Adenylate energy charge measurements in soil

Adenosine 5'-triphosphate (ATP), adenosine 5'-diphosphatc (ADP) and adenosine 5'-monophosphate (AMP) were extracted from soil with either a solution of trichloroacetic acid, paraquat and phosphate (TCA reagent) or a mixture of chloroform, sodium hydrogen carbonate, phosphate and adenosine (NaHCO₃ reagent). Standard enzymic procedures were used to convert ADP and AMP to ATP, which was measured by the fire-fly luciferin-luciferase system. The measured quantities of nucleotides were corrected for incomplete extraction using the percentage recoveries of added ATP, ADP and AMP. The adenylate energy charge ratio (AEC) was calculated from the formula AEC = ([ATP] + 0.5 [ADP])/([ATP] + [ADP] + [AMP]).Measurements were made on a grassland soil, following a conditioning incubation at 15°C and 50% WHC for 7 days. Additional measurements were made on the same soil after a further 50- or 100-day incubation at 25°C and 50% WHC, with or without an amendment of 1100 μg ryegrass Cg⁻¹ soil, added at the end of the conditioning incubation. Biomass-ATP concentration, measured in TCA extracts, changed little, even on prolonged incubation, and was maintained at a level comparable to that observed in earlier work (about 10 p mol ATP g⁻¹ biomass C). AEC values in TCA soil extracts were high (0.8–0.9) for all soil treatments and independent of substrate addition or length of incubation.In contrast, AEC was low (0.4) in fresh soil extracted with NaHCO₃ reagent, but increased to 0.6 when ryegrass was incubated with the soil for 50 days. Although the total adenine nucleotide pool (i.e. [ATP] + [ADP] + [AMP]) was similar as measured in NaHCO₃ and in TCA soil extracts, both energy charge and ATP content were lower in the NaHCO₃ extracts. It was therefore concluded that the main reason for the lower AECs observed with the NaHCO₃ reagent was that microbial ATPases were still active during extraction and caused appreciable hydrolysis of microbial ATP to ADP and AMP. In contrast, the TCA reagent rapidly inactivates ATPases and is therefore preferable for extracting adenine nucleotides from soil.The results indicate that the soil microbial biomass, although a mainly dormant population, maintains both AEC and ATP at levels characteristic of exponentially growing organisms in vitro, even during prolonged incubation without fresh substrate. It was also concluded that roots make a negligible contribution to total ATP extracted from fresh sieved soil.     

Estimation of the adenylate energy charge in unamended and amended agricultural soils

To determine relatively low concentrations of adenine nucleotides in agricultural soils a NaHCO₃-based extradant was developed and compared with the trichloroacetic acid-paraquat-phosphate extradant. The new medium, consisting of chloroform, sodium bicarbonate, phosphate and adenosine (pH 8.0) gave soil extracts which could be investigated without further neutralization and dilution. ATP was measured directly in the soil extracts by the luciferin-luciferase system. ADP and AMP were estimated after their enzymatic conversion to ATP by standard methods. The quantities of nucleotides corrected for recovery of standards were used to calculate the adenylate energy charge (AEC) from the formula AEC = [ATP] + 1/2[ADP]/[ATP] + [ADP] + [AMP], The AEC was estimated in six unplanted soils from agricultural fields. A very similar energy charge of 0.3-0.4 was found in all soils sampled which indicates a low metabolic activity of the soil population. Two other soils with a pronounced difference in biomass-C content were used to investigate the influence of different amendments on the AEC. In an experiment with low glucose supplements up to 500 μg C g^?1 soil, the soil with the low biomass-C (a cambisol) showed a distinct increase of the AEC from 0.34 to 0.50, whereas the soil with the high biomass-C content (a phaeozem) increased its AEC only slightly from 0.32 to 0.37. In another experiment with high glucose supplements the phaeozem reached its maximum AEC value of 0.56 after the addition of 4000 μg Cg^?1 soil. An amendment with 8000 μg C g^?1 soil gave no further increase. In the combisol the addition of 1000 μg C g^?1 soil increased the AEC to 0.61. Higher supplements gave only a slight further increase to a maximum value of 0.67 after the addition of 8000 μg C g^?1 soil. The same AEC value was reached when the cambisol was amended with a mixture of organic substrates at a concentration of 10,000 μg C g^?1 soil.     

