Mass spectrometric characterization of amino sugar aldononitrile acetate derivatives used for isotope enrichment assessment of microbial residues

Identifying the transformation of amino sugars in soils is essential in understanding microbial contribution to soil organic matter turnover and sequestration. Using a recently developed method, combining gas chromatograph/mass spectrometry (GC/MS) with laboratory incubation of substrates containing ¹³C or ¹⁵N isotopes, we were able to trace isotopic changes in amino sugar compounds. This allows us to quantitatively evaluate C or N enrichment in amino sugars during transformation in soils using the fragment (F) abundance ratio of m/z F + n to F (n is original skeleton atom number in each fragment). However, there is still lack of detail structural and substitutional information for each ion fragment. In order to improve the interpretation and increase our ability to study amino sugar turnover, we grew labeled amino sugars in lab-cultured organisms. We spectrometrically investigated the ion structures and original skeleton C number (mass shift n) in major ion fragments based on applying multiple representative isotope labels. Our results categorically confirm that previously made assumptions were correct regarding the substitutional number “n” of the glucosamine (He et al., 2006). Our study also added valuable structural information for aldononitrile acetate derivatized glucosamine and muramic acid upon electron impact ionization in MS.     

Carbohydrate and amino acid composition of dissolved organic matter leached from soil

Low molecular weight organic substances (LMWOS) in soil and soil solution include mainly amino acids, carboxylic acids, and carbohydrates. Due to their high bioavailability they play a crucial role in the cycles of C and nutrients in soils. The variety of soil processes that involve LMWOS requires identifying their composition to elucidate reactions and transformations. In most studies, LMWOS are extracted under artificial conditions, e.g. batch experiments, which may overestimate the actual concentrations. This study measures the composition of carbohydrates and amino acids in solution of a Haplic Luvisol leached in a column experiment. A combined system for simultaneous leaching and blowout of CO₂ was used to estimate LMWOS decomposition. ¹⁴C-labeled glucose was added as a highly sensitive tracer to control the efficiency of the LMWOS extraction by leaching and to estimate LMWOS decomposition during leaching. High performance liquid chromatography (HPLC), optimized for soil extracts, was used to analyze LMWOS composition. For HPLC optimization, different preparations of leached solutions (filtration vs. centrifugation, and drying vs. no-drying) were compared. For sugar determination, drying had no influence on the solution concentrations. In contrast, amino acid concentrations significantly decreased by drying LMWOS eluted substances. Combining the HPLC identification of eluted substances with ¹⁴C tracer application revealed that about 5% of the glucose could be leached unchanged within 786 min (13.1 h), whereas about 84% remained in the soil, 9% were decomposed to CO₂, and 2% were transformed to other LMWOS and recovered in the soil solution. The total amino acid concentration (TAC) in soil solution was about 8.2 μmol l⁻¹, dominated by alanine (14.4% of TAC), glycine (13.4%), glutamic acid (9.9%), serine (9.4%), and leucine (9.3%). The total carbohydrate concentration was about 2.4 μM, dominated by glucose (29.9%), glucuronic acid (26.8%), and galacturonic acid (17.3%). Ratios of hexoses to pentoses, amino sugars glucosamine to galactosamine, and neutral sugars to uronic acids were determined. All three parameters pointed to the dominant influence of plants as the source of LMWOS in the leached soil solution. Within the small contribution of microorganisms, bacteria dominated over fungi. These used biomarker ratios as well as LMWOS concentrations differed widely from the ones obtained with conventional batch extraction. More research is necessary to evaluate the application of these biomarkers to soil solutions.     

Amino sugars and muramic acid—biomarkers for soil microbial community structure analysis

Characterizing functional and phylogenetic microbial community structure in soil is important for understanding the fate of microbially-derived compounds during the decomposition and turn-over of soil organic matter. This study was conducted to test whether amino sugars and muramic acid are suitable biomarkers to trace bacterial, fungal, and actinomycetal residues in soil. For this aim, we investigated the pattern, amounts, and dynamics of three amino sugars (glucosamine, mannosamine and galactosamine) and muramic acid in the total microbial biomass and selectively cultivated bacteria, fungi, and actinomycetes of five different soils amended with and without glucose. Our results revealed that total amino sugar and muramic acid concentrations in microbial biomass, extracted from soil after chloroform fumigation varied between 1 and 27 mg kg⁻¹ soil. In all soils investigated, glucose addition resulted in a 50-360% increase of these values. In reference to soil microbial biomass-C, the total amino sugar- and muramic acid-C concentrations ranged from 1-71 g C kg⁻¹ biomass-C. After an initial lag phase, the cultivated microbes revealed similar amino sugar concentrations of about 35, 27 and 17 g glucosamine-C kg⁻¹ TOC in bacteria, fungi, and actinomycetes, respectively. Mannosamine and galactosamine concentrations were lower than those for glucosamine. Mannosamine was not found in actinomycete cultures. The highest muramic acid concentrations were found in bacteria, but small amounts were also found in actinomycete cultures. The concentrations of the three amino sugars studied and muramic acid differed significantly between bacteria and the other phylogenetic microbial groups under investigation (fungi and actinomycetes). Comparison between the amino sugar and muramic acid concentrations in soil microbial biomass, extracted after chloroform fumigation, and total concentrations in the soil showed that living microbial biomass contributed negligible amounts to total amino sugar contents in the soil, being at least two orders of magnitude greater in the soils than in the soil inherent microbial biomass. Thus, amino sugars are significantly stabilized in soil.     

