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Water-stable aggregates, glomalin-related soil protein, and carbohydrates in a chronosequence of sandy hydromorphic soils
Soil Biology & Biochemistry 42 (2010) 1505e1511
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Water-stable aggregates, glomalin-related soil protein, and carbohydrates in a chronosequence of sandy hydromorphic soils Marie Spohn*, Luise Giani Soil Science Group, Department of Biology and Environmental Sciences, Carl von Ossietzky University Oldenburg, 26121 Oldenburg, Germany
a r t i c l e i n f o
a b s t r a c t
Article history: Received 5 January 2010 Received in revised form 30 April 2010 Accepted 17 May 2010 Available online 31 May 2010
The conversion of pasture to cropland leads to a decline of aggregation in topsoils and to a decrease of aggregate-binding agents such as carbohydrates and glomalin-related soil protein (GRSP). Till now, studies on soil aggregation focused either on carbohydrates or on GRSP as a binding agent in aggregates. In this study we analyse the development of the relationship between carbohydrates, GRSP, TOC and aggregatestability following land-use change. Furthermore, we discuss the contents of carbohydrates, GRSP and TOC in each of the aggregate fractions. For these purposes, a chronosequence of sites, which were converted from pasture to cropland at different periods in history, was established. To get further insight into the impact of different types of land-use, also soils under forest, either afforested or permanent, were studied. The mean-weight diameter (MWD) of water-stable aggregates, the carbohydrate, and the GRSP content were determined in 49 soils. It was found that the MWD of the water-stable aggregates decreased monoexponentially (R2 ¼ 0.66) by 66% during the first 46 years after conversion of the soils from pasture to cropland. During the same period, the carbohydrate content decreased very rapidly after the land use change by 64% and the GRSP content decreased more slowly by 57%. The MWD of the forest soils were in the same range as those of the permanent pasture soils although they exhibit significantly higher TOC contents, which indicate that other stabilization mechanisms are dominant in forest soils, less important in the chronosequence soils. TOC, carbohydrates and the GRSP contents were sigmoidally correlated with the MWD. Among the four water-stable aggregate fractions TOC and carbohydrates exhibited high contents in the macroaggregates and were less present in the microaggregates. GRSP, in contrast, was more equally distributed among the four water-stable aggregate fractions. Ó 2010 Elsevier Ltd. All rights reserved.
Keywords: Water-stable aggregates Glomalin-related soil protein Carbohydrates Chronosequence Sandy hydromorphic soils
1. Introduction Soil aggregation supports soil fertility as it reduces erosion and mediates soil aeration, water infiltration, and retention (Oades, 1984). Furthermore, soil aggregation protects soil organic matter (SOM) from getting mineralized as it physically reduces the accessibility of organic compounds for microorganisms and exoenzymes and the oxygen diffusion (Lützow et al., 2006). Aggregation is mediated by soil organic matter (SOM), biota, ionic bridging, clay, and carbonates (Bronick and Lal, 2005). A variety of binding mechanisms works simultaneously at different spatial scales to stabilize aggregates. Tisdall and Oades (1982) stated that primarily, mineral particles are bound together by persistent binding agents like biotic debris while these microaggregates, in turn, are bound together to macroaggregates by transient and temporal binding agents (polysaccharides, roots, and fungal hyphae). This * Corresponding author. Tel.: þ49 (0)441 798 4504. E-mail address: [email protected] (M. Spohn). 0038-0717/$ e see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.soilbio.2010.05.015
aggregate hierarchy theory has been used by many authors to explain correlations between a reduction of aggregation and a loss of SOC (Six et al., 2000, 2004). More recently, it is assumed that microaggregates are formed within macroaggregates and protect SOM more effectively than macroaggregates (Oades, 1984; Puget et al., 2000; Six et al., 2000). Aggregation is influenced by land use and land use change in the way that the proportion of water stable macroaggregates is reduced in the order forest > pasture/grassland > arable land and is further diminished with the duration of arable use (Ashagrie et al., 2007; Haynes et al., 1991; Jastrow, 1996; John et al., 2005). Microaggregates, however, seem to be less influenced by land use (Oades, 1984; Besnard et al., 1996; Puget et al., 2000). Polysaccharides from bacteria, fungi or root mucilage are described as a labile SOM fraction which acts as an important binding agent for aggregation (Jolivet et al., 2006; Oades, 1984; Puget et al., 1999). More recently, also the heat-stable protein glomalin which is produced by arbuscular mychorriza has been shown to cause soil aggregation (Bedini et al., 2009; Harner et al., 2004; Wright and Upadhyaya, 1998).
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Glomalin promotes soil aggregation especially in coarse textured soils and its content in soils decreases with the duration of the arable use of the soils (Preger et al., 2007; Rillig et al., 2003). Till now, studies on soil aggregation focused either on carbohydrates or on GRSP as a binding agent in aggregates. Hence, the main objective of this study was to study the development of the relationship between carbohydrates, GRSP, TOC and aggregatestability following land-use change. Furthermore, we discuss the contents of carbohydrates, GRSP and TOC in each of the aggregate factions. For these purposes, a chronosequence of sites, which were converted from pasture to cropland at different periods in history, was established. To get further insight into the impact of different types of land-use, also soils under forest, either afforested or permanent, were studied. 2. Material and methods 2.1. Study site, geo-data sources and processing
soil. The mixed samples were homogenized, oven dried (70 C, 96 h), and subsequently crushed to pass a screen (2 mm-opening). 70 C was chosen as a drying temperature in order to prevent microbial degradation during the drying process. 2.3. Water-stable aggregate fractionation Water-stable aggregates were fractionated according to Kemper and Rosenau (1986). 70 g of oven dried soil were soaked in H2O dest. on a 2000 mm sieve for 10 min to allow slaking. Subsequently, the sieve was moved up and down in water by about 3 cm with 50 repetitions. The fraction < 2000 mm was passed on to a 630 mm sieve. This procedure was repeated with a 200 and a 63 mm sieve. Each fraction was recollected from the sieve, dried at 70 C, and weighed. The mean weight diameter (MWD) of each sample was calculated by
MWD ¼
The study was conducted in a region called Artland, 40 km north of Osnabrück in Northwest Germany (52 360 -52 410 N, 7 540 -8 140 E). The area was formed in the Pleistocene, and Gleyic Podzols, Haplic Podzols, and Haplic Gleysols represent its main soil groups today. It is characterized by an annual mean precipitation of 800 mm and an annual mean air temperature of 8.7 C. Historical topographic maps which contain precise information on land use were used to attribute former uses of arable land, pasture, and forest in order to establish a chronosequence. Sites, which were converted from pasture to arable land at defined periods of time were selected. As a zero point of the chronosequence, permanent pasture sites were chosen. In addition, sites under forest which have either been used as forest at least since 1789 (permanent forest) or were afforested between 1851 and 1900 (old forest) were selected. All sites have a size of 0.8 to 1.7 ha. For this study, we exclusively sampled Gleyic Podzols and Haplic Gleysols as we expected sandy hydromorphic soils to react very sensitively to land use changes (Spohn and Giani, in press). All sampled soils are situated in the four sampling subregions Lohne, Gröhnloh, Mühlen, and Nortrup and exhibit very similar textures, Alox, Feox contents and pH (Table 1). 2.2. Sampling The upper 20 cm of 49 arable, pasture, and forest sites were sampled in late October 2008 and February 2009. This sampling depth was chosen as a common sampling depth in studies dealing with SOM responses to land use change (Mann, 1986). On every site samples were taken each 10 m along two diagonal transects, thus the number of samples depends on the size of the site. The minimal amount of subsamples was 20. Samples of each site were combined to gain one mixed sample per site. On the forest sites, litter and the organic layer were removed in order to sample solely the mineral
