The Use of Fluorogenic Substrates To Measure Fungal Presence and Activity in Soil (original) (raw)
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Comparison of fungal and microbial biomass of three soils under different land use systems
Nepalese Journal of Agricultural Sciences , 2015
The study was undertaken to compare fungal and microbial biomass of three soils from different land use systems (i.e. garden, arable land and catch crop). Chloroform-fumigation extraction and adenosine 5"-triphophate (ATP) measurement were used to assess microbial biomass while determination of ergosterol concentration was done to assess living fungal biomass in the soils. Catch crop soil revealed to have considerably higher microbial activity and carbon turnover in the soil. The mean fungal biomass was significantly higher in the catch crop soil (1.68 μg g-1) than garden (1.19 μg g-1) and arable land (0.67 μg g-1). The considerable high ergosterol: microbial biomass C ratio in the catch crop soil suggest that fungal part of microbial biomass was high in this soil compared to the rest which did not differ appreciably. The garden soil had significantly higher mean microbial biomass C of 765 μg g-1 soil as compared to arable land (345 μg g-1) and catch crop (379 μg g-1) soils. The average ATP concentration in the garden (6.7 nmol g-1 dry soil) and catch crop (6.6 nmol g-1 dry soil) soils were considerably larger than that of arable land soil (3.8 nmol g-1 dry soil). The variation in fungal and microbial biomass in the soils may be due to differences in microbial species assemblage, plant species and environmental conditions.
Fluorescein diacetate hydrolysis as a measure of fungal biomass in soil
Current Microbiology, 2001
Abstract. The fatty acid methyl esters of lipids extracted from an agricultural soil in the preharvest period of soybean or middle growth cycle from wheat were characterized and quantified by gas-liquid chromatography. The fatty acids 18:26 and 16:15 were used as markers of ...
Microbiology, 2006
Tundra, chernozem (virgin and arable), soddy-podzolic (coniferous forest, meadow, and arable), and grey forest (larch forest) soils were used to separate the contributions of fungi and bacteria to substrateinduced respiration (SIR) with the help of antibiotics. For soils with a high content of organic matter (tundra and chernozem: 12 and 8%, respectively), the procedure of selective inhibition of SIR has been optimized. This procedure consists in application of high concentrations of streptomycin (50-120 mg/g of soil) and cycloheximide (50-80 mg/g of soil) and decreasing the weight of the analyzed soil sample. Soils under study have shown the predominant contribution of fungi (63-82%) to the total SIR. The fungal-bacterial ratio in the soils of natural ecosystems (0-5 cm, without litter) was 4.3, 2.2, 1.5, and 1.5 for tundra soil, virgin chernozem, coniferous (soddy-podzolic soil), and larch (grey forest soil) forests, respectively. The lower layers of soddy-podzolic (5 − 10 cm) and grey forest (48-58 cm) soils showed a decrease in the fungal and increase in the bacterial component in the total SIR.