Microbial biomass dynamics in recently air-dried and rewetted soils compared to others stored air-dry for up to 103 years

Our aim was to compare the soil microbial biomass concentration and its activity (measured as CO₂-C evolved) following the rewetting and aerobic incubation of soils which have previously been stored air-dry for different periods. Some of the soils have been stored in the Rothamsted sample archive for 103 years, others were comparable freshly sampled soils following air-drying and rewetting and other soils were stored air-dry for 2 years then rewetted for the work described here. Following air-drying, soil ATP concentrations were variable in recently air-dried soil, comprising about 10-35% of the initial ATP concentrations in fresh soil. Following rewetting, the percentage recovery of ATP increased in all soils by 7 days, then declined to between 73% and 87% of the original ATP concentration in the air-dried soils by day 12. Storage of air-dried soils decreased the ability of the microbial biomass to restore its ATP concentrations. For example, the ATP concentration in a soil sampled from stubbed (i.e. tree seedling, saplings and bushes cut frequently to ground level) grassland of the Broadbalk continuous wheat experiment at Rothamsted then air-dried for 2 years was only about 14% of that in the fresh soil at 2 days after rewetting. In other soils from the Hoosfield Barley Experiment, also at Rothamsted, previously given NPK or FYM since 1852, and sampled then stored air-dry for between 13 and 83 years, from 52% to 57% of the ATP in the comparable fresh soils was measured at two days after rewetting. The soil ATP concentration then changed little more up to 12 days. One of the most interesting findings was that while the microbial biomass ATP concentration in the above NPK soils only ranged from about 2 to 4 μmol ATP g⁻¹ biomass C, in the FYM soil the microbial biomass ATP concentrations (range 11.5-13.6 μmol ATP g⁻¹ biomass C) were the same as we repeatedly measure in fresh moist aerobic soil. We do not yet know the reasons for this. More than twice as much CO₂-C was evolved from the long-term stored soils than from freshly sampled ones. However, the specific respiration of the microbial biomass did not change much after the first 12 years of storage, indicating that loss of viability mainly occurred in the earlier years.     

Adenylates as an estimate of microbial biomass C in different soil groups

Adenylate (i.e. adenosine tri- (ATP), di- (ADP) and monophosphates (AMP)) and microbial biomass C data were collected over a wide range of sites including forest floor layers and forest, grassland and arable soils. Microbial biomass C was measured by fumigation extraction and adenylates after alkaline Na₃PO₄/DMSO/EDTA extraction and HPLC detection. Our aims were (1) to test whether the sum of adenylates is a better estimate for microbial biomass than the determination of ATP, (2) to compare our conversion values with those proposed by others, and (3) to analyse whether soil properties or land use form affect the relationships between ATP, adenylates and microbial biomass C. A close relationship was found between microbial biomass C and ATP (r=0.96), but also with the sum of adenylates (r=0.96) within all appropriately conditioned soil samples (n=112). In the mineral soil (n=98), the geometric means of the ATP-to-microbial biomass C ratio and the adenylates-to-microbial biomass C ratio were 7.4 and 11.4 μmol g⁻¹, respectively. The mean ratios did not differ significantly between the different texture classes and land use forms. In the forest floor, the ATP-to-microbial biomass C ratio and the adenylates-to-microbial biomass C ratio were both roughly two-thirds of those of the mineral soil. The average adenylate energy charge (AEC) of all soil samples was 0.79 and showed a strong negative relationship with the soil pH (r=−0.69). However, the AEC is presumably only indirectly affected by the soil pH.     