n X
Xi Wi ;
i¼1
where Xi is the mean diameter of each size fraction and Wi is the proportion of the total sample weight in the corresponding size fraction (Kemper and Rosenau, 1986). 2.4. Carbohydrate extraction and determination The extraction procedure for the carbohydrates was slightly modified from the method described by Martens and Frankenberger (1990). Briefly, 2 g of oven dried soil were treated with 20 ml 0.125 M H2SO4 in a water bath at 80 C for 7 h and were vortexed for 10 s every 30 min during the extraction procedure. Subsequently, the samples were treated with EDTA (ethylene diaminetetraacetate) in order to prevent co-precipitation of the sugars with cations. The assay was titrated to pH 3.5e4.0 with 5 M KOH and centrifuged. The carbohydrates in the supernatant were determined following Waffenschmidt and Jaenicke (1987) by the 2,20 Bicinchoninate-assay (BCA-assay). 20 ml of the supernatant were added to 2 ml of the BCA-assay working reagent and incubated at 70 C for 60 min. After cooling down, to room temperature the samples were measured photometrically at 560 nm (UVmini1240, Shimadzu). D-Glucose was used as a standard. 2.5. Glomalin extraction and determination Glomalin was extracted following the method of Wright and Upadhyaya (1996, 1998), but with 50 mM diphosphate (adjusted to pH 8.0) as extractant as suggested by Halvorson and Gonzales (2006). Briefly, 2 g of oven dried soil were extracted by a total volume of 50 ml diphosphate at 128 C during four extraction cycles, each lasting for 1 h. The pooled extracts were centrifuged to remove soil particles and protein concentration was determined by
Table 1 Sampling subregions, number of sites per subregion, and particle size distribution, pH and oxalate extractable Fe and Al of the investigated Gleyic Podzols and Haplic Gleysols. Standard deviations calculated from triplicates of one sample per site are given in brackets. Sampling subregion
Number of sites
Soilgroup
Texture Sand [%]
Silt [%]
Clay [%]
Lohne Gröhnloh Mühlen Nortup
24 12 8 5
Gleyic Podzol Gleyic Podzol Haplic Gleysol Haplic Gleysol
85.0 80.5 80.0 80.0
10.6 (6.4) 14.8 (7.0) 9.6 (1.7) 16.0 (3.9)
4.4 (4.1) 4.6 (1.9) 9.9 (2.0) 4.0 (2.1)
a b
Exclusive forest sites: pHH2O 3.4 (0.3). Exclusive forest sites: Feox 11.9 (1.0) g kg1.
(8.2) (8.4) (3.4) (3.5)
pHH2Oa
Feoxb [g kg1]
Alox [g kg1]
5.5 (1.1) 5.6 (0.9) 5.6 (0.3) 5.7 (1.3)
0.97 (0.47) 1.63 (1.45) 0.40 (0.27) 0.23 (0.27)
2.76 (0.43) 1.00 (0.40) 0.46 (0.23) 0.48 (0.27)
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the Bradford assay (Bradford, 1976) using BSA (bovine serum albumin) as a standard. 2.6. TOC analysis Samples were analyzed for TOC and total N with a CHNS analyzer (Flash EA 112 series, Thermo Electron Cooperation) after being ground to fine powder with a ball mill (MM301, Retsch) for 5 min and dried at 105 C.
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displayed an average MWD of 1.44 (0.47) mm, and there was no significant difference between old and permanent forest sites (Table 2). The loss of the MWD of the aggregates from the chronosequence sites is caused by a loss of the proportion of water stable macroaggregates (aggregates > 200 mm) (Fig. 2). The proportion of waterstable macroaggregates ranged from 31.5 to 73.5% in the permanent pasture sites and decreased to 9.0 (7.5) % in course of the first 46 years of cultivation. The proportion of aggregates <63 mm remained smaller than 2% in all analysed samples.
2.7. Statistics 3.2. Carbohydrates All biochemical and physical analysis were performed in triplicates. Mean values and standard deviations were calculated using Excel. The MWD and the concentrations of carbohydrates and GRSP in time were described by a monoexponential function using the software Origin 6.0. Regression analysis was conducted using SPSS. In order to calculated differences in soil aggregation, carbohydrate and GRSP contents following land use change, the mean value of sites of the same age was subtracted from the mean value of five permanent pasture sites that represent the zero point of the chronosequence. To asses whether differences between groups of sites of the same duration of land use were significant, t-tests were performed using SPSS, where p < 0.05 was considered as the threshold value for significance. T-tests were also used to asses whether differences in the TOC, carbohydrate and GRSP contents in the different aggregate size fractions were significant. 3. Results 3.1. Water-stable aggregates In the permanent pasture sites, the average MWD of the waterstable aggregates was 1.31 (0.27) mm. It decreased to 0.45 (0.16) mm during the first 46 years after conversion of the sites (Fig. 1). This results in a loss of 66% of the initial MWD. The decrease of the MWD was approximated with a monoexponential function (R2 ¼ 0.66, p < 0.001). Although showing significantly higher TOC contents (123.3 and 180.2 g TOC kg1), two very wet permanent pasture sites exhibited MWDs, which are in the same range as those of the regular permanent pasture sites (Table 2). Forest sites
The diluted acid soluble carbohydrate content in the permanent pasture sites ranged from 11.7 to 36.6 with an average of 23.9 (8.5) mmol glucose-equivalent kg1 (Fig. 3). After the conversion of the soils from pasture to cropland the carbohydrate concentration decreased immediately in the first decade after the land use change to an average concentration of 8.6 (1.74) mmol glucose-equivalent kg1 and remained stable at this level (Fig. 3). This results in a loss of 64.2% of the initial carbohydrate content during the first 46 years after conversion. The decrease of the carbohydrate content was approximated with a monoexponential function (R2 ¼ 0.59, p < 0.001). The two very wet permanent pasture sites (which are close to Histic Gleysols) exhibited carbohydrate contents of 54.0 and 34 7 mmol glucose-equivalent kg1 (Table 2). Old Forest sites displayed an average carbohydrate content of 26.0 (8.6) mmol glucose-equivalent kg1, and the two permanent forest soils had an average carbohydrate content of 55.1 and 38.6 mmol glucoseequivalent kg1 (Table 2). 3.3. GRSP and TOC In the permanent pasture sites, the average GRSP content was 7.0 (2.1) g kg1. It decreased to 3.0 (0.4) g kg1 during the first 46 years after conversion of the sites (Fig. 4). This results in a loss of 57% of the initial GRSP content. The decrease of the GRSP content was described by a monoexponential function (R2 ¼ 0.71, p < 0.001). The very wet permanent pasture soil displayed 20.7 and 17.7 g kg1 GRSP (Table 2). The GRSP content of the old forest soils ranged from 6.6 to 25.9 g kg1 with an average of 11.3 (7.3) g kg1 GRSP. The permanent forest soils exhibited 12.6 and 30.6 g kg1 GRSP. While the TOC content decreased during the first 46 years after conversion by 64% (from 35.4 (12.1) to 12.88 (5.9) g kg1), the GRSP/TOC ratio increased from 0.21 (0.08) to 0.29 (0.12) g g1 (Fig. 5). 3.4. MWD, TOC, carbohydrates and GRSP The MWD of the chronosequence, the wet permanent pasture, and the forest sites was significantly correlated with the TOC, the carbohydrates and the GRSP content (Fig. 6). All three correlations were approximated by sigmoidal functions. The correlation was high for the TOC (R2 ¼ 0.62, p < 0.001) and the GRSP (R2 ¼ 0.67, p < 0.001), and less strong for the carbohydrates (R2 ¼ 0.58, p < 0.001). 3.5. TOC, carbohydrates, and GRSP in water-stable aggregate fractions
Fig. 1. Relationship between the MWD of the water-stable aggregates and the time after conversion of the soils from pasture to cropland from 0 to 46 years. Bars depict standard deviations calculated from triplicates of one sample. R2 ¼ 0.72, p < 0.001.
