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Spatial distribution of glomalin-related soil protein and its relationships with root mycorrhization, soil aggregates, carbohydrates, activity of protease and β-glucosidase in the rhizosphere of Citrus unshiu
Soil Biology & Biochemistry 45 (2012) 181e183
Contents lists available at SciVerse ScienceDirect
Soil Biology & Biochemistry journal homepage: www.elsevier.com/locate/soilbio
Short communication
Spatial distribution of glomalin-related soil protein and its relationships with root mycorrhization, soil aggregates, carbohydrates, activity of protease and b-glucosidase in the rhizosphere of Citrus unshiu Qiang-Sheng Wu a, *, Xin-Hua He b, c, d, Ying-Ning Zou a, Kai-Ping He a, Ya-Hong Sun a, Ming-Qin Cao a a
College of Horticulture and Gardening, Yangtze University, Jingzhou, Hubei 434025, China State Centre of Excellence for Ecohydrology, Edith Cowan University, Joondalup, WA 6027, Australia Centre for Ecosystem Management/School of Natural Sciences, Edith Cowan University, Joondalup, WA 6027, Australia d School of Plant Biology, University of Western Australia, Crawley, WA 6009, Australia b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 14 June 2011 Received in revised form 18 August 2011 Accepted 9 October 2011 Available online 20 October 2011
Relationships between the spatial distributions of glomalin-related soil protein (GRSP) and soil aggregates, carbohydrates or relevant enzymes are poorly studied. We found that two categories of GRSP, the easily extractable Bradford-reactive soil protein (EE-BRSP) and total BRSP (T-BRSP), respectively ranged between 0.3e0.6 and 0.5e0.8 mg/g DW soil, and these two BRSPs decreased with the increase of soil depth (0e40 cm) in the rhizosphere of a 22-year-old Citrus unshiu orchard. Both EE-BRSP and T-BRSP were significantly positively correlated with mycorrhization, 0.25e0.50 mm soil water-stable aggregates, water-extractable or hydrolyzable carbohydrates, and b-glucosidase, but significantly negatively correlated with protease. Our results demonstrate that the spatial distribution of GRSP is significantly affected by mycorrhization, soil carbohydrate, b-glucosidase and protease. Ó 2011 Elsevier Ltd. All rights reserved.
Keywords: b-glucosidase Citrus Glomalin Glomalin-related soil protein Mycorrhization Protease
As an insoluble N-linked glycoprotein with w37% carbon (C) and 3e5% nitrogen (N), glomalin contributes to soil structure stabilization and C pools (Wright and Upadhyaya, 1998; Lovelock et al., 2004). Glomalin is mainly produced by arbuscular mycorrhizal fungi (AMF) hyphae and quantified from soils as glomalin-related soil protein (GRSP). The GRSP is often related to soil C pool (Treseder and Turner, 2007), though glomalin has limited contribution to C budgets in short-term experiments (Driver et al., 2005). Less information is available to the spatial distributions of GRSP in the soil profile and its relationships with root and soil carbohydrates, though glomalin is used to evaluate soil quality (He et al., 2010). Soil b-glucosidase is a useful soil quality indicator related to C cycling (Stott et al., 2010), whereas soil protease is highly correlated with soil N transformation (Rejsek et al., 2008). However, it is unclear if soil b-glucosidase or protease is related to GRSP, which contains both C and N. Citrus strongly depends on AMF in the field (Wu and Xia, 2006). To better understand the relationships between the formation of
* Corresponding author. Tel./fax: þ86 716 8066262. E-mail address: [email protected] (Q.-S. Wu). 0038-0717/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.soilbio.2011.10.002
GRSP and relevant soil variables, we first assessed the spatial distribution of GRSP, carbohydrates, water-stable aggregates (WSA), b-glucosidase and protease in the rhizosphere of Citrus unshiu (cv. Guoqing-1 grafted on Poncirus trifoliata), and then correlated GRSP with these variables. The experiment was conducted in a campus orchard that has w400 22-year-old citrus trees belonging to Yangtze University, China. Sixteen similar plants were randomly selected from four blocks (4-trees 4-blocks). Soils (Xanthi-udic ferralsol) and roots were collected at 0e10, 10e20, 20e30 and 30e40 cm depth within a 2 m radius of tree canopy on 26 May 2010. Samples from 4-trees/ block were mixed as one composite sample. Part of fresh samples was for mycorrhization and enzyme analysis. The rest was air-dried and ground (4 mm) for other analysis. The GRSP is assayed by the Bradford method as Bradfordreactive soil proteins (BRSPs), which include the easily extractable BRSP (EE-BRSP) and total BRSP (T-BRSP) (Rosier et al., 2006). Determination of EE-BRSP and T-BRSP was carried out following Bedini et al. (2009), WSA following Yan (1988), soluble sugar following Yemm and Willis (1954), soil hot water-extractable and hydrolyzable carbohydrates following Li et al. (2002), b-glucosidase activity following Stott et al. (2010), soil protease following
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Q.-S. Wu et al. / Soil Biology & Biochemistry 45 (2012) 181e183
80 a
70
70 b
60 50
b
60
a b
40
50
c
b
40 30
30 c
20
20
Root soluble sugar concentration
Root mycorrhizal colonization
80
10
10
0
0 0-10
10-20
20-30
30-40
Soil depth (cm) Fig. 1. Spatial distribution of root mycorrhizal colonization (%, open bars) and root soluble sugar concentration (mg/g DW, solid bars) in the rhizosphere of 22-year-old Citrus unshiu. Data (means SE, n ¼ 4) followed by different letters (a, b, c) above the bars are significantly different among soil depths for each variable at P < 0.05.