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.     

Carbohydrate dynamics in particle-size fractions of sandy spodosols following forest conversion to maize cropping

In southwest France, much of the forested land on sandy spodosols has been converted to continuous maize cropping in the last few decades. To evaluate the impacts of this change on soil organic matter properties, we compared total organic C and neutral and amino sugars content in whole soil and particle size separates of two forested, and five related sites that had been either clear-cut for 12 and 18 mo, or cultivated to maize for 4-22 yr. Soil carbohydrates accounted for 4-7% of the total organic C across all sites. Soil organic C contents of clear-cut and cultivated sites were only 57-79% of the average value measured in forested sites. Accordingly, carbohydrate content of clear-cut and cultivated sites were only 35-66% of the value in forested sites. Ordering the sites in a chronosequence indicated that both total organic C and carbohydrate contents decreased with an increase in time elapsed since clear-cutting and maize cultivation. The only exception was a partial recovery of carbohydrate content in the site that had been under continuous maize for 22 yr. The clay+silt fraction (0-50 μm) was enriched in carbohydrates, mainly of microbial origin, whereas the sand size fractions (50-200 and 200-2000 μm) contained fewer carbohydrates which were mainly of plant origin. Monosaccharide analysis of particle size separates revealed significant differences in carbohydrate composition between sites. Relative to forested sites, the coarse and fine sand fractions in clear-cut and cultivated sites were depleted in carbohydrates and were relatively enriched in plant-derived carbohydrates. Carbohydrate content of the clay+silt fraction drastically decreased upon clear-cutting. Amino sugar content was consistently lower in clear-cut and cultivated sites than in forested sites, indicating that microbial populations were negatively affected by clear-cutting and cultivation. The fungal population appeared more sensitive than bacteria to these land-use changes as indicated by a greater decline in glucosamine than in muramic acid contents.     

Dynamics of the nitrous oxide reducing community during adaptation to Zn stress in soil

Laboratory studies show that the nitrous oxide (N₂O) reduction rate in soil is strongly inhibited by trace metal contamination; however, this effect appears transient. Here we assess if this recovery is due to microbial adaptation associated with shifts in community composition. Soils were spiked with zinc chloride (0-5000 mg Zn kg⁻¹) in a factorial design with 3 application rates of organic matter (OM), i.e. 0, 2 and 4 g milled hay kg⁻¹, to accelerate growth and, potentially, adaptation rate. The soil treatments were incubated outdoors with free drainage during 1 year and periodically sampled. The potential N₂O reduction rate, measured in an anaerobic laboratory assay, was inhibited by Zn during the first 2 months after spiking with 50% inhibition at 500-1000 mg Zn kg⁻¹. After 6 months exposure, the N₂O reduction rate recovered to at least 80% of the rate in the control treatment in the series receiving OM up to the largest Zn dose, but strong inhibition remained in the series which did not receive OM. In this series recovery was only observed after 12 months exposure. Soil pore water Zn concentrations did not explain the recovery of the N₂O reduction rate in the control series suggesting that recovery is due to adaptation and not to reduced Zn bioavailability. The faster recovery in the series receiving OM was partially, but not fully related to the effects of OM on Zn bioavailability. The recovery at all Zn and OM treatments co-varied with a recovery of nosZ gene abundance from about 1 × 10⁷ copies g⁻¹ soil in the soil treatments with decreased activity to 5 × 10⁸ copies g⁻¹ soil in the other soil treatments. The nosZ gene DGGE profile of the soil microbial communities revealed minor changes in the nosZ containing community. This study strongly suggests that the transient effects of trace metal inhibition of N₂O reduction is due to the development of a Zn tolerant denitrifying community.     

