Soil microbial carbon and phosphorus as influenced by phosphorus fertilization and tillage in a maize-soybean rotation in south-western Quebec Aiguo Liu1,2, Chantal Hamel3, Tim Spedding4, Tie-Quan Zhang5, Rene Mongeau6, Georges R. Lamarre7, and Gilles Tremblay8 1
College of Resources and Environmental Science, Shanxi Agricultural University, Taigu, Shanxi, China; 2Pacific Research Center, Agriculture and Agri-Food Canada, 6947 Highway 7, P.O. Box 1000, Agassiz, British Columbia, Canada V0M 1A0 (e-mail:
[email protected]); 3Semiarid Prairie Agricultural Research Centre, Agriculture and Agri-Food Canada, Box. 1030, Airport Road, Swift Current, Saskatchewan, Canada S9H 3X2; 4Department of Natural Resource Sciences, Macdonald Campus, McGill University, 21111 Lakeshore Road, Ste-Anne-deBellevue, Quebec, Canada H9X 3V9; 5Greenhouse and Processing Crops Research Centre, Agriculture and Agri-Food Canada, 2585 County Rd 20E, Harrow, Ontario, Canada N0R 1G0; 6Ministe`re de l’Agriculture, des Peˆcheries et de l’Alimentation, 870 rue Cure´ St-Georges, St-Jean-sur-Richelieu, Quebec, Canada J2X 2Z8; 7 Ministe`re de l’Agriculture, des Peˆcheries et de l’Alimentation, 177 St-Joseph, Ste-Martine Que´bec, Canada J0S 1V0; and 8CE´ROM Centre de recherche sur les grains, 335 chemin des Vingt-cinq Est, Saint-Bruno-de-Montarville, Que´bec, Canada 3V5 4P6. Received 22 February 2007, accepted 29 October 2007. Liu, A., Hamel, C., Spedding, T., Zhang, T.-Q., Mongeau, R., Lamarre, G. R. and Tremblay, G. 2008. Soil microbial carbon and phosphorus as influenced by phosphorus fertilization and tillage in a maize-soybean rotation in south-western Quebec. Can. J. Soil Sci. 88: 2130. The management of the soil microbial P pool could improve system sustainability. The long-term impact of inorganic P inputs (0, 40 and 80 kg P2O5 ha1) and tillage (conventional and ridge tillage) on soil microbial biomass P (SMB-P) was defined in the soybean phase of a 10-yr-old maize-soybean rotation, on a Gleysolic clay-loam. Soil microbial biomass C (SMB-C) and SMB-P were determined four times in the growing season. Yearly applications of 40 kg P2O5 ha1 increased soil organic carbon level, partly explaining the increase in SMB-P measured at this rate. Results suggest that P-mediated modification of soil microbial community structure also contributed to increase SMB-P at this P rate. The increase of application of P rate (80 kg P2O5 ha1) produced the largest soybean yield, but generally decreased SMB-P. Our results and those of others suggest that balanced soil fertility (corresponding to fertilizer recommendations in our case) promotes soil microbial development. The use of ridge tillage did not increase the soil organic carbon level, but did increase SMB-P. The SMB-P pool was large (equivalent to 24.4 kg P2O5 ha 1) in the 0- to 20-cm soil layer, but unrelated to yield. Improving the ability of crops to access this pool of soil P would increase the value of its management. Key words: Conventional tillage, conservation tillage, P fertilization, soil microbial biomass C, P, and C to P ratio Liu, A., Hamel, C., Spedding, T., Zhang, T.-Q., Mongeau, R., Lamarre, G. R. et Tremblay, G. 2008. Influence des engrais phosphate´s et du travail du sol sur le carbone et le phosphore microbienne du sol dans un assolement maı¨ s-soja du sud-ouest du Que´bec. Can. J. Soil Sci. 88: 2130. Ge´rer le re´servoir de P microbienne du sol pourrait ame´liorer la pe´rennite´ des syste`mes agricoles. Les auteurs ont pre´cise´ l’incidence a` long terme des apports de P inorganique (0, 40 et 80 kg de P2O5 par hectare) et du travail du sol (classique et en billons) sur le P issu de la biomasse microbienne (PBM), dans la fraction soja d’un assolement maı¨ s-soja de dix ans re´alise´ sur un gleysol de type loam argileux. Le C de la biomasse microbienne (PBM) et le PBM ont e´te´ dose´s a` quatre moments durant la saison ve´ge´tative. L’application annuelle de 40 kg de P2O5 par hectare augmente la teneur en carbone organique du sol, ce qui explique en partie la hausse du PBM releve´e a` ce taux d’application. Les re´sultats laissent croire que la modification apporte´e par le P a` la structure de la population microbienne du sol contribue aussi a` augmenter le PBM a` ce taux. La majoration du taux d’application d’engrais P (80 kg de P2O5 par hectare) engendre le meilleur rendement du soja, mais on assiste ge´ne´ralement a` une diminution du PBM. Ces re´sultats et ceux d’autres chercheurs donnent a` penser qu’une bonification e´quilibre´e du sol (identique aux recommandations de fertilisation, dans le cas qui nous concerne) favorise le de´veloppement de la microflore tellurique. Le travail du sol en billons n’accroıˆ t pas la concentration de carbone organique mais bien celle de PBM. Les vingt premiers centime`tres de sol renferment beaucoup de PBM (l’e´quivalent de 24,4 kg de P2O5 par hectare), mais ce facteur ne pre´sente aucun lien avec le rendement. La gestion des cultures serait plus profitable si on amenait ces dernie`res a` mieux exploiter le re´servoir de P pre´sent dans le sol. Mots cle´s: Travail classique du sol, travail re´duit du sol, amendement P, C de la biomasse microbienne, P, ratio C:P
