- Email: [email protected]
β-Glucosidase activity in pasture soils
Applied Soil Ecology 20 (2002) 157–162
Short communication
-Glucosidase activity in pasture soils Benjamin L. Turner a,∗ , David W. Hopkins b , Philip M. Haygarth a , Nick Ostle c a
Institute of Grassland and Environmental Research, North Wyke Research Station, Okehampton, Devon EX20 2SB, UK b Department of Environmental Science, University of Stirling, Stirling FK9 4LA, UK c Centre for Ecology and Hydrology, Merlewood Research Station, Grange-Over-Sands, Cumbria LA11 6JU, UK Received 8 January 2002
Abstract -Glucosidase is involved in the degradation of cellulose in soils and has potential for monitoring biological soil quality. We assayed -glucosidase activity in 29 permanent grassland soils from England and Wales with contrasting physico-chemical and biological properties (clay content 22–68%; total carbon 29–80 mg g−1 ; microbial carbon 412–3412 g g−1 ). Substrate induced -glucosidase activity ranged between 1.12 and 6.12 mol para-nitrophenol g−1 soil h−1 and was positively correlated with concentrations of clay, total carbon and microbial carbon. This suggests that substrate-induced -glucosidase activity is an integrative measure of physico-chemical and biological soil properties and may have applications in monitoring biological soil quality. © 2002 Elsevier Science B.V. All rights reserved. Keywords: -Glucosidase; Soil; Microbial biomass; Carbon; Grassland; Soil quality
1. Introduction Of the extracellular enzymes in soils, those involved in the degradation of soil organic matter are of particular interest. -Glucosidase (EC 3.2.1.21; obsolete name gentiobiase or cellobiase) is one such enzyme, being involved in the enzymatic degradation of cellulose, the main component of plant polysaccharides. Cellulose consists of polymer chains of -1,4, linked glucose units and its enzymatic degradation Abbreviations: Cmic , soil microbial carbon; Ctotal , total soil carbon; Nmic , soil microbial nitrogen; Ntotal , total soil nitrogen; pNP, para-nitrophenol; pNPG, para-nitrophenyl--d-glucopyranoside ∗ Corresponding author. Present address: United States Department of Agriculture–Agricultural Research Service, Northwest Irrigation and Soils Research Laboratory, 3793 N. 3600 E., Kimberly, ID 83341, USA. Tel.: +1-208-423-6524; fax: +1-208-423-6555. E-mail address: [email protected] (B.L. Turner).
is initiated by endo--1,4-glucanase (EC 3.1.2.4), which breaks cellulose chains into smaller units, and cellobiohydrolase (EC 3.1.2.91), which cleaves the dimer cellobiose (two -1,4 linked glucose units) from the reducing ‘ends’ of the molecules. -Glucosidase completes the hydrolysis process by catalysing the cleavage of cellobiose to release two moles of glucose per mole of cellobiose and, therefore, regulates the supply of an important energy source for microorganisms unable to directly take up cellobiose. Indeed, -glucosidase activity may be the rate-limiting step in cellulose degradation (Alef and Nannipieri, 1995). -Glucosidase is derived predominantly from soil microbial heterotrophs, in particular members of the mucorales (fungi), such as Actinomucor or Mortierella (Hayano and Tubaki, 1985). Its synthesis in such organisms is induced by the products of cellulose breakdown, including cellobiose, glucose
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B.L. Turner et al. / Applied Soil Ecology 20 (2002) 157–162
and their metabolites (Stewart and Leatherwood, 1976). There is currently great interest in the use of extracellular enzymes as biological indicators of soil quality, because they are relatively simple to determine, have microbial ecological significance, are sensitive to environmental stress and respond rapidly to changes in land management (Dick, 1997; Yakovchenko et al., 1996). -Glucosidase activity may be a particularly useful enzyme for soil quality monitoring because of its central role in soil organic matter cycling, which is generally regarded as an important component of soil quality. Research has shown that -glucosidase is the most abundant and easily detected of the three enzymes involved in cellulose degradation in soil, and is rarely substrate limited, thus making it ideal to examine the importance of physico-chemical controls on the turnover of soil organic matter (Debosz et al., 1999; Eivazi and Tabatabai, 1988). Indeed, it provides an early indication of changes in organic matter status and turnover (Bandick and Dick, 1999; Debosz et al., 1999; Monreal and Bergstrom, 2000). The adoption of soil enzyme activities in soil quality monitoring requires information on activities from a wide range of soil types and land uses under steady-state conditions, in addition to a mechanistic understanding of how soil properties control these activities (Trasar-Cepeda et al., 2000). The aims of this work were (i) to determine -glucosidase activity in a wide range of soils with contrasting physico-chemical and biological properties, but under similar vegetation/land use, and (ii) to investigate the factors controlling -glucosidase in those soils.