The TOC and carbohydrate contents of the water-stable aggregate fractions of a representative cropland, pasture, and forest site decreased in the order > 630 mm-, >2000 mm-, >200 mm-, >63 mmfraction (Fig. 7). This distribution was the same for all analysed samples, independently of land use. The ratio between the concentrations in the macroaggregates and the concentration in the
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Table 2 TOC, GRSP and carbohydrate content, MWD of the water-stable aggregates, GRSP/TOC, and proportion of macroaggregates in wet permanent pasture, old forest, and permanent forest soils. Standard deviations calculated from triplicates of one sample are given in brackets. Site and land use
TOC [g kg1]
GRSP [g kg1]
Carbohydrates [mmol gluc.-eq kg1]
MWD [mm]
GRSP/TOC [g g1]
Proportion of macroaggregates [%]
Wet permanent pasture
180.1 (52.0) 123.3 (60.4)
20.7 (0.7) 17.7 (0.4)
54.0 (3.1) 34.7 (0.0)
1.80 (0.1) 2.16 (0.2)
0.12 0.14
91.57 86.34
79.3 (3.6) 57.5 (7.9) 75.2 (10.1) 49.6 (5.8) 63.6 (2.9) 77.5 (14.3)
6.6 (0.2) 7.8 (0.1) 10.4 (0.4) 8.7 (0.3) 8.4 (0.7) 25.9 (0.2)
24.4 20.1 28.5 17.1 39.3 17.9
0.72 1.12 1.89 1.23 1.07 1.59
(0.0) (0.1) (0.1) (0.0) (0.0) (0.3)
0.08 0.14 0.14 0.18 0.13 0.33
31.23 42.74 85.05 52.66 66.99 54.50
187.4 (6.2) 128.5 (15.9)
30.6 (1.0) 12.6 (0.2)
38.6 (1.4) 55.1 (7.4)
2.15 (0.0) 1.60 (0.0)
0.16 0.10
67.04 86.64
Old forest
Permanent forest
(3.6) (0.1) (1.7) (1.1) (4.8) (0.6)
microaggregates for all chronosequence soils was 0.52 (0.21) for the TOC, 0.45 (0.04) for the carbohydrates and 1.05 (0.20) for the GRSP. This ratio of the GRSP concentrations is significantly different from both the ratio of TOC and carbohydrates, while the ratios of TOC and carbohydrates are not significantly different from each other. Thus, the GRSP is more equally distributed among the aggregate fractions than the TOC and the carbohydrates. 4. Discussion 4.1. Soil aggregation and land use We found that the MWD of the water-stable aggregates was reduced by 66% during the first 46 years after the conversion from pasture to cropland (Fig. 1). The decay of the MWD was caused by a breakdown of macroaggregates (>200 mm), which resulted in an increase of the proportion of microaggregates (63e200 mm) in the cause of cultivation (Fig. 2). This finding is consistent with the aggregate hierarchy theory (Tisdall and Oades, 1982), which states that macroaggregates are formed by microaggregates being sticked together by relatively easily biodegradable compounds. The MWD of the wet permanent pasture and forest soils were in the same range as those of the regular permanent pasture soils although they exhibited significantly higher TOC contents (Table 2). Therefore, we
Fig. 2. Relationship between the proportion of (A) the water-stable macroaggregates (aggregates > 2000 and >630 mm) and (B) the water-stable microaggregates (aggregates > 200 and > 63 mm) and the time after conversion of the soils from pasture to cropland from 0 to 46 year. R2 ¼ 0.61, p < 0.001.
conclude other stabilization mechanisms are dominant in these soils which are less important in the chronosequence soils, as for example the relatively low pH in the forest soils, and the moisture in the wet permanent pasture sites (Spohn and Giani, in press). Our results go in line with Besnard et al. (1996) who found the amount of aggregated soil is reduced by 40% during the first 35 years after the conversion of forest to cropland in a humic acid loam in France. Similarly, Ashagrie et al. (2007) reported that 26 years after conversion of natural forest to cropland in a Rhodic Nitrosol in the Tropics, the proportion of macroaggregates is reduced by 30% Bongiovanni and Lobartini (2006) found that the content of macroaggregates was 1.7 times lower after 50 years of cropland use than in a natural forest site. John et al. (2005) showed that in the arable soils aggregates <1000 mm were most abundant, while in grassland an forest soils aggregates > 1000 mm dominated. 4.2. Carbohydrates and land use The diluted acid soluble carbohydrate contents reported in this study are fairly diverse in forest and pasture sites. On an average they decreased rapidly after the conversion of the pasture sites to cropland (Fig. 3). This fast decline of the carbohydrate contents in comparison to the TOC (Fig. 5) or the GRSP (Fig. 4) clearly shows the lability of carbohydrates. Only a few studies address the influence
Fig. 3. Relationship between the carbohydrate content of the chronosequence sites and the time after conversion of the soils from pasture to cropland from 0 to 46 years. Bars depict standard deviations calculated from triplicates of one sample. R2 ¼ 0.59, p < 0.001.
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Fig. 4. Relationship between the GRSP content of the chronosequence sites and the time after conversion of the soils from pasture to cropland from 0 to 46 year. Bars depict standard deviations calculated from triplicates of one sample. R2 ¼ 0.71, p < 0.001.
of land-use change on carbohydrate contents so far. Jolivet et al. (2006), who studied carbohydrates in cultivated forest and clearcut sites reported that in the cultivated soils only 57e79% of the carbohydrate content of the forest sites was exhibited. Zhang et al. (1999) found a 54% decrease in amino sugar content after native grassland was converted to agricultural land. In a comparison of a cultivated and a forest site, it has been shown that the cropland soil contained only 45% of the hot water soluble carbohydrates of forest soil (Bongiovanni and Lobartini, 2006). 4.3. GRSP and land use The term glomalin-related soil protein (GRSP) in this study refers to an operational defined heat-stable soil protein fraction that is extracted and determined via the Bradford assay (Rillig, 2004). The GRSP contents of the chronosequence sites laid within the range of 2 to 14 mg g1 (Fig. 4), which is reported for most soils (Wright and Upadhyaya, 1998). However, much higher GRSP concentrations
Fig. 5. TOC content and GRSP/TOC ratio of the chronosequence sites as a function of the time after conversion of the soils from pasture to cropland from 0 to 46 years. R2 ¼ 0.20, p < 0.05.