Adenylate energy charge and ATP-to-microbial biomass C ratio in soils differing in the intensity of disturbance

We carried out an 8-days' incubation experiment with three different intensities of soil disturbance to analyse the effects on the ATP-to-microbial biomass C ratio and on the adenylate energy charge (AEC=(ATP+0.5×ADP)/(AMP+ADP+ATP). Single mixing of soil at 50% water holding capacity with a spatula during weighing of the samples into extraction jars at the end of the 8-days' incubation or 8-times repeated daily mixing for 2 min triggered the immediate formation of ATP, increasing both AEC and the ATP-to-microbial biomass C ratio. The energy for this extra ATP produced seems to be mainly derived from an accelerated turnover of C within the microbial biomass. In contrast, 8-days' continuous mixing led to a significant decrease in AEC and ATP-to-microbial biomass C ratio.     

Long‐term effects of leguminous cover crops on microbial indices and their relationships in soils of a coconut plantation of a humid tropical region

In this study, leguminous crops like Atylosia scarabaeoides, Centrosema pubescens, Calopogonium mucunoides, and Pueraria phaseoloides. grown as soil cover individually in the interspaces of a 19‐yr‐old coconut plantation in S. Andaman (India) were assessed for their influence on various microbial indices (microbial biomass C, biomass N, basal respiration, ergosterol, levels of ATP, AMP, ADP) in soils (0–50 cm) collected from these plots after 10 years. The effects of these cover crops on . CO₂ (metabolic quotient), adenylate energy charge (AEC), and the ratios of various soil microbial properties viz., biomass C : soil organic C, biomass C : N, biomass N : total N, ergosterol : biomass C, and ATP : biomass C were also examined. Cover cropping markedly enhanced the levels of organic matter and microbial activity in soils after the 10‐yr‐period. Microbial biomass C and N, basal respiration, . CO₂, ergosterol and levels of ATP, AMP, ADP in the cover‐cropped plots significantly exceeded the corresponding values in the control plot. While the biomass C : N ratio tended to decrease, the ratios of biomass N : total N, ergosterol : biomass C, and ATP : biomass C increased significantly due to cover cropping. Greater ergosterol : biomass C ratio in the cover‐cropped plots indicated a decomposition pathway dominated by fungi, and high . CO₂ levels in these plots indicated a decrease in substrate use efficiency probably due to the dominance of fungi. The AEC levels ranged from 0.80 to 0.83 in the cover‐cropped plots, thereby reflecting greater microbial proliferation and activity. The ratios of various microbial and chemical properties could be assigned to three different factors by principal components analysis. The first factor (PC1) with strong loadings of ATP : biomass C ratio, AEC, and . CO₂ reflected the specific metabolic activity of soil microbes. The ratios of ergosterol : biomass C, soil organic C : total N, and biomass N : total N formed the second factor (PC2) indicating a decomposition pathway dominated by fungi. The biomass C : N and biomass C : soil organic C ratios formed the third principal component (PC3), reflecting soil organic matter availability in relation to nutrient availability. Overall, the study suggested that Pueraria phaseoloides. or Atylosia scarabaeoides were better suited as cover crops for the humid tropics due to their positive contribution to soil organic C, N, and microbial activity.     

Adenylates in the soil microbial biomass at different temperatures

Five soils from temperate sites (Germany; 2 arable and 3 grassland) were incubated aerobically at 5, 10, 15, 20, 25, 35, and 40 °C for 8 days. Soils were analysed for soil microbial biomass C, biomass N, AMP, ADP, and ATP to determine whether the increase in the ATP-to-microbial biomass C ratio with increasing temperature was either due to an increase in the adenylate energy charge (AEC) or de novo synthesis of ATP, or both. Around 80% of the variance in microbial biomass C and biomass N was explained by differences in soil properties, only 7% by the temperature treatments. Averaging the data of all 5 soils for each incubation temperature, the microbial biomass C content decreased with increasing temperature from 15 to 40 °C continuously by 2.5 μg g⁻¹ soil °C⁻¹ after 8-days' incubation. However, this decrease was not accompanied by a similar decrease in microbial biomass N. The average microbial biomass C/N ratio was 6.8. Between 54 and 76% of the variance in AMP, ADP, ATP and the sum of adenylates was explained by differences in soil properties and between 14 (ADP) and 27% (ATP) by the temperature treatments. However, temperature effects on AMP and ADP were variable and inconsistent. In contrast, ATP and consequently also the sum of adenylates increased continuously from 5 to 30 °C followed by a decline to 40 °C. The AEC showed similarly a small, but significant increase with increasing temperature from 0.73 to 0.85 at 30 °C. Consequently, the majority of the variance, i.e. roughly 60% in AEC values, but also in ATP-to-microbial biomass C ratios was explained by the incubation temperature. The mean ATP-to-microbial biomass C ratio increased from 4.7 μmol g⁻¹ at 5 °C to a 2.5 fold maximum of 12.0 μmol g⁻¹ at 35 °C. This increase was linear with a rate of 0.26 μmol ATP g⁻¹ microbial biomass C °C⁻¹. The energy for the extra ATP produced during temperature increase is probably derived from an accelerated turnover of endocellular C reserves in the microbial biomass.     