2. Methods 2.1. Soil collection and description Twenty-nine permanent lowland grassland soils were sampled during October 1998 from sites around England and Wales to give a wide range of organic matter and textural characteristics. Intact turves were taken to 10 cm depth from each site and transported back to the laboratory in plastic trays. Water-holding capacity was determined by saturating each turf and allowing drainage to field capacity under cover for 48 h at ambient temperature. Duplicate cores were
dried at 105 ◦ C to determine moisture content. Each soil was sieved (4 mm) to remove large roots, stones and macrofauna, before being left to equilibrate for 1 week at 10–15 ◦ C. 2.2. β-Glucosidase assay -Glucosidase activity was assayed by the method of Eivazi and Tabatabai (1988), using the substrate analogue para-nitrophenyl--d-glucopyranoside (pNPG). The concentrations of buffer and terminator solutions were increased from those used in the original method to account for the greater buffering capacity of our soils. Moist soil (1.00 g) was weighed into screw-cap glass test tubes (three replicate samples per soil) and incubated for 1 h in a water bath at 37 ◦ C with 4 ml of 0.05 M modified universal buffer (pH 6.0) and 1 ml of 25 mM pNPG (5 mM final concentration) dissolved in buffer. The reaction was terminated by adding 1 ml of 0.5 M CaCl2 and 4 ml of 0.2 M Tris–hydroxymethyl(aminomethane), adjusted to pH 12 with NaOH. The mixture was centrifuged for 10 min at 1500 × g and the absorbance measured at 410 nm. Values were corrected for a blank (substrate added immediately after the addition of CaCl2 and Tris–NaOH) and for adsorption of released para-nitrophenol ( pNP) in the soil (Vuorinen, 1993). -Glucosidase activity is expressed as mol pNP released g−1 dry soil h−1 . 2.3. Determination of other soil properties Microbial carbon (Cmic ) and microbial nitrogen (Nmic ) were determined by chloroform fumigation and extraction with K2 SO4 (Brookes et al., 1985; Vance et al., 1987). Total soil carbon (Ctotal ) and total soil nitrogen (Ntotal ) were determined simultaneously using a Carlo–Erba model NA2000 nitrogen analyser. Soil textural information was obtained by wet sieving followed by analysis using a micromeritics Sedigraph 5100 with a micromeritics Mastertech 51 automatic sampler. Soil pH was determined in a 1:2.5 soil-to-deionised water ratio. An index of soil C quality was provided by the Cmic -to-Ctotal ratio (%) (i.e. the proportion of the Ctotal in the microbial biomass). The ratio of Cmic -to-Ctotal varies between <1 and 5% and has proved to be a relatively sensitive indicator of the state and changes in soil organic matter
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Table 2 Correlation coefficients (r values) for relationships between soil properties in 29 permanent lowland grassland soils from England and Wales
Clay pH Total C Total N Total C-to-N ratio Microbial C Microbial N Microbial C-to-N ratio Microbial C-to-total C ratio -Glucosidase activity
Clay
pH
Total C
Total N
1.00 0.20 0.79∗∗∗ 0.73∗∗∗ 0.12 0.63∗∗∗ 0.54∗∗ 0.29 0.16 0.64∗∗∗
1.00 0.25 0.15 0.27 −0.14 0.01 −0.34 −0.43∗ 0.05
1.00 0.96∗∗∗ −0.05 0.74∗∗∗ 0.73∗∗∗ 0.19 0.15 0.77∗∗∗
1.00 −0.32 0.78∗∗∗ 0.83∗∗∗ 0.08 0.26 0.81∗∗∗
Total C-to-N ratio
1.00 −0.07 −0.26 0.30 −0.42∗ −0.27
Microbial C
Microbial N
1.00 0.87∗∗∗ 0.34 0.76∗∗∗ 0.89∗∗∗
1.00 −0.11 0.61∗∗∗ 0.77∗∗∗
Microbial C-to-N ratio
Microbial C-to-total C ratio
1.00 0.33 0.29
1.00 0.51∗∗
Significant relationships are indicated in bold letters. Significance levels are denoted by (*), (**) and (***), representing significance at the 5, 1 and 0.1% levels, respectively.
