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Earthworm population dynamics as influenced by the quality of exogenous organic matter
ARTICLE IN PRESS Pedobiologia 52 (2008) 139—150
www.elsevier.de/pedobi
Earthworm population dynamics as influenced by the quality of exogenous organic matter Ben L.M. Leroya,, Olaf Schmidtb, Annemie Van den Bosschea, Dirk Reheulc, Maurice Moensd,e a
Department of Soil Management and Soil Care, Ghent University, Coupure Links 653, 9000 Gent, Belgium School of Biology and Environmental Science, Agriculture and Food Science Centre, University College Dublin, Belfield, Dublin 4, Ireland c Department of Plant Production, Ghent University, Coupure Links 653, 9000 Gent, Belgium d Institute for Agricultural and Fisheries Research, Burg. Van Gansberghelaan 96, 9820 Merelbeke, Belgium e Department of Crop Protection, Ghent University, Coupure Links 653, 9000 Gent, Belgium b
Received 28 January 2008; received in revised form 2 July 2008; accepted 3 July 2008
KEYWORDS Compost; Lumbricidae; Organic fertilizers; Mustard extraction method; Hand sorting; Slurry
Summary We conducted a replicated field plot experiment to investigate the influence of five exogenous organic materials (farmyard manure, cattle slurry and three types of compost), as well as mineral fertilization and two unfertilized control treatments (with and without a crop) on the number, biomass and species composition of earthworms (Lumbricidae) in an arable soil. A crucial feature of the experimental design was that the same mass of exogenous organic C was applied in each of the five organic treatments (1500–4000 kg C ha1), making it possible to interpret observed effects in terms of the quality of the organic amendments. Earthworms were sampled twice per year using a combination of mustard extraction and handsorting. Two and a half years after the first of four organic matter applications, the farmyard manure and cattle slurry treatments had the largest earthworm abundance of about 800–900 individuals m2 with 120–140 g biomass m2. The unamended controls had the smallest earthworm number (about 150 individuals m2), while the three compost treatments had intermediate values (400–500 individuals m2). Since the mass of exogenous organic C applied was the same in the five organic treatments and since final soil organic C contents under these treatments were statistically similar (p40.05), we hypothesize that the observed large differences in earthworm abundance between the farmyard manure and cattle slurry treatments on the one hand and the three compost treatments on the other hand were caused by differences in the chemical properties, and hence nutritional value for earthworms, of the applied amendments. We propose that manure and slurry provided
Corresponding author. Tel.: +32 9 264 60 57; fax: +32 9 264 62 47.
E-mail address: [email protected] (B.L.M. Leroy). 0031-4056/$ - see front matter & 2008 Elsevier GmbH. All rights reserved. doi:10.1016/j.pedobi.2008.07.001
ARTICLE IN PRESS 140
B.L.M. Leroy et al. earthworms with larger amounts of available C sources for direct consumption (including polysaccharides and proteinaceous materials) than the composted organic materials which were more humified and stabilized as a result of extended microbial aerobic degradation. & 2008 Elsevier GmbH. All rights reserved.
Introduction It is well known that earthworms play a major role in soil ecosystem functioning, most notably by influencing nutrient cycling processes in terrestrial ecosystems (Lee, 1985; Blair et al., 1995; Edwards and Bohlen, 1996). Since there is a resurgence in interest in using organic fertilizers rather than inorganic fertilizers to supply nutrients for crop production, management of earthworm populations is becoming more important for sustaining soil productivity and fertility in agro-ecosystems (Whalen et al., 1998). It is widely believed that all organic fertilizers support larger earthworm populations by providing a nutrient-rich food resource (Bohlen et al., 1995; Whalen et al., 1998). Inorganic fertilizers may also contribute indirectly to an increase in earthworm abundance by increasing the quantity of crop residues returned to the soil (Edwards et al., 1995). The influence of conventional fertilizers, such as farmyard manure and slurry, on the dynamics of earthworm populations in agro-ecosystems is well documented (Lee, 1985; Edwards and Bohlen, 1996). A number of studies have examined the long-term impact of organic and inorganic fertilizer applications on earthworm populations. For example, Tiwari (1993) found that earthworm biomass in ploughed fallow plots of a former pineapple orchard in India was three times greater in plots receiving manure than in inorganically fertilized plots, while a combination of manure and inorganic fertilizer supported the greatest earthworm biomass. On a continuously cropped, North American corn agro-ecosystem receiving fertilizers for 6 years, Whalen et al. (1998) recorded a significantly greater earthworm number and biomass in manureamended plots compared to inorganic fertilizertreated plots. In contrast, there has been little research comparing the effects of various kinds or qualities of organic amendments (e.g., manure, slurry, biosolids, composts) on earthworm populations in agro-ecosystems (Bu ¨nemann et al., 2006). Yet, it is known that different organic materials, including uncomposted and composted green wastes and manures, affect soil-dwelling invertebrate pests such as nematodes differently (Litterick et al.,
2004). We conducted a field plot experiment to assess if eight different fertilizer regimes, of which five were organic amendments used in Belgian agriculture, would cause changes in the earthworm abundance and the species composition. Preliminary results from that comparative study suggested that farmyard manure, cattle slurry and three different types of compost all caused similarly large increases in earthworm numbers 12 months after initial application of these amendments (Leroy et al., 2007). Here, we report longer term results from the same field experiment and we focus on the novel question of how organic fertilizers with a different composition, and hence quality, can influence earthworm abundance differently. We could address this question because the same mass of organic C was applied with each organic material.
Materials and methods Experimental design The experimental field, located in Melle (experimental site of Ghent University, Belgium, 501590 N, 031490 E, 11 m above sea level) is a sandy loam soil with the following granulometric composition: 11.7% 0–2 mm, 52.0% 2–50 mm and 36.3% 450 mm. Prior to the experiment, the field was cropped with monoculture maize for 8 years with mineral fertilization only. Table 1 summarizes the available meteorological data, covering a large part of the experimental period at Melle. The field experiment, started in 2005, was a randomized complete block design with four replicates comparing eight treatments (Figure 1): FYM, cattle slurry+crop residues (CSL), vegetable, fruit and garden waste compost (VFG), two types of farm compost (CMC1 and CMC2), mineral fertilizer (MIN N), and two treatments without fertilization (one with a crop (NF+) and one without (NF), to assess the impact of the presence or absence of a crop). The two types of farm compost differed in the composition of the starting materials. CMC1 was composed of mostly woody, C-rich materials resulting in a final C/N ratio of ca. 20–40;
ARTICLE IN PRESS Earthworms and quality of exogenous organic matter Table 1.
141
Total precipitation (mm) and average monthly air temperature (1C) at Melle Month 1
Total 2
3
4
5
6
7
8
9
10
11
12
24.0 70.3 55.6 46.0
68.2 52.4 0.0 50.0
59.3 112.6 108.2 59.0
39.2 45.7 102.8 65.0
118.6 21.6 177.7 72.0
98.4 292.7 84.7 74.0
43.8 19.2 84.3 72.0
40.5 106.3 63.0 72.0
82.1 79.4 – 64.0
50.6 107.6 – 59.0
temperature (1C) 5.2 2.8 6.8 1.9 2.8 5.1 7.4 6.4 8.2 2.4 3.1 5.2
9.7 9.1 12.8 8.4
12.2 14.2 14.3 12.1
18.2 16.3 17.5 15.1
18.2 21.8 17.3 16.8
16.3 16.9 16.9 16.7
16.0 18.0 14.1 14.4
14.1 13.8 10.2 10.3
6.8 9.6 – 6.2
3.9 5.8 – 3.2
Precipitation (mm) 2005 36.7 63.4 2006 16.2 96.6 2007 78.6 108.4 42.0 Norma 51.0 Average air 2005 2006 2007 Norma a
724.8 1020.6 –
Norm: average over the last 30 years.
Figure 1. Layout of the field experiment.