Specific respiration rates, adenylates, and energy budgets of soil microorganisms after addition of transgenic Bt-maize straw

An arable soil was incubated with straw (stem+leaves) of two transgenic Bt-maize varieties (Novelis: event MON810 and Valmont: event Bt176) and the two corresponding near-isogenic varieties (Nobilis and Prelude). The aim was to evaluate the use of these substrates for microbial growth and maintenance in soil during early decomposition. The addition of Bt-maize straw increased CO₂ production rates and the specific respiration rates CO₂-C/microbial biomass C and CO₂-C/ATP significantly compared with the addition of non-Bt maize straw. This extra energy in the Bt-maize straw could not be used for microbial biomass or ATP and ADP production, and was lost for maintenance. In addition, increased death rates of microbial biomass occurred in the soils treated with the Bt-maize straw from day 3 to 21. Generally, most of the energy was stored in microbial biomass, whereas only 10% of energy was stored in ATP, and only 1-2% in ADP. The AEC (adenylate energy charge: (ATP+0.5×ADP)/(AMP+ADP+ATP)) was not affected by any treatment. The reasons for the lower efficiency of microbial substrate use after adding Bt-maize straw cannot be fully explained by the present experiment. However, a risk assessment has to look at the impact of transgenic plant material on soil microorganisms at different maturity stages.     

干土效应对土壤生物组成及矿化与硝化作用的影响

将经过风干、过筛后的2种旱地红壤加水培养,并和新鲜土培养条件相比较,研究干土效应对土壤生物组成及矿化与硝化作用的影响.试验共4个处理(1)农田旱地风干土加水培养(RU);(2)农田旱地新鲜土培养(FU);(3)苗圃旱地风干土加水培养(RN);(4)苗圃旱地新鲜土培养(FN).结果表明红壤风干土加水预培养5 d后,细菌、放线菌、真菌数量比新鲜土显著增加(p＜0.01),细菌数量增加最为明显,农田旱地和苗圃旱地风干土处理分别是新鲜土的6.26倍和6.84倍,红壤风干土加水培养处理的微生物量碳、氮也随之增加.培养28 d后土壤中微生物数量趋于稳定,与预培养5 d时的数量相当或稍有下降,但风干后加水培养处理的微生物数量仍保持大于新鲜土的趋势(农田旱地的放线菌除外),微生物量碳、氮也存在同样的趋势.风干土加水培养后微生物数量的迅速增加,使得氮素矿化速度加快,由此导致NH+4-N量显著增加(p＜0.01),培养28 d后,NH+4-N量较预培养5 d时有所增加,且明显高于新鲜土培养处理;NO-3-N含量也增加,但新鲜土处理显著高于风干土处理.土壤风干处理对土壤自由生活线虫的影响比较大,农田旱地和苗圃旱地风干土加水培养28 d后,其自由生活线虫数量仅为新鲜土的16.0%和30.1%,显示风干土加水培养难以恢复土壤微型动物的数量.28 d的矿化和硝化培养试验结果显示,风干土加水培养处理的净矿化量和矿化率均高于新鲜土处理,苗圃旱地风干土处理的增量达到了显著水平(p＜0.05),但是硝化作用却刚好相反,农田旱地和苗圃旱地的净硝化量及硝化率均是新鲜土处理显著高于风干土处理(p＜0.05),其原因是对硝化作用起重要作用的硝化菌(氨氧化细菌和亚硝酸氧化细菌)数量在经历了风干过程后很难恢复到新鲜土水平.     

