundamental themes in ecology. However, the diversity of native arbuscular mycorrhizal fungi (AMF) species in African tropical agro-ecosystems remains weakly known. This research was carried out to assess the morphological diversity of indigenous AMF species associated with rhizosphere of cowpea (Vigna unguiculata (L.) Walp.) in different agro-ecological zones (AEZ) of Benin and to examine the effects of soil chemical properties, climatic factors and agricultural practices on this diversity. Results showed that, in Benin, cowpea was grown by farmers in very exhausted soils, where available phosphorus and potassium were deficient. The indigenous AMF spore density was on average 202 spores per 100 g dry soil and there was no difference in the density among the agro-ecological zones (P = 0.56). Fifteen AMF morphospecies belonging to eight genera (Gigaspora, Scutellospora, Racocetra, Acaulospora, Funneliformis, Rhizophagus, Glomus and Claroideoglomus) were detected. The computed species richness estimators indicated that a limited number of additional undetected morphospecies are probably present in cowpea fields. According to analysis of similarity (ANOSIM), the arbuscular mycorrhizas community composition of the Sudanian zone and Guinean zone were not significantly different (r = −0.01; P = 0.517). The diversity of AMF morphospecies in the fields was weak with prevalence of Glomeraceae (92%). Furthermore, AMF diversity and evenness indices were negatively correlated with annual rainfall (P < 0.01) and with available phosphorus (P < 0.05). However, no significant correlation was observed between AMF diversity indices and soil organic carbon. Ultimately, this study tends to confirm that soil management practices (tillage, soil disturbance) have negative effect on AMF diversity.     

The effect of mixing ground leaf litters to soil on the development of pitch pine ectomycorrhizal and soil arthropod communities in natural soil microcosm systems

The addition of leaf litter to soil influences both the nutrients and polyphenols of soil. It is likely that contrasting nutrient and polyphenolic composition of different plant litters may affect plant growth, mycorrhizal and soil arthropod communities. We report results from a microcosm experiment of effects of incorporation of three single leaf litter species and a mixture of all three on pitch pine seedling growth, their ectomycorrhizal community and soil arthropod community. The three litter species (pine, oak and huckleberry) represent co-dominant species within the New Jersey pine barrens ecosystem. We show that the leaf litters have different composition of nutrients and polyphenols, with rooting matrix containing pine litter having lower inorganic nitrogen content (1.6 μg g⁻¹) than oak (19.9 μg g⁻¹) and huckleberry (4.4 μg g⁻¹), but oak litter having the highest extractable phosphorus (13.3 cf. 0-0.08 μg g⁻¹) and total phenol content and lowest condensed tannin content. These differences were imparted to rooting matrix of homogenized humic (Oa) layer of pine barrens soil to which milled leaf litter was added and used in the microcosms. Pitch pine seedlings grew significantly better in un-amended rooting matrix (0.33±0.02 g) than any of the litter treatments (0.15±0.02-0.17±0.01 g) and tissue P concentrations tracked phosphate concentrations in the rooting matrix. Total P accumulation into plant tissue was higher in oak than control, attributable to a significantly higher (P<0.05) accumulation in roots (3.3±0.19 mg g⁻¹) compared to other species (1.1±0.04-2.3±0.08 mg g⁻¹). No relationship was seen between tissue N concentration and soil N, but seedlings growing in huckleberry litter amended soil accumulated less N than control. The effect of leaf litters on the ectomycorrhizal community composition were determined by PCA (first two axes accounted for 81% of the variance) and stepwise multiple regression analysis. These analyses showed that huckleberry leaf litter had a significant impact on mycorrhizal community composition with morphotypes Cg and DB being more abundant in the presence of huckleberry litter (178±13 cf. 68±15-106±15 for Cg and 141±11 cf. 88±23-111±18 for DB) and its influence of elevating nitrate nitrogen, organic nitrogen, total phenols and protein precipitation content of the rooting matrix. Mycorrhizal morphotypes BS and SB were significantly more abundant in the community where these soil factors were low in the absence of leaf litter addition. Total ectomycorrhizal abundance was negatively related to hydrolysable tannin concentration in the rooting matrix (r²=0.132, P<0.05). There was no influence of leaf litter type on mite density (dominated by non-burrowing phthiracarids), but collembolan density (dominated by Folsomia spp) showed a greater than threefold reduction in population density in the presence of leaf litter (F=6.47, P<0.05). Collembolan density was positively correlated with mycorrhizal morphotypes GS and SB (P<0.05) and negatively related to morphotypes DB (P<0.05) and soil extractable NH₄-N (P<0.05), suggesting a possible selection of fungal species in their diet and a relationship between collembola and nitrification.     