Changes in microfungal communities, fungal activities and humic substances (HS) in agricultural soils kept under different fertilization regimes were observed and their causal relationships were investigated in a long-term field experiment. Fertilization did not change the abundance of HS-utilizing microfungi and, except for organic amendment alone, total culturable microfungi were also unaffected by this factor. Organic fertilization increased activities of manganese peroxidase (MnP) and proteinase, but decreased endo-1,4-β-glucanase activity compared to the corresponding control without organic fertilization. In soils treated with mineral fertilizers, the activities of MnP, endo-1,4-β-glucanase and proteinase were higher than in control without any mineral treatment. Both the aromaticity of fulvic acid and the molar mass of humic acid was lower in soil with organic fertilization, which may be a result of oxidative degradation mediated by higher MnP activity observed in treatments with organic fertilization. Abbreviations ABTS 2,2´-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) OC oxidizable (organic) carbon CFU colony forming units OF organic fertilization FA fulvic acid SGF silica-gel supplemented with fulvic acid HA humic acid SI Sørenson index HS humic substances SOM soil organic matter MEA malt extract agar QSI quantitative Sørenson index MF mineral fertilization RDA redundancy analysis MnP manganese peroxidase Conversion of humus plays an important role in maintaining global carbon cycle and is essential for the long-term sustainability of ecosystems. HS are present in water and sediments, but the main part is deposited in soils, where they constitute an important, relatively stable source of carbon, making up to 70–80 % of the organic carbon present in the biosphere (Schlesinger 1997). Management of plant cultures and their fertilization affect not only plants, but also soil microorganisms (Marschner et al. 2003) and can thus cause changes in the deposition of HS. Increased input of organic matter to the soil together with sufficient availability of mineral nutrients after MF may result in an increased rate of decomposition processes and microbial growth. Despite the importance of HS, the response of HS in respect to various fertilization regimes has received only limited attention. It has been shown that increased resistance of HA to microbial degradation is associated with increased SOM contents (Filip and Kubát 2001). Although various microorganisms can de-polymerize HS (Gramss et al. 1999) or change their physico-chemical features (ezáová and Gryndler 2006), they are considered to be very stable in soil (Hurst et al. 1962). In spite of the apparent significant association between SOM and microfungi (Hršelová et al. 1999), only few studies on the influence of various fertilization regimes on these microorganisms are available. Here we analyzed soil samples from a field experiment run for 47 years where plots were treated by different levels of MF and OF. The objective was to contribute to an increased understanding of changes in microfungal communities, fungal activities and HS in agricultural soils caused by fertilization.
The utility of ergosterol as a bioindicator of fungi in temperate soils
Soil Biology and Biochemistry, 2000
In this paper we evaluate the utility of ergosterol as a measure of fungal biomass in temperate soils. We summarise published ®ndings and compare them with data from our own broad-scale assessment of the relationship between ergosterol and ATP in a range of temperate soils. Two hundred and ninety ®ve plots (three cores taken from each 10 Â 10 m plot) in seven ecotypes were sampled. Soils ranged from entirely mineral to entirely organic (0.01±46% C org and sites comprised two primary successions, one on shingle ridge on the south coast of England and one in the slack of a dune blow-out on the south coast of Wales, various meadow, pasture (some restored after opencast mining) and ancient woodland soils throughout England and acid forest soils in Central Europe. We found a strong relationship between ergosterol and ATP r 2 0X80), which was largely unaected by the key soil properties of C org , C/N ratio, moisture and pH. The sources and implications of the 20% of residual variance were explored by assuming that the error was compounded from three sources: the inaccuracies in methods of analysis of ergosterol and ATP, the failings of each of the variables to estimate their underlying populations (i.e., fungal and total biomass, respectively) e var), and the non-equivalence of these populations (i.e., their incomplete overlap) e pop). By partitioning the residual variance into components corresponding to the levels of sampling, we estimated that the sum of the systematic portions of e var and e pop formed as much as three quarters of the 20% of residual variance in the ATP±ergosterol correlation, leaving just 5% mostly due to random error. Despite this close relationship, the attainment of a universal conversion factor between ergosterol and fungal biomass, applicable to all temperate soils, remains elusive and problematic. Many problems are caused by a lack of comparability between the various measures of fungal and total biomass used and the reliability, or otherwise, of extrapolations based on measures of axenic cultures (in contrast to in-situ measurements). The issue is further complicated by the non-linearity of the relationship between fungal biomass and fungal surface area; ergosterol is more correctly an index of the latter since it is a principal membrane sterol. We conclude that ergosterol is likely to be a reliable indicator of the extent of fungal membranes in temperate soils, if not an accurate measure of fungal biomass.