Reaction of microorganisms to rewetting in continuous cereal and legume rotation soils of semi-arid Sub-Saharan Africa

A 20-day incubation experiment with continuous cereal (CC) versus cereal legume (CL) rotation soils of two semi-arid Sub-Saharan sites (Fada-Kouaré in Burkina Faso, F, and Koukombo in Togo, K) were carried out to investigate the effects of rewetting on soil microbial properties. Site- and system-specific reactions of soil microorganisms were observed on cumulative CO₂ production, adenylates (ATP, ADP, and AMP), microbial biomass C and N, ergosterol, muramic acid and glucosamine. Higher values of all parameters were found in the CL rotation soils and in both soils from Fada-Kouaré. While the inorganic N concentration showed only a system-specific response to rewetting, the adenylate energy charge (AEC) showed only a site-specific response. ATP recovered within 6 h after rewetting from ADP and AMP due to rehydration of microorganisms and not due to microbial growth. Consequently, no N seemed to be immobilized by microorganisms and all NO₃ in the soil was immediately available to the plants. The fungal cell-membrane component ergosterol was three (CC) and five (CL) times larger at Fada than in the respective soils at Koukombo. The concentrations of the bacterial cell-wall component muramic acid were by 20% and of mainly fungal glucosamine by 10% larger in the CL rotation soils than in the CC soils. This indicates long-shifts in the microbial community structure.     

Relation between respiration,ATP content,and Adenylate Energy Charge (AEC) after incubation at different temperatures and after drying and rewetting

Temperature, drying, and rewetting are important climatic factors that control microbial properties. In the present study we looked at the respiration rates, adenosine 5′‐triphosphate (ATP) content, and adenylate energy charge (AEC) as a measure for energy status of microbial biomass in the upper 5 cm of mineral soils of three beech forests at different temperatures and after rewetting. The soils differed widely in pH (4.0 to 6.0), microbial biomass C (92 to 916 μg (g DW)^—1) and ATP content (2.17 to 7.29 nmol ATP (g DW)^—1). The soils were incubated for three weeks at 7 °C, 14 °C, and 21 °C. After three weeks the microbial properties were determined, retaining temperature conditions. The temperature treatment did not significantly affect AEC or ATP content, but respiration rates increased significantly with increasing temperature. In a second experiment the soils were dried for 12 hours at 40 °C. Afterwards the soils were rewetted and microbial properties were monitored for 72 hours. After the drying, respiration rates dropped below the detection limit, but within one hour after rewetting respiration rates increased above control level. Drying reduced AEC by 16 % to 44 % and ATP content by 47 % to 78 %, respectively. Rewetting increased AEC and ATP content significantly as compared to dry soil, but after 72 hours the level of the controls was still not reached. The level of AEC values indicated dormant cells, but ATP content increased. These results indicate that the microbial carbon turnover was not directly linked to microbial growth or microbial energy status. Furthermore our results indicate that AEC may describe an average energy status but does not reflect phases of growing, dormant, or dying cells in the complex microbial populations of soils.     

Assessing the effects of air-drying and rewetting pre-treatment on soil microbial biomass,basal respiration,metabolic quotient and soluble carbon under Mediterranean conditions