Cao et al. (1982), and AM colonization following Phillips and Hayman (1970). Significantly higher root AM mycorrhization of C. unshiu ranked at 0e10 > 10e20 z 20e30 > 30e40 cm soil depth (Fig. 1), which is consistent with results from C. aurantium and P. trifoliata C. limon (Levy et al., 1983). Root AM colonization was significantly negatively correlated with 0.5e1.0 mm size soil WSA and protease (r ¼ 0.57e0.93, P < 0.05), but significantly positively correlated with root soluble sugar, soil hot water-extractable and hydrolyzable carbohydrates, EE-BRSP, T-BRSP, and soil b-glucosidase (r ¼ 0.67e0.94, P < 0.01). Both EE-BRSP and T-BRSP were highest at 0e10 and 10e20 cm and higher at 20e30 cm than at 30e40 cm soil depth (Table 1), and EE-BRSP was significantly positively correlated with T-BRSP (r ¼ 0.89, P < 0.01). A range of 0.3e0.6 mg EE-BRSP/g soil in the rhizosphere of C. unshiu (Table 1) is consistent with those (0.3e0.6 mg/g) found in the Mu Us sandland, China (Bai et al., 2009) and in a long-term maize monoculture in Italy (Bedini et al., 2007), but lower than those (3.87e3.94 mg/g) in Manawatu and Ngamoko grassland soils in New Zealand (Rillig et al., 2006). A range of 0.5e0.8 mg T-BRSP/g soil (Table 1) is comparable with that in Texas grass lands (<2.0 mg, Wright and Upadhyaya, 1998), but is much lower than that found in grassland soils (9.3e12.0 mg, Rillig et al., 2006; Bedini et al., 2007), Mexican montane rainforest soils (1.0e12.2 mg, Violi et al., 2008) and Costa Rica tropical rainforest soils (3.9 mg, Lovelock et al., 2004). This may be due to a higher proportion of Glomus species in specific habitats (Treseder and Turner, 2007) or different land uses (Wang et al., 2011). Thus, BRSPs vary with soil types and/or host species. On the other hand, the concentration of glomalin may be dependent on hyphae turnover, since glomalin is released to soil when hyphae eventually die
or senesce (Driver et al., 2005). In addition, BRSPs were significantly positively correlated with AM colonization (P < 0.01, Table 2). Percentages of >2.00 mm WSA and total WSA were similar among soil depths, whereas total WSA was significantly higher in 1.00e2.00 mm > 0.50e1.00 mm > 2.00 mm > 0.25e0.50 mm size for the same soil depth (Table 1). Glomalin stabilizes soil aggregates through a ‘gluing’ action (Wright and Upadhyaya, 1998; Bedini et al., 2009). It is therefore reasonable that BRSPs are significantly positively correlated with the finest 0.25e0.50 mm WSA (P < 0.05e0.01) (Table 2). In addition, EE-BRSP was significantly negatively correlated with the 0.50e1.00 mm WSA (P < 0.05) (Table 2). In contrast, T-BRSP was positively correlated with 2.00e4.00 and 1.00e2.00 mm WSA under the pot-grown AM P. trifoliata seedlings (Wu et al., 2008). This inconsistency with no correlation between BRSPs and 1.00e2.00 or >2.00 mm WSA in this field study may be due to a better development of AM and glomalin under optimal glasshouse conditions. Root soluble sugar, soil hot water-extractable and hydrolyzable carbohydrates were highest at 0e10 cm and higher at 10e20/ 20e30 cm than at 30e40 cm soil depth (Fig. 1; Table 1). Both EEBRSP and T-BRSP were significantly positively correlated with root soluble sugar, soil hot water-extractable or hydrolyzable carbohydrates (P < 0.05 or 0.01) (Table 2), implying that a certain amount of root and soil C is from these GRSPs. Indeed, w40% of