(Anderson and Domsch, 1989; Mazzarino et al., 1993; Sparling, 1992). We considered that, assuming stable conditions, greater Cmic -to-Ctotal ratios indicated less recalcitrant soil organic matter.
3. Results Properties of the 29 soils are presented in Table 1. Total C concentrations ranged from 28.9 to 80.4 mg
C g−1 , Ntotal concentrations from 2.85 to 8.70 mg N g−1 , clay contents from 22 to 68%, and pH values from 4.4 to 6.8. The concentrations of Cmic (between 412 and 3412 g C g−1 soil) and Nmic (between 57 and 346 g N g−1 soil) have been previously reported (Turner et al., 2001). Microbial C as a fraction of the Ctotal (Cmic -to-Ctotal ratio) ranged between 1.1 and 4.7% (Table 1). Strong positive correlations (P < 0.001) existed between Ctotal , Ntotal and clay content (Table 2).
Fig. 1. Relationships between -glucosidase activity (mol pNP g−1 dry soil h−1 ) in 29 permanent lowland grassland soils from England and Wales and (a) clay content (%); (b) total soil C (mg g−1 dry soil); (c) microbial C (mg g−1 dry soil); (d) microbial C-to-total soil C ratio (%).
B.L. Turner et al. / Applied Soil Ecology 20 (2002) 157–162
Similarly, Cmic and Nmic were strongly positively correlated with clay content, Ctotal and Ntotal . Soil pH was not influential in these relationships, but was significantly negatively correlated with the Cmic -to-Ctotal ratio (P < 0.05). The Cmic -to-Ctotal ratio was poorly correlated with clay content, indicating the insensitivity of this parameter to soil texture. The activity of -glucosidase varied amongst the 29 soils between 1.12 and 6.12 mol pNP g−1 dry soil h−1 (Table 1). The activity was most strongly correlated with Cmic (r = 0.89), but was also strongly positively correlated with clay content, Ctotal and Ntotal , Nmic and the Cmic -to-Ctotal ratio (Table 2 and Fig. 1). 4. Discussion The values of -glucosidase activity reported here are similar to those in the literature for soils with similar organic matter contents. For example, in the A horizons of oak woodland soils of NW Spain, -glucosidase activity varied between 0.67 and 4.58 mol pNP g−1 dry soil h−1 , with activity in the O horizons between 2.05 and 29.6 mol pNP g−1 dry soil h−1 (Trasar-Cepeda et al., 2000). Eivazi and Tabatabai (1988) measured -glucosidase activities of 0.42–3.78 mol pNP g−1 dry soil h−1 for seven soils with organic C contents between 5 and 55 mg C g−1 soil, whilst Bandick and Dick (1999) measured activities between 0.29 and 2.11 mol pNP g−1 dry soil h−1 in soils with organic matter contents between 23 and 41 mg C g−1 soil under various arable rotations and fertiliser/manure applications. Most of the variation in -glucosidase activity in the soils was explained by Cmic and Ctotal , which appears logical because -glucosidase is synthesised by soil microorganisms in response to the presence of suitable substrate. Therefore, its activity would be expected to reflect labile organic C turnover in the soil, confirmed by the strong correlations between -glucosidase activity and the Cmic -to-Ctotal ratio. The strong relationship with Cmic also suggests that the activity of extracellular immobilised enzymes is negligible or unimportant, which would agree with the results of Kiss et al. (1972), who stated that cellobiose degradation in soils was due to enzymes released from proliferating organisms rather than the accumulated enzyme fraction. Alternatively,
161