Soil biochemical properties are useful indicators of soil quality as they are very sensitive to disturbance. Sample storage or pre-treatments could affect the results in these assays, which are normally determined on fresh samples, kept cold or frozen. The objectives of this study were to (i) evaluate the effect of air-drying or incubation of rewetted air-dried soil samples on microbial biomass carbon (MBC), basal soil respiration (BSR), qCO₂ and water soluble carbon (WSC), in soils from different locations, with different degradation status and sampling seasons, and (ii) assess if air-drying or incubation of rewetted air-dried soil samples is an accurate sample storage and pre-treatment procedure for these soil properties in soil quality evaluations under semiarid Mediterranean conditions. Our results showed that air-drying does not have the same effects on MBC, BSR, qCO₂ and WSC depending on the geographical situation and sampling date. It seems that the warmest and driest place and season show less variation when using air-dried soil samples, with values representative of those obtained under field-moist conditions. Short incubations (4, 8 and 12 days at 23 °C) provoked a general decrease in all properties, probably due to labile organic compounds depletion. Hence, air-dried soils can be used as part of soil quality analysis to estimate these biochemical properties in summer time in the semiarid region of South-East Spain, because they have not suffered severe affections. Moreover, MBC could also be determined using air-dried soil in the driest zones during all year. In contrast, estimations with incubated soil samples are not, in any case, representative of field-moist soil values.     

Microbial biomass and activity indices after organic substrate addition to a selenium-contaminated soil

High concentrations of Se in soil might have negative effects on microorganisms. For this reason, the effect of organic substrate addition (glucose + maize straw) on Se volatilisation in relation to changes in microbial biomass and activity indices was investigated using an artificially Se-contaminated soil. Microbial biomass N was reduced on average by more than 50% after substrate addition, but adenylate energy charge (AEC) and metabolic quotient qCO₂ were both increased. The Se content decreased by nearly 30% only with the addition of the organic substrate at 25°C. No significant Se loss occurred without substrate at 25°C or with substrate at 5°C. In the two treatments with substrate addition, the substrate-derived CO₂ evolution was about 30% lower with Se addition than without. In contrast, Se had no effect on any of the other soil microbial indices analysed, i.e. microbial biomass C, microbial biomass N, adenosine triphosphate (ATP), AEC, ATP-to-microbial biomass C, and qCO₂.     

Interactions between soil properties and moisture content in crust formation, runoff and interrill erosion from tilled loess soils

Soil-surface seals and crusts resulting from aggregate breakdown reduce the soil infiltration rate and may induce erosion by increasing runoff. The cultivated loess areas of northwestern Europe are particularly prone to these processes.Surface samples of ten tilled silty loamy loess soils, ranging in clay content from 120 to 350 g kg⁻¹ and in organic carbon from 10 to 20 g kg⁻¹, were packed into 0.5 m² plots with 5% slopes and subjected to simulated rainfall applied at 30 mm h⁻¹. The 120 minutes rainfall events were applied to initially field-moist soil, air-dried soil and rewetted soil to investigate the effect of soil moisture content prior to rainfall. Runoff and eroded sediments were collected at 5 minutes intervals. Aggregate stability of the soils was assessed by measuring particle-size distribution after different treatments.All soils formed seals. Runoff rates were between 70 and 90% by the end of the rainfall event for field-moist plots. There were large differences between soil runoff rates for the air-dried and rewetted plots. Interrill erosion was associated with runoff, and sediment concentration in runoff readily reached a steady-state value. Measurements of aggregate stability for various treatments were in good agreement with sealing, runoff and erosion responses to rainfall. Runoff and erosion were lower for air-dried plots than for field-moist plots, and were either intermediate or lowest for rewetted plots, depending on soil characteristics. Soils with a high clay content had the lowest erosion rate when they were rewetted, whereas the soil with a high organic-carbon content had the lowest erosion rate in air-dry conditions. The results indicate the complexity of the effect of initial moisture content, and the interactions between soil properties and climate.     

Phosphatase activity does not limit the microbial use of low molecular weight organic-P substrates in soil