glomalin-stored C was from mycorrhizal roots (Phillips and Fahey, 2005) and up to 5% soil C was accumulated with glomalin deposition, when AM hyphae senesced (Treseder and Turner, 2007). Activity of soil b-glucosidase or protease was significantly decreased or increased with the increase of soil depth (Table 1). Both EE-BRSP and T-BRSP were significantly positively correlated with soil b-glucosidase (P < 0.01, Table 2), suggesting that GRSP, likes b-glucosidase, may contribute to the release of glucose maintaining metabolically active microbial biomass in the soil (Martinez-Salgado et al., 2010). Meanwhile, EE-BRSP and T-BRSP were significantly negatively correlated with soil protease (P < 0.01, Table 2), inconsistently with previous results on T-BRSP, obtained in four land use types (farmland, grassland, orchard and abandoned land) (Tang et al., 2009). Since GRSP can be rapidly degraded by soil protease (Bai et al., 2009), such inconsistency suggests that GRSP degradation varies with ecological habitats. Such variations between BRSPs and soil protease might also be attributed to glomalin itself, because glomalin is mixture of compounds and not a specific single molecule (Rillig and Steinberg, 2002), and the soil distribution of these various compounds possibly influences the activity of soil protease, especially at 30e40 cm depth. These results confirm that glomalin released by living mycorrhizal hyphae not only contributes to rhizospheric C but also to N since glomalin is a N-linked glycoprotein (Rillig and Steinberg,
Table 1 Spatial changes of soil water-stable aggregates, soil carbohydrates, BRSPs and soil related enzymes in the rhizosphere of Citrus unshiu cv. Guoqing 1 fruit tree grafted on Poncirus trifoliata. Soil depth (cm)
0.25e0.50 mm 0.50e1.00 mm 1.00e2.00 mm >2.00 mm
Soil water-stable aggregates (%)
0e10 10e20 20e30 30e40
7.4 3.5a, c 7.7 0.7a, d 6.2 3.0ab, d 4.7 2.6b, d
32.7 2.1a, a 30.7 2.4b, b 29.9 2.7b, b 27.8 1.8g, b
Carbohydrates (mg/g DW soil) BRSPs (mg/g DW soil) Soil enzymes
35.1 1.2g, a 34.5 1.3g, a 42.6 1.3a, a 37.4 1.8b, a
15.5 3.3a, 14.2 4.3a, 14.0 3.1a, 15.5 1.0a,
b c c c
Total
Hot waterextractable carbohydrate
Hydrolyzable carbohydrate
EE-BRSP
T-BRSP
b-glucosidase Protease (mg Glycine/ (mg sugar g DW soil) reduced/g DW soil)
90.6 3.4a 87.2 4.5a 92.7 5.1a 84.6 1.9a
3.7 0.2a 2.9 0.3b 2.9 0.2b 2.7 0.3b
16.4 0.8a 11.1 1.9b 10.9 0.5b 7.8 1.1g
0.6 0.1a 0.6 0.1ab 0.5 0.0b 0.3 0.0g
0.8 0.1a 0.8 0.1a 0.6 0.1b 0.5 0.0g
0.2 0.1g 0.4 0.1b 0.4 0.1b 0.9 0.1a
1.5 0.0a 1.0 0.1b 0.8 0.1g 0.6 0.1d
Note: data were means SE (n ¼ 4) followed by the same letter among soil depths for the same particle size within a column (a, b, g, d) or among particle sizes for the same soil depth within a row (a, b, c, d) are not significantly different at P < 0.05.
Q.-S. Wu et al. / Soil Biology & Biochemistry 45 (2012) 181e183
183
Table 2 Pearson correlation coefficients between EE-BRSP or T-BRSP and other variables (n ¼ 16 for the four soil depths).
EE-BRSP T-BRSP
Mycorrhizal colonization
Soil water-stable aggregate (mm) 0.25e0.50
0.50e1.00
1.00e2.00
>2.00
Total
0.876** 0.850**
0.677** 0.502*
0.614* 0.405
0.296 0.439
0.182 0.125
0.261 0.327
Root soluble sugar
Soil hot waterextractable carbohydrate
Soil hydrolyzable carbohydrate
Soil protease
Soil b-glucosidase
0.882** 0.874**
0.494* 0.489*
0.759** 0.778**
0.889** 0.834**
0.761** 0.836**
Note: * P < 0.05. ** P < 0.01.