the relationship between -glucosidase activity and clay content might reflect the potential for enzyme immobilisation in the soil and, therefore, the dominance of immobilised extracellular enzymes, as suggested by several studies (Busto and Perez-Mateos, 2000; Hayano and Katami, 1977; Lensi et al., 1991). Clearly, the ecological significance of -glucosidase activity measurements cannot be understood without information on the location of these enzymes in soil. Our results suggest that, in the absence of substrate-limitation (a scenario unlikely to occur in cellulose-loaded pasture soils), substrate-induced -glucosidase activity is regulated by processes that control general heterotroph activity. Clay content is clearly the key factor. By providing physical protection for organic matter, microbes, nutrients and enzymes, clay content has both direct and indirect effects on soil biological quality and, most probably, the ability of soil to resist and recover from changes/perturbations. The close relationships between -glucosidase activity and these soil properties (microbial biomass, soil organic matter and soil texture) suggests that -glucosidase activity can provide a meaningful integrative measure of physico-chemical and biological soil quality parameters and, therefore, has a potentially important role in monitoring soil biological quality.
Acknowledgements The authors thank the National Soil Resources Institute for access to the National Soil Inventory database. Ben Turner was funded by the UK Natural Environment Research Council.
References Alef, K., Nannipieri, P., 1995. -Glucosidase activity. In: Alef, K., Nannipieri, P. (Eds.), Methods in Applied Soil Microbiology and Biochemistry. Academic Press, London, UK, pp. 24–28. Anderson, T.H., Domsch, K.H., 1989. Ratios of microbial biomass carbon to total organic carbon in arable soils. Soil Biol. Biochem. 21, 471–479. Bandick, A.K., Dick, R.P., 1999. Field management effects on soil enzyme activities. Soil Biol. Biochem. 31, 1471–1479. Brookes, P.C., Landman, A., Pruden, G., Jenkinson, D.S., 1985. Chloroform fumigation and the release of soil nitrogen: a rapid
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direct extraction method to measure microbial biomass nitrogen in soil. Soil Biol. Biochem. 17, 837–842. Busto, M.D., Perez-Mateos, M., 2000. Characterization of -dglucosidase extracted from soil fractions. Eur. J. Soil Sci. 51, 193–200. Debosz, K., Rasmussen, P.H., Pedersen, A.R., 1999. Temporal variations in microbial biomass C and cellulolytic enzyme activity in arable soils: effects of organic matter input. Appl. Soil Ecol. 13, 209–218. Dick, R.P., 1997. Soil enzyme activities as integrative indicators of soil health. In: Pankhurst, C.E., Doube, B.M., Gupta, V.V.S.R. (Eds.), Biological Indicators of Soil Health. CAB International, Wallingford, UK, pp. 121–156. Eivazi, F., Tabatabai, M.A., 1988. Glucosidases and galactosidases in soils. Soil Biol. Biochem. 20, 601–606. Hayano, K., Katami, K., 1977. Extraction of -glucosidase activity from pea-field soil. Soil Biol. Biochem. 9, 349–351. Hayano, K., Tubaki, K., 1985. Origin and properties of -glucosidase activity of tomato-field soil. Soil Biol. Biochem. 17, 553–557. Kiss, S., Drˇagan-Bularda, M., Khaziev, F.H., 1972. Influence of chloromycetin on the activities of some oligases of soils. Lucrãri Conferintã Nationalã Stiintã Solului, Iasi 1970, 451–462. Lensi, R., Lescure, C., Steinberg, C., Savoie, J.M., Faurie, G., 1991. Dynamics of residual enzyme activities, denitrification potential, and physico-chemical properties in a ␥-sterilized soil. Soil Biol. Biochem. 23, 367–373.