Surface runoff phosphorus (P) loss in relation to phosphatase activity and soil P fractions in Florida sandy soils under citrus production

Phosphorus losses by surface runoff from agricultural lands have been of public concern due to increasing P contamination to surface waters. Five representative commercial citrus groves (C1-C5) located in South Florida were studied to evaluate the relationships between P fractions in soils, surface runoff P, and soil phosphatase activity. A modified Hedley P sequential fractionation procedure was employed to fractionate soil P. Soil P consisted of mainly organically- and Ca/Mg-bound P fractions. The organically-bound P (biological P, sum of organic P in the water, NaHCO₃ and NaOH extracts) was dominant in the acidic sandy soils from the C2 and C3 sites (18% and 24% of total soil P), whereas the Ca/Mg-bound P (HCl-extractable P) accounted for 45-60% of soil total P in the neutral and alkaline soils (C1, C4 and C5 soils). Plant-available P (sum of water and NaHCO₃ extractable P fractions) ranged from 27 to 61 mg P kg⁻¹ and decreased in the order of C3>C4>C1>C2>C5. The mean total P concentrations (TP) in surface runoff water samples ranged from 0.51 to 2.64 mg L⁻¹. Total P, total dissolved P (TDP), and PO₄³⁻-P in surface runoff were significantly correlated with soil biological P and plant-available P forms (p<0.01), suggesting that surface runoff P was directly derived from soil available P pools, including H₂O- and NaHCO₃- extractable inorganic P, water-soluble organic P, and NaHCO₃- and NaOH-extractable organic P fractions, which are readily mineralized by soil microorganisms and/or enzyme mediated processes. Soil neutral (55-190 mg phenol kg⁻¹ 3 h⁻¹) and natural (measured at soil pH) phosphatase activities (77-295 mg phenol kg⁻¹ 3 h⁻¹) were related to TP, TDP, and PO₄³⁻-P in surface runoff, and plant-available P and biological P forms in soils. These results indicate that there is a potential relationship between soil P availability and phosphatase activities, relating to P loss by surface runoff. Therefore, the neutral and natural phosphatase activities, especially the natural phosphatase activity, may serve as an index of surface runoff P loss potential and soil P availability.     

Modification and application of a soil ATP determination method

Accurate estimation of microbial adenosine 5′-triphosphate (ATP) is a pre-requisite to quantify the impact of varying environments on microbial activity of soil. We investigated the effectiveness of a high efficiency soil ATP determination method (PA) [Webster, J.J., Hampton, G.J., Leach, F.R., 1984. ATP in soil: a new extractant and extraction procedure. Soil Biology & Biochemistry 4, 335-342] in 10 Ontario (Canada) soils collected along a 100 m transect and spanning a textural class gradient ranging from a sandy loam to clay loam with increasing organic matter. Modifications of the method involved using an extract of autoclaved soil to make the standard curve, as it was found that the light emitted by ATP luciferin-luciferase bioluminescence reaction in the pure extractant was different from that in the extracts. Replacing Tricine with Tris buffer in the assay significantly improved the light emission. On an average, the internal standard calibration method (ISM) measured a smaller amount of extracted ATP (1199 ng ATP g⁻¹ soil) and a lower recovery of ATP spike (82.4±7.2%) than did the standard curve method (SCM) (1246 ng ATP g⁻¹ soil and 91.2±4.5%, respectively) (P<0.05 for both comparisons). However, the average total estimated ATP was higher with ISM (1474±102 ng ATP g⁻¹ soil) than with SCM (1373±88 ng ATP g⁻¹ soil) (P<0.07). While the recovery rates determined using SCM were consistent among the soils tested, the rates measured using ISM was negatively correlated with soil clay and organic matter content, implying that the latter assay was affected by the soil properties. Our results confirmed that the recovery rates obtained by the PA method were the highest among those reported, when only SCM was used.     

Long-term accumulation and material balance of organic matter in the soil of an effluent infiltration basin