Fig. 6. Sigmoidal relationship between (A) TOC, (B) carbohydrates, (C) GRSP content and the MWD of the chronosequence, the wet permanent pasture and the forest sites. R2 ¼ 0.62, R2 ¼ 0.58 and R2 ¼ 0.67, for all p < 0.001.
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(>100 mg g1) have been reported from old tropical soils (Rillig et al., 2001) and from undisturbed volcanic soils from Japan (>60 mg g1, Nichols and Wright, 2004). A humoferric podzol under oak forest in Ireland was reported to contain 69 mg g1 (Nichols and Wright, 2004). We found that the Gleyic Podzols and Haplic Gleysols under forest displayed from 6.6 to 30.6 g kg1 GRSP (Table 2). Nichols and Wright (2004) stated that generally, acidic soils have higher GRSP concentrations than calcareous soils. This is consistent with the relatively high GRSP concentrations reported in this study. It has been shown that GRSP contents react sensitively towards land use change. The A horizon of a cropland displayed only 62% of GRSP respect to natural forests (Rillig, 2004). Preger et al. (2007) observed in a chronosequence of sandy soils from the South-African Highveld that after 53 years after conversion from grassland to cropland only 67% of the initial Bradford reactive soil protein were present. Similarly, we found that the GRSP content decreased from 7.0 to 3.0 g kg1 monoexponentially (R2 ¼ 0.71). Preger et al. (2007) discovered that GRSP is enriched with respect to the TOC in the course of arable use of the soils from a GRSP/TOC ratio of 0.14 to approximately 0.27. We found that the GRSP/TOC ratio increased from 0.21 to 0.29 (Fig. 5). In contrast to Preger et al. (2007), we observed that the increase of the GRSP/TOC ratio took place after 25 years of cultivation while in the tropic soils, an increase of the ratio was reported to occur already 3 years after the conversion. The reason for this more rapid enrichment is most probably is the higher mineralization rate of the SOM in the tropics. In general, our findings confirm that the GRSP, although not being a pure but an operational defined C-fraction is either more slowly biodegraded than the majority of the other compounds that form the TOC, or is produced in high amounts even in the old C-depleted cropland soils.
4.4. Binding agents in aggregates
Fig. 7. Distribution of (A) TOC, (B) carbohydrates, and (C) the GRSP among the aggregate fractions for a cropland, a permanent pasture and a forest soil. Bars depict standard deviations calculated from triplicates of one sample.
It has been found that the carbohydrate content increased with the aggregate size (Puget, 1999). Gijsman and Thomans (1995), however, found a sigmoidal relation between aggregate stability and hot water extractable carbohydrates. At about 78% of aggregate stability, a plateau was reached, and further increase of the carbohydrate content did not enhance aggregation. Wright and Upadhyaya (1998) found a logarithmic correlation between the aggregate stability and glomalin in different soils under different types of land use in some parts of the USA and in Scotland. Their strongest correlation was obtained with the immunoreactive easily extractable glomalin fraction (R2 ¼ 0.86). Correspondingly, Harner et al. (2004) described a sigmoidal correlation between the GRSP and the aggregate stability from soils of a floodplain (R2 ¼ 0.55). In this study, a sigmoidal correlation between the MWD and TOC (R2 ¼ 0.62), the GRSP (R2 ¼ 0.67), and the carbohydrates (R2 ¼ 0.58) was found (Fig. 6). Therefore, we conclude that both the carbohydrates and the GRSP contribute to the aggregate stability of the sandy soils under study. The correlation between the TOC and the MWD indicates that most probably also other compounds which form the SOM contribute to the aggregation of the soils. The sigmoidal shape of the graph describing the relation between the organic matter compounds and the MWD shows that sandy soils have a limited intrinsic capacity for aggregation. This means that after a threshold value is reached a further increase in GRSP, carbohydrates or TOC does not enhance aggregation. The analyses of the distribution of TOC, carbohydrates, and GRSP among the different soil fractions showed, that the relative distribution of the carbohydrates was the same as for the TOC, decreasing in the order >630, >2000, >200, >63 mm, independently of land use (Fig. 7). The GRSP content, however, was more equally distributed among the four water-stable aggregate fractions.
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Acknowledgements We thank the Studienstiftung des deutschen Volkes for granting a PhD-scholarship. References Ashagrie, Y., Zech, W., Guggenberger, G., Mamo, T., 2007. Soil aggregation, and total and particulate organic matter following conversion of native forests to continuous cultivation in Ethiopia. Soil and Tillage Research 94, 101e108. Bedini, S., Pellegrino, E., Avio, L., Pellegrini, S., Bazzoffi, P., Argese, E., Giovannetti, M., 2009. Changes in soil aggregation and glomalin-related soil protein content as affected by the arbuscular mycorrhizal fungal species Glomus mosseae and Glomus intraradices. Soil Biology and Biochemistry 41, 1491e1496. Bongiovanni, M.D., Lobartini, J.C., 2006. Particulate organic matter, carbohydrate, humic acid contents in soil macro- and microaggregates as affected by cultivation. Geoderma 136, 660e665. Bradford, M.M., 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry 7, 248e254. Besnard, E., Chenu, C., Balesdent, J., Puget, P., Arrouays, D., 1996. Fate of particulate organic matter in soil aggregates during cultivation. European Journal of Soil Science 47, 495e503. Bronick, C.J., Lal, R., 2005. Soil structure and management: a review. Geoderma 124, 3e22. Gijsman, A.J., Thomans, R.J., 1995. Aggregate size distribution and stability of an Oxisol under legume-based and pure grass pastures in the Eastern Colombian savannas. Australian Journal of Soil Research 33, 153e165. Halvorson, J.J., Gonzales, J.M., 2006. Bradford reactive soil protein in Appalachian soils: distribution and response to incubation, extraction reagent and tannins. Plant and Soil 286, 339e356. Harner, M.J., Ramsey, P.W., Rillig, M.C., 2004. Protein accumulation and distribution in floodplain soils and river foam. Ecology Letters 7, 829e836. Haynes, R.J., Swift, R.S., Stephen, R.C., 1991. Influenced of mixed cropping rotations (pasture-arable) on organic matter content, water stable aggregation and clod porosity in a group of soils. Soil and Tillage Research 12, 77e87. Jastrow, J.D., 1996. Soil aggregate formation and the accrual of particulate and mineral-associated organic matter. Soil Biology & Biochemistry 28, 665e676. John, B., Yamashita, T., Ludwig, B., Flessa, H., 2005. Storage of organic carbon in aggregate and density fractions of silty soil under different types of land use. Geoderma 128, 63e79. Jolivet, C., Angers, D.A., Chantigny, M.H., Andreux, F., Arrouays, D., 2006. Carbohydrate dynamics in particle-size fractions of sandy spodosols following forest conversion to maize cropping. Soil Biology & Biochemistry 38, 2834e2842. Kemper, W.D., Rosenau, K., 1986. Size distribution of aggregates. In: Klute, A. (Ed.), Methods of Soil Analysis, Part 1. ASA, Madison, WI, pp. 425e442. Lützow, M., Kögel-Knabner, I., Ekschmitt, K., Matzner, E., Guggenberger, G., Marschner, B., Flessa, H., 2006. Stabilization of organic matter in temperate