Mazzarino, M.J., Szott, L., Jiminez, M., 1993. Dynamics of soil total-C and total-N, microbial biomass, and water-soluble-C in tropical agroecosystems. Soil Biol. Biochem. 25, 205–214. Monreal, C.M., Bergstrom, D.W., 2000. Soil enzymatic factors expressing the influence of land use, tillage system and texture on soil biochemical quality. Can. J. Soil Sci. 80, 419–428. Sparling, G.P., 1992. Ratio of microbial biomass carbon to soil organic carbon as a sensitive indicator of changes in soil organic matter. Aust. J. Soil Res. 30, 195–207. Stewart, B.J., Leatherwood, J.M., 1976. Depressed synthesis of cellulase by Cellulomonas. J. Bacteriol. 128, 609–615. Trasar-Cepeda, C., Leirós, M.C., Gil-Sotres, F., 2000. Biochemical properties of acid soils under climax vegetation (Atlantic oakwood) in an area of the European temperate-humid zone (Galacia, NW Spain): specific parameters. Soil Biol. Biochem. 32, 747–755. Turner, B.L., Bristow, A.W., Haygarth, P.M., 2001. Rapid estimation of microbial biomass in grassland soils by ultra-violet absorbance. Soil Biol. Biochem. 33, 913–919. Vance, E.D., Brookes, P.C., Jenkinson, D.S., 1987. An extraction method for measuring soil microbial biomass C. Soil Biol. Biochem. 19, 703–707. Vuorinen, A.H., 1993. Requirement of p-nitrophenol standard for each soil. Soil Biol. Biochem. 25, 295–296. Yakovchenko, V., Sikora, L.J., Kaufman, D.D., 1996. A biologically based indicator of soil quality. Biol. Fertility Soils 21, 245–251.
Short communication
-Glucosidase activity in pasture soils Benjamin L. Turner a,∗ , David W. Hopkins b , Philip M. Haygarth a , Nick Ostle c a
Institute of Grassland and Environmental Research, North Wyke Research Station, Okehampton, Devon EX20 2SB, UK b Department of Environmental Science, University of Stirling, Stirling FK9 4LA, UK c Centre for Ecology and Hydrology, Merlewood Research Station, Grange-Over-Sands, Cumbria LA11 6JU, UK Received 8 January 2002
Abstract -Glucosidase is involved in the degradation of cellulose in soils and has potential for monitoring biological soil quality. We assayed -glucosidase activity in 29 permanent grassland soils from England and Wales with contrasting physico-chemical and biological properties (clay content 22–68%; total carbon 29–80 mg g−1 ; microbial carbon 412–3412 g g−1 ). Substrate induced -glucosidase activity ranged between 1.12 and 6.12 mol para-nitrophenol g−1 soil h−1 and was positively correlated with concentrations of clay, total carbon and microbial carbon. This suggests that substrate-induced -glucosidase activity is an integrative measure of physico-chemical and biological soil properties and may have applications in monitoring biological soil quality. © 2002 Elsevier Science B.V. All rights reserved. Keywords: -Glucosidase; Soil; Microbial biomass; Carbon; Grassland; Soil quality
1. Introduction Of the extracellular enzymes in soils, those involved in the degradation of soil organic matter are of particular interest. -Glucosidase (EC 3.2.1.21; obsolete name gentiobiase or cellobiase) is one such enzyme, being involved in the enzymatic degradation of cellulose, the main component of plant polysaccharides. Cellulose consists of polymer chains of -1,4, linked glucose units and its enzymatic degradation Abbreviations: Cmic , soil microbial carbon; Ctotal , total soil carbon; Nmic , soil microbial nitrogen; Ntotal , total soil nitrogen; pNP, para-nitrophenol; pNPG, para-nitrophenyl--d-glucopyranoside ∗ Corresponding author. Present address: United States Department of Agriculture–Agricultural Research Service, Northwest Irrigation and Soils Research Laboratory, 3793 N. 3600 E., Kimberly, ID 83341, USA. Tel.: +1-208-423-6524; fax: +1-208-423-6555. E-mail address: [email protected] (B.L. Turner).