Subtropical plantations are large carbon sinks: Evidence from two monoculture plantations in South China

Quantifying the net carbon (C) storage of forest plantations is required to assess their potential to offset fossil fuel emissions. In this study, a biometric approach was used to estimate net ecosystem productivity (NEP) for two monoculture plantations in South China: Acacia crassicarpa and Eucalyptus urophylla. This approach was based on stand-level net primary productivity (NPP, based on direct biometric inventory) and heterotrophic respiration (R_h). In comparisons of R_h determination based on trenching vs. tree girdling, both trenching and tree girdling changed soil temperature and soil moisture relative to undisturbed control plots, and we assess the effects of corrections for disturbances of soil moisture and soil moisture on the estimation of soil CO₂ efflux partitioning. Soil microbial biomass and dissolved organic carbon were significantly lower in trenched plots than in tree girdled plots for both plantations. Annual soil CO₂ flux in trenched plots (R_h-t) was significantly lower than in tree-girdled plots (R_h-g) in both plantations. The estimates of R_h-t and R_h-g, expressed as a percentage of total soil respiration, were 58 ± 4% and 74 ± 6%, respectively, for A. crassicarpa, and 64 ± 3% and 78 ± 5%, respectively, for E. urophylla. By the end of experiment, the difference in soil CO₂ efflux between the trenched plots and tree-girdled plots had become small for both plantations. Annual R_h (mean of the annual R_h-t and R_h-g) and net primary production (NPP) were 470 ± 25 and 800 ± 118 g C m⁻² yr⁻¹, respectively, for A. crassicarpa, and 420 ± 35 and 2380 ± 187 g C m⁻² yr⁻², respectively, for E. urophylla. The two plantations in the developmental stage were large carbon sinks: NEP was 330 ± 76 C m⁻² yr⁻¹ for A. crassicarpa and 1960 ± 178 g C m⁻² yr⁻¹ for E. urophylla.     

Community composition and activities of denitrifying bacteria from adjacent agricultural soil, riparian soil, and creek sediment in Oregon, USA

We examined denitrifying bacteria from wet soils and creek sediment in an agroecosystem in Oregon, USA that received inputs of nitrogen (N) fertilizer. Our objective was to determine the variation in denitrifying community composition and activities across three adjacent habitats: a fertilized agricultural field planted to perennial ryegrass, a naturally vegetated riparian area, and creek sediment. Using C₂H₂ inhibition, denitrifying enzyme and N₂O-reductase activities were determined in short-term incubations of anaerobic slurries. A key gene in the denitrification pathway, N₂O reductase (nosZ), served as a marker for denitrifiers. Mean denitrifying enzyme activity (DEA) was similar among habitats, ranging from 0.5 to 1.8 μg N g⁻¹ dry soil h⁻¹. However, the ratio of N₂O production, without C₂H₂, to DEA was substantially higher in riparian soil (0.64±0.02; mean±standard error, n=12) than in agricultural soil (0.19±0.02) or creek sediment (0.32±0.03). Mean N₂O-reductase activity ranged from 0.5 to 3.2 μg N g⁻¹ dry soil h⁻¹, with greater activity in agricultural soil than in riparian soil. Denitrifying community composition differed significantly among habitats based on nosZ terminal-restriction fragment length polymorphisms. The creek sediment community was unique. Communities in the agricultural and riparian soil were more closely related but distinct. A number of unique nosZ genotypes were detected in creek sediment. Sequences of nosZ obtained from riparian soil were closely related to nosZ from Bradyrhizobium japonicum. Although nosZ distribution and N₂O-reductase activity differed among habitats, relationships between activity and community composition appeared uncoupled across the agroecosystem.     