Plant roots and soil microorganisms contain significant quantities of low molecular weight (MW) phosphorylated nucleosides and sugars. Consequently, upon death these can represent a significant input of organic-P to the soil. Some of these organic-P substrates must first be dephosphorylated by phosphatases before being assimilated by the soil microbial community while others can be taken up directly from soil solution. To determine whether sorption or phosphatase activity was limiting the bioavailability of low MW organic-P in soil we compared the microbial uptake and C mineralization of a range of ¹⁴C-labeled organic-P substrates [glucose-6-phosphate, adenosine monophosphate (AMP), adenosine diphosphate (ADP) and adenosine triphosphate (ATP)] to that of the parent compounds (adenosine and glucose). In a fertile grassland soil we showed that at low organic-P substrate concentrations (<0.5 mM) phosphatase activity did not limit microbial uptake or mineralization in comparison to their non-phosphorylated counterparts. However, at high substrate concentrations (1-10 mM) the mineralization of the organic-P compounds was significantly lower than that of the non-phosphorylated compounds suggesting that phosphatase activity or microbial transporter capacity limited bioavailability. Sorption to the solid phase followed the series glucose<adenosine<G-6-P<AMP<ADP=ATP. However, sorption of the organic-P compounds to the solid phase did not appear to greatly affect bioavailability. The high adenosine mineralization capacity of the microbial biomass suggests that nucleosides may represent a significant source of C and N to the soil microbial biomass. We conclude that at low organic-P substrate concentrations typical of those in soil, neither phosphatase activity nor sorption greatly limits their bioavailability.     

Correlation between four methods to estimate total microbial biomass in stored,air-dried and glucose-amended soils

Correlation between the microbial volume, chloroform fumigation (CO₂-C flush), substrateinduced respiration (SIR) and ATP content methods to estimate microbial biomass was assessed on three New Zealand soils (two grassland, one arable) under three different treatments (stored, air-dried and glucose-amended). There were significant, positive correlations between all methods, r = 0.69–0.88, which were improved, r = 0.71–0.96, if the data for air-dried or glucose-amended soils were excluded from the analyses. The best agreement was between CO₂-C flush and ATP and the worst between CO₂-C flush and microbial volume. Exclusion of air-dried soil data improved these correlations.Estimates of microbial biomass for each soil often differed significantly between the four methods, when conversion factors cited in the literature were used. Ratios (i.e. conversion factors) between CO₂-C flush and ATP or SIR, or SIR and volume, were different to those cited in the literature, and only similar if specific data were excluded.We recommend that a minimum of two and preferably three methods be used to quantify the microbial population of soil, and that emphasis should be placed on the relative differences within and between soils using data which have not been converted to biomass C. Conversion of data to biomass C may result in substantial errors.     

Impact of cycloheximide addition on adenylates in soil

Cycloheximide inhibits specifically the ribosomal protein synthesis of eukaryotic cells, i.e. the metabolism of soil fungi. We measured cycloheximide effects on adenylates in 20 different soils (0-10 cm depth) from arable, grass and forest land with a large variety of soil properties. The aims were (1) to assess the interactions between cycloheximide effects and soil properties and (2) to prove the relationship between cycloheximide effects on ATP and the ergosterol-to-microbial biomass C ratio, which is an indicator for the fungal proportion of the total microbial biomass. The adenylates ATP, ADP and AMP were measured 6 h after adding either 10 mg cycloheximide per gram soil in combination with 24 mg talcum per gram soil or 24 mg talcum per gram soil solely. The medians of the relative increases in AMP and ADP were 45 and 25% and the medians of the relative decreases in ATP and adenylates were −36 and −12%. These changes in adenylate composition lead to a cycloheximide-induced relative decrease in the adenylate energy charge level of 15%. The relative decrease in ATP content after cycloheximide addition was significantly correlated with the ATP-to-microbial biomass C ratio, but not with the ergosterol-to-microbial biomass C ratio. The absolute increase in ADP and the absolute decrease in ATP were affected by the clay content according to principal component analysis. The reduction of the ATP-to-microbial biomass C ratio indicates that this ratio had the potential of being an important ecotoxicological indicator of direct toxic effects of organic pollutants on soil microorganisms.     