2002; Lovelock et al., 2004). Therefore, both soil b-glucosidase and protease should be considered for studying soil C pools. In conclusion, our results demonstrate that the spatial distribution of GRSP is positively affected by mycorrhization, root and soil carbohydrates and soil b-glucosidase, but negatively by soil protease. Acknowledgements This study was supported by the Key Project of Chinese Ministry of Education (211107), the Science-Technology Research Project of Hubei Provincial Department of Education, China (Q20111301), and the National Natural Science Foundation of China (30800747). We are grateful to the Chief Editor RG Joergensen and two anonymous reviewers for their language editing and valuable comments on this manuscript. References Bai, C.M., He, X.L., Tang, H.L., Shan, B.Q., Zhao, L.L., 2009. Spatial distribution of arbuscular mycorrhizal fungi, glomalin and soil enzymes under the canopy of Astragalus adsurgens Pall. in the Mu Us sandland, China. Soil Biology and Biochemistry 41, 941e947. Bedini, S., Avio, L., Argese, E., Giovannetti, M., 2007. Effects of long-term land use on arbuscular mycorrhizal fungi and glomalin-related soil protein. Agriculture, Ecosystems and Environment 120, 463e466. 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 affect by the arbuscular mycorrhizal fungi species Glomus mosseae and Glomus intraradices. Soil Biology and Biochemistry 41, 1491e1496. Cao, C.J., Zhang, Z.M., Zhou, L.K., 1982. Comprasions of determined methods of several soil protease activities. Chinese Journal of Soil Science 13, 39e40 (in Chinese with English abstract). Driver, J.D., Holben, W.E., Rillig, M.C., 2005. Characterization of glomalin as a hyphal wall component of arbuscular mycorrhizal fungi. Soil Biology and Biochemistry 37, 101e106. He, X.L., Li, Y.P., Zhao, L.L., 2010. Dynamics of arbuscular mycorrhizal fungi and glomalin in the rhizosphere of Artemisia ordosica Krasch. in Mu Us sandland, China. Soil Biology and Biochemistry 42, 1313e1319. Levy, Y., Dodd, J., Krikun, J., 1983. Effect of irrigation, water salinity and rootstock on the vesicular distribution of vesicular-arbuscular mycorrhiza in citrus roots. New Phytologist 95, 397e403. Li, X.G., Cui, Z.J., Wang, L.Y., 2002. Effect of straw on soil organic carbon constitution and structural stability. Acta Pedologica Sinina 39, 421e428 (in Chinese with English abstract). Lovelock, C.E., Wright, S.F., Clark, D.A., Ruess, R.W., 2004. Soil stocks of glomalin produced by arbuscular mycorrhizal fungi across a tropical rainforest landscape. Journal of Ecology 92, 278e287.
Martinez-Salgado, M.M., Gutiérrez-Romero, V., Jannsens, M., Ortega-Blu, R., 2010. Biological soil quality indicators: a review. In: Mendez-Vilas, A. (Ed.), Current Research, Technology and Education Topics in Applied Microbiology and Microbial Biotechnology. World Scientific Publishing Co. Pte. Ltd., Singapore, pp. 319e328. Phillips, J.M., Hayman, D.S., 1970. Improved produres for clearing roots and staining parasitic and vesicular-arbuscular mycorrhizal fungi for rapid assessment of infection. Transactions British Mycological Society 55, 158e161. Phillips, R.P., Fahey, T.J., 2005. Patterns of rhizosphere carbon flux in sugar maple (Acer saccharum) and yellow birch (Betula allegheniensis) sapling. Global Change Biology 11, 983e995. Rejsek, K., Formanek, P., Pavelka, M., 2008. Estimation of protease activity in soils at low temperatures by casein amendment and with substitution of buffer by demineralized water. Amino Acids 35, 411e417. Rillig, M.C., Hoye, A.T., Carran, A., 2006. Minimal direct contribution of arbuscular mycorrhizal fungi to DOC leaching in grassland through losses of glomalinrelated soil protein. Soil Biology and Biochemistry 38, 2967e2970. Rillig, M.C., Steinberg, P.D., 2002. Glomalin production by an arbuscular mycorrhizal fungus: a mechanism of habitat modification. Soil Biology and Biochemistry 34, 1371e1374. Rosier, C.L., Hoye, A.T., Rillig, M.C., 2006. Glomalin-related soil protein: assessment of current detection