The fate of organic matter (OM) in large-scale infiltration basins used for wastewater treatment by the soil aquifer treatment (SAT) system was investigated. Measured changes in the organic matter concentrations in the soil profiles of the infiltration basins and detailed long-term records of OM concentrations in the recharged effluent and in the observation wells and recovery wells water, were used to calculate OM material balances in the SHAFDAN wastewater treatment plant, serving the City of Tel-Aviv, Israel, since 1977. The average annual total organic matter (TOM) load delivered by the effluents to the soil was ~ 5 kg m^− 2 y^− 1. Soil OM concentrations increased from 0.11% in the pristine soil to ~ 0.8% and ~ 0.6%, in the 0-0.15 m and 0.15-0.30 m soil layers, respectively, after ~ 20 y of effluent recharge, but did not change significantly in the 1.80-2.10 m deep layer. The OM accumulation rates in the top two soil layers were fast initially, then declined slowly and the OM concentrations approached a steady state following 10-15 y of effluent recharge. This suggests that stabilization of the ‘active biofilm’ layer in the infiltration basins' soils is a relatively slow process. Material-balance calculations showed, that accumulated OM in the top 0-2.1 m soil layer amounted to only ~ 4% of the TOM added by the effluents during ~ 20 y of recharge. Along the flow pathway of the effluent through the vertical 50-100 m thick soil-sediment column, DOC concentrations decreased by 70-90% (from ~ 18.9 mg L^− 1 to ~ 3.7 mg L^− 1). Continued flow in the aquifer from the observation wells to the recovery wells further decreased DOC concentrations by about 50% (from ~ 3.7 to ~ 1.5 mg L^− 1).     

Fate of low molecular weight organic substances in an arable soil: From microbial uptake to utilisation and stabilisation

Microbial uptake and utilisation are the main transformation pathways of low molecular weight organic substances (LMWOS) in soil, but details on transformations are strongly limited. As various LMWOS classes enter biochemical cycles at different steps, we hypothesize that the percentage of their carbon (C) incorporation into microbial biomass and consequently stabilisation in soil are different.Representatives of the three main groups of LMWOS: amino acids (alanine, glutamate), sugars (glucose, ribose) and carboxylic acids (acetate, palmitate) – were applied at naturally-occurring concentrations into a loamy arable Luvisol in a field experiment. Incorporation of ¹³C from these LMWOS into extractable microbial biomass (EMB) and into phospholipid fatty acids (PLFAs) was investigated 3 d and 10 d after application. The microbial utilisation of LMWOS for cell membrane construction was estimated by replacement of PLFA-C with ¹³C.35–80% of initially applied LMWOS-¹³C was still present in the composition of soil organic matter after 10 days of experiment, with 10–24% of ¹³C incorporation into EMB at day three and 1–15% at day 10. Maximal incorporation of ¹³C into EMB was observed from sugars and the least from amino acids. Strong differences in microbial utilisation between LMWOS were observed mainly at day 10. Thus, despite similar initial rapid uptake by microorganisms, further metabolism within microbial cells accounts for the specific fate of C from various LMWOS in soils.¹³C from each LMWOS was incorporated into each PLFA. This reflects the ubiquitous utilisation of all LMWOS by all functional microbial groups. The preferential incorporation of palmitate into PLFAs reflects its role as a direct precursor for fatty acids. Higher ¹³C incorporation from alanine and glucose into specific PLFAs compared to glutamate, ribose and acetate reflects the preferential use of glycolysis-derived substances in the fatty acids synthesis.Gram-negative bacteria (16:1ω7c and 18:1ω7c) were the most abundant and active in LMWOS utilisation. Their high activity corresponds to a high demand for anabolic products, e.g. to dominance of pentose-phosphate pathway, i.e. incorporation of ribose-C into PLFAs. The ¹³C incorporation from sugars and amino acids into filamentous microorganisms was lower than into all prokaryotic groups. However, for carboxylic acids, the incorporation was in the same range (0.1–0.2% of the applied carboxylic acid ¹³C) as that of gram-positive bacteria. This may reflect the dominance of fungi and other filamentous microorganisms for utilisation of acidic and complex organics.Thus, we showed that despite similar initial uptake, C from individual LMWOS follows deviating metabolic pathways which accounts for the individual fate of LMWOS-C over 10 days. Consequently, stabilisation of C in soil is mainly connected with its incorporation into microbial compounds of various stability and not with its initial microbial uptake.     

Rice root-derived carbon input and its effect on decomposition of old soil carbon pool under elevated CO2

Rice (Oryza sativa) was grown in sunlit, semi-closed growth chambers (4×3×2 m, L×W×H) at 650 μl l⁻¹ CO₂ (elevated CO₂) to determine: (1) rice root-derived carbon (C) input into the soil under elevated CO₂ in one growing season, and (2) the effect of the newly input C on decomposition of the more recalcitrant native soil organic C. The initial δ¹³C value of the experimental soil was −25.8‰, which was 6‰ less depleted in ¹³C than the plants grown under elevated CO₂. Significant changes in δ¹³C of the soil organic C were detected after one growing season. The amount of new soil C input was estimated to be 0.9 t ha⁻¹ (or 2.1%) at 30 kg N ha⁻¹ and 1.8 t ha⁻¹ (4.1%) at 90 kg N ha⁻¹. Changes in soil δ¹³C suggested that the surface 5 cm of soil received more C input from plants than soils below. Laboratory incubation (25 °C) of soils from different horizons indicated that increased availability of the labile plant-derived C in the soil reduced decomposition of the native soil organic C. Provided the retardant effect of the new C on old soil organic C holds in the field in the longer-term, paddy soils will likely sequester more C from the atmosphere if more plant C enters the soil under elevated atmospheric CO₂.     