Abbreviations: AMF, arbuscular mycorrhizal; MBP, microbial biomass P; SMB-C, soil microbial biomass C; SMB-P, soil microbial biomass P 21
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CANADIAN JOURNAL OF SOIL SCIENCE
Soil microorganisms are a critical link in N, P and S cycling, and a pool of nutrients; some of them directly promote plant growth (Paul and Clark 1996). Management of soil microbial biomass could improve the sustainability of cropping systems. However, the effects of cropping practices such as tillage and fertilization on the soil microbial biomass, and in particular the soil microbial biomass P (SMB-P) pool, need to be better understood before it can be wisely managed. Soil tillage is perceived as a key factor affecting soil microorganisms (Lupwayi et al. 2001). Tillage reduces soil organic matter level by enhancing wind and water erosion loss (Pimentel and Kounang 1998) and exposing occluded organic compounds to oxidation (Baldock and Skjemstad 2000), and soil microbial biomass is usually related to soil organic matter level (Wardle 1998). Many researchers have shown that soil microbial biomass and activity respond to changes in tillage practice (Fyles et al. 1988; Angers et al. 1993; Bossio et al. 1998; Wardle et al. 1999), with conservation tillage increasing soil microbial biomass C (SMB-C) on the long-term (Salinas-Garcia et al. 1997a, b). Tillage also dilutes soil organic matter in the topsoil layer with impact on soil microbial communities. SMB-P and SMB-C in the 0- to 5-cm soil layer were reduced by about one-half when a pasture was ploughed (Aslam et al. 1999). The effect of P fertilization on soil microorganisms is still not well known. Phosphorus availability can increase microbial biomass and activity in natural settings (Sundareshwar et al. 2003) and inorganic P fertilization was reported to influence microbial activities (Acton and Gregorich 1995; Belay et al. 2002). According to the rules of biological stoichiometry (Elser et al. 2003), the relative amounts of available nutrients in the environment is a selective force shaping biodiversity in microbial communities. A study of the literature revealed that soil microbial biomass C:N:P ratio is well constrained around 60:7:1, at the global scale, although significant variations are seen in the ratios measured in grassland versus forest ecosystems, reflecting dissimilarities in soil microbial community structure (Cleveland and Liptzin 2007). Variation in microbial C:N:P ratios are attributed to the relative abundance of fungi and bacteria in the different microbial communities. Fungi tend to be richer in structural elements and their C:N and C:P ratio is generally higher that that of bacteria. Phosphorus is an essential element for microbial growth and function in addition to being essential to crop plants production, but there are only a few reports on the effects of P fertilizer on soil microbial biomass. Belay et al. (2002) reported that the sole application of inorganic phosphorus fertilizer resulted in the reduction of total soil organic C, but did not change the size of the SMB-P pool or the number of bacteria, actinomycetes and fungi. Under semiarid conditions, the soil microbial community was found to be highly plastic and 37 yr of wheat production without P fertilization did not
influence soil microbial biomass. Rather, variations in the biomass and structure of the soil microbial community were attributed to changes in soil osmotic potential (Hamel et al. 2006). As a habitat for microorganisms, soil chemical and physical conditions do influence microbial growth and community structure. These conditions change with variation in weather and plant development, and soil microbial biomass tends to vary seasonally (Patra et al. 1990; Perrott et al. 1990; Murphy et al. 1998). Agricultural activities further alter the soil environment in various ways. The soil microbial community is in constant change (Bossio et al. 1998; Ajwa et al. 1999; Spedding et al. 2004) and should be seen as a succession driven by variations in the soil environment. As such, the influence of cropping practices on the soil microbial biomass should be examined over