CMC2 was made from green, N-rich materials and had a final C/N ratio of 10–20. Experimental plots (8 m 6 m) were located next to each other within blocks, thus with no distance between the differently treated plots. Furthermore, there were no above or below ground barriers between the plots, since this would have interfered with tillage, fertilization and crop protection operations. Organic amendments and fertilizers were applied on four occasions (Table 2). The first application took place on 21 April 2005 and fodder beet was sown the day after. The second fertilization was on
6 October 2005 followed by the sowing of winter wheat the next day. One month after the harvest of the winter wheat, the organic amendments and fertilizers were applied for the third time (7 September 2006) and a catch crop (Phacelia) was sown. On 2 May 2007, the fourth application took place and red cabbage was planted 3 weeks later. Amounts of organic amendments were adjusted to supply all plots, receiving organic amendments, with equal amounts of organic C. This amount of organic C was 4000 kg C ha1 in application one and two, 1500 kg C ha1 in application three and 2000 kg C ha1 in application four. In all cases, the
ARTICLE IN PRESS 142
B.L.M. Leroy et al.
Table 2. The amounts of organic fertilizers and their C and N content, and extra mineral N applied on 21 April 2005, 6 October 2005, 7 September 2006 and 2 May 2007 (abbreviations of treatments, see Fig. 1) C content (g kg1 fresh matter)
Treatment (fertilizer)
21/04/05 (4000 kg C ha1) FYM 62.2 VFG 179.4 CMC1 71.4 CMC2 59.7 CSL 26.7a +Crop residues 378.0 (straw) MIN N –
N content (g kg1 fresh matter)
Applied amount (kg ha1)
Extra mineral N to be added (kg ha1)
4.7 15.2 1.7 2.8 3.9a 5.5
64,329 22,303 56,007 67,058 77,382b 4704
105 114 165 165
–
–
165
1
06/10/05 (4000 kg C ha ) FYM VFG CMC1 CMC2 CSL +Crop residues (beet leaves) MIN N
–
23/03/06
26/04/06
106.8 175.7 71.8 77.9 20.4a 49.1
6.8 14.5 3.1 7.2 2.8a 3.8
37,453 22,770 55,718 51,348 74,698b 53,636
81 88 91 89
97 98 99 97
74
94
–
–
–
91
98
6.7 15.5 4.1 3.4 3.8a –
14,398 8188 19,330 23,757 56,754b –
66 67 86 86
6.1 9.1 3.6 6.0 3.2a
19,125 14,329 10,417 21,986 71,287b –
1
07/09/06 (1500 kg C ha ) FYM 104.2 VFG 183.2 CMC1 77.6 CMC2 63.1 CSL 26.4a MIN N –
– 86
1
02/05/07 (2000 kg C ha ) FYM 104.6 VFG 139.6 CMC1 192.0 CMC2 91.0 CSL 28.1a MIN N – a b
–
106 103 170 162 – 162
g l1. l ha1.
organic amendments were incorporated to a depth of 20 cm using a rotary tiller. To correct for differences in the plant available N content of the different organic amendments, extra mineral N (ammonium nitrate 27%) was applied on organically amended plots where needed to achieve equal levels of plant available N in all treatments and thus to support equal crop growth. For the fodder beet, the catch crop and the red cabbage, the mineral N was applied together with the organic amendments, while for the winter wheat, the extra mineral N was splitapplied on 23 March 2006 and 26 April 2006. For the calculation of the amount of extra mineral N to be added, the mineralization rates of the soil and of
the organic fertilizers, both determined by laboratory incubation as described by De Neve and Hofman (1996), were taken into account together + with the amount of mineral N (NO 3 and NH4) present in the soil at the time of fertilization. On the CSL plots, part of the organic C was applied as crop residues (except before the catch crop and the red cabbage), since applications of 4000 kg C ha1 as slurry, would have provided excessive amounts of mineral N. The first time (21 April 2005), the crop residues used were straw, while the second time (6 October 2005), it was fodder beet leaves. At each fertilization, except before the Phacelia, on the minerally fertilized plots and on the plots receiving less than 300 kg ha1 K2O and 100 kg ha1
ARTICLE IN PRESS
a a a a a a a a 36.771.5 34.771.3 35.071.9 35.871.9 36.071.2 35.070.7 35.173.0 35.870.4 146.278.4 abc 158.6714.6 bc 141.479.3 abc 150.1712.5 c 127.175.6 a 127.776.0 a 131.478.0 ac 132.873.5 ac Different letters indicate a significant difference per parameter according to Tukey’s HSD test (po0.05).
6.1070.10 6.2270.07 5.9870.05 6.0470.10 6.0870.06 5.8270.18 5.8670.17 5.9770.05 15/10/07 FYM VFG CMC1 CMC2 CSL MIN N NF+ NF
ab b abc ab ab c ac abc
1.1670.07 1.1970.03 1.1170.05 1.1370.03 1.0970.06 0.9670.04 0.9870.05 0.9870.04
a a a a ac b bc bc
0.10370.004 0.10970.005 0.09570.007 0.10370.002 0.09570.002 0.08370.004 0.08570.004 0.08470.003
ab b a ab a c c c
28.673.9 26.072.5 23.271.8 25.473.1 25.072.0 25.072.5 24.672.2 24.770.2
a a a a a a a a
2.2270.45 2.1670.57 1.8970.64 2.0470.64 2.1470.39 1.7970.45 1.6770.24 1.5070.18
a a a a a a a a
24.375.6 a 18.571.8 a 19.5710.3 a 20.275.5 a 24.672.1 a 18.978.5 a 11.672.3 a 16.172.4 a
32.7 122.4 19.7 1.60
Mg (mg per 100 g dry soil) Ca (mg per 100 g dry soil) K (mg per 100 g dry soil) Na (mg per 100 g dry soil) P (mg per 100 g dry soil)
20.8 0.086 1.01 5.90
Earthworms were sampled five times during the experimental period using a mustard powder suspension, following Gunn (1992) and Lawrence and Bowers (2002), and subsequent handsorting. Due to heavy rains (Table 1) and resultant unfavourable soil conditions in the weeks before the start of the experiment, the first sampling in 2005 could only be accomplished 6 weeks after the start of the experiment (31/05/05 vs. 21/04/05) and it was restricted to the unfertilized plots (NF+ and NF). The results of this sampling were considered representative of the initial situation because the field had a uniform management history and the eight plots assessed were randomized in the field (Figure 1). The other samplings were carried out in spring and autumn of 2006 and 2007, since in
26/10/04
Earthworm sampling and analysis
Total N (%)
Soil chemical properties of the experimental field (0–20 cm) were assessed at the start of the experiment and after two and a half years of different fertilizer regimes (Table 3). The pH was measured potentiometrically in a 1:2.5 soil:KCl extract. In soils where the pH-KCl is lower than 6.5, the amount of organic C is equal to the amount of total C as no free carbonates are normally present. In these soil samples, total C, and hence organic C, and total N were measured by dry combustion at 850 1C using an elemental analyser (Vario MAX CNS, Elementar). K, Na, Ca, Mg and available P were assessed by extraction of the soil with ammoniumlactate-acetic acid (extraction ratio 1:20) in dark polyethylene bottles, shaken for 4 h and the suspension was filtered in dark polyethylene bottles that were stored cool (4 1C) until analysis. The P concentration was measured colorimetrically by the Mo blue method (Scheel, 1936) at 700 nm with a photometer (Universal Photometer, Vitatron). The concentration of K and Na was measured with a flame photometer (Elex 6361, Eppendorf) in an airpropane flame at 768 and 589 nm, respectively, while for the Ca concentration, an air-acetylene flame was used at 623 nm. The concentration of Mg was determined by atomic absorption spectrometry (SpectrAA Atomic Absorption Spectrometer, Varian) at 285 nm.
C (%)
Soil chemistry
pH-KCl
P2O5 out of the organic amendments, an extra amount of K2O (300 kg ha1) and P2O5 (100 kg ha1) was applied to achieve equal minimum levels of plant available K2O and P2O5.
143 Table 3. Soil chemical properties of the experimental field (0–20 cm), before the start of the experiment (26 October 2004) and at the end of the experimental period (15 October 2007) (abbreviations of treatments, see Fig. 1)
Earthworms and quality of exogenous organic matter
ARTICLE IN PRESS 144 temperate regions earthworms are most active during spring and autumn when temperatures are moderate and soil moisture levels adequate (Bohlen et al., 1995): sampling two on 16–17 May 2006, sampling three on 2–3 October 2006, sampling four on 4–5 June 2007 and sampling five on 24–25 September 2007. The day before each sampling, 6 g of mustard powder was mixed with 15 ml of water to produce a paste; immediately before sampling, this mustard paste was mixed with 0.8 l of water. A metal frame (20 cm 20 cm) was placed on the soil and all surface plant litter within the frame removed. The mustard solution was then poured evenly across the frame. After 10 min, while collecting the emerging earthworms, this treatment was repeated. After another 10 min, the soil in the same quadrant was excavated to a depth of 20 cm, using another metal frame, spread on a white plastic sheet and handsorted to recover remaining earthworms. Two samples were taken per replicate plot. All earthworms were taken to the lab and washed. They were transferred into a 4% formalin solution and the total weight of the preserved earthworms (with gut content) per sample was recorded. Finally, the earthworms were stored until species identification of the adults.