Sulphur immobilization and arylsulphatase activity in two calcareous arable and fallow soils as affected by glucose additions

In this study we examined the effects of glucose-C on the activities of fungi and bacteria determined by the method of substrate-induced respiration (SIR) in combination with the selective inhibition technique, the immobilized-S and the arylsulphatase (ARS) activity in two calcareous arable and fallow soils. The amounts of glucose-C were added at six doses: 0, 125, 250, 500, 750 and 1000 mg kg^− 1 soil to the soils and then incubated for one week with a Na₂³⁵SO₄ solution (518.9 kBq kg^− 1 dry soil and 20 mg S kg^− 1 dry soil) prior to analysis. At the highest dose of 1000 mg kg^− 1 soil, fungal activity increased by 59.1% (of the dose 0) versus 45.5% for bacterial activity in the arable soil, while in the fallow soil the increases were more marked and corresponded to 69.9% and 71.1%, respectively. Largest increase in immobilized-S was observed in the arable soil (300.7%) compared with the fallow soil (153.1%). In contrast, the ARS activity increased by 16.4% in the arable soil versus 32.1% in the fallow soil. These results indicate that glucose proportionately affected more the intensities of immobilized-S than those of ARS. Strong positive correlation coefficients were found between fungal activities and immobilized-S in the arable soil (r = 0.96, P < 0.01) and in the fallow soil (r = 0.98, P < 0.001). However, non-significant correlations were observed between fungal activities and ARS in both studied soils. As to bacterial activities, positive significant correlation coefficients were found with immobilized-S in the arable soil (r = 0.95, P < 0.01) and in the fallow soil (r = 0.90, P < 0.05) as well as with ARS activities in the arable soil (r = 0.83, P < 0.05) and in the fallow soil (r = 0.97, P < 0.01). Overall, we also found positive and significant correlation coefficients of immobilized-S with ARS activities in the arable soil (r = 0.86, P < 0.05) and in the fallow soil (r = 0.83, P < 0.05). Accordingly, the results showed a presence of extracellular arylsulphatase activity of 38.7 mg p-nitrophenol kg^− 1 soil h^− 1 in the arable soil and of 63.5 mg p-nitrophenol kg^− 1 soil h^− 1 in the fallow soil. It was concluded that fallowing maintained larger activities of fungi, bacteria and arylsulphatase compared with the arable soil.     

Changes in soil biological and biochemical characteristics in a long-term field trial on a sub-tropical inceptisol

Soil organic carbon (SOC), microbial biomass carbon (MBC), their ratio (MBC/SOC) which is also known as microbial quotient, soil respiration, dehydrogenase and phosphatase activities were evaluated in a long-term (31 years) field experiment involving fertility treatments (manure and inorganic fertilizers) and a maize (Zea mays L.)-wheat (Triticum aestivum L.)-cowpea (Vigna unguiculata L.) rotation at the Indian Agricultural Research Institute near New Delhi, India. Applying farmyard manure (FYM) plus NPK fertilizer significantly increased SOC (4.5-7.5 g kg⁻¹), microbial biomass (124-291 mg kg⁻¹) and microbial quotient from 2.88 to 3.87. Soil respiration, dehydrogenase and phosphatase activities were also increased by FYM applications. The MBC response to FYM+100% NPK compared to 100% NPK (193 vs. 291 mg kg⁻¹) was much greater than that for soil respiration (6.24 vs. 6.93 μl O₂ g⁻¹ h⁻¹) indicating a considerable portion of MBC in FYM plots was inactive. Dehydrogenase activity increased slightly as NPK rates were increased from 50% to 100%, but excessive fertilization (150% NPK) decreased it. Acid phosphatase activity (31.1 vs. 51.8 μg PNP g⁻¹ h⁻¹) was much lower than alkali phosphatase activity (289 vs. 366 μg PNP g⁻¹ h⁻¹) in all treatments. Phosphatase activity was influenced more by season or crop (e.g. tilling wheat residue) than fertilizer treatment, although both MBC and phosphatase activity were increased with optimum or balanced fertilization. SOC, MBC, soil respiration and acid phosphatase activity in control (no NPK, no manure) treatment was lower than uncultivated reference soil, and soil respiration was limiting at N alone or NP alone treatments.     