and quantification tools. Soil Biology and Biochemistry 38, 2205e2211. Stott, D.E., Andrews, S.S., Liebig, M.A., Wienhold, B.J., Karlen, D.L., 2010. Evaluation of b-glucosidase activity as a soil quality indicator for the soil management assessment framework. Soil Science Society of America Journal 74, 107e119. Tang, H.L., Liu, L., Wang, L., Ba, C.J., 2009. Effect of land use type on profile distribution of glomalin. Chinese Journal of Eco-Agriculture 17, 1137e1142 (in Chinese with English abstract). Treseder, K.K., Turner, K.M., 2007. Glomalin in ecosystems. Soil Science Society of America Journal 71, 1257e1266. Violi, H.A., Barrientos-Priego, A.F., Wright, S.F., Escamilla-Prado, E., Morton, J.B., Menge, J.A., Lovatt, C.J., 2008. Disturbance changes arbuscular mycorrhizal fungi phenology and soil glomalin concentrations but not fungal spore composition in montane rainforests in Veracruz and Chiapas, Mexico. Forest Ecology and Management 254, 276e290. Wang, P., Liu, J.H., Xia, R.X., Wu, Q.S., Wang, M.Y., Dong, T., 2011. Arbuscular mycorrhizal development, glomalin-related soil protein (GRSP) content, and rhizospheric phosphatase activity in citrus orchards under different types of soil management. Journal of Plant Nutrition and Soil Science 174, 65e72. Wright, S.F., Upadhyaya, A., 1998. A survey of soils for aggregate stability and glomalin, a glycoproteins produced by hyphae of arbuscular mycorrhizal fungi. Plant and Soil 198, 97e107. Wu, Q.S., Xia, R.X., Zou, Y.N., 2008. Improved soil structure and citrus growth after inoculation with three arbuscular mycorrhizal fungi under drought stress. European Journal of Soil Biology 44, 122e128. Wu, Q.S., Xia, R.X., 2006. Arbuscular mycorrhizal fungi influence growth, osmotic adjustment and photosynthesis of citrus under well-watered and water stress conditions. Journal of Plant Physiology 163, 417e425. Yan, C.R., 1988. Research Methods of Soil Fertility (in Chinese). Agricultural Press, Beijing. Yemm, E.W., Willis, A.J., 1954. The estimation of carbohydrates in plant extracts by anthrone. Biochemical Journal 57, 508e514.
Contents lists available at SciVerse ScienceDirect
Soil Biology & Biochemistry journal homepage: www.elsevier.com/locate/soilbio
Short communication
Spatial distribution of glomalin-related soil protein and its relationships with root mycorrhization, soil aggregates, carbohydrates, activity of protease and b-glucosidase in the rhizosphere of Citrus unshiu Qiang-Sheng Wu a, *, Xin-Hua He b, c, d, Ying-Ning Zou a, Kai-Ping He a, Ya-Hong Sun a, Ming-Qin Cao a a
College of Horticulture and Gardening, Yangtze University, Jingzhou, Hubei 434025, China State Centre of Excellence for Ecohydrology, Edith Cowan University, Joondalup, WA 6027, Australia Centre for Ecosystem Management/School of Natural Sciences, Edith Cowan University, Joondalup, WA 6027, Australia d School of Plant Biology, University of Western Australia, Crawley, WA 6009, Australia b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 14 June 2011 Received in revised form 18 August 2011 Accepted 9 October 2011 Available online 20 October 2011
Relationships between the spatial distributions of glomalin-related soil protein (GRSP) and soil aggregates, carbohydrates or relevant enzymes are poorly studied. We found that two categories of GRSP, the easily extractable Bradford-reactive soil protein (EE-BRSP) and total BRSP (T-BRSP), respectively ranged between 0.3e0.6 and 0.5e0.8 mg/g DW soil, and these two BRSPs decreased with the increase of soil depth (0e40 cm) in the rhizosphere of a 22-year-old Citrus unshiu orchard. Both EE-BRSP and T-BRSP were significantly positively correlated with mycorrhization, 0.25e0.50 mm soil water-stable aggregates, water-extractable or hydrolyzable carbohydrates, and b-glucosidase, but significantly negatively correlated with protease. Our results demonstrate that the spatial distribution of GRSP is significantly affected by mycorrhization, soil carbohydrate, b-glucosidase and protease. Ó 2011 Elsevier Ltd. All rights reserved.