Statistical analysis Earthworm data of the four dates at which all plots were sampled (May 2006–September 2007) were analysed by analysis of variance (ANOVA) for repeated measures using StatView v. 5.0 (SAS Institute Inc.). Sources of variability were partitioned into between-subjects factors (blocks; treatments) and within-subjects factors (sampling date). Treatment means were compared by Tukey’s HSD post-hoc tests. The two subsamples from each plot per sampling date were pooled for ANOVA because nesting of factors could not be combined with the repeated measures procedure. Final soil chemical data were analysed by one-way ANOVA and treatment means compared with Tukey’s HSD test.
Results Soil chemistry The four applications of all organic amendments, except CSL, significantly (po0.05) increased the soil organic C content compared to unamended treatments (Table 3). The organically amended
B.L.M. Leroy et al. plots had a similar final organic C content. All organically amended plots had a significantly higher total N content than the unamended plots, but significant differences (po0.05) could also be observed between the CMC1 and the CSL plots on the one hand and the VFG plots with the highest total N content on the other. Both the organic C and the total N content were similar on the unamended plots (MIN N, NF+ and NF). The other soil chemical properties showed small differences or trends among the treatments. Significant differences among treatments (po0.05) could only be observed for the pH-KCl and the Ca content.
Earthworms In all organically amended plots, the earthworm number and biomass was higher than in the unamended plots for the spring 2006 sampling, being on average at least twice as high compared to the initial situation (Figure 2A). However, no clear differences in the earthworm abundance among the five organic amendments could be observed. The unfertilized plots (NF+ and NF) had the lowest earthworm number and biomass and the mineral fertilization (MIN N) was intermediate. From the sampling in autumn 2006 onwards, the FYM and CSL plots exceeded on all sampling occasions, the compost and unamended plots in earthworm number, reaching densities of 800 individuals m2 (Figure 2B–D). Earthworm numbers were similar in the three compost treatments (VFG, CMC1 and CMC2), with an average of about 400–500 individuals m2. The differences in earthworm number among the three unamended plots (MIN N, NF+ and NF), which could be observed in spring 2006 (Figure 2A), disappeared at the following sampling dates, except for the sampling in spring 2007 (Figure 2C). The relatively high earthworm number in the unamended plots at the last sampling (Figure 2D) was surprising, with values reaching 300–350 individuals m2, twice as high as the initial situation. Except for these higher earthworm numbers, Figure 3A shows that since the sampling in autumn 2006, the earthworm numbers remained relatively constant in all plots. The eight treatments can be clearly divided in three groups: FYM and CSL plots, compost plots and unamended plots. For the earthworm biomass, it took until spring 2007 to cluster the fertilizer regimes in a similar fashion as for the earthworm number (Figure 3B). The differences were, however, less pronounced and the earthworm biomass showed a higher
ARTICLE IN PRESS Earthworms and quality of exogenous organic matter
600 500 400 300 200 100 0
YM
F
FG
V
C1
CM
C2
SL
NN
I C M CM Treatment
F+
N
600 500 400 300 200 100 0
F
V
CM
SL
I C M CM Treatment
NN
N
F+
300
0
M
G
VF
C1
CM
+ N C2 CSL IN NF M CM Treatment
-
NF
A. rosea A. icterica A. chlorotica A. caliginosa L castaneus L. rubellus L. terrestris Juvenile Biomass 150
1000 900
NF
Number of earthworms
700
C2
400
100
m-2
Number of earthworms m-2
800
C1
500
FY
140 130 120 110 100 90 80 70 60 50 40 30 20 10 0
FG
600
NF
900
YM
700
200
A. rosea A. icterica A. chlorotica A. caliginosa L castaneus L. rubellus L. terrestris Juvenile Biomass 150
1000
800
Earthworm biomass (g m-2)
700
140 130 120 110 100 90 80 70 60 50 40 30 20 10 0
900
800 700 600 500 400 300 200 100 0
M
FY
G
VF
C1
CM
+ N C2 CSL IN NF M CM Treatment
-
140 130 120 110 100 90 80 70 60 50 40 30 20 10 0
Earthworm biomass (g m-2)
800
A. rosea A. icterica A. chlorotica A. caliginosa L castaneus L. rubellus L. terrestris Juvenile 150 Biomass
1000
Earthworm biomass (g m-2)
Number of earthworms m-2
900
150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0
Number of earthworms m-2
1000
Earthworm biomass (g m-2)
A. rosea A. icterica A. chlorotica A. caliginosa L castaneus L. rubellus L. terrestris Juvenile Biomass
145
NF
Figure 2. Earthworm number, subdivided in adult species and juveniles, and total biomass (treatment means7S.D.), and the initial situation on 31 May 2005 (full line ¼ number; dotted line ¼ biomass) (abbreviations of treatments, see Figure 1): (A) sampling date: 16–17 May 2006; (B) sampling date: 2–3 October 2006; (C) sampling date: 4–5 June 2007; (D) sampling date: 24–25 September 2007.
variability compared to the earthworm number. In autumn 2006 (Figure 2B), the earthworm biomass was highest in the FYM plots, while in spring 2007, the FYM and CSL plots had a higher earthworm biomass than the other treatments, as was the case for the earthworm number (Figure 2C). In the MIN N
plots, the biomass was comparable to the compost plots, while the two unfertilized plots had the lowest biomass. In autumn 2007, the highest biomass was again recorded on the FYM and CSL plots, while a rise in the earthworm biomass (as was seen for the earthworm number) in the unfertilized
ARTICLE IN PRESS 146
B.L.M. Leroy et al.
1000 900
FYM VFG CMC1 CMC2 CSL MIN N NF+ NF-
Earthworm number m-2
800 700 600 500 400 300 200 100 0
5 5 6 6 7 6 7 /06 /06 /05 /07 4/0 /30/0 /5/05 1/10 /16/0 /24/0 /29/0 /4/06 0/10 2/15 /20/0 /28/0 /3/07 /8/07 1/14 9 6 3 1 1 8 1 4 5 2 1 7 9 1
4/2
Date 160
Earthworm biomass (g m-2)
140 120
FYM VFG CMC1 CMC2 CSL MIN N NF+ NF-
100 80 60 40 20 0 5 6 5 6 6 7 7 /06 /06 /05 /07 4/0 /30/0 /5/05 1/10 /16/0 /24/0 /29/0 /4/06 0/10 2/15 /20/0 /28/0 /3/07 /8/07 1/14 2 / 6 1 9 3 4 1 5 8 1 2 7 1 4 1 9 Date
Figure 3. Dynamics of the earthworm abundance since the start of the experiment (abbreviations of treatments, see Figure 1): (A) abundance (number m2), (B) biomass (g m2).
plots was observed (Figure 2D), with values only slightly lower than in the compost plots. Repeated measures ANOVA of the combined data (four sampling dates in 2006 and 2007) confirmed a highly significant effect of fertilization treatment on earthworm number and biomass (Table 5). The small but significant block effect suggests that blocking in this field was successful, removing some field-related variability. Significant sampling date effects probably reflect systematically higher earthworm numbers and biomass across all treatments in autumn 2007, and the significant
date treatment interaction for numbers was caused by the rapid increase in autumn 2006 in FYM and CSL treatments (Table 4). Post-hoc treatment comparisons, across dates, confirmed that the FYM and CSL plots supported significantly higher earthworm abundances than the plots treated with composts and significantly higher earthworm biomass than the two farm composts (Table 5). In the samples taken in spring 2006, the amount of juveniles varied between 63% and 82% of the total earthworm number, while for the later
ARTICLE IN PRESS Earthworms and quality of exogenous organic matter Table 4. Summary results of repeated measures analysis of variance (ANOVA) of earthworm abundance and biomass Source of variation
F-ratioa
DF
Abundance
Biomass
Between-subjects Block (B) Treatment (T) Error
3 7 21
3.1* 21.3***
3.7* 22.0***
Within-subjects Sampling date (D) DB DT Error
3 9 21 63
21.5*** 0.8 ns 2.4**
6.9*** 1.0 ns 1.0 ns
a Significance levels: ***po0.001; **po0.01; *po0.05; ns: not significant (p40.05).