Concentration of sugars, phenolic acids, and amino acids in forest soils exposed to elevated atmospheric CO2 and O3

Concentrations of soluble soil sugars, soluble phenolic acids, and free amino acids were measured in three forest communities at the FACTS-II Aspen FACE Site near Rhinelander, WI, in order to better understand how elevated atmospheric CO₂ and O₃ are influencing soil nutrient availability and cycling. Sugars, phenolic acids, and amino acids are mostly derived from plant and microbial processes, and have the potential to be influenced by changes in carbon inputs. We hypothesized that concentrations in the soil would parallel increases seen in biological activity, due to greater net primary productivity under elevated CO₂ and seasonal patterns of root growth. Chemical analysis of soils revealed marginally significant increases of total soluble sugars and total soluble phenolic acids in the elevated CO₂ treatment (+27 mg kg⁻¹, +0.02 μmol g⁻¹), but there were no significant differences in concentrations due to elevated O₃ or CO₂+O₃. Total free amino acid concentrations were not affected by any of the treatments, but significant shifts in individual amino acids were observed. Elevated CO₂ and the interaction treatment (elevated CO₂+O₃) increased aspartic acid concentrations, while elevated O₃ treatment decreased the concentration of valine. Concentrations of sugars increased throughout the growing season, while phenolic acids were constant and amino acids decreased. The birch-aspen community had the highest concentration of phenolic acids and sugars overall, while maple-aspen had the lowest. These findings suggest that concentrations of soluble sugars, soluble phenolic acids, and free amino acids in the soil are strongly influenced by soil properties, plant and microbial activity, plant community composition, and to a lesser degree, changes in atmospheric CO₂ and O₃.     

Enzyme activities and microbial biomass in coastal soils of India

Soil salinity is a serious problem for agriculture in coastal regions, wherein salinity is temporal in nature. We studied the effect of salinity, in summer, monsoon and winter seasons, on microbial biomass carbon (MBC) and enzyme activities (EAs) of the salt-affected soils of the coastal region of the Bay of Bengal, Sundarbans, India. The average pH of soils collected from different sites, during different seasons varied from 4.8 to 7.8. The average organic C (OC) and total N (TN) content of the soils ranged between 5.2-14.1 and 0.6-1.4 g kg⁻¹, respectively. The electrical conductivity of the saturation extract (ECe) of soils, averaged over season, varied from 2.2 to 16.3 dSm⁻¹. The ECe of the soils increased five fold during the summer season (13.8 dSm⁻¹) than the monsoon season (2.7 dSm⁻¹). The major cation and anion detected were Na⁺ and Cl⁻, respectively. Seasonality exerted considerable effects on MBC and soil EAs, with the lowest values recorded during the summer season. The activities of β-glucosidase, urease, acid phosphatase and alkaline phosphatase were similar during the winter and monsoon season. The dehydrogenase activity of soils was higher in monsoon than in winter. Average MBC, dehydrogenase, β-glucosidase, urease, acid phosphatase and alkaline phosphatase activities of the saline soils ranged from 125 to 346 mg kg⁻¹ oven dry soil, 6-9.9 mg triphenyl formazan (TPF) kg⁻¹ oven dry soil h⁻¹, 18-53 mg p-nitro phenol (PNP) kg⁻¹ oven dry soil h⁻¹, 38-86 mg urea hydrolyzed kg⁻¹ oven dry soil h⁻¹, 213-584 mg PNP kg⁻¹ oven dry soil h⁻¹ and 176-362 mg PNP g⁻¹ oven dry soil h⁻¹, respectively. The same for the non-saline soils were 274-446 mg kg⁻¹ oven dry soil, 8.8-14.4 mg TPF kg⁻¹ oven dry soil h⁻¹, 41-80 mg PNP kg⁻¹ oven dry soil h⁻¹, 89-134 mg urea hydrolyzed kg⁻¹ oven dry soil h⁻¹, 219-287 mg PNP kg⁻¹ oven dry soil h⁻¹ and 407-417 mg PNP kg⁻¹ oven dry soil h⁻¹, respectively. About 48%, 82%, 48%, 63%, 40% and 48% variation in MBC, dehydrogenase activity, β-glucosidase activity, urease activity, acid phosphatase activity and alkaline phosphatase activity, respectively, could be explained by the variation in ECe of saline soils. Suppression of EAs of the coastal soils during summer due to salinity rise is of immense agronomic significance and needs suitable interventions for sustainable crop production.     