the growing season, as the expression of effects can be masked by point-in-time environmental conditions. This study aimed at filling the gap in knowledge on the effect of P fertilization and tillage on the soil microbial biomass as a P pool for crops. We tested the hypothesis that inorganic P fertilization and conservation tillage would increase the SMB-P pool. For this, we analyzed the variation in the C and P content of soil microbial biomass over the growing season, in a 10-yrold field experiment. MATERIALS AND METHODS Experimental Site The experimental site was located at L’Acadie, southwestern Quebec (45819?N, 73821?W). Mean annual air temperature is 7.28C at this location, and the average annual precipitation is 960 mm. The soil is a clay-loam of the St-Blaise series (Dark Grey Gleysol). The site was under conventional tillage prior to treatment application. Soil nutrient status in the 0- to 10-cm and 10- to 20cm soil layers on the control plots is given in Table 1. Experimental Design A maize-soybean rotation experiment was set up in 1992 to test the long-term effects of tillage and P fertilization practices on yield and soil quality. The experiment included three levels of P applications, 0 kg P2O5 ha1 (P0), 40 kg P2O5 ha1 (P40), and 80 kg P2O5 ha1 (P80), and two tillage management systems, a conventional and a conservation tillage system described below. The experiment was arranged in a split plot design with four replicates. The main plots (25 m by 2.28 m) received the tillage treatments, and the subplots received the fertilizer treatments. Subplots consisted in six 25-m long rows, 76 cm apart, thus subplot size was 4.58 m25 m. Plots with conservation tillage treatment were under ridge tillage with permanent ridges from 1991 to 1995, and then under no-till on the permanent ridges i.e., crops were seeded on the permanent ridges with a no-till seeder. The conventional tillage treatment consisted of
LIU ET AL. * TILLAGE, P FERTILIZATION AND SOIL MICROBIAL BIOMASS
23
Table 1. Soil nutrient status in the 0- to 10-cm and 10- to 20-cm depths of the study site Mehlich 3 extractable nutrients Depth (cm)
Organic C (g kg1)
Total N (g kg1)
P
K
Ca
Mg
Zn
Cu
Fe
010 1020
18.190.87z 16.790.94
1.2890.09 1.1990.08
6894.0 6293.7
10595.1 8294.0
34229147 26769115
396918.6 332915.6
4.390.29 5.390.36
0.7090.05 0.6090.04
188910.0 190910.1
z
All values represent means9SEM.
moldboard ploughing (conventional tillage) to 20-cm depth after harvest in fall and two passes of harrow cultivator to 10-cm depth for seedbed preparation in May, prior to seeding. Maize (cv. Pioneer 38W36) was seeded at a rate of 75 000 plants ha1 in 1992, 1993, 1994, 1996, 1998, 2000, while soybean (cv. Hogata) was seeded at a rate of 450 000 plants ha1 in 1995, 1997, 1999 and 2001. The P fertilizer treatments (P0, P40 and P80) were applied every year. Potassium fertilizer was not applied at least since 1991, but 160 kg N ha1 was applied to maize crops in 1992, 1993, 1994, 1996, 1998, and 2000. The conventional seeder placed the fertilizer 5 cm below the soil surface, whereas the no-till seeder placed it 2.5 cm below the soil surface. Yearly, preseeding applications of Round-Up† (a.i. glyphosate) were used on all the plots including conventional and conservation tillage treatments to control weeds. The contact herbicides Basagran forte† (a.i. Imazamox) and Pursuit† (a.i. Imazethapyr), in soybean, and Bladex† (a.i. Cyanazine) and Dual† (a.i. Metolachlor), in maize, were used post-emergence. Only the two middle rows of each plot were used for grain yield determination after harvesting with a combine. Seed yield data are expressed as kg ha1 after adjustment to 13% moisture. Soil sampling Sampling was undertaken at four dates during the 2001 growing season. These were: pre-planting (May 02), 6 wk after planting (Jun. 18), late-July (Jul. 23) and preharvest at crop maturity (Oct. 03). At each sampling date, a soil probe (2.0-cm diameter) was used to obtain cores from the 0- to 20-cm soil layer. Cores were divided into two segments, 010 cm and 1020 cm, and each was placed in a separate sample bag. Ten soil cores (2.0 cm in diameter) for each depth, taken 5 cm from the plant rows at random throughout the sub-plot, were combined to make one composite sample for each depth and each sub-plot. Samples were then placed in a cooler containing ice in the field, brought to the laboratory and stored at 4oC before analysis. SMB-C SMB-C was determined on duplicate fresh field moist samples kept at 4oC for a maximum of 3 d prior to analysis. Visible plant debris and roots were removed with tweezers. SMB-C was determined by chloroform fumigation extraction as outlined by Voroney et al.