Table 5. Multiple treatment mean comparisons (Tukey–Kramer) of earthworm abundance and biomass (treatments sharing the same letter are not significantly different at po0.05) Treatment
Number
Biomass
FYM CSL VFG CMC1 CMC2 MIN N NF+ NF
a a
a a b b b b
c c c c
d d
b b
c c c
d d d d
e e e
samplings, 70–94% of the earthworms sampled were juveniles. The earthworm species found were Lumbricus terrestris as anecic species, Lumbricus castaneus and Lumbricus rubellus as epigeic species and Aporrectodea caliginosa, Allolobophora chlorotica, Allolobophora icterica and Allolobophora rosea as endogeic species. The relative distribution of these earthworm species among the sampling dates and the treatments did not show clear differences or trends, with L. rubellus, A. caliginosa and A. chlorotica being the most abundant species (Figure 2A–D).
Discussion and conclusions The soil organic C content had increased in all plots receiving organic amendments and was
147 significantly higher (except from CSL) than that in the unamended plots after the two and a half year experimental period (Table 3). These results agree with previous studies of similar or longer duration (3–6 years) comparing effects of fresh and composted organic matter treatments with that of mineral fertilizer controls on soil organic C contents (e.g., Canali et al., 2004; Senesi and Plaza, 2007). The slightly lower soil C content under CSL was probably due to the composition or quality of this organic amendment. Cattle slurry, as a fresh product, is composed of more labile organic C than the other organic materials which contain more stable organic C due to the incorporation of straw in farmyard manure, or the composting process above ground during several months for the composts. The increase in earthworm abundance in the plots receiving organic amendments can be linked with the higher organic C content of these plots. Addition of organic matter is believed to be one of the major management variables affecting earthworm abundance. By providing food sources and increasing the overall levels of soil organic matter, organic amendments usually increase earthworm abundance (Bohlen et al., 1995). This increase as a response to organic amendments can be particularly pronounced in disturbed habitats of low organic matter content (Lowe and Butt, 2002; Curry, 2004), as was true for this experimental field which had been under monoculture maize receiving only mineral fertilizer for 8 years. Our experiment provides convincing evidence that compost amendments, incorporated into the soil to 20 cm, can enhance earthworm abundance significantly. Few comparable data on compost effects on earthworms are available from other agro-ecosystems, but Thomson and Hoffmann (2007) recently reported increased earthworm abundance (exotic Lumbricidae) under plots surface-mulched with compost made of domestic prunings for two growing seasons compared to bare plots in an Australian vineyard. The organic matter that provides the food base for the earthworm community is vitally important in determining their distribution and abundance (Edwards and Bohlen, 1996). For instance, Hendrix et al. (1992) reported a significant correlation between earthworm abundance and soil organic carbon content over a range of sites in Georgia, USA. However, from the results of the present research, we can assume that other factors than solely the soil organic C content influence the earthworm abundance. Although the final organic C content of all organically amended plots was statistically similar, ranging from 1.09% in the CSL
ARTICLE IN PRESS 148 plots to 1.19% in the VFG plots (Table 3), clear and significant differences in earthworm abundance (number and biomass) were observed, with the highest abundances found in the FYM and CSL plots (Figure 3A and B). We hypothesize that this seemingly contradicting result of a disconnection between the soil organic carbon content on the one hand and the earthworm abundance on the other can be explained by the chemical properties of the organic amendments used. The chief biochemical process of composting is humification, which is defined as the prolonged stabilization of organic substances against biodegradation. Composts are thus chemically stabilized and biologically matured amendments (Senesi and Plaza, 2007). As a result of extended microbial degradation, composts have lower total and watersoluble organic C contents, higher relative contents of humic acid like substances and a higher degree of polycondensation and polymerization than their raw starting materials. Animal manures, by contrast, are rich in incompletely humified materials including polysaccharides and S- and N-containing groups, most notably proteinaceous materials (Senesi and Plaza, 2007). These chemical differences between organic amendments also translate into different chemical characteristics of the organic matter in soil treated with these materials. For example, a recent Canadian study reported significantly higher humification indices (ratio of organic C content in unhumified fraction to fulvic and humic acids) and higher amino sugar contents (especially glucosamine) in two soil aggregate size fractions (0.25–2.00 and 42.00 mm) of loam soil treated for 3 years with fresh paper mill sludge (40 t ha1 year1) compared to soil amended with the same but composted sludge (Bipfubusa et al., 2008). In the present experiment, four applications of relatively fresh farmyard manure or cattle slurry that had not been subjected to composting resulted in much larger earthworm abundance increases than in the plots amended with any of the three composts (Figure 3A and B). We hypothesize that the nutritional value for earthworms of both the cattle slurry and farmyard manure, and hence also of the soil organic matter in plots treated with these raw materials, were higher than that of the composts and the soil OM in the compost plots, in which the applied organic matter was more decomposed and stabilized, due to the aerobic composting for several weeks or months. Our interpretation of field population data concurs with current ecological theory that has revised the traditional understanding of the trophic ecology of earthworms (Curry and Schmidt, 2007). Experimental evidence suggests firstly that
B.L.M. Leroy et al. (endogeic) earthworms are limited by available C in soil, irrespective of the total C content of soil (Tiunov and Scheu, 2004) and secondly that earthworms do not depend on micro-organisms as a food source but consume organic matter directly (Salamon et al., 2006). Since the same amounts of organic C were added in our organic treatments, it is plausible that higher amounts of nutritionally available C in the uncomposted organic treatments supported larger earthworm populations. It is also likely that earthworm abundances were not related to microbial biomass which can sometimes be elevated in soils treated with composts. One example is the study by Canali et al. (2004) in which an orchard soil receiving poultry manure for 6 years had a smaller microbial biomass and lower microbially mineralizable C content than the same soil amended with composted manure. However, in the present experiment, soil microbial biomass was not significantly different among the five organic treatments (Leroy, 2008), suggesting that availability of organic matter, and not microbes, determined the size of lumbricid community. From the third earthworm sampling onwards (autumn 2006), earthworm abundance remained fairly stable, but biomass continued to increase in response to repeated organic matter application. This pattern is similar to Irish data (Schmidt and Curry, 2001), which suggested that earthworm populations stabilize only 2–3 years after regular organic matter inputs to cultivated soil commenced. It would, however, be necessary to continue monitoring earthworm abundance longer to establish whether the treatment differences would become more pronounced in the present experiment. Based on the sampling in spring 2006, preliminary results suggested that the mineral fertilization and the presence or absence of a crop also influenced the earthworm abundance, since differences (although not significant, p40.05) in earthworm biomass and number were recorded between the MIN N, NF+ and NF plots (Figure 2A, also Leroy et al., 2007). These differences seemed to disappear in the next samplings, except in spring 2007, when the earthworm number and biomass in the MIN N plots tended to be again slightly higher than in the unfertilized plots. This is possibly due to a better crop performance on the minerally fertilized plots compared to the NF+ plots (Leroy, 2008), and hence a higher root and crop biomass. This higher biomass results in a higher amount of above and below ground crop litter, which is an important source of organic matter on which earthworms feed (Curry, 2004). Generally, the effects of the application of moderate levels of mineral fertilizer on earthworm
ARTICLE IN PRESS Earthworms and quality of exogenous organic matter abundance are considered positive in the literature, reflecting increased litter quality and quantity (Bostro ¨m, 1988; Muldowney et al., 2003). Again, future sampling would be required to establish if this effect of mineral fertilization on earthworm abundance would persist in the long term.
Acknowledgements The authors wish to thank Franky Van Peteghem and Jean-Pierre Van Maerke for their much appreciated assistance with the field work, and Luc Deboosere, Olle Victoor, Mathieu Schatteman, Tina Coddens and Sophie Schepens for their practical assistance with the earthworm sampling and in the lab. The authors thank AgriVet for the use of its experimental fields.