Keywords: b-glucosidase Citrus Glomalin Glomalin-related soil protein Mycorrhization Protease
As an insoluble N-linked glycoprotein with w37% carbon (C) and 3e5% nitrogen (N), glomalin contributes to soil structure stabilization and C pools (Wright and Upadhyaya, 1998; Lovelock et al., 2004). Glomalin is mainly produced by arbuscular mycorrhizal fungi (AMF) hyphae and quantified from soils as glomalin-related soil protein (GRSP). The GRSP is often related to soil C pool (Treseder and Turner, 2007), though glomalin has limited contribution to C budgets in short-term experiments (Driver et al., 2005). Less information is available to the spatial distributions of GRSP in the soil profile and its relationships with root and soil carbohydrates, though glomalin is used to evaluate soil quality (He et al., 2010). Soil b-glucosidase is a useful soil quality indicator related to C cycling (Stott et al., 2010), whereas soil protease is highly correlated with soil N transformation (Rejsek et al., 2008). However, it is unclear if soil b-glucosidase or protease is related to GRSP, which contains both C and N. Citrus strongly depends on AMF in the field (Wu and Xia, 2006). To better understand the relationships between the formation of
* Corresponding author. Tel./fax: þ86 716 8066262. E-mail address: [email protected] (Q.-S. Wu). 0038-0717/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.soilbio.2011.10.002
GRSP and relevant soil variables, we first assessed the spatial distribution of GRSP, carbohydrates, water-stable aggregates (WSA), b-glucosidase and protease in the rhizosphere of Citrus unshiu (cv. Guoqing-1 grafted on Poncirus trifoliata), and then correlated GRSP with these variables. The experiment was conducted in a campus orchard that has w400 22-year-old citrus trees belonging to Yangtze University, China. Sixteen similar plants were randomly selected from four blocks (4-trees 4-blocks). Soils (Xanthi-udic ferralsol) and roots were collected at 0e10, 10e20, 20e30 and 30e40 cm depth within a 2 m radius of tree canopy on 26 May 2010. Samples from 4-trees/ block were mixed as one composite sample. Part of fresh samples was for mycorrhization and enzyme analysis. The rest was air-dried and ground (4 mm) for other analysis. The GRSP is assayed by the Bradford method as Bradfordreactive soil proteins (BRSPs), which include the easily extractable BRSP (EE-BRSP) and total BRSP (T-BRSP) (Rosier et al., 2006). Determination of EE-BRSP and T-BRSP was carried out following Bedini et al. (2009), WSA following Yan (1988), soluble sugar following Yemm and Willis (1954), soil hot water-extractable and hydrolyzable carbohydrates following Li et al. (2002), b-glucosidase activity following Stott et al. (2010), soil protease following
182
Q.-S. Wu et al. / Soil Biology & Biochemistry 45 (2012) 181e183
80 a
70
70 b
60 50
b
60
a b
40
50
c
b
40 30
30 c
20
20
Root soluble sugar concentration
Root mycorrhizal colonization
80
10
10
0
0 0-10
10-20
20-30
30-40
Soil depth (cm) Fig. 1. Spatial distribution of root mycorrhizal colonization (%, open bars) and root soluble sugar concentration (mg/g DW, solid bars) in the rhizosphere of 22-year-old Citrus unshiu. Data (means SE, n ¼ 4) followed by different letters (a, b, c) above the bars are significantly different among soil depths for each variable at P < 0.05.
Cao et al. (1982), and AM colonization following Phillips and Hayman (1970). Significantly higher root AM mycorrhization of C. unshiu ranked at 0e10 > 10e20 z 20e30 > 30e40 cm soil depth (Fig. 1), which is consistent with results from C. aurantium and P. trifoliata C. limon (Levy et al., 1983). Root AM colonization was significantly negatively correlated with 0.5e1.0 mm size soil WSA and protease (r ¼ 0.57e0.93, P < 0.05), but significantly positively correlated with root soluble sugar, soil hot water-extractable and hydrolyzable carbohydrates, EE-BRSP, T-BRSP, and soil b-glucosidase (r ¼ 0.67e0.94, P < 0.01). Both EE-BRSP and T-BRSP were highest at 0e10 and 10e20 cm and higher at 20e30 cm than at 30e40 cm soil depth (Table 1), and EE-BRSP was significantly positively correlated with T-BRSP (r ¼ 0.89, P < 0.01). A range of 0.3e0.6 mg EE-BRSP/g soil in the rhizosphere of C. unshiu (Table 1) is consistent with those (0.3e0.6 mg/g) found in the Mu Us sandland, China (Bai et al., 2009) and in a long-term maize monoculture in Italy (Bedini et al., 2007), but lower than those (3.87e3.94 mg/g) in Manawatu and Ngamoko grassland soils in New Zealand (Rillig et al., 2006). A range of 0.5e0.8 mg T-BRSP/g soil (Table 1) is comparable with that in Texas grass lands (<2.0 mg, Wright and Upadhyaya, 1998), but is much lower than that found in grassland soils (9.3e12.0 mg, Rillig et al., 2006; Bedini et al., 2007), Mexican montane rainforest soils (1.0e12.2 mg, Violi et al., 2008) and Costa Rica tropical rainforest soils (3.9 mg, Lovelock et al., 2004). This may be due to a higher proportion of Glomus species in specific habitats (Treseder and Turner, 2007) or different land uses (Wang et al., 2011). Thus, BRSPs vary with soil types and/or host species. On the other hand, the concentration of glomalin may be dependent on hyphae turnover, since glomalin is released to soil when hyphae eventually die