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soils: mechanisms and their relevance under different soil conditions. European Journal of Soil Science 57, 426e445. Mann, L.K., 1986. Changes in soil carbon storage after cultivation. Soil Science 142 (5), 279e288. Martens, D.A., Frankenberger, W.T., 1990. Quantification of soil saccharides by spectrophotometric methods. Soil Biology and Biochemistry 22, 1173e1175. Nichols, K.A., Wright, S.F., 2004. Contributions of fungi to soil organic matter in agroecosystems. In: Magdoff, F., Weil, R.R. (Eds.), Soil Organic Matter in Sustainable Agriculture. CRC Press, Boca Raton, pp. 179e198. Oades, J.M., 1984. Soil organic matter and structural stability: mechanism and implications for management. Plant and Soil 76, 319e337. Preger, A.C., Rillig, M.C., Johns, A.R., Du Preez, C.C., Lobe, I., Amelung, W., 2007. Losses of Glomalin-related soil protein under prolonged arable cropping: a chronosequence study in sandy soils of the South African Highveld. Soil Biology & Biochemistry 39, 445e453. Puget, P., Angers, D.A., Chenu, C., 1999. Nature of carbohydrates associated with waterstable aggregates of two cultivated soils. Soil Biology & Biochemistry 31, 55e63. Puget, P., Chenu, C., Balesdent, J., 2000. Dynamics of soil organic matter associated with particle-size fractions of water-stable aggregates. European Journal of Soil Science 51, 595e605. Rillig, M.C., 2004. Arbuscular mycorrhizae, glomalin and soil quality. Canadian Journal of Soil Science 84, 355e365. Rillig, M.C., Ramsey, P.W., Morris, S., Eldor, P.A., 2003. Glomalin, an arbuscularmycorrhizal fungal soil protein, responds to land use change. Plant and Soil 253, 293e299. Rillig, M.C., Wright, S.F., Nichols, K.A., Schmidt, W.F., Torn, M.S., 2001. Large contribution of arbuscular mycorrhizal fungi to soil carbon pools in tropical forest soils. Plant and Soil 233, 167e177. Six, J., Bossuyt, H., Degryze, S., Denef, K., 2004. A history of research on the link between (micro)aggregates, soil biota, and soil organic matter. Soil & Tillage Research 79, 7e31. Six, J., Paustian, K., Elliot, E.T., Combrink, C., 2000. Soil structure and organic matter: I. Distribution of aggregate-size classes and aggregate-associated carbon. Soil Science Society of America Journal 64, 681e689. Spohn, M., Giani, L. Total, hot water extractable, and oxidation-resistant organic carbon in sandy hydromorphic soils e analysis of a 220-year chronosequence. Plant and Soil, in press, DOI: 10.1007/s11104-010-0322-5. Tisdall, J.M., Oades, J.M., 1982. Organic matter and water-stable aggregates in soils. Journal of Soil Science 33, 141e163. Waffenschmidt, S., Jaenicke, L., 1987. Assay of reducing sugars in nanomole range with 2,20 Bicinchoninate. Analytical Biochemistry 165, 337e340. Wright, S.F., Upadhyaya, A., 1996. Extraction of an abundant and unusual protein from soil and comparison with hyphal protein of mycorrhizal fungi. Soil Science 161, 575. Wright, S.F., Upadhyaya, A., 1998. A survey of soils for aggregate stability and glomalin, a glycoprotein produced by hyphae of arbuscular mycorrhizal fungi. Plant and Soil 198, 97e107. Zhang, X., Amelung, W., Yuan, Y., Samson-Liebig, S., Brown, L., Zech, W., 1999. Landuse effects on amino sugars in particle size fractions of an Argiudoll. Applied Soil Ecology 11, 271e275.
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Water-stable aggregates, glomalin-related soil protein, and carbohydrates in a chronosequence of sandy hydromorphic soils Marie Spohn*, Luise Giani Soil Science Group, Department of Biology and Environmental Sciences, Carl von Ossietzky University Oldenburg, 26121 Oldenburg, Germany
a r t i c l e i n f o
a b s t r a c t
Article history: Received 5 January 2010 Received in revised form 30 April 2010 Accepted 17 May 2010 Available online 31 May 2010
The conversion of pasture to cropland leads to a decline of aggregation in topsoils and to a decrease of aggregate-binding agents such as carbohydrates and glomalin-related soil protein (GRSP). Till now, studies on soil aggregation focused either on carbohydrates or on GRSP as a binding agent in aggregates. In this study we analyse the development of the relationship between carbohydrates, GRSP, TOC and aggregatestability following land-use change. Furthermore, we discuss the contents of carbohydrates, GRSP and TOC in each of the aggregate fractions. For these purposes, a chronosequence of sites, which were converted from pasture to cropland at different periods in history, was established. To get further insight into the impact of different types of land-use, also soils under forest, either afforested or permanent, were studied. The mean-weight diameter (MWD) of water-stable aggregates, the carbohydrate, and the GRSP content were determined in 49 soils. It was found that the MWD of the water-stable aggregates decreased monoexponentially (R2 ¼ 0.66) by 66% during the first 46 years after conversion of the soils from pasture to cropland. During the same period, the carbohydrate content decreased very rapidly after the land use change by 64% and the GRSP content decreased more slowly by 57%. The MWD of the forest soils were in the same range as those of the permanent pasture soils although they exhibit significantly higher TOC contents, which indicate that other stabilization mechanisms are dominant in forest soils, less important in the chronosequence soils. TOC, carbohydrates and the GRSP contents were sigmoidally correlated with the MWD. Among the four water-stable aggregate fractions TOC and carbohydrates exhibited high contents in the macroaggregates and were less present in the microaggregates. GRSP, in contrast, was more equally distributed among the four water-stable aggregate fractions. Ó 2010 Elsevier Ltd. All rights reserved.
Keywords: Water-stable aggregates Glomalin-related soil protein Carbohydrates Chronosequence Sandy hydromorphic soils
1. Introduction Soil aggregation supports soil fertility as it reduces erosion and mediates soil aeration, water infiltration, and retention (Oades, 1984). Furthermore, soil aggregation protects soil organic matter (SOM) from getting mineralized as it physically reduces the accessibility of organic compounds for microorganisms and exoenzymes and the oxygen diffusion (Lützow et al., 2006). Aggregation is mediated by soil organic matter (SOM), biota, ionic bridging, clay, and carbonates (Bronick and Lal, 2005). A variety of binding mechanisms works simultaneously at different spatial scales to stabilize aggregates. Tisdall and Oades (1982) stated that primarily, mineral particles are bound together by persistent binding agents like biotic debris while these microaggregates, in turn, are bound together to macroaggregates by transient and temporal binding agents (polysaccharides, roots, and fungal hyphae). This * Corresponding author. Tel.: þ49 (0)441 798 4504. E-mail address: [email protected] (M. Spohn). 0038-0717/$ e see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.soilbio.2010.05.015
aggregate hierarchy theory has been used by many authors to explain correlations between a reduction of aggregation and a loss of SOC (Six et al., 2000, 2004). More recently, it is assumed that microaggregates are formed within macroaggregates and protect SOM more effectively than macroaggregates (Oades, 1984; Puget et al., 2000; Six et al., 2000). Aggregation is influenced by land use and land use change in the way that the proportion of water stable macroaggregates is reduced in the order forest > pasture/grassland > arable land and is further diminished with the duration of arable use (Ashagrie et al., 2007; Haynes et al., 1991; Jastrow, 1996; John et al., 2005). Microaggregates, however, seem to be less influenced by land use (Oades, 1984; Besnard et al., 1996; Puget et al., 2000). Polysaccharides from bacteria, fungi or root mucilage are described as a labile SOM fraction which acts as an important binding agent for aggregation (Jolivet et al., 2006; Oades, 1984; Puget et al., 1999). More recently, also the heat-stable protein glomalin which is produced by arbuscular mychorriza has been shown to cause soil aggregation (Bedini et al., 2009; Harner et al., 2004; Wright and Upadhyaya, 1998).