References Bipfubusa, M., Angers, D.A., N’Dayegamiye, A., Antoun, H., 2008. Soil aggregation and biochemical properties following the application of fresh and composted organic amendments. Soil Sci. Soc. Am. J. 72, 160–166. Blair, J.M., Parmelee, R.W., Lavelle, P., 1995. Influences of earthworms on biogeochemistry. In: Hendrix, P.F. (Ed.), Earthworm Ecology and Biogeography in North America. Lewis, Boca Raton, FL, pp. 127–158. Bohlen, P.J., Edwards, W.M., Edwards, C.A., 1995. Earthworm community structure and diversity in experimental agricultural watersheds in Northeastern Ohio. Plant Soil 170, 233–239. Bostro ¨m, U., 1988. Ecology of earthworms in arable land: population dynamics and activity in four cropping systems. Report No. 34, Department of Ecology and Environmental Research, Swedish University of Agricultural Sciences, Uppsala, Sweden. Bu ¨nemann, E.K., Schwenke, G.D., Van Zwieten, L., 2006. Impact of agricultural inputs on soil organisms – a review. Aust. J. Soil Res. 44, 379–406. Canali, S., Trinchera, A., Intrigliolo, F., Pompili, L., Nisini, L., Mocali, S., Torrisi, B., 2004. Effect of long term addition of composts and poultry manure on soil quality of citrus orchards in Southern Italy. Biol. Fertil. Soils 40, 206–210. Curry, J.P., 2004. Factors affecting the abundance of earthworms in soils. In: Edwards, C.A. (Ed.), Earthworm Ecology. CRC Press, Boca Raton, pp. 91–113. Curry, J.P., Schmidt, O., 2007. The feeding ecology of earthworms – a review. Pedobiologia 50, 463–477. De Neve, S., Hofman, G., 1996. Modelling N mineralization of vegetable crop residues during laboratory incubations. Soil Biol. Biochem. 28, 1451–1457.
149 Edwards, C.A., Bohlen, P.J., 1996. Biology and Ecology of Earthworms. Chapman & Hall, London. Edwards, C.A., Bohlen, P.J., Linden, D.R., Subler, S., 1995. Earthworms in agroecosystems. In: Hendrix, P.F. (Ed.), Earthworm Ecology and Biogeography in North America. Lewis, Boca Raton, FL, pp. 185–213. Gunn, A., 1992. The use of mustard to estimate earthworm populations. Pedobiologia 36, 65–67. Hendrix, P.F., Mueller, P.R., Bruce, R.R., Langdale, G.W., Parmelee, R.W., 1992. Abundance and distribution of earthworms in relation to landscape factors on the Georgia Piedmont USA. Soil Biol. Biochem. 24, 1357–1361. Lawrence, A.P., Bowers, M.A., 2002. A test of the ‘‘hot’’ mustard extraction method of sampling earthworms. Soil Biol. Biochem. 34, 549–552. Lee, K.E., 1985. Earthworms: Their Ecology and Relationships with Soils and Land Use. Academic Press, Sydney. Leroy, B.L.M., 2008. Soil food web, C and N transformations and soil structure: interactions and feedback mechanisms as a function of the quality of exogenous organic matter. Ph.D. Thesis, Ghent University, Gent, Belgium (ISBN 978-90-5989-241-5). Leroy, B.L.M., Van den Bossche, A., De Neve, S., Reheul, D., Moens, M., 2007. The quality of exogenous organic matter: short term influence on earthworm abundance. Eur. J. Soil Biol. 43 (Suppl. 1), S196–S200. Litterick, A.M., Harrier, L., Wallace, P., Watson, C.A., Wood, M., 2004. The role of uncomposted materials, composts, manures, and compost extracts in reducing pest and disease incidence and severity in sustainable temperate agricultural and horticultural crop production – a review. Crit. Rev. Plant Sci. 23, 453–479. Lowe, C.N., Butt, K.R., 2002. Influence of organic matter on earthworm production and behavior: a laboratorybased approach with applications for soil restoration. Eur. J. Soil Biol. 38, 173–176. Muldowney, J., Curry, J.P., O’Keefe, J., Schmidt, O., 2003. Relationships between earthworm populations, grassland management and badger density in Co. Kilkenny, Ireland. Pedobiologia 47, 913–919. Salamon, J.A., Alphei, J., Ruf, A., Schaefer, M., Scheu, S., Schneider, K., Suhrig, A., Maraun, M., 2006. Transitory dynamic effects in the soil invertebrate community in a temperate deciduous forest: effects of resource quality. Soil Biol. Biochem. 38, 209–221. Scheel, K.C., 1936. Colorimetric determination of phosphoric acid in fertilizers with the Aulfrich photometer. Z. Anal. Chem. 105, 256–259. Schmidt, O., Curry, J.P., 2001. Population dynamics of earthworms (Lumbricidae) and their role in nitrogen turnover in wheat and wheat–clover cropping systems. Pedobiologia 45, 174–187. Senesi, N., Plaza, C., 2007. Role of humification processes in recycling organic wastes of various nature and sources as soil amendments. Clean-Soil Air Water 35, 26–41.
ARTICLE IN PRESS 150 Thomson, L.J., Hoffmann, A.A., 2007. Effects of ground cover (straw and compost) on the abundance of natural enemies and soil macro invertebrates in vineyards. Agric. For. Entomol. 9, 173–179. Tiunov, A.V., Scheu, S., 2004. Carbon availability controls the growth of detritivores (Lumbricidae) and their effect on nitrogen mineralization. Oecologia 138, 83–90.
B.L.M. Leroy et al. Tiwari, S.C., 1993. Effects of organic manure and NPK fertilization on earthworm activity in an Oxisol. Biol. Fertil. Soils 16, 293–295. Whalen, J.K., Parmelee, R.W., Edwards, C.A., 1998. Population dynamics of earthworm communities in corn agroecosystems receiving organic or inorganic fertilizer amendments. Biol. Fertil. Soils 27, 400–407.
www.elsevier.de/pedobi
Earthworm population dynamics as influenced by the quality of exogenous organic matter Ben L.M. Leroya,, Olaf Schmidtb, Annemie Van den Bosschea, Dirk Reheulc, Maurice Moensd,e a
Department of Soil Management and Soil Care, Ghent University, Coupure Links 653, 9000 Gent, Belgium School of Biology and Environmental Science, Agriculture and Food Science Centre, University College Dublin, Belfield, Dublin 4, Ireland c Department of Plant Production, Ghent University, Coupure Links 653, 9000 Gent, Belgium d Institute for Agricultural and Fisheries Research, Burg. Van Gansberghelaan 96, 9820 Merelbeke, Belgium e Department of Crop Protection, Ghent University, Coupure Links 653, 9000 Gent, Belgium b
Received 28 January 2008; received in revised form 2 July 2008; accepted 3 July 2008
KEYWORDS Compost; Lumbricidae; Organic fertilizers; Mustard extraction method; Hand sorting; Slurry
Summary We conducted a replicated field plot experiment to investigate the influence of five exogenous organic materials (farmyard manure, cattle slurry and three types of compost), as well as mineral fertilization and two unfertilized control treatments (with and without a crop) on the number, biomass and species composition of earthworms (Lumbricidae) in an arable soil. A crucial feature of the experimental design was that the same mass of exogenous organic C was applied in each of the five organic treatments (1500–4000 kg C ha1), making it possible to interpret observed effects in terms of the quality of the organic amendments. Earthworms were sampled twice per year using a combination of mustard extraction and handsorting. Two and a half years after the first of four organic matter applications, the farmyard manure and cattle slurry treatments had the largest earthworm abundance of about 800–900 individuals m2 with 120–140 g biomass m2. The unamended controls had the smallest earthworm number (about 150 individuals m2), while the three compost treatments had intermediate values (400–500 individuals m2). Since the mass of exogenous organic C applied was the same in the five organic treatments and since final soil organic C contents under these treatments were statistically similar (p40.05), we hypothesize that the observed large differences in earthworm abundance between the farmyard manure and cattle slurry treatments on the one hand and the three compost treatments on the other hand were caused by differences in the chemical properties, and hence nutritional value for earthworms, of the applied amendments. We propose that manure and slurry provided
Corresponding author. Tel.: +32 9 264 60 57; fax: +32 9 264 62 47.