(1993). Briefly, 15 g of field-moist soil was extracted with 45 mL of 0.5 M K2SO4 after shaking for 1 h at 150 rev. min 1 and filtered through P2 filter paper (fine porosity). These extracts are referred to as non-fumigated. A 50-mL glass beaker containing 15 g of fieldmoist soil was placed in a desiccator lined with moist paper towel and then fumigated with non-alcohol based chloroform (Caledon Laboratories, Georgetown, Canada) for 24 h at room temperature in the dark. After 24 h, the paper towel and chloroform were removed from the desiccator, which was then evacuated for 5 min. The soil was extracted and filtered in the same manner as outlined for the non-fumigated soil. These extracts are referred to as fumigated. Both fumigated and non-fumigated extracts were split into two parts and each part was frozen until C analysis. A third soil subsample was used to measure gravimetric soil moisture content after drying for 24 h at 1058C. For carbon analysis, 5 mL of sample was diluted with 15 mL of de-ionized water and solution pH was adjusted to 2.5 00.5 with 2 N HCl. Organic carbon in fumigated and non-fumigated extracts underwent combustion at 680oC and was detected with a non-dispersive infrared gas analyzer using a Shimadzu TOC-V organic carbon analyzer (Shimadzu Corporation, Kyoto, Japan). SMBC was calculated as the difference in organic carbon in fumigated and non-fumigated samples divided by an extraction efficiency factor of 0.45 (Jenkinson et al. 2004). SMB-P SMB-P was determined using the procedure described by Brookes et al. (1982, 1985). Three soil sub-samples of approximately 12 g each were used; the first part, for non-fumigated soil, the second one for correction of inorganic P fixed during 0.5 M NaHCO3 extraction by added spike P, and the third part for fumigated soil. All extractions were done in triplicate for each soil. Thus, a flask containing 2 g of soil (oven-dry basis) was immediately shaken with 40 mL of 0.5 M NaHCO3 (pH 8.5) for 30 min at 150 rev. min 1 on an orbital shaker and filtered through Whatman 42 filter paper. A second flask containing 2 g of soil was spiked with 25 mg P g1 before extraction with the 0.5 M NaHCO3 solution. A third flask containing approximately 2 g of soil was fumigated with chloroform for 24 h and then extracted with NaHCO3 following the procedure above. A 1-mL aliquot of each NaHCO3 extract was neutralized
24
CANADIAN JOURNAL OF SOIL SCIENCE
with 0.375 mL of 10% H2SO4, vortexed and diluted with 10 mL of double-distilled water prior to analysis. Inorganic P concentration in unfumigated and fumigated NaHCO3 extracts was measured on a microplate system using the ammonium molybdate-malachite green assay (Jeannotte et al. 2004). Absorbance was measured at 600 nm using a Bio-Tek Model EL 309 microplate reader (Bio-Tek Instruments, Winooski, VT). The microbial biomass P (MBP) concentration (mg P g1 soil) was calculated as: MBP (DIPfumigated B DIPunfumigated )= (KEP % recovery)
(1)
where DIPfumigated is the dissolved inorganic P concentration (mg P g1 soil) in NaHCO3 extracts of fumigated soil, DIPunfumigated is the DIP concentration (mg P g1 soil) in NaHCO3 extracts of unfumigated soil, KEP is 0.4, accounting for the efficiency of P extraction from lysed microbial cells (Brookes et al. 1982; Hedley and Stewart 1982), and percent covery is the proportion of spike recovered in each unfumigated soil sample. Soil Chemical Measurements Soil chemical measurements were made on the field samples taken on 2001 May 02. Total organic C was measured with a Carlo-Erba. Total N was determined by the Kjeldahl method (McGill et al. 1993). After soil samples were digested, nitrogen in these solutions was analyzed colorimetrically using the Lachat Quik-Chem AE flow-injection autoanalyzer (Lachat Instruments, Milwaukee, WI). Soil available P was extracted with the Mehlich-3 extractant (Mehlich 1984), which is highly correlated with plant P uptake in many soils of Quebec (Tran and Simard 1993). Statistical Analysis Data were analyzed with a mixed model using the PROC MIXED function of SAS software (SAS for Windows, Version 9.1, Cary, NC). The first two variables, tillage and P fertilizer arranged as a split-plot design, were fixed levels, and soil depth, random levels;