is initiated by endo--1,4-glucanase (EC 3.1.2.4), which breaks cellulose chains into smaller units, and cellobiohydrolase (EC 3.1.2.91), which cleaves the dimer cellobiose (two -1,4 linked glucose units) from the reducing ‘ends’ of the molecules. -Glucosidase completes the hydrolysis process by catalysing the cleavage of cellobiose to release two moles of glucose per mole of cellobiose and, therefore, regulates the supply of an important energy source for microorganisms unable to directly take up cellobiose. Indeed, -glucosidase activity may be the rate-limiting step in cellulose degradation (Alef and Nannipieri, 1995). -Glucosidase is derived predominantly from soil microbial heterotrophs, in particular members of the mucorales (fungi), such as Actinomucor or Mortierella (Hayano and Tubaki, 1985). Its synthesis in such organisms is induced by the products of cellulose breakdown, including cellobiose, glucose
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and their metabolites (Stewart and Leatherwood, 1976). There is currently great interest in the use of extracellular enzymes as biological indicators of soil quality, because they are relatively simple to determine, have microbial ecological significance, are sensitive to environmental stress and respond rapidly to changes in land management (Dick, 1997; Yakovchenko et al., 1996). -Glucosidase activity may be a particularly useful enzyme for soil quality monitoring because of its central role in soil organic matter cycling, which is generally regarded as an important component of soil quality. Research has shown that -glucosidase is the most abundant and easily detected of the three enzymes involved in cellulose degradation in soil, and is rarely substrate limited, thus making it ideal to examine the importance of physico-chemical controls on the turnover of soil organic matter (Debosz et al., 1999; Eivazi and Tabatabai, 1988). Indeed, it provides an early indication of changes in organic matter status and turnover (Bandick and Dick, 1999; Debosz et al., 1999; Monreal and Bergstrom, 2000). The adoption of soil enzyme activities in soil quality monitoring requires information on activities from a wide range of soil types and land uses under steady-state conditions, in addition to a mechanistic understanding of how soil properties control these activities (Trasar-Cepeda et al., 2000). The aims of this work were (i) to determine -glucosidase activity in a wide range of soils with contrasting physico-chemical and biological properties, but under similar vegetation/land use, and (ii) to investigate the factors controlling -glucosidase in those soils.