E-mail address: [email protected] (B.L.M. Leroy). 0031-4056/$ - see front matter & 2008 Elsevier GmbH. All rights reserved. doi:10.1016/j.pedobi.2008.07.001
ARTICLE IN PRESS 140
B.L.M. Leroy et al. earthworms with larger amounts of available C sources for direct consumption (including polysaccharides and proteinaceous materials) than the composted organic materials which were more humified and stabilized as a result of extended microbial aerobic degradation. & 2008 Elsevier GmbH. All rights reserved.
Introduction It is well known that earthworms play a major role in soil ecosystem functioning, most notably by influencing nutrient cycling processes in terrestrial ecosystems (Lee, 1985; Blair et al., 1995; Edwards and Bohlen, 1996). Since there is a resurgence in interest in using organic fertilizers rather than inorganic fertilizers to supply nutrients for crop production, management of earthworm populations is becoming more important for sustaining soil productivity and fertility in agro-ecosystems (Whalen et al., 1998). It is widely believed that all organic fertilizers support larger earthworm populations by providing a nutrient-rich food resource (Bohlen et al., 1995; Whalen et al., 1998). Inorganic fertilizers may also contribute indirectly to an increase in earthworm abundance by increasing the quantity of crop residues returned to the soil (Edwards et al., 1995). The influence of conventional fertilizers, such as farmyard manure and slurry, on the dynamics of earthworm populations in agro-ecosystems is well documented (Lee, 1985; Edwards and Bohlen, 1996). A number of studies have examined the long-term impact of organic and inorganic fertilizer applications on earthworm populations. For example, Tiwari (1993) found that earthworm biomass in ploughed fallow plots of a former pineapple orchard in India was three times greater in plots receiving manure than in inorganically fertilized plots, while a combination of manure and inorganic fertilizer supported the greatest earthworm biomass. On a continuously cropped, North American corn agro-ecosystem receiving fertilizers for 6 years, Whalen et al. (1998) recorded a significantly greater earthworm number and biomass in manureamended plots compared to inorganic fertilizertreated plots. In contrast, there has been little research comparing the effects of various kinds or qualities of organic amendments (e.g., manure, slurry, biosolids, composts) on earthworm populations in agro-ecosystems (Bu ¨nemann et al., 2006). Yet, it is known that different organic materials, including uncomposted and composted green wastes and manures, affect soil-dwelling invertebrate pests such as nematodes differently (Litterick et al.,
2004). We conducted a field plot experiment to assess if eight different fertilizer regimes, of which five were organic amendments used in Belgian agriculture, would cause changes in the earthworm abundance and the species composition. Preliminary results from that comparative study suggested that farmyard manure, cattle slurry and three different types of compost all caused similarly large increases in earthworm numbers 12 months after initial application of these amendments (Leroy et al., 2007). Here, we report longer term results from the same field experiment and we focus on the novel question of how organic fertilizers with a different composition, and hence quality, can influence earthworm abundance differently. We could address this question because the same mass of organic C was applied with each organic material.
Materials and methods Experimental design The experimental field, located in Melle (experimental site of Ghent University, Belgium, 501590 N, 031490 E, 11 m above sea level) is a sandy loam soil with the following granulometric composition: 11.7% 0–2 mm, 52.0% 2–50 mm and 36.3% 450 mm. Prior to the experiment, the field was cropped with monoculture maize for 8 years with mineral fertilization only. Table 1 summarizes the available meteorological data, covering a large part of the experimental period at Melle. The field experiment, started in 2005, was a randomized complete block design with four replicates comparing eight treatments (Figure 1): FYM, cattle slurry+crop residues (CSL), vegetable, fruit and garden waste compost (VFG), two types of farm compost (CMC1 and CMC2), mineral fertilizer (MIN N), and two treatments without fertilization (one with a crop (NF+) and one without (NF), to assess the impact of the presence or absence of a crop). The two types of farm compost differed in the composition of the starting materials. CMC1 was composed of mostly woody, C-rich materials resulting in a final C/N ratio of ca. 20–40;
ARTICLE IN PRESS Earthworms and quality of exogenous organic matter Table 1.
141
Total precipitation (mm) and average monthly air temperature (1C) at Melle Month 1
Total 2
3
4
5
6
7
8
9
10
11
12
24.0 70.3 55.6 46.0
68.2 52.4 0.0 50.0
59.3 112.6 108.2 59.0
39.2 45.7 102.8 65.0
118.6 21.6 177.7 72.0
98.4 292.7 84.7 74.0
43.8 19.2 84.3 72.0
40.5 106.3 63.0 72.0
82.1 79.4 – 64.0
50.6 107.6 – 59.0
temperature (1C) 5.2 2.8 6.8 1.9 2.8 5.1 7.4 6.4 8.2 2.4 3.1 5.2
9.7 9.1 12.8 8.4
12.2 14.2 14.3 12.1
18.2 16.3 17.5 15.1
18.2 21.8 17.3 16.8
16.3 16.9 16.9 16.7
16.0 18.0 14.1 14.4
14.1 13.8 10.2 10.3
6.8 9.6 – 6.2
3.9 5.8 – 3.2
Precipitation (mm) 2005 36.7 63.4 2006 16.2 96.6 2007 78.6 108.4 42.0 Norma 51.0 Average air 2005 2006 2007 Norma a
724.8 1020.6 –
Norm: average over the last 30 years.
Figure 1. Layout of the field experiment.
CMC2 was made from green, N-rich materials and had a final C/N ratio of 10–20. Experimental plots (8 m 6 m) were located next to each other within blocks, thus with no distance between the differently treated plots. Furthermore, there were no above or below ground barriers between the plots, since this would have interfered with tillage, fertilization and crop protection operations. Organic amendments and fertilizers were applied on four occasions (Table 2). The first application took place on 21 April 2005 and fodder beet was sown the day after. The second fertilization was on
6 October 2005 followed by the sowing of winter wheat the next day. One month after the harvest of the winter wheat, the organic amendments and fertilizers were applied for the third time (7 September 2006) and a catch crop (Phacelia) was sown. On 2 May 2007, the fourth application took place and red cabbage was planted 3 weeks later. Amounts of organic amendments were adjusted to supply all plots, receiving organic amendments, with equal amounts of organic C. This amount of organic C was 4000 kg C ha1 in application one and two, 1500 kg C ha1 in application three and 2000 kg C ha1 in application four. In all cases, the
ARTICLE IN PRESS 142
B.L.M. Leroy et al.
Table 2. The amounts of organic fertilizers and their C and N content, and extra mineral N applied on 21 April 2005, 6 October 2005, 7 September 2006 and 2 May 2007 (abbreviations of treatments, see Fig. 1) C content (g kg1 fresh matter)
Treatment (fertilizer)
21/04/05 (4000 kg C ha1) FYM 62.2 VFG 179.4 CMC1 71.4 CMC2 59.7 CSL 26.7a +Crop residues 378.0 (straw) MIN N –
N content (g kg1 fresh matter)
Applied amount (kg ha1)
Extra mineral N to be added (kg ha1)
4.7 15.2 1.7 2.8 3.9a 5.5
64,329 22,303 56,007 67,058 77,382b 4704
105 114 165 165
–
–
165
1
06/10/05 (4000 kg C ha ) FYM VFG CMC1 CMC2 CSL +Crop residues (beet leaves) MIN N
–
23/03/06
26/04/06
106.8 175.7 71.8 77.9 20.4a 49.1
6.8 14.5 3.1 7.2 2.8a 3.8
37,453 22,770 55,718 51,348 74,698b 53,636
81 88 91 89
97 98 99 97
74
94
–
–
–
91
98
6.7 15.5 4.1 3.4 3.8a –
14,398 8188 19,330 23,757 56,754b –
66 67 86 86
6.1 9.1 3.6 6.0 3.2a
19,125 14,329 10,417 21,986 71,287b –
1
07/09/06 (1500 kg C ha ) FYM 104.2 VFG 183.2 CMC1 77.6 CMC2 63.1 CSL 26.4a MIN N –
– 86
1
02/05/07 (2000 kg C ha ) FYM 104.6 VFG 139.6 CMC1 192.0 CMC2 91.0 CSL 28.1a MIN N – a b
–
106 103 170 162 – 162
g l1. l ha1.