sampling time was considered as a repeated-measure. Statistical significance was determined at the 5% probability level. When interactions between factors were significant, the means of combinations of each level of these factors were compared. RESULTS Soil Organic C, total N and Extractable P Ten years of tillage treatments did not modify soil organic C, total N and extractable P levels (Table 2). Soil organic C was greater in both, the 0- to 10-cm and 10- to 20-cm soil layers, when P fertilizer was applied. Total N in the 0- to 10-cm soil layer was increased with the high P fertilization rate (P80) as compared with P0. Mehlich-3 extractible P level increased with P fertilization rates. SMB-C The SMB-C concentration was greater in the top 0- to 10-cm soil layer than at 10- to 20-cm at all sampling times (Figs. 1 and 2). There were interactions between tillage and sampling time, and between P rate and sampling time in both soil layers (Table 3). The SMB-C in the 0- to 10-cm soil layer was larger under soil conservation tillage than conventional tillage in May and October, but not in June and July, a period of vigorous plant growth. In the 10- to 20-cm soil layer, conventional tillage reduced SMB-C in June, compared with conservation tillage (Fig. 1). At most sampling times, P fertilization (P40 and P80) increased SMB-C concentration in the 0- to 10-cm soil layers, although there was no difference between treatments in June or between the P0 and P80 treatments in October. In the 10- to 20-cm soil layer, the P40 treatment had more SMB-C in May and July compared with P0 (Fig. 2). The P80 treatment had more SMB-C in May than the P0, but a lower SMB-C concentration in June than P0 and P40 treatments (Fig. 2). In the top 0- to 10-cm soil layer, SMB-C was greater in July than at all other sampling times (Figs. 1 and 2). In the 10- to 20-cm soil layer, SMB-C decreased from July to October (Figs. 1 and 2).
Table 2. Effects of tillage practice and P fertilization on total organic C, total N and Mehlich-3 extractable P in the 0- to 10-cm and 10- to 20-cm soil depths Total organic C (mg g 1) Factor
Total N (mg g 1)
Mehlich-3 extractable P (mg g 1)
Level
010 cm
1020 cm
010 cm
1020 cm
010 cm
1020 cm
Tillage system
Conventional Conservation
19.8 91.12a 21.2 91.35a
19.191.11a 20.090.85a
1.28 90.08a 1.29 90.09a
1.21900.7a 1.1990.06a
45.2 92.11a 46.4 93.22a
44.893.26a 45.294.21a
P level (kg P2O5 ha1)
0 40 80
18.1 90.83b 21.4 91.23a 22.1 91.42a
16.790.73b 20.991.26a 21.191.31a
1.18 90.06b 1.30 90.07ab 1.38 90.11a
1.1290.08a 1.2490.08a 1.2590.09a
28.5 91.94c 47.2 92.76b 61.7 94.56a
26.492.25c 45.193.62b 63.694.18a
ac All values represent means9SEM, and different letters following the mean values in the same column within the same factor indicate significant difference at P B0.05.
LIU ET AL. * TILLAGE, P FERTILIZATION AND SOIL MICROBIAL BIOMASS 260
260
0-10cm
0-10cm
240
220
220
200
200
180
180 Conventional Conservational
160 140 10-20cm 240 220
SMB-C (µg g-1)
SMB-C (µg g-1)
240
P0 P40 P80
160 140 260
10-20cm
P0 P40 P80
240 220
200
200
180
180
160
Conventional Conservational
140
160 140
May
June
July
Oct
May
June July Sampling time
Sampling time
Fig. 1. Effects of tillage system on SMB-C in the 0- to 10-cm and 10- to 20-cm soil layers at different sampling times. All values represent means9SEM (n 12).
Oct
Fig. 2. Effects of P fertilizer rates on SMB-C in the 0- to 10-cm and 10- to 20-cm soil layers at different sampling times. All values represent means9SEM (n 8).