2. Methods 2.1. Soil collection and description Twenty-nine permanent lowland grassland soils were sampled during October 1998 from sites around England and Wales to give a wide range of organic matter and textural characteristics. Intact turves were taken to 10 cm depth from each site and transported back to the laboratory in plastic trays. Water-holding capacity was determined by saturating each turf and allowing drainage to field capacity under cover for 48 h at ambient temperature. Duplicate cores were
dried at 105 ◦ C to determine moisture content. Each soil was sieved (4 mm) to remove large roots, stones and macrofauna, before being left to equilibrate for 1 week at 10–15 ◦ C. 2.2. β-Glucosidase assay -Glucosidase activity was assayed by the method of Eivazi and Tabatabai (1988), using the substrate analogue para-nitrophenyl--d-glucopyranoside (pNPG). The concentrations of buffer and terminator solutions were increased from those used in the original method to account for the greater buffering capacity of our soils. Moist soil (1.00 g) was weighed into screw-cap glass test tubes (three replicate samples per soil) and incubated for 1 h in a water bath at 37 ◦ C with 4 ml of 0.05 M modified universal buffer (pH 6.0) and 1 ml of 25 mM pNPG (5 mM final concentration) dissolved in buffer. The reaction was terminated by adding 1 ml of 0.5 M CaCl2 and 4 ml of 0.2 M Tris–hydroxymethyl(aminomethane), adjusted to pH 12 with NaOH. The mixture was centrifuged for 10 min at 1500 × g and the absorbance measured at 410 nm. Values were corrected for a blank (substrate added immediately after the addition of CaCl2 and Tris–NaOH) and for adsorption of released para-nitrophenol ( pNP) in the soil (Vuorinen, 1993). -Glucosidase activity is expressed as mol pNP released g−1 dry soil h−1 . 2.3. Determination of other soil properties Microbial carbon (Cmic ) and microbial nitrogen (Nmic ) were determined by chloroform fumigation and extraction with K2 SO4 (Brookes et al., 1985; Vance et al., 1987). Total soil carbon (Ctotal ) and total soil nitrogen (Ntotal ) were determined simultaneously using a Carlo–Erba model NA2000 nitrogen analyser. Soil textural information was obtained by wet sieving followed by analysis using a micromeritics Sedigraph 5100 with a micromeritics Mastertech 51 automatic sampler. Soil pH was determined in a 1:2.5 soil-to-deionised water ratio. An index of soil C quality was provided by the Cmic -to-Ctotal ratio (%) (i.e. the proportion of the Ctotal in the microbial biomass). The ratio of Cmic -to-Ctotal varies between <1 and 5% and has proved to be a relatively sensitive indicator of the state and changes in soil organic matter
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Table 2 Correlation coefficients (r values) for relationships between soil properties in 29 permanent lowland grassland soils from England and Wales
Clay pH Total C Total N Total C-to-N ratio Microbial C Microbial N Microbial C-to-N ratio Microbial C-to-total C ratio -Glucosidase activity
Clay
pH
Total C
Total N
1.00 0.20 0.79∗∗∗ 0.73∗∗∗ 0.12 0.63∗∗∗ 0.54∗∗ 0.29 0.16 0.64∗∗∗
1.00 0.25 0.15 0.27 −0.14 0.01 −0.34 −0.43∗ 0.05
1.00 0.96∗∗∗ −0.05 0.74∗∗∗ 0.73∗∗∗ 0.19 0.15 0.77∗∗∗
1.00 −0.32 0.78∗∗∗ 0.83∗∗∗ 0.08 0.26 0.81∗∗∗
Total C-to-N ratio
1.00 −0.07 −0.26 0.30 −0.42∗ −0.27
Microbial C
Microbial N
1.00 0.87∗∗∗ 0.34 0.76∗∗∗ 0.89∗∗∗
1.00 −0.11 0.61∗∗∗ 0.77∗∗∗
Microbial C-to-N ratio
Microbial C-to-total C ratio
1.00 0.33 0.29
1.00 0.51∗∗
Significant relationships are indicated in bold letters. Significance levels are denoted by (*), (**) and (***), representing significance at the 5, 1 and 0.1% levels, respectively.
(Anderson and Domsch, 1989; Mazzarino et al., 1993; Sparling, 1992). We considered that, assuming stable conditions, greater Cmic -to-Ctotal ratios indicated less recalcitrant soil organic matter.
3. Results Properties of the 29 soils are presented in Table 1. Total C concentrations ranged from 28.9 to 80.4 mg
C g−1 , Ntotal concentrations from 2.85 to 8.70 mg N g−1 , clay contents from 22 to 68%, and pH values from 4.4 to 6.8. The concentrations of Cmic (between 412 and 3412 g C g−1 soil) and Nmic (between 57 and 346 g N g−1 soil) have been previously reported (Turner et al., 2001). Microbial C as a fraction of the Ctotal (Cmic -to-Ctotal ratio) ranged between 1.1 and 4.7% (Table 1). Strong positive correlations (P < 0.001) existed between Ctotal , Ntotal and clay content (Table 2).