Adenosine triphosphate and microbial biomass in soil

Two methods for measuring adenosine 5'-triphosphate (ATP) in soil were compared, one based on extraction with NaHCO₃-CHCl₃ and thel other on extraction by a trichloracetic acid-phosphate-paraquat reagent. Recoveries of added ATP were greater with the NaHCO₃-CHCl₃ reagent but the extraction of “native” soil ATP by NaHCO₃-CHCl₃ was only about a third of that by TCA-phosphate-paraquat.Microbial biomass C and ATP were measured in 8 contrasting English soils, using the fumigation method to measure biomass C and the TCA-phosphate-paraquat method to measure ATP. Except in one acid woodland soil, the ratio (ATP content of the soil)/(biomass C content of the soil) was relatively constant, with a mean of 7.3 mg ATP g^?1 biomass C for the different soils. This value is very similar to that obtained earlier in a range of 11 grassland and arable soils from Australia. Taking the English and Australian grassland and arable soils together, there is a close (r = 0.975) linear relationship between ATP and microbial biomass C that holds over a wide range of soils and climates. From this relationship, the soil biomass contains 7.25 mg ATP g^?1 biomass C, equivalent to an ATP-to-C ratio of 138, or to 6.04 μmoles ATP g^?1 dry biomass.The acid woodland soil (pH 3.9) contained much less biomass C, as measured by the fumigation method, than would have been expected from this relationship. This, and other evidence, suggests that the fumigation method for measuring microbial biomass C breaks down in strongly acid soils.The ATP content of the biomass did not depend on the P status of the soil, as indicated by NaHCO₃-extractable P.     

Decomposition of Zn-rich Arabidopsis halleri Litter in Low and High Metal Soil in the Presence and Absence of EDTA

Hyperaccumulating plants are increasingly investigated in combination with EDTA addition to soil for phytoremediation of heavy metal contaminated soils. A 60-day incubation experiment was carried out to investigate the effects of heavy metal release during the decomposition of Zn-rich (15.7 mg g^?1 dry weight) Arabidopsis halleri litter on C mineralization, microbial biomass C, biomass N, ATP, and adenylate energy charge (AEC). These effects were investigated in two soils with different Zn, Cu, and Pb levels, with and without EDTA addition to soil. The sole addition of Zn-rich A. halleri litter to the two soils did not increase the contents of NH₄NO₃ extractable Zn, only with the combined additions of EDTA and litter was there a considerable increase, being equivalent to three times the added amount in the low metal soil and to 50% in the high metal soil. Litter amendment increased the CO₂ evolved; being equivalent to 44% of the added C in the two soils, but EDTA addition had no significant effect on CO₂ evolution. Litter amendment resulted also in an 18% increase in microbial biomass C, 27% increase in ATP and 6% increase in AEC in the two soils, but EDTA had again no effect on these indices at both metal levels. In contrast, the sole addition of litter had no effect on microbial biomass N, but EDTA addition increased microbial biomass N on average by 49%. The application of EDTA for chelate-assisted phytoextraction should in the future consider the risk of groundwater pollution, which is intensified by resistance of EDTA to microbial decomposition.     

Interactions between soil organic matter status, cropping history, method of quantification and sample pretreatment and their effects on measured aggregate stability

 The effects of sample pretreatment (field-moist, air-dried or tension rewetted) on aggregate stability measured by wet sieving or turbidimetry were compared for a group of soil samples ranging in organic C content from 20 to 40 g C kg^–1. Concentrations of total N, total and hot-water-extractable carbohydrate and microbial biomass C were linearly related to those of organic C. Aggregate stability measured by wet sieving using air-dried or field-moist samples and that measured by turbidimetry, regardless of sample pretreatment, increased curvilinearly with increasing soil organic C content. However, when tension-rewetted samples were used for wet sieving, aggregate stability was essentially unaffected by soil organic C content. Measurements of aggregate stability (apart from wet sieving using rewetted soils) were closely correlated with one another and with organic C, total and extractable carbohydrate and microbial biomass C content of the soils. The short-term effects of aggregate stability were also studied. Soils from under long-term arable management and those under long-term arable followed by 1 or 3 years under pasture had similar organic C contents, but aggregate stability measured by turbidimetry and by wet sieving using air-dried or field-moist samples increased with increasing years under pasture. Light fraction C, microbial biomass and hot-water-extractable carbohydrate concentrations also increased. It was concluded that both total and labile soil organic C content are important in relation to water-stable aggregation and that the use of tension-rewetted samples to measure stability by wet sieving is unsatisfactory since little separation of values is achieved. Received: 6 January 1999