Table 3. Summary of analysis of variance for SMB-C, SMB-P AND SMB-C:P in 0-to 10-cm and 10- to 20-cm layers 010 cm Parameter SMB-C
SMB-P
SMB-C:P
25
1020 cm
Source
DF
F value
PrP
DF
F value
PrP
Tillage (T) P level (P) Sampling Time (S) TP TS P S TP S Tillage (T) P level (P) Sampling time (S) TP TS P S TP S
1 2 3 2 3 6 6 1 2 3 2 3 6 6
9.16 19.5 3.35 0.21 7.58 2.34 0.88 6.35 6.55 25.8 0.29 2.76 0.32 0.68
0.0035 B0.0001 0.0240 0.8150 0.0002 0.0411 0.5181 0.0142 0.0024 B0.0001 0.7487 0.0490 0.9236 0.6678
1 2 3 2 3 6 6 1 2 3 2 3 6 6
4.23 4.93 3.05 0.87 6.56 10.8 1.05 4.01 5.74 25.7 0.30 4.01 2.37 1.02
0.0437 0.0101 0.0346 0.4222 0.0006 B0.0001 0.3998 0.0493 0.0049 B0.0001 0.7446 0.0107 0.0382 0.4163
Tillage (T) P level (P) Sampling time (S) TP TS P S TP S
1 2 3 2 3 6 6
9.83 3.23 23.2 0.11 2.85 2.31 0.10
0.0026 0.0459 B0.0001 0.9002 0.0440 0.0438 0.9959
1 2 3 2 3 6 6
4.11 3.66 28.3 0.01 4.42 2.52 0.05
0.0467 0.0310 B0.0001 0.9915 0.0058 0.0295 0.9996
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CANADIAN JOURNAL OF SOIL SCIENCE
SMB-P Tillage effects on SMB-P were larger in the top 0- to 10cm than in the 0- to 20-cm soil layer (Figs. 3 and 4). In the 0- to 10-cm soil layer, SMB-P was larger under conservation than conventional tillage in June and October; in the 10- to 20-cm layer, SMB-P was also larger under conservation tillage in July (Fig. 3). In general, SMB-P had a quadratic response to P fertilizer application rates in the 0- to 10-cm soil layer, increasing from P0 to P40, but decreasing thereafter (Fig. 4). There was a time lag in SMB-P response to P fertilization in the 10- to 20-cm soil layer, where fertilization effects were present only in July and October. The largest effects of fertilization on SMB-P were seen with P40, which increased SMB-P, and the influence of P40 was large in both soil layers. SMB-P increased from May to June, remained high from June to July, and decreased in October in the 0- to 10-cm and 10- to 20-cm soil layers (Figs. 3 and 4). SMB-C:P Ratio The mean SMB-C:P ratio was smaller in the top 0- to 10-cm than that in the 10- to 20-cm layer (Figs. 5 and 6). The SMB-C:P ratio varied during the season and was smaller in June and July than in May, in both soil layers (Figs. 5 and 6). In the topsoil layer, the depressing effect of conservation tillage on SMB-C:P ratio was significant
in June and October, whereas it was significant in July for the 10- to 20-cm soil layer (Fig. 5). In the top 0- to 10-cm soil layer, P40 reduced the SMB-C:P ratio as compared with P0 at the first two sampling times, i.e., May and June, while in the 10- to 20-cm soil layer this effect occurred in July and October (Fig. 6). The P80 rate did not decrease the SMB-C:P ratio as compared with P0 at any sampling times and any soil depths. The SMB-C:P ratio sharply decreased in both soil layers from May to June, remained low in July, and increased slightly in October (Figs. 5 and 6). Soybean Grain Yield Similar soybean grain yields were measured under conventional and conservation tillage systems. Soybean grain yield was greater at P80 than at any other P rates, but P40 did not increase yield (Table 4). DISCUSSION It appears that the SMB-P pool is plastic and can be increased by P fertilizer application, at least to some extent. In our case, the application of 40 kg P2O5 ha1 for 10 yr increased the SMB-P pool by 14.6% as compared with the unfertilized control, raising it to 10.6 kg P ha1 in the whole rooting depth (020 cm), which is equivalent to 24.4 kg P2O5 ha1. The SMB-P 7
7
0-10cm
0-10cm
6
6
5
5 4
4
Conventional Conservational
2 1 7
10-20cm
6 5
SMB-P (µg g-1)
SMB-P (µg g-1)
3
3 2 7
P0 P40 P80
10-20cm
P0 P40 P80
6 5
4
4 3
3
2
Conventional Conservational
1
2 May
June July Sampling time
Oct
Fig. 3. Effects of tillage system on SMB-P in the 0- to 10-cm and 10- to 20-cm soil layers at different sampling times. All values represent means9SEM (n 12).