Fig. 1. Relationships between -glucosidase activity (mol pNP g−1 dry soil h−1 ) in 29 permanent lowland grassland soils from England and Wales and (a) clay content (%); (b) total soil C (mg g−1 dry soil); (c) microbial C (mg g−1 dry soil); (d) microbial C-to-total soil C ratio (%).
B.L. Turner et al. / Applied Soil Ecology 20 (2002) 157–162
Similarly, Cmic and Nmic were strongly positively correlated with clay content, Ctotal and Ntotal . Soil pH was not influential in these relationships, but was significantly negatively correlated with the Cmic -to-Ctotal ratio (P < 0.05). The Cmic -to-Ctotal ratio was poorly correlated with clay content, indicating the insensitivity of this parameter to soil texture. The activity of -glucosidase varied amongst the 29 soils between 1.12 and 6.12 mol pNP g−1 dry soil h−1 (Table 1). The activity was most strongly correlated with Cmic (r = 0.89), but was also strongly positively correlated with clay content, Ctotal and Ntotal , Nmic and the Cmic -to-Ctotal ratio (Table 2 and Fig. 1). 4. Discussion The values of -glucosidase activity reported here are similar to those in the literature for soils with similar organic matter contents. For example, in the A horizons of oak woodland soils of NW Spain, -glucosidase activity varied between 0.67 and 4.58 mol pNP g−1 dry soil h−1 , with activity in the O horizons between 2.05 and 29.6 mol pNP g−1 dry soil h−1 (Trasar-Cepeda et al., 2000). Eivazi and Tabatabai (1988) measured -glucosidase activities of 0.42–3.78 mol pNP g−1 dry soil h−1 for seven soils with organic C contents between 5 and 55 mg C g−1 soil, whilst Bandick and Dick (1999) measured activities between 0.29 and 2.11 mol pNP g−1 dry soil h−1 in soils with organic matter contents between 23 and 41 mg C g−1 soil under various arable rotations and fertiliser/manure applications. Most of the variation in -glucosidase activity in the soils was explained by Cmic and Ctotal , which appears logical because -glucosidase is synthesised by soil microorganisms in response to the presence of suitable substrate. Therefore, its activity would be expected to reflect labile organic C turnover in the soil, confirmed by the strong correlations between -glucosidase activity and the Cmic -to-Ctotal ratio. The strong relationship with Cmic also suggests that the activity of extracellular immobilised enzymes is negligible or unimportant, which would agree with the results of Kiss et al. (1972), who stated that cellobiose degradation in soils was due to enzymes released from proliferating organisms rather than the accumulated enzyme fraction. Alternatively,
161
the relationship between -glucosidase activity and clay content might reflect the potential for enzyme immobilisation in the soil and, therefore, the dominance of immobilised extracellular enzymes, as suggested by several studies (Busto and Perez-Mateos, 2000; Hayano and Katami, 1977; Lensi et al., 1991). Clearly, the ecological significance of -glucosidase activity measurements cannot be understood without information on the location of these enzymes in soil. Our results suggest that, in the absence of substrate-limitation (a scenario unlikely to occur in cellulose-loaded pasture soils), substrate-induced -glucosidase activity is regulated by processes that control general heterotroph activity. Clay content is clearly the key factor. By providing physical protection for organic matter, microbes, nutrients and enzymes, clay content has both direct and indirect effects on soil biological quality and, most probably, the ability of soil to resist and recover from changes/perturbations. The close relationships between -glucosidase activity and these soil properties (microbial biomass, soil organic matter and soil texture) suggests that -glucosidase activity can provide a meaningful integrative measure of physico-chemical and biological soil quality parameters and, therefore, has a potentially important role in monitoring soil biological quality.
Acknowledgements The authors thank the National Soil Resources Institute for access to the National Soil Inventory database. Ben Turner was funded by the UK Natural Environment Research Council.
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