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Glomalin promotes soil aggregation especially in coarse textured soils and its content in soils decreases with the duration of the arable use of the soils (Preger et al., 2007; Rillig et al., 2003). Till now, studies on soil aggregation focused either on carbohydrates or on GRSP as a binding agent in aggregates. Hence, the main objective of this study was to study the development of the relationship between carbohydrates, GRSP, TOC and aggregatestability following land-use change. Furthermore, we discuss the contents of carbohydrates, GRSP and TOC in each of the aggregate factions. For these purposes, a chronosequence of sites, which were converted from pasture to cropland at different periods in history, was established. To get further insight into the impact of different types of land-use, also soils under forest, either afforested or permanent, were studied. 2. Material and methods 2.1. Study site, geo-data sources and processing
soil. The mixed samples were homogenized, oven dried (70 C, 96 h), and subsequently crushed to pass a screen (2 mm-opening). 70 C was chosen as a drying temperature in order to prevent microbial degradation during the drying process. 2.3. Water-stable aggregate fractionation Water-stable aggregates were fractionated according to Kemper and Rosenau (1986). 70 g of oven dried soil were soaked in H2O dest. on a 2000 mm sieve for 10 min to allow slaking. Subsequently, the sieve was moved up and down in water by about 3 cm with 50 repetitions. The fraction < 2000 mm was passed on to a 630 mm sieve. This procedure was repeated with a 200 and a 63 mm sieve. Each fraction was recollected from the sieve, dried at 70 C, and weighed. The mean weight diameter (MWD) of each sample was calculated by
MWD ¼
The study was conducted in a region called Artland, 40 km north of Osnabrück in Northwest Germany (52 360 -52 410 N, 7 540 -8 140 E). The area was formed in the Pleistocene, and Gleyic Podzols, Haplic Podzols, and Haplic Gleysols represent its main soil groups today. It is characterized by an annual mean precipitation of 800 mm and an annual mean air temperature of 8.7 C. Historical topographic maps which contain precise information on land use were used to attribute former uses of arable land, pasture, and forest in order to establish a chronosequence. Sites, which were converted from pasture to arable land at defined periods of time were selected. As a zero point of the chronosequence, permanent pasture sites were chosen. In addition, sites under forest which have either been used as forest at least since 1789 (permanent forest) or were afforested between 1851 and 1900 (old forest) were selected. All sites have a size of 0.8 to 1.7 ha. For this study, we exclusively sampled Gleyic Podzols and Haplic Gleysols as we expected sandy hydromorphic soils to react very sensitively to land use changes (Spohn and Giani, in press). All sampled soils are situated in the four sampling subregions Lohne, Gröhnloh, Mühlen, and Nortrup and exhibit very similar textures, Alox, Feox contents and pH (Table 1). 2.2. Sampling The upper 20 cm of 49 arable, pasture, and forest sites were sampled in late October 2008 and February 2009. This sampling depth was chosen as a common sampling depth in studies dealing with SOM responses to land use change (Mann, 1986). On every site samples were taken each 10 m along two diagonal transects, thus the number of samples depends on the size of the site. The minimal amount of subsamples was 20. Samples of each site were combined to gain one mixed sample per site. On the forest sites, litter and the organic layer were removed in order to sample solely the mineral
n X
Xi Wi ;
i¼1
where Xi is the mean diameter of each size fraction and Wi is the proportion of the total sample weight in the corresponding size fraction (Kemper and Rosenau, 1986). 2.4. Carbohydrate extraction and determination The extraction procedure for the carbohydrates was slightly modified from the method described by Martens and Frankenberger (1990). Briefly, 2 g of oven dried soil were treated with 20 ml 0.125 M H2SO4 in a water bath at 80 C for 7 h and were vortexed for 10 s every 30 min during the extraction procedure. Subsequently, the samples were treated with EDTA (ethylene diaminetetraacetate) in order to prevent co-precipitation of the sugars with cations. The assay was titrated to pH 3.5e4.0 with 5 M KOH and centrifuged. The carbohydrates in the supernatant were determined following Waffenschmidt and Jaenicke (1987) by the 2,20 Bicinchoninate-assay (BCA-assay). 20 ml of the supernatant were added to 2 ml of the BCA-assay working reagent and incubated at 70 C for 60 min. After cooling down, to room temperature the samples were measured photometrically at 560 nm (UVmini1240, Shimadzu). D-Glucose was used as a standard. 2.5. Glomalin extraction and determination Glomalin was extracted following the method of Wright and Upadhyaya (1996, 1998), but with 50 mM diphosphate (adjusted to pH 8.0) as extractant as suggested by Halvorson and Gonzales (2006). Briefly, 2 g of oven dried soil were extracted by a total volume of 50 ml diphosphate at 128 C during four extraction cycles, each lasting for 1 h. The pooled extracts were centrifuged to remove soil particles and protein concentration was determined by
Table 1 Sampling subregions, number of sites per subregion, and particle size distribution, pH and oxalate extractable Fe and Al of the investigated Gleyic Podzols and Haplic Gleysols. Standard deviations calculated from triplicates of one sample per site are given in brackets. Sampling subregion
Number of sites
Soilgroup
Texture Sand [%]
Silt [%]
Clay [%]
Lohne Gröhnloh Mühlen Nortup
24 12 8 5
Gleyic Podzol Gleyic Podzol Haplic Gleysol Haplic Gleysol
85.0 80.5 80.0 80.0
10.6 (6.4) 14.8 (7.0) 9.6 (1.7) 16.0 (3.9)
4.4 (4.1) 4.6 (1.9) 9.9 (2.0) 4.0 (2.1)
a b
Exclusive forest sites: pHH2O 3.4 (0.3). Exclusive forest sites: Feox 11.9 (1.0) g kg1.
(8.2) (8.4) (3.4) (3.5)
pHH2Oa
Feoxb [g kg1]
Alox [g kg1]
5.5 (1.1) 5.6 (0.9) 5.6 (0.3) 5.7 (1.3)
0.97 (0.47) 1.63 (1.45) 0.40 (0.27) 0.23 (0.27)
2.76 (0.43) 1.00 (0.40) 0.46 (0.23) 0.48 (0.27)
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the Bradford assay (Bradford, 1976) using BSA (bovine serum albumin) as a standard. 2.6. TOC analysis Samples were analyzed for TOC and total N with a CHNS analyzer (Flash EA 112 series, Thermo Electron Cooperation) after being ground to fine powder with a ball mill (MM301, Retsch) for 5 min and dried at 105 C.
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displayed an average MWD of 1.44 (0.47) mm, and there was no significant difference between old and permanent forest sites (Table 2). The loss of the MWD of the aggregates from the chronosequence sites is caused by a loss of the proportion of water stable macroaggregates (aggregates > 200 mm) (Fig. 2). The proportion of waterstable macroaggregates ranged from 31.5 to 73.5% in the permanent pasture sites and decreased to 9.0 (7.5) % in course of the first 46 years of cultivation. The proportion of aggregates <63 mm remained smaller than 2% in all analysed samples.