or senesce (Driver et al., 2005). In addition, BRSPs were significantly positively correlated with AM colonization (P < 0.01, Table 2). Percentages of >2.00 mm WSA and total WSA were similar among soil depths, whereas total WSA was significantly higher in 1.00e2.00 mm > 0.50e1.00 mm > 2.00 mm > 0.25e0.50 mm size for the same soil depth (Table 1). Glomalin stabilizes soil aggregates through a ‘gluing’ action (Wright and Upadhyaya, 1998; Bedini et al., 2009). It is therefore reasonable that BRSPs are significantly positively correlated with the finest 0.25e0.50 mm WSA (P < 0.05e0.01) (Table 2). In addition, EE-BRSP was significantly negatively correlated with the 0.50e1.00 mm WSA (P < 0.05) (Table 2). In contrast, T-BRSP was positively correlated with 2.00e4.00 and 1.00e2.00 mm WSA under the pot-grown AM P. trifoliata seedlings (Wu et al., 2008). This inconsistency with no correlation between BRSPs and 1.00e2.00 or >2.00 mm WSA in this field study may be due to a better development of AM and glomalin under optimal glasshouse conditions. Root soluble sugar, soil hot water-extractable and hydrolyzable carbohydrates were highest at 0e10 cm and higher at 10e20/ 20e30 cm than at 30e40 cm soil depth (Fig. 1; Table 1). Both EEBRSP and T-BRSP were significantly positively correlated with root soluble sugar, soil hot water-extractable or hydrolyzable carbohydrates (P < 0.05 or 0.01) (Table 2), implying that a certain amount of root and soil C is from these GRSPs. Indeed, w40% of glomalin-stored C was from mycorrhizal roots (Phillips and Fahey, 2005) and up to 5% soil C was accumulated with glomalin deposition, when AM hyphae senesced (Treseder and Turner, 2007). Activity of soil b-glucosidase or protease was significantly decreased or increased with the increase of soil depth (Table 1). Both EE-BRSP and T-BRSP were significantly positively correlated with soil b-glucosidase (P < 0.01, Table 2), suggesting that GRSP, likes b-glucosidase, may contribute to the release of glucose maintaining metabolically active microbial biomass in the soil (Martinez-Salgado et al., 2010). Meanwhile, EE-BRSP and T-BRSP were significantly negatively correlated with soil protease (P < 0.01, Table 2), inconsistently with previous results on T-BRSP, obtained in four land use types (farmland, grassland, orchard and abandoned land) (Tang et al., 2009). Since GRSP can be rapidly degraded by soil protease (Bai et al., 2009), such inconsistency suggests that GRSP degradation varies with ecological habitats. Such variations between BRSPs and soil protease might also be attributed to glomalin itself, because glomalin is mixture of compounds and not a specific single molecule (Rillig and Steinberg, 2002), and the soil distribution of these various compounds possibly influences the activity of soil protease, especially at 30e40 cm depth. These results confirm that glomalin released by living mycorrhizal hyphae not only contributes to rhizospheric C but also to N since glomalin is a N-linked glycoprotein (Rillig and Steinberg,
Table 1 Spatial changes of soil water-stable aggregates, soil carbohydrates, BRSPs and soil related enzymes in the rhizosphere of Citrus unshiu cv. Guoqing 1 fruit tree grafted on Poncirus trifoliata. Soil depth (cm)
0.25e0.50 mm 0.50e1.00 mm 1.00e2.00 mm >2.00 mm
Soil water-stable aggregates (%)
0e10 10e20 20e30 30e40
7.4 3.5a, c 7.7 0.7a, d 6.2 3.0ab, d 4.7 2.6b, d
32.7 2.1a, a 30.7 2.4b, b 29.9 2.7b, b 27.8 1.8g, b
Carbohydrates (mg/g DW soil) BRSPs (mg/g DW soil) Soil enzymes
35.1 1.2g, a 34.5 1.3g, a 42.6 1.3a, a 37.4 1.8b, a
15.5 3.3a, 14.2 4.3a, 14.0 3.1a, 15.5 1.0a,
b c c c
Total
Hot waterextractable carbohydrate
Hydrolyzable carbohydrate
EE-BRSP
T-BRSP
b-glucosidase Protease (mg Glycine/ (mg sugar g DW soil) reduced/g DW soil)
90.6 3.4a 87.2 4.5a 92.7 5.1a 84.6 1.9a
3.7 0.2a 2.9 0.3b 2.9 0.2b 2.7 0.3b
16.4 0.8a 11.1 1.9b 10.9 0.5b 7.8 1.1g
0.6 0.1a 0.6 0.1ab 0.5 0.0b 0.3 0.0g
0.8 0.1a 0.8 0.1a 0.6 0.1b 0.5 0.0g
0.2 0.1g 0.4 0.1b 0.4 0.1b 0.9 0.1a
1.5 0.0a 1.0 0.1b 0.8 0.1g 0.6 0.1d
Note: data were means SE (n ¼ 4) followed by the same letter among soil depths for the same particle size within a column (a, b, g, d) or among particle sizes for the same soil depth within a row (a, b, c, d) are not significantly different at P < 0.05.