Belowground carbon allocation patterns in a dry Mediterranean ecosystem: A comparison of two models

Total belowground C allocation (TBCA) accounts for a large fraction of gross primary production, it may overtake aboveground net primary production, and contributes to the primary source of detrital C in the mineral soil. Here, we measure soil respiration, water erosion, litterfall and estimate annual changes in C stored in mineral soil, litter and roots, in three representative land uses in a Mediterranean ecosystem (late-successional forest, abandoned agricultural field, rain-fed olive grove), and use two C balance approaches (steady-state and non-steady-state) to estimate TBCA. Both TBCA approaches are compared to assess how different C fluxes (outputs and inputs) affect our estimates of TBCA within each land use. In addition, annual net primary productivity is determined and C allocation patterns are examined for each land use. We hypothesized that changes in C stored in mineral soil, litter and roots will be slight compared to soil respiration, but will still have a significant effect on the estimates of TBCA. Annual net primary productivity was 648 ± 31.5, 541 ± 42.3 and 324 ± 22.3 g C m⁻² yr⁻¹ for forest, abandoned agricultural field and olive grove, respectively. Across land uses, more than 60% of the C was allocated belowground. Soil respiration (F_S) was the largest component in the TBCA approaches across all land uses. Annual C losses through water erosion were negligible compared to F_S (less than 1%) and had little effect on the estimates of TBCA. Annual changes in C stored in the soil, litter layer and roots were low compared to F_S (16, 24 and 10% for forest, abandoned agricultural field and olive grove, respectively), but had a significant effect on the estimates of TBCA. In our sites, an assumption that Δ[C_S + C_R + C_L]/Δt = 0 will underestimate TBCA, particularly in the abandoned agricultural field, where soil C storage may be increasing more rapidly. Therefore, the steady-state model is unsuited to these Mediterranean ecosystems and the full model is recommended.     

8-羟基喹啉(pH8.3)分光光度法测定酸性土壤中的可溶性铝

8-羟基喹啉(pH8.3)分光光度法对酸性土壤中的可溶性铝测定的研究表明,Al标准曲线的线性范围为0 ~ 50 mmol/L,相关系数R2 = 0.9999,表观摩尔吸光系数为1.995 104 L/(mol·cm),回收率在98.7 % ~ 106.8 %之间,该方法重现性好,选择性高。     

Kinetics of microbial growth and degradation of organic substrates in subsoil as affected by an inhibitor, benzotriazole: Model based analyses of experimental results

Deep transport of degradable compounds through soils may occur if the metabolic activity in the soil profile is low; either by natural causes (low temperature during ice melt) or by toxic pollutants. De-icing chemicals (for roads and airports) represents a severe challenge to the soil's purifying capacity; rapid infiltration of contaminated water occurs in near-frozen soil, the contamination includes toxic compounds. Degradation experiments were conducted with toluene, and three compounds frequently used for de-icing; acetate, formate and glycol. The substrates were added to a subsoil (0.93 μmol substrate-carbon (C) g⁻¹ soil, with ample amounts of nitrogen (N) and phosphorous (P)); and their mineralization was monitored in the presence of a toxic compound, benzotriazol (BTA) at various concentrations. BTA is commonly used as an additive in commercial de-icing fluids. A second and third dose of substrate was added after complete degradation of the previous one. The mineralization curves of the three consecutive doses were used to estimate kinetic parameters by fitting to a Monod-model. The model parameters estimated for each substance were the initial biomass C of the organisms growing on each substrate, C_b0, their maximum substrate uptake rates, V_max, their apparent substrate affinity, K_S, and growth yield, Y. The C_b0 values for pristine soil were 4.9, 20.5 and 10 nmol C g⁻¹ soil for formate, glycol and acetate, respectively, and 1-2 orders of magnitude lower for toluene. The K_S values were 1.1, 0.6, 2.5 and 0.13 mM for formate, glycol, acetate, and toluene, respectively. The high K_S values probably reflect diffusion limitations. The estimated yields (Y) in the absence of BTA were 0.032, 0.53 and 0.42 g biomass-C g⁻¹ substrate-C for growth on formate, glycol and acetate, respectively. BTA invariably reduced the growth yield for organisms growing on the different substrates, and the yield reduction increased with increasing BTA concentrations (more than 50% reduction at 400 mg BTA l⁻¹). The degradation of the four substrates showed major differences in BTA-sensitivity, and there were strikingly weak signs (if any) of increasing BTA tolerance during growth in the presence of BTA (analyses of second and third dose experiments). The modelling of the consecutive substrate doses corroborated previous investigations of BTA effects on mineralization and community PLFA [Jia et al., 2006. Organic compounds that reach subsoil may threaten groundwater quality; effect of benzotriazole on degradation kinetics and microbial community composition. Soil Biology & Biochemistry 38, 2543-2556]. The results and the estimated Monod parameters are useful for predictive modelling of transport and degradation of pollutants as well as natural substances in sub-soils.     