2.7. Statistics 3.2. Carbohydrates All biochemical and physical analysis were performed in triplicates. Mean values and standard deviations were calculated using Excel. The MWD and the concentrations of carbohydrates and GRSP in time were described by a monoexponential function using the software Origin 6.0. Regression analysis was conducted using SPSS. In order to calculated differences in soil aggregation, carbohydrate and GRSP contents following land use change, the mean value of sites of the same age was subtracted from the mean value of five permanent pasture sites that represent the zero point of the chronosequence. To asses whether differences between groups of sites of the same duration of land use were significant, t-tests were performed using SPSS, where p < 0.05 was considered as the threshold value for significance. T-tests were also used to asses whether differences in the TOC, carbohydrate and GRSP contents in the different aggregate size fractions were significant. 3. Results 3.1. Water-stable aggregates In the permanent pasture sites, the average MWD of the waterstable aggregates was 1.31 (0.27) mm. It decreased to 0.45 (0.16) mm during the first 46 years after conversion of the sites (Fig. 1). This results in a loss of 66% of the initial MWD. The decrease of the MWD was approximated with a monoexponential function (R2 ¼ 0.66, p < 0.001). Although showing significantly higher TOC contents (123.3 and 180.2 g TOC kg1), two very wet permanent pasture sites exhibited MWDs, which are in the same range as those of the regular permanent pasture sites (Table 2). Forest sites
The diluted acid soluble carbohydrate content in the permanent pasture sites ranged from 11.7 to 36.6 with an average of 23.9 (8.5) mmol glucose-equivalent kg1 (Fig. 3). After the conversion of the soils from pasture to cropland the carbohydrate concentration decreased immediately in the first decade after the land use change to an average concentration of 8.6 (1.74) mmol glucose-equivalent kg1 and remained stable at this level (Fig. 3). This results in a loss of 64.2% of the initial carbohydrate content during the first 46 years after conversion. The decrease of the carbohydrate content was approximated with a monoexponential function (R2 ¼ 0.59, p < 0.001). The two very wet permanent pasture sites (which are close to Histic Gleysols) exhibited carbohydrate contents of 54.0 and 34 7 mmol glucose-equivalent kg1 (Table 2). Old Forest sites displayed an average carbohydrate content of 26.0 (8.6) mmol glucose-equivalent kg1, and the two permanent forest soils had an average carbohydrate content of 55.1 and 38.6 mmol glucoseequivalent kg1 (Table 2). 3.3. GRSP and TOC In the permanent pasture sites, the average GRSP content was 7.0 (2.1) g kg1. It decreased to 3.0 (0.4) g kg1 during the first 46 years after conversion of the sites (Fig. 4). This results in a loss of 57% of the initial GRSP content. The decrease of the GRSP content was described by a monoexponential function (R2 ¼ 0.71, p < 0.001). The very wet permanent pasture soil displayed 20.7 and 17.7 g kg1 GRSP (Table 2). The GRSP content of the old forest soils ranged from 6.6 to 25.9 g kg1 with an average of 11.3 (7.3) g kg1 GRSP. The permanent forest soils exhibited 12.6 and 30.6 g kg1 GRSP. While the TOC content decreased during the first 46 years after conversion by 64% (from 35.4 (12.1) to 12.88 (5.9) g kg1), the GRSP/TOC ratio increased from 0.21 (0.08) to 0.29 (0.12) g g1 (Fig. 5). 3.4. MWD, TOC, carbohydrates and GRSP The MWD of the chronosequence, the wet permanent pasture, and the forest sites was significantly correlated with the TOC, the carbohydrates and the GRSP content (Fig. 6). All three correlations were approximated by sigmoidal functions. The correlation was high for the TOC (R2 ¼ 0.62, p < 0.001) and the GRSP (R2 ¼ 0.67, p < 0.001), and less strong for the carbohydrates (R2 ¼ 0.58, p < 0.001). 3.5. TOC, carbohydrates, and GRSP in water-stable aggregate fractions
Fig. 1. Relationship between the MWD of the water-stable aggregates and the time after conversion of the soils from pasture to cropland from 0 to 46 years. Bars depict standard deviations calculated from triplicates of one sample. R2 ¼ 0.72, p < 0.001.
The TOC and carbohydrate contents of the water-stable aggregate fractions of a representative cropland, pasture, and forest site decreased in the order > 630 mm-, >2000 mm-, >200 mm-, >63 mmfraction (Fig. 7). This distribution was the same for all analysed samples, independently of land use. The ratio between the concentrations in the macroaggregates and the concentration in the
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Table 2 TOC, GRSP and carbohydrate content, MWD of the water-stable aggregates, GRSP/TOC, and proportion of macroaggregates in wet permanent pasture, old forest, and permanent forest soils. Standard deviations calculated from triplicates of one sample are given in brackets. Site and land use
TOC [g kg1]
GRSP [g kg1]
Carbohydrates [mmol gluc.-eq kg1]
MWD [mm]
GRSP/TOC [g g1]
Proportion of macroaggregates [%]
Wet permanent pasture
180.1 (52.0) 123.3 (60.4)
20.7 (0.7) 17.7 (0.4)
54.0 (3.1) 34.7 (0.0)
1.80 (0.1) 2.16 (0.2)
0.12 0.14
91.57 86.34
79.3 (3.6) 57.5 (7.9) 75.2 (10.1) 49.6 (5.8) 63.6 (2.9) 77.5 (14.3)
6.6 (0.2) 7.8 (0.1) 10.4 (0.4) 8.7 (0.3) 8.4 (0.7) 25.9 (0.2)
24.4 20.1 28.5 17.1 39.3 17.9
0.72 1.12 1.89 1.23 1.07 1.59
(0.0) (0.1) (0.1) (0.0) (0.0) (0.3)
0.08 0.14 0.14 0.18 0.13 0.33
31.23 42.74 85.05 52.66 66.99 54.50
187.4 (6.2) 128.5 (15.9)
30.6 (1.0) 12.6 (0.2)
38.6 (1.4) 55.1 (7.4)
2.15 (0.0) 1.60 (0.0)
0.16 0.10
67.04 86.64
Old forest
Permanent forest
(3.6) (0.1) (1.7) (1.1) (4.8) (0.6)
microaggregates for all chronosequence soils was 0.52 (0.21) for the TOC, 0.45 (0.04) for the carbohydrates and 1.05 (0.20) for the GRSP. This ratio of the GRSP concentrations is significantly different from both the ratio of TOC and carbohydrates, while the ratios of TOC and carbohydrates are not significantly different from each other. Thus, the GRSP is more equally distributed among the aggregate fractions than the TOC and the carbohydrates. 4. Discussion 4.1. Soil aggregation and land use We found that the MWD of the water-stable aggregates was reduced by 66% during the first 46 years after the conversion from pasture to cropland (Fig. 1). The decay of the MWD was caused by a breakdown of macroaggregates (>200 mm), which resulted in an increase of the proportion of microaggregates (63e200 mm) in the cause of cultivation (Fig. 2). This finding is consistent with the aggregate hierarchy theory (Tisdall and Oades, 1982), which states that macroaggregates are formed by microaggregates being sticked together by relatively easily biodegradable compounds. The MWD of the wet permanent pasture and forest soils were in the same range as those of the regular permanent pasture soils although they exhibited significantly higher TOC contents (Table 2). Therefore, we
Fig. 2. Relationship between the proportion of (A) the water-stable macroaggregates (aggregates > 2000 and >630 mm) and (B) the water-stable microaggregates (aggregates > 200 and > 63 mm) and the time after conversion of the soils from pasture to cropland from 0 to 46 year. R2 ¼ 0.61, p < 0.001.