Q.-S. Wu et al. / Soil Biology & Biochemistry 45 (2012) 181e183
183
Table 2 Pearson correlation coefficients between EE-BRSP or T-BRSP and other variables (n ¼ 16 for the four soil depths).
EE-BRSP T-BRSP
Mycorrhizal colonization
Soil water-stable aggregate (mm) 0.25e0.50
0.50e1.00
1.00e2.00
>2.00
Total
0.876** 0.850**
0.677** 0.502*
0.614* 0.405
0.296 0.439
0.182 0.125
0.261 0.327
Root soluble sugar
Soil hot waterextractable carbohydrate
Soil hydrolyzable carbohydrate
Soil protease
Soil b-glucosidase
0.882** 0.874**
0.494* 0.489*
0.759** 0.778**
0.889** 0.834**
0.761** 0.836**
Note: * P < 0.05. ** P < 0.01.
2002; Lovelock et al., 2004). Therefore, both soil b-glucosidase and protease should be considered for studying soil C pools. In conclusion, our results demonstrate that the spatial distribution of GRSP is positively affected by mycorrhization, root and soil carbohydrates and soil b-glucosidase, but negatively by soil protease. Acknowledgements This study was supported by the Key Project of Chinese Ministry of Education (211107), the Science-Technology Research Project of Hubei Provincial Department of Education, China (Q20111301), and the National Natural Science Foundation of China (30800747). We are grateful to the Chief Editor RG Joergensen and two anonymous reviewers for their language editing and valuable comments on this manuscript. References Bai, C.M., He, X.L., Tang, H.L., Shan, B.Q., Zhao, L.L., 2009. Spatial distribution of arbuscular mycorrhizal fungi, glomalin and soil enzymes under the canopy of Astragalus adsurgens Pall. in the Mu Us sandland, China. Soil Biology and Biochemistry 41, 941e947. Bedini, S., Avio, L., Argese, E., Giovannetti, M., 2007. Effects of long-term land use on arbuscular mycorrhizal fungi and glomalin-related soil protein. Agriculture, Ecosystems and Environment 120, 463e466. 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 affect by the arbuscular mycorrhizal fungi species Glomus mosseae and Glomus intraradices. Soil Biology and Biochemistry 41, 1491e1496. Cao, C.J., Zhang, Z.M., Zhou, L.K., 1982. Comprasions of determined methods of several soil protease activities. Chinese Journal of Soil Science 13, 39e40 (in Chinese with English abstract). Driver, J.D., Holben, W.E., Rillig, M.C., 2005. Characterization of glomalin as a hyphal wall component of arbuscular mycorrhizal fungi. Soil Biology and Biochemistry 37, 101e106. He, X.L., Li, Y.P., Zhao, L.L., 2010. Dynamics of arbuscular mycorrhizal fungi and glomalin in the rhizosphere of Artemisia ordosica Krasch. in Mu Us sandland, China. Soil Biology and Biochemistry 42, 1313e1319. Levy, Y., Dodd, J., Krikun, J., 1983. Effect of irrigation, water salinity and rootstock on the vesicular distribution of vesicular-arbuscular mycorrhiza in citrus roots. New Phytologist 95, 397e403. Li, X.G., Cui, Z.J., Wang, L.Y., 2002. Effect of straw on soil organic carbon constitution and structural stability. Acta Pedologica Sinina 39, 421e428 (in Chinese with English abstract). Lovelock, C.E., Wright, S.F., Clark, D.A., Ruess, R.W., 2004. Soil stocks of glomalin produced by arbuscular mycorrhizal fungi across a tropical rainforest landscape. Journal of Ecology 92, 278e287.
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