organic amendments were incorporated to a depth of 20 cm using a rotary tiller. To correct for differences in the plant available N content of the different organic amendments, extra mineral N (ammonium nitrate 27%) was applied on organically amended plots where needed to achieve equal levels of plant available N in all treatments and thus to support equal crop growth. For the fodder beet, the catch crop and the red cabbage, the mineral N was applied together with the organic amendments, while for the winter wheat, the extra mineral N was splitapplied on 23 March 2006 and 26 April 2006. For the calculation of the amount of extra mineral N to be added, the mineralization rates of the soil and of
the organic fertilizers, both determined by laboratory incubation as described by De Neve and Hofman (1996), were taken into account together + with the amount of mineral N (NO 3 and NH4) present in the soil at the time of fertilization. On the CSL plots, part of the organic C was applied as crop residues (except before the catch crop and the red cabbage), since applications of 4000 kg C ha1 as slurry, would have provided excessive amounts of mineral N. The first time (21 April 2005), the crop residues used were straw, while the second time (6 October 2005), it was fodder beet leaves. At each fertilization, except before the Phacelia, on the minerally fertilized plots and on the plots receiving less than 300 kg ha1 K2O and 100 kg ha1
ARTICLE IN PRESS
a a a a a a a a 36.771.5 34.771.3 35.071.9 35.871.9 36.071.2 35.070.7 35.173.0 35.870.4 146.278.4 abc 158.6714.6 bc 141.479.3 abc 150.1712.5 c 127.175.6 a 127.776.0 a 131.478.0 ac 132.873.5 ac Different letters indicate a significant difference per parameter according to Tukey’s HSD test (po0.05).
6.1070.10 6.2270.07 5.9870.05 6.0470.10 6.0870.06 5.8270.18 5.8670.17 5.9770.05 15/10/07 FYM VFG CMC1 CMC2 CSL MIN N NF+ NF
ab b abc ab ab c ac abc
1.1670.07 1.1970.03 1.1170.05 1.1370.03 1.0970.06 0.9670.04 0.9870.05 0.9870.04
a a a a ac b bc bc
0.10370.004 0.10970.005 0.09570.007 0.10370.002 0.09570.002 0.08370.004 0.08570.004 0.08470.003
ab b a ab a c c c
28.673.9 26.072.5 23.271.8 25.473.1 25.072.0 25.072.5 24.672.2 24.770.2
a a a a a a a a
2.2270.45 2.1670.57 1.8970.64 2.0470.64 2.1470.39 1.7970.45 1.6770.24 1.5070.18
a a a a a a a a
24.375.6 a 18.571.8 a 19.5710.3 a 20.275.5 a 24.672.1 a 18.978.5 a 11.672.3 a 16.172.4 a
32.7 122.4 19.7 1.60
Mg (mg per 100 g dry soil) Ca (mg per 100 g dry soil) K (mg per 100 g dry soil) Na (mg per 100 g dry soil) P (mg per 100 g dry soil)
20.8 0.086 1.01 5.90
Earthworms were sampled five times during the experimental period using a mustard powder suspension, following Gunn (1992) and Lawrence and Bowers (2002), and subsequent handsorting. Due to heavy rains (Table 1) and resultant unfavourable soil conditions in the weeks before the start of the experiment, the first sampling in 2005 could only be accomplished 6 weeks after the start of the experiment (31/05/05 vs. 21/04/05) and it was restricted to the unfertilized plots (NF+ and NF). The results of this sampling were considered representative of the initial situation because the field had a uniform management history and the eight plots assessed were randomized in the field (Figure 1). The other samplings were carried out in spring and autumn of 2006 and 2007, since in
26/10/04
Earthworm sampling and analysis
Total N (%)
Soil chemical properties of the experimental field (0–20 cm) were assessed at the start of the experiment and after two and a half years of different fertilizer regimes (Table 3). The pH was measured potentiometrically in a 1:2.5 soil:KCl extract. In soils where the pH-KCl is lower than 6.5, the amount of organic C is equal to the amount of total C as no free carbonates are normally present. In these soil samples, total C, and hence organic C, and total N were measured by dry combustion at 850 1C using an elemental analyser (Vario MAX CNS, Elementar). K, Na, Ca, Mg and available P were assessed by extraction of the soil with ammoniumlactate-acetic acid (extraction ratio 1:20) in dark polyethylene bottles, shaken for 4 h and the suspension was filtered in dark polyethylene bottles that were stored cool (4 1C) until analysis. The P concentration was measured colorimetrically by the Mo blue method (Scheel, 1936) at 700 nm with a photometer (Universal Photometer, Vitatron). The concentration of K and Na was measured with a flame photometer (Elex 6361, Eppendorf) in an airpropane flame at 768 and 589 nm, respectively, while for the Ca concentration, an air-acetylene flame was used at 623 nm. The concentration of Mg was determined by atomic absorption spectrometry (SpectrAA Atomic Absorption Spectrometer, Varian) at 285 nm.
C (%)
Soil chemistry
pH-KCl
P2O5 out of the organic amendments, an extra amount of K2O (300 kg ha1) and P2O5 (100 kg ha1) was applied to achieve equal minimum levels of plant available K2O and P2O5.
143 Table 3. Soil chemical properties of the experimental field (0–20 cm), before the start of the experiment (26 October 2004) and at the end of the experimental period (15 October 2007) (abbreviations of treatments, see Fig. 1)
Earthworms and quality of exogenous organic matter
ARTICLE IN PRESS 144 temperate regions earthworms are most active during spring and autumn when temperatures are moderate and soil moisture levels adequate (Bohlen et al., 1995): sampling two on 16–17 May 2006, sampling three on 2–3 October 2006, sampling four on 4–5 June 2007 and sampling five on 24–25 September 2007. The day before each sampling, 6 g of mustard powder was mixed with 15 ml of water to produce a paste; immediately before sampling, this mustard paste was mixed with 0.8 l of water. A metal frame (20 cm 20 cm) was placed on the soil and all surface plant litter within the frame removed. The mustard solution was then poured evenly across the frame. After 10 min, while collecting the emerging earthworms, this treatment was repeated. After another 10 min, the soil in the same quadrant was excavated to a depth of 20 cm, using another metal frame, spread on a white plastic sheet and handsorted to recover remaining earthworms. Two samples were taken per replicate plot. All earthworms were taken to the lab and washed. They were transferred into a 4% formalin solution and the total weight of the preserved earthworms (with gut content) per sample was recorded. Finally, the earthworms were stored until species identification of the adults.
Statistical analysis Earthworm data of the four dates at which all plots were sampled (May 2006–September 2007) were analysed by analysis of variance (ANOVA) for repeated measures using StatView v. 5.0 (SAS Institute Inc.). Sources of variability were partitioned into between-subjects factors (blocks; treatments) and within-subjects factors (sampling date). Treatment means were compared by Tukey’s HSD post-hoc tests. The two subsamples from each plot per sampling date were pooled for ANOVA because nesting of factors could not be combined with the repeated measures procedure. Final soil chemical data were analysed by one-way ANOVA and treatment means compared with Tukey’s HSD test.
Results Soil chemistry The four applications of all organic amendments, except CSL, significantly (po0.05) increased the soil organic C content compared to unamended treatments (Table 3). The organically amended
B.L.M. Leroy et al. plots had a similar final organic C content. All organically amended plots had a significantly higher total N content than the unamended plots, but significant differences (po0.05) could also be observed between the CMC1 and the CSL plots on the one hand and the VFG plots with the highest total N content on the other. Both the organic C and the total N content were similar on the unamended plots (MIN N, NF+ and NF). The other soil chemical properties showed small differences or trends among the treatments. Significant differences among treatments (po0.05) could only be observed for the pH-KCl and the Ca content.