May
June July Sampling time
Oct
Fig. 4. Effects of P fertilizer rates on SMB-P in the 0- to 10-cm and 10- to 20-cm soil layers at different sampling times. All values represent means9SEM (n 8).
LIU ET AL. * TILLAGE, P FERTILIZATION AND SOIL MICROBIAL BIOMASS 80 0-10cm
27
0-10cm
70
70
60
60 50
50
40
20 10-20cm 70
SMB-C:P ratio
SMB-C:P ratio
40 Conventional Conservational
30
P0 P40 P80
30 10-20cm
70 60
60 50
50
40
40
30
P0 P40 P80
Conventional Conservational
20
30 May
June July Sampling time
Oct
May
June July Sampling time
Oct
Fig. 5. Effects of tillage system on SMB-C:P ratio in the 0- to 10-cm and 10- to 20-cm soil layers at different sampling times. All values represent means9SEM (n 12).
Fig. 6. Effects of P fertilizer rates on SMB-C:P ratio in the 0- to 10-cm and 10- to 20-cm soil layers at different sampling times. All values represent means9SEM (n 8).
pool sequesters a biologically significant amount of P, but in our study, SMB-P was unrelated to soybean yield. Observation of a larger effect of P fertilizer rate on SMB-P than SMB-C leading to lower topsoil SMB-C:P ratio, even at pre-seeding in the absence of plants, suggests that 10 yr of different P fertilization regimes modified the soil microbial community, increasing the relative abundance of bacteria, which generally have a larger P requirement than fungi (Cleveland and Liptzin 2007). Although soil microbial biomass is usually limited by C and N (Joergensen and Scheu 1999), greater P availability may favour some microbial groups and change the structure of a community (Grover 2004). In this experiment, 40 kg P2O5 per hectare in a soil marginally low in P increased soil microbial components, especially SMB-P. While increase in SMB-C can be attributed to the enhancing effect of fertilization on soil organic matter level, SMB-P enrichment is attributed to changes in the structure of the soil microbial community. The increase in SMB-P with P40 that was observed in our study concurs with previous findings showing increased SMB-P with the application of rock phosphate in an acidic forest soil (He et al. 1997). But in our study, not only did the amount of soil available P influence soil microbial biomasses, but also whether soil nutrients were balanced or not was a determinant factor. SMB-C
and SMB-P were increased at the intermediate P fertilization level, consisting of yearly application of 40 kg P2O5 ha1, and the recommended N fertilization rate of 160 kg ha1 applied to corn crops only, in alternate years. Absence of, or excessive, P fertilization was applied with the N rate kept constant, impairing the soil macronutrient balance. Consequently, soil conditions were less conducive to microbial growth with the P0 and P80 treatments. Our results agree with those of Lukito et al. (1998), who found that SMB-C and SMB-P increased with increased P application up to rates beyond which soil microbial biomasses were unchanged
Table 4. Effects of tillage practice and P fertilization on soybean grain yield in 2001 Factor Tillage management P level (kg P2O5 ha 1)
Level
Soybean grain yield (kg ha1)
Conventional Conservational 0 40 80
1077951.5a 1124958.1a 1033961.9b 1040923.3b 1229980.7a
a, b All values represent means9SEM, and different letters following the mean values in the same column within the same factor indicate significant difference at PB0.05.