Shifts in amino sugar and ergosterol contents after addition of sucrose and cellulose to soil

An incubation experiment with organic soil amendments was carried out with the aim to determine whether formation and use of microbial tissue (biomass and residues) could be monitored by measuring glucosamine and muramic acid. Living fungal tissue was additionally determined by the cell-membrane component ergosterol. The organic amendments were fibrous maize cellulose and sugarcane sucrose adjusted to the same C/N ratio of 15. In a subsequent step, spherical cellulose was added without N to determine whether the microbial residues formed initially were preferentially decomposed. In the non-amended control treatment, ergosterol remained constant at 0.44 μg g⁻¹ soil throughout the 67-day incubation. It increased to a highest value of 1.9 μg g⁻¹ soil at day 5 in the sucrose treatment and to 5.0 μg g⁻¹ soil at day 33 in the fibrous cellulose treatment. Then, the ergosterol content declined again. The addition of spherical cellulose had no further significant effects on the ergosterol content in these two treatments. The non-amended control treatment contained 48 μg muramic acid and 650 μg glucosamine g⁻¹ soil at day 5. During incubation, these contents decreased by 17% and 19%, respectively. A 33% increase in muramic acid and an 8% increase in glucosamine were observed after adding sucrose. Consequently, the ratio of fungal C to bacterial C based on bacterial muramic acid and fungal glucosamine was lowered in comparison with the other two treatments. No effect on the two amino sugars was observed after adding cellulose initially or subsequently during the second incubation period. This indicates that the differences in quality between sucrose and cellulose had a strong impact on the formation of microbial residues. However, the amino sugars did not indicate a preferential decomposition of microbial residues as N sources.     

Variation in oxalic acid production and mycelial compatibility within field populations of Sclerotinia sclerotiorum

This study was aimed at detecting mycelial compatibility groups and variations in oxalic acid production in Sclerotinia sclerotiorum. For this purpose, 121 isolates of this plant pathogen recovered from lettuce, soybean and sunflower field crops, and grouped in 46 MCGs were tested for their ability to release oxalic acid and other organic acids to the medium. Oxalic acid production on liquid media was measured spectrophotometrically and release of organic acids was estimated by isolate abilities to discolour solid media amended with bromophenol blue. There were significant differences among MCGs in both oxalic acid and organic acids releasing, ranging the mean production of oxalic acid between 18 and 110 μg oxalic acid mg⁻¹ dry wt. When isolates were grouped by their hosts, those obtained from soybean presented the highest release of oxalic acid (71 μg oxalic acid mg⁻¹ dry wt), while those from sunflower showed the highest release of other acids to the medium. Solid medium discoloration was not correlated with oxalic acid concentration in liquid medium (Spearman R=−0.085; P=0.126).     

Laccase and peroxidase isoenzymes during leaf litter decomposition of Quercus ilex in a Mediterranean ecosystem

The dynamics of leaf litter decomposition of Quercus ilex (L.) were investigated over a 2 year period by determining the activities and isoenzyme distribution of laccases and peroxidases. The analysis of isoenzymes was performed by isoelectric focusing on high stability pH gradients with high resolving power. The preparation of zymograms was carried out using the leaf litter extract without previous concentration. During litter decomposition, laccase and peroxidase activities changed as well as the type and number of enzyme isoforms. The activities of both enzymes were low (≤0.017 and ≤0.031 mmol o-tolidine oxidized h⁻¹ g⁻¹ d.w. for laccase and peroxidase, respectively) in first year and increased in October-January of the second year of litter decay. The highest activities measured after 15-18 months of litter exposure (0.37±0.03 and 0.19±0.02 mmol o-tolidine oxidized h⁻¹ g⁻¹ d.w. for laccase and peroxidase, respectively), showed that litter chemical composition affected the growth of ligninolytic microbial community. The activation energy for laccase and peroxidase reactions also changed during decomposition: the highest values (55±6 kJ mol⁻¹ for laccase and 60±6 kJ mol⁻¹ for peroxidase) occurred in autumn-winter, even if spatial changes were evidenced. Some enzyme isoforms (pI=5.3 and 5.5 for laccase and pI=5.0 and 5.1 for peroxidase, respectively), contributed more than others to the overall laccase and peroxidase activity, suggesting that some ligninolytic species bloomed in particular seasons of the year, even if other species with similar functional activities colonized the litter.