Microbial biomass and activity at various soil depths in a Brazilian oxisol after two decades of no-tillage and conventional tillage

The advantages of no-tillage (NT) over conventional tillage (CT) systems in improving soil quality are generally accepted, resulting from benefits in soil physical, chemical and biological properties. However, most evaluations have only considered surface soil layers (maximum 0-30 cm depth), and values have not been corrected to account for changes in soil bulk density. The objective of this study was to estimate a more realistic contribution of the NT to soil fertility, by evaluating C- and N-related soil parameters at the 0-60 cm depth in a 20-year experiment established on an oxisol in southern Brazil, with a soybean (summer)/wheat (winter) crop succession under NT and CT. At full flowering of the soybean crop, soil samples were collected at depths of 0-5, 5-10, 10-20, 20-30, 30-40, 40-50 and 50-60 cm. For the overall 0-60 cm layer, correcting the values for soil bulk density, NT significantly increased the stocks of C (18%) and N (16%) and microbial biomass C (35%) and N (23%) (MB-C and -N) in comparison to CT. Microbial basal respiration and microbial quotient (qMic) were also significantly increased under NT. When compared with CT, NT resulted in gains of 0.8 Mg C ha⁻¹ yr⁻¹ (67% of which was in the 0-30 cm layer) and 70 kg N ha⁻¹ yr⁻¹ (73% in the 0-30 cm layer). In the 0-5-cm layer, MB-C was 82% higher with NT than with CT; in addition, the 0-30 cm layer accumulated 70% of the MB-C with NT, and 58% with CT. In comparison to CT, the NT system resulted in total inputs of microbial C and N estimated at 38 kg C ha⁻¹ yr⁻¹ and 1.5 kg N ha⁻¹ yr⁻¹, respectively. Apparently, N was the key nutrient limiting C and N stocks, and since adoption of NT resulted in a significant increase of N in soils which were deficient in N, efforts should be focused on increasing N inputs on NT systems.     

Soil C and N pools in patchy shrublands of the Negev and Chihuahuan Deserts

Patchy distribution of vegetation within semi-arid shrublands is normally mirrored in the soil beneath perennial shrubs (macrophytic patches), compared to inter-shrub areas (microphytic patches). To determine impacts of (1) litterfall inputs within vegetation patches and (2) rainfall distribution on soil C and N, we investigated soil C and N pools and associated soil properties in two semi-arid shrublands, in the Negev Desert of Israel (Lehavim), which receives >90% of annual rainfall during winter and in the Chihuahuan Desert, USA (FHMR) that experiences a bimodal (Summer-Winter) annual rainfall pattern. We also evaluated grazing effects on soil C and N pools at Lehavim. More distinct differences in soil properties existed between patch types at the Negev site, where the soils contained higher soil organic C and N, amino acids and sugars, asparaginase activity and plant-available N than those at FHMR. Soil organic C (0-5 cm) in macrophytic patches was 39 g/kg at Lehavim and 13 g/kg at FHMR, and asparaginase activity was as high as 70 μg N/g 2 h in macrophytic patches at Lehavim, two times higher than at FHMR. The soil (0-5 cm) δ¹³C was −15 to −18‰ at Lehavim and −18 to −19‰ at FHMR, with significantly lower δ¹³C in macrophytic patches at both sites. The δ¹³C suggested that considerable macrophytic patch soil C was derived from cyanobacteria at Lehavim and C4 grasses at FHMR. Plant litter δ¹⁵N was 0.9‰ at Lehavim and 0.6‰ at FHMR, suggesting that much plant N was derived from N fixation. Concentrations of inorganic soil N (NH₄⁺+NO₃⁻) were up to 37 mg N/kg at Lehavim and <9 mg N/kg at FHMR. Grazing at Lehavim resulted in lower soil CH, AA, and AS. We conclude that differences between the sites are due largely to (i) higher amounts of litterfall C and N inputs within macrophytic patches at Lehavim and (ii) the different precipitation patterns, with summer precipitation at FHMR promoting increased organic matter mineralization compared to Lehavim, which experiences Winter precipitation only. Furthermore, greater differences in soil properties between patch types at Lehavim compared to FHMR can likely be attributed to the increasing importance of physical processes of resource dispersion at the more humid site in Arizona.