Effect of pollutants on the ergosterol content as indicator of fungal biomass
Journal of Microbiological Methods, 2002
Ergosterol content was determined in 20 white-rot fungi isolates and the values ranged from 2380 to 13 060 Ag g À 1 fungal biomass. Significant changes of ergosterol content according the physiological stage for Bjerkandera adusta 4312 and Coriolopsis gallica 8260 were found, showing the highest values during the stationary phase. However, in the case of Phanerochaete chrysosporium 3642, no changes were detected during growth. The effect of pollutants, such as heavy metals and fungicides, on the ergosterol content of C. gallica was determined. Heavy metals (Cu 80 ppm, Zn 50 ppm or Cd 10 ppm) and fungicides (thiram 3 ppm or pentachlorophenol 1.5 ppm) at concentrations that reduce the metabolic activity between 18% and 53% (pollutant-stressed cultures) did not affect the ergosterol content. Only the fungicide zineb (25 ppm) reduced significantly the ergosterol content in biomass basis. In soil experiments with Cu (80 ppm) or thiram (10 ppm) after 15 and 30 days of incubation, the ergosterol content in soil was linearly correlated to the fungal biomass C in both polluted and control soil cultures. The ergosterol content was independent of the presence or the absence of pollutants. Thus, these results indicate that ergosterol can be a useful indicator for fungal biomass in polluted soils, and can be applied for monitoring bioremediation processes. D
Sources of error in direct microscopic methods for estimation of fungal biomass in soil
Soil Biology & Biochemistry, 1995
of error in direct microscopic measurement of fungal hyphae in soil were examined and fungal biomass estimates and associated variability obtained by the direct counting method and the ergosterol technique were compared. Nested random effects ANOVA indicated that the major source of variance in the direct microscopic counting method were the people examining the prepared microscope slides, accounting for 83% of the total variance. Sampling variability accounted for approximately 14% of the total variance. Fungal biomass values calculated from soil ergosterol concentrations were close to the range of values derived from hyphal length estimates but coefficients of variation were much lower for soil ergosterol determinations (6-13%) than for hyphal length estimates (16-32%). For one soil sample, we compared total hyphal length and fungal biomass estimates from our lab to those of another lab. Values obtained by the other laboratory were outside the range of values and 95% confidence intervals reached in our lab. Comparison of fungal hyphal length estimates from undisturbed prairie soil and an adjacent cultivated soil indicated that the undisturbed soil contained more than twice as much fungal biomass. Results of our study indicate: (1) extreme caution must be used when comparing hyphal length and fungal biomass estimates made by different laboratories using the direct counting method; and (2) soil ergosterol determinations can provide information on fungal biomass that may be useful in comparing direct count estimates by different labs.
Biology and Fertility of Soils, 2002
The degree of contact between crop residues and the soil matrix, as determined by the method of residue incorporation, affects decomposition dynamics both under natural and experimental conditions. In microcosm experiments we tested the hypothesis that poor residuesoil contact reduces the decomposition of structural plant constituents through delayed colonization by microorganisms degrading cellulose and hemicellulose. Barley straw or red clover foliage was either confined in buried mesh bags or homogeneously mixed into a loamy topsoil or a silty subsoil to create poor or intimate residue-soil contact in microbiologically rich and less rich environments, respectively. Soil type had no effect on decomposition of the easily degradable clover residues, but cumulative mineralization of barley straw C after 52 days at 15°C was less in the subsoil than in the topsoil by 12% of initial C. For clover material, poor soil contact increased cumulative C mineralization by 5% of initial C in the loamy topsoil but had no effect in the silty subsoil. For the more slowly degradable, cellulose-and hemicellulose-rich straw, on the other hand, poor soil contact reduced C mineralization by 6% of initial C. The results from the loamy topsoil were confirmed in a second experiment in a sandy topsoil. The reduced decomposition of straw with poor soil contact could not be explained by less favourable abiotic conditions, N deficiency nor exclusion of larger animals by mesh bags. Reduced strawsoil contact delayed measured increases in fungal ergos-terol concentration, ratio of fungal to bacterial substrateinduced respiration, number of cellulase-producing, colony-forming bacterial units and activity of cellulases and hemicellulase on the residues. Thus, the results supported our hypothesis and underscore the importance of ensuring representative conditions for the soil microflora when decomposition dynamics are studied in microcosms experiments designed to mimic field conditions.