conclude other stabilization mechanisms are dominant in these soils which are less important in the chronosequence soils, as for example the relatively low pH in the forest soils, and the moisture in the wet permanent pasture sites (Spohn and Giani, in press). Our results go in line with Besnard et al. (1996) who found the amount of aggregated soil is reduced by 40% during the first 35 years after the conversion of forest to cropland in a humic acid loam in France. Similarly, Ashagrie et al. (2007) reported that 26 years after conversion of natural forest to cropland in a Rhodic Nitrosol in the Tropics, the proportion of macroaggregates is reduced by 30% Bongiovanni and Lobartini (2006) found that the content of macroaggregates was 1.7 times lower after 50 years of cropland use than in a natural forest site. John et al. (2005) showed that in the arable soils aggregates <1000 mm were most abundant, while in grassland an forest soils aggregates > 1000 mm dominated. 4.2. Carbohydrates and land use The diluted acid soluble carbohydrate contents reported in this study are fairly diverse in forest and pasture sites. On an average they decreased rapidly after the conversion of the pasture sites to cropland (Fig. 3). This fast decline of the carbohydrate contents in comparison to the TOC (Fig. 5) or the GRSP (Fig. 4) clearly shows the lability of carbohydrates. Only a few studies address the influence
Fig. 3. Relationship between the carbohydrate content of the chronosequence sites and the time after conversion of the soils from pasture to cropland from 0 to 46 years. Bars depict standard deviations calculated from triplicates of one sample. R2 ¼ 0.59, p < 0.001.
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Fig. 4. Relationship between the GRSP content of the chronosequence sites and the time after conversion of the soils from pasture to cropland from 0 to 46 year. Bars depict standard deviations calculated from triplicates of one sample. R2 ¼ 0.71, p < 0.001.
of land-use change on carbohydrate contents so far. Jolivet et al. (2006), who studied carbohydrates in cultivated forest and clearcut sites reported that in the cultivated soils only 57e79% of the carbohydrate content of the forest sites was exhibited. Zhang et al. (1999) found a 54% decrease in amino sugar content after native grassland was converted to agricultural land. In a comparison of a cultivated and a forest site, it has been shown that the cropland soil contained only 45% of the hot water soluble carbohydrates of forest soil (Bongiovanni and Lobartini, 2006). 4.3. GRSP and land use The term glomalin-related soil protein (GRSP) in this study refers to an operational defined heat-stable soil protein fraction that is extracted and determined via the Bradford assay (Rillig, 2004). The GRSP contents of the chronosequence sites laid within the range of 2 to 14 mg g1 (Fig. 4), which is reported for most soils (Wright and Upadhyaya, 1998). However, much higher GRSP concentrations
Fig. 5. TOC content and GRSP/TOC ratio of the chronosequence sites as a function of the time after conversion of the soils from pasture to cropland from 0 to 46 years. R2 ¼ 0.20, p < 0.05.
Fig. 6. Sigmoidal relationship between (A) TOC, (B) carbohydrates, (C) GRSP content and the MWD of the chronosequence, the wet permanent pasture and the forest sites. R2 ¼ 0.62, R2 ¼ 0.58 and R2 ¼ 0.67, for all p < 0.001.
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(>100 mg g1) have been reported from old tropical soils (Rillig et al., 2001) and from undisturbed volcanic soils from Japan (>60 mg g1, Nichols and Wright, 2004). A humoferric podzol under oak forest in Ireland was reported to contain 69 mg g1 (Nichols and Wright, 2004). We found that the Gleyic Podzols and Haplic Gleysols under forest displayed from 6.6 to 30.6 g kg1 GRSP (Table 2). Nichols and Wright (2004) stated that generally, acidic soils have higher GRSP concentrations than calcareous soils. This is consistent with the relatively high GRSP concentrations reported in this study. It has been shown that GRSP contents react sensitively towards land use change. The A horizon of a cropland displayed only 62% of GRSP respect to natural forests (Rillig, 2004). Preger et al. (2007) observed in a chronosequence of sandy soils from the South-African Highveld that after 53 years after conversion from grassland to cropland only 67% of the initial Bradford reactive soil protein were present. Similarly, we found that the GRSP content decreased from 7.0 to 3.0 g kg1 monoexponentially (R2 ¼ 0.71). Preger et al. (2007) discovered that GRSP is enriched with respect to the TOC in the course of arable use of the soils from a GRSP/TOC ratio of 0.14 to approximately 0.27. We found that the GRSP/TOC ratio increased from 0.21 to 0.29 (Fig. 5). In contrast to Preger et al. (2007), we observed that the increase of the GRSP/TOC ratio took place after 25 years of cultivation while in the tropic soils, an increase of the ratio was reported to occur already 3 years after the conversion. The reason for this more rapid enrichment is most probably is the higher mineralization rate of the SOM in the tropics. In general, our findings confirm that the GRSP, although not being a pure but an operational defined C-fraction is either more slowly biodegraded than the majority of the other compounds that form the TOC, or is produced in high amounts even in the old C-depleted cropland soils.
4.4. Binding agents in aggregates
Fig. 7. Distribution of (A) TOC, (B) carbohydrates, and (C) the GRSP among the aggregate fractions for a cropland, a permanent pasture and a forest soil. Bars depict standard deviations calculated from triplicates of one sample.
It has been found that the carbohydrate content increased with the aggregate size (Puget, 1999). Gijsman and Thomans (1995), however, found a sigmoidal relation between aggregate stability and hot water extractable carbohydrates. At about 78% of aggregate stability, a plateau was reached, and further increase of the carbohydrate content did not enhance aggregation. Wright and Upadhyaya (1998) found a logarithmic correlation between the aggregate stability and glomalin in different soils under different types of land use in some parts of the USA and in Scotland. Their strongest correlation was obtained with the immunoreactive easily extractable glomalin fraction (R2 ¼ 0.86). Correspondingly, Harner et al. (2004) described a sigmoidal correlation between the GRSP and the aggregate stability from soils of a floodplain (R2 ¼ 0.55). In this study, a sigmoidal correlation between the MWD and TOC (R2 ¼ 0.62), the GRSP (R2 ¼ 0.67), and the carbohydrates (R2 ¼ 0.58) was found (Fig. 6). Therefore, we conclude that both the carbohydrates and the GRSP contribute to the aggregate stability of the sandy soils under study. The correlation between the TOC and the MWD indicates that most probably also other compounds which form the SOM contribute to the aggregation of the soils. The sigmoidal shape of the graph describing the relation between the organic matter compounds and the MWD shows that sandy soils have a limited intrinsic capacity for aggregation. This means that after a threshold value is reached a further increase in GRSP, carbohydrates or TOC does not enhance aggregation. The analyses of the distribution of TOC, carbohydrates, and GRSP among the different soil fractions showed, that the relative distribution of the carbohydrates was the same as for the TOC, decreasing in the order >630, >2000, >200, >63 mm, independently of land use (Fig. 7). The GRSP content, however, was more equally distributed among the four water-stable aggregate fractions.
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