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CANADIAN JOURNAL OF SOIL SCIENCE
or reduced. Kouno et al. (1999), using a different approach, also confirmed the importance of nutrient balance for optimal soil microbial growth. Increasing rates of N fertilizer up to 200 mg N kg 1 soil increased SMB-C, but higher rates decreased SMB-C; SMB-P, in turn, increased up to 400 mg N kg1, where a plateau was reached. Our experiment is very different from that of Kouno et al. (1999), which was a laboratory incubation experiment, and used variable rates of N rather than P fertilizer, but both studies support the validity of the nutrient balance principles, and conclude that microbial biomass is negatively affected by insufficient or excessive amounts of water-soluble N or P. The importance of nutrient balance as a determinant of soil microbial biomass in agricultural systems was reported (Goyal et al. 1992; Belay et al. 2002). These reports, in contrast to that of Lukito et al. (1998), indicate that the impact of a balanced fertilization on soil microbial biomass was probably indirect, and occurred through better plant production and rhizodeposition under balanced nutrient availability. Our results also suggest that at least part of the effect of P fertilization rate on soil microorganisms was mediated by plant contribution of C to microorganisms. Too high soil P fertility is known to reduce root exudation (Graham et al. 1981) and plant C allocation to arbuscular mycorrhizal (AMF) networks (Smith and Read 1997). These are two important C sources for soil microorganisms. Plants commonly lose 10 to 30% of their photosynthate-C into the soil through root excretion and turnover (Bowen and Rovira 1999), and AMF materials were shown to represent 20% of the dry biomass of mycorrhizal soybean roots (Bethlenfalvay et al. 1982). The inhibition of AMF at too low and too high soil P fertility is well documented (Bethlenfalvay et al. 1982; Liu et al. 2000; Azcon et al. 2003). Plant stimulation of soil microbial biomasses through higher C export below ground is supported by the increase in soil organic C observed in soils receiving the P40 and P80 treatments, compared with the unfertilized control. The organic C and total N levels in soils fertilized with P40 were similar to those measured in soils fertilized with P80, where crop yield was the largest. The increase in SMB-P and SMB-C:P due to the P40 treatment occurred earlier in the top 0- to 10-cm and later in the 10- to 20-cm soil layer, suggesting the involvement of growing roots in this effects. Soil microbial biomasses were often larger in conservation than in conventional tillage management. This is consistent with the positive effect of conservation tillage on AMF proliferation in soil (Kabir 2005). Increased soil microbial biomass under conservation tillage is often attributed to an increase in soil organic C, which contains substrates for microbial growth (Franzluebbers and Arshad 1996; Kandeler et al. 1999; Jastrow et al. 2007). However, the soil organic C level was not significantly affected by tillage in this experiment. It may be that the lack of tillage disturbance made
the soil environment under the conservation tillage system a more favorable habitat for soil microbes, but this remains to be determined. We found a decrease in SMB-C:P during times of vigorous plant growth, in June and July, suggesting a modification of the soil microbial community structure during the season, through preferential stimulation of bacteria by the developing crop. This is surprising as we know that the growth of AMF, which are obligate biotrophs, is synchronized with crop development in agricultural fields (Kabir et al. 1998), and that these fungi are very abundant in cultivated soil where they account for about 25% of the soil microbial biomass (Olsson et al. 1999). AMF may differ from other fungal taxonomic groups as they are specialized in P uptake and transport. They are known to accumulate and transport P in specialized vacuoles (Uetake et al. 2002) and, therefore, their C:P ratio may be lower than other fungi. The importance of AMF biomass as a pool of P for crops needs to be clarified. CONCLUSION The soil microbial P pool was equivalent to 24.4 kg P2O5 ha 1 in the Dark Grey Gleysol under study. Conservation tillage increased this pool, as did a balanced fertilizer regime, but the absence of P or excessive P fertilization reduced the SMB-P pool. P fertilization apparently had a direct effect on SMB-P when a balanced P rate was used, but the negative effect of a high P rate seemed largely mediated by modification of plant C partitioning to the soil. Ten years of balanced fertilization increased soil organic C level and the higher P rate increased crop productivity. SMB-P was higher in period of active plant growth, but was unrelated to yield. The value of managing the SMB-P pool would be clearer if the mechanisms by which crops access SMB-P were better understood. ACKNOWLEDGEMENT Authors gratefully acknowledge the thorough revision and numerous invaluable comments of the associate editor and the reviewers. Acton, D. F. and Gregorich, L. J. 1995. The health of our soils: toward sustainable agriculture in Canada. Agriculture AgriFood Canada, CDR Unit, Ottawa, ON. 138 pp. Ajwa, H. A., Dell, C. J. and Rice, C. W. 1999. Changes in enzyme activities and microbial biomass of tallgrass prairie soil as related to burning and nitrogen fertilization. Soil Biol. Biochem. 31: 769777. Angers, D., Bissonnette, N., Le´ge`re, A. and Samson, N. 1993. Microbial and biochemical changes induced by rotation and tillage in soil under barley production. Can. J. Soil Sci. 73: 3950. Aslam, T., Choudhary, M. A. and Saggar, S. 1999. Tillage impacts on soil microbia biomass C, N and P, earthworms and agronomy after two years of cropping following permanent pasture in New Zealand. Soil Tillage Res. 51: 103111.
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