Determinination of soil fungal biomass from soil ergosterol analyses
Soil Biology and Biochemistry, 2000
Determination of fungal biomass (FB) and FB-carbon (FB-C) from soil ergosterol concentration is dicult because of unknown ergosterol-to-fungal biomass (E-to-FB) conversion factors and inecient ergosterol extraction methods. We applied a microwave-assisted extraction (MAE) and high performance liquid chromatographic (HPLC) procedure to measure ergosterol in soil samples. The E-to-FB conversion factors were determined in six species of fungi grown in vitro. The MAE method was fast and extracted up to nine times more soil ergosterol than a classical re¯uxing saponi®cation method. Soil ergosterol was separated and quanti®ed rapidly (<10 min) by HPLC. Alternaria alternata, Chaetomium globosum, Fusarium oxysporum, Penicillum chrysogenum, Rhizopus stolonifer and Trichoderma harzianum isolated from soil and plant matrices were grown in batch. Ergosterol and biomass content were determined in mycelia harvested during the stationary and exponential phases of growth. Total mycelial ergosterol ranged from 180 to 2178 mg, and total dry biomass ranged from 17 to 595 mg. Total ergosterol and fungal dry biomass were strongly associated (r 2 = 0.95). The C content in mycelial mats averaged 43% (21.1, SD), and was similar among fungal species and growth phases. The analyses of variance showed that the E-to-FB ratio was similar among fungal species or growth phase. An average ergosterol concentration of 4 mg mg À1 dry biomass was determined for the six species of fungi, which gave a conversion factor of 250 mg dry biomass mg À1 ergosterol. The MAE method recovered an average of 62% (211%, SD) of the ergosterol added in mycelial mats to soils prior to extraction, and its recovery was independent of soil properties. The E-to-FB ratio and percent recovery of mycelial ergosterol helped establish for the ®rst time relationships determining soil FB and FB-C from soil ergosterol concentration. The amount of FB ranged from 155 to 4745 mg g À1 and that for FB-C ranged from 67 to 2040 mg g À1 for dierent soils, and was higher in samples taken from native undisturbed land than in samples taken from adjacent cultivated ®elds. Measurement of soil ergosterol concentration is a useful estimate content of the living soil FB. 7
Geoderma, 2016
The interrelations between the microbial biomass, total N, organic P, and organic S in the accumulation of soil organic C (SOC) and the contribution of fungal and bacterial residues to SOC, based on amino sugar data, were investigated in the current study. The soils had been developed under humid temperate, arid subtropical , and tropical climatic conditions and were used as arable, grass, and forest land. They covered a wide range in soil pH, in salinity as well as in the contents of clay and SOC. An increased microbial biomass C/N ratio due to nutrient limitation is not reflected by any increase in SOM elemental C/N/P/S ratio within the total nutrient pool or within the organic fraction. Increased SOC/total N and SOC/organic S ratios reduced the contribution of microbial biomass C to SOC, whereas neutral soil pH and high clay contents had positive effects. An increased formation of microbial biomass C also enlarged the contribution of microbial residue C to SOC, which accounted on average for 48% SOC in the neutral arable and moderate acidic grassland soils and for 30% in the saline arable and strongly acidic forest soils. The fungal C to bacterial C ratio increased with increasing acidity from 0.9 in the saline arable soils to 4.5 in the strongly acidic forest soils. Consequently, the relationship of the fungal C to bacterial C ratio was not simply related to the contribution of microbial residues to SOC. The lower the ergosterol to fungal GlcN ratio, the higher the contribution of microbial residues to SOC. This means the higher the contribution of arbuscular mycorrhizal fungi and the lower the contribution of saprotrophic fungi to the microbial community, the more fungal residues can be sequestered to SOC.