Earthworms In all organically amended plots, the earthworm number and biomass was higher than in the unamended plots for the spring 2006 sampling, being on average at least twice as high compared to the initial situation (Figure 2A). However, no clear differences in the earthworm abundance among the five organic amendments could be observed. The unfertilized plots (NF+ and NF) had the lowest earthworm number and biomass and the mineral fertilization (MIN N) was intermediate. From the sampling in autumn 2006 onwards, the FYM and CSL plots exceeded on all sampling occasions, the compost and unamended plots in earthworm number, reaching densities of 800 individuals m2 (Figure 2B–D). Earthworm numbers were similar in the three compost treatments (VFG, CMC1 and CMC2), with an average of about 400–500 individuals m2. The differences in earthworm number among the three unamended plots (MIN N, NF+ and NF), which could be observed in spring 2006 (Figure 2A), disappeared at the following sampling dates, except for the sampling in spring 2007 (Figure 2C). The relatively high earthworm number in the unamended plots at the last sampling (Figure 2D) was surprising, with values reaching 300–350 individuals m2, twice as high as the initial situation. Except for these higher earthworm numbers, Figure 3A shows that since the sampling in autumn 2006, the earthworm numbers remained relatively constant in all plots. The eight treatments can be clearly divided in three groups: FYM and CSL plots, compost plots and unamended plots. For the earthworm biomass, it took until spring 2007 to cluster the fertilizer regimes in a similar fashion as for the earthworm number (Figure 3B). The differences were, however, less pronounced and the earthworm biomass showed a higher
ARTICLE IN PRESS Earthworms and quality of exogenous organic matter
600 500 400 300 200 100 0
YM
F
FG
V
C1
CM
C2
SL
NN
I C M CM Treatment
F+
N
600 500 400 300 200 100 0
F
V
CM
SL
I C M CM Treatment
NN
N
F+
300
0
M
G
VF
C1
CM
+ N C2 CSL IN NF M CM Treatment
-
NF
A. rosea A. icterica A. chlorotica A. caliginosa L castaneus L. rubellus L. terrestris Juvenile Biomass 150
1000 900
NF
Number of earthworms
700
C2
400
100
m-2
Number of earthworms m-2
800
C1
500
FY
140 130 120 110 100 90 80 70 60 50 40 30 20 10 0
FG
600
NF
900
YM
700
200
A. rosea A. icterica A. chlorotica A. caliginosa L castaneus L. rubellus L. terrestris Juvenile Biomass 150
1000
800
Earthworm biomass (g m-2)
700
140 130 120 110 100 90 80 70 60 50 40 30 20 10 0
900
800 700 600 500 400 300 200 100 0
M
FY
G
VF
C1
CM
+ N C2 CSL IN NF M CM Treatment
-
140 130 120 110 100 90 80 70 60 50 40 30 20 10 0
Earthworm biomass (g m-2)
800
A. rosea A. icterica A. chlorotica A. caliginosa L castaneus L. rubellus L. terrestris Juvenile 150 Biomass
1000
Earthworm biomass (g m-2)
Number of earthworms m-2
900
150 140 130 120 110 100 90 80 70 60 50 40 30 20 10 0
Number of earthworms m-2
1000
Earthworm biomass (g m-2)
A. rosea A. icterica A. chlorotica A. caliginosa L castaneus L. rubellus L. terrestris Juvenile Biomass
145
NF
Figure 2. Earthworm number, subdivided in adult species and juveniles, and total biomass (treatment means7S.D.), and the initial situation on 31 May 2005 (full line ¼ number; dotted line ¼ biomass) (abbreviations of treatments, see Figure 1): (A) sampling date: 16–17 May 2006; (B) sampling date: 2–3 October 2006; (C) sampling date: 4–5 June 2007; (D) sampling date: 24–25 September 2007.
variability compared to the earthworm number. In autumn 2006 (Figure 2B), the earthworm biomass was highest in the FYM plots, while in spring 2007, the FYM and CSL plots had a higher earthworm biomass than the other treatments, as was the case for the earthworm number (Figure 2C). In the MIN N
plots, the biomass was comparable to the compost plots, while the two unfertilized plots had the lowest biomass. In autumn 2007, the highest biomass was again recorded on the FYM and CSL plots, while a rise in the earthworm biomass (as was seen for the earthworm number) in the unfertilized
ARTICLE IN PRESS 146
B.L.M. Leroy et al.
1000 900
FYM VFG CMC1 CMC2 CSL MIN N NF+ NF-
Earthworm number m-2
800 700 600 500 400 300 200 100 0
5 5 6 6 7 6 7 /06 /06 /05 /07 4/0 /30/0 /5/05 1/10 /16/0 /24/0 /29/0 /4/06 0/10 2/15 /20/0 /28/0 /3/07 /8/07 1/14 9 6 3 1 1 8 1 4 5 2 1 7 9 1
4/2
Date 160
Earthworm biomass (g m-2)
140 120
FYM VFG CMC1 CMC2 CSL MIN N NF+ NF-
100 80 60 40 20 0 5 6 5 6 6 7 7 /06 /06 /05 /07 4/0 /30/0 /5/05 1/10 /16/0 /24/0 /29/0 /4/06 0/10 2/15 /20/0 /28/0 /3/07 /8/07 1/14 2 / 6 1 9 3 4 1 5 8 1 2 7 1 4 1 9 Date
Figure 3. Dynamics of the earthworm abundance since the start of the experiment (abbreviations of treatments, see Figure 1): (A) abundance (number m2), (B) biomass (g m2).
plots was observed (Figure 2D), with values only slightly lower than in the compost plots. Repeated measures ANOVA of the combined data (four sampling dates in 2006 and 2007) confirmed a highly significant effect of fertilization treatment on earthworm number and biomass (Table 5). The small but significant block effect suggests that blocking in this field was successful, removing some field-related variability. Significant sampling date effects probably reflect systematically higher earthworm numbers and biomass across all treatments in autumn 2007, and the significant
date treatment interaction for numbers was caused by the rapid increase in autumn 2006 in FYM and CSL treatments (Table 4). Post-hoc treatment comparisons, across dates, confirmed that the FYM and CSL plots supported significantly higher earthworm abundances than the plots treated with composts and significantly higher earthworm biomass than the two farm composts (Table 5). In the samples taken in spring 2006, the amount of juveniles varied between 63% and 82% of the total earthworm number, while for the later
ARTICLE IN PRESS Earthworms and quality of exogenous organic matter Table 4. Summary results of repeated measures analysis of variance (ANOVA) of earthworm abundance and biomass Source of variation
F-ratioa
DF
Abundance
Biomass
Between-subjects Block (B) Treatment (T) Error
3 7 21
3.1* 21.3***
3.7* 22.0***
Within-subjects Sampling date (D) DB DT Error
3 9 21 63
21.5*** 0.8 ns 2.4**
6.9*** 1.0 ns 1.0 ns
a Significance levels: ***po0.001; **po0.01; *po0.05; ns: not significant (p40.05).
Table 5. Multiple treatment mean comparisons (Tukey–Kramer) of earthworm abundance and biomass (treatments sharing the same letter are not significantly different at po0.05) Treatment
Number
Biomass
FYM CSL VFG CMC1 CMC2 MIN N NF+ NF
a a
a a b b b b
c c c c
d d
b b
c c c
d d d d
e e e
samplings, 70–94% of the earthworms sampled were juveniles. The earthworm species found were Lumbricus terrestris as anecic species, Lumbricus castaneus and Lumbricus rubellus as epigeic species and Aporrectodea caliginosa, Allolobophora chlorotica, Allolobophora icterica and Allolobophora rosea as endogeic species. The relative distribution of these earthworm species among the sampling dates and the treatments did not show clear differences or trends, with L. rubellus, A. caliginosa and A. chlorotica being the most abundant species (Figure 2A–D).
Discussion and conclusions The soil organic C content had increased in all plots receiving organic amendments and was
147 significantly higher (except from CSL) than that in the unamended plots after the two and a half year experimental period (Table 3). These results agree with previous studies of similar or longer duration (3–6 years) comparing effects of fresh and composted organic matter treatments with that of mineral fertilizer controls on soil organic C contents (e.g., Canali et al., 2004; Senesi and Plaza, 2007). The slightly lower soil C content under CSL was probably due to the composition or quality of this organic amendment. Cattle slurry, as a fresh product, is composed of more labile organic C than the other organic materials which contain more stable organic C due to the incorporation of straw in farmyard manure, or the composting process above ground during several months for the composts. The increase in earthworm abundance in the plots receiving organic amendments can be linked with the higher organic C content of these plots. Addition of organic matter is believed to be one of the major management variables affecting earthworm abundance. By providing food sources and increasing the overall levels of soil organic matter, organic amendments usually increase earthworm abundance (Bohlen et al., 1995). This increase as a response to organic amendments can be particularly pronounced in disturbed habitats of low organic matter content (Lowe and Butt, 2002; Curry, 2004), as was true for this experimental field which had been under monoculture maize receiving only mineral fertilizer for 8 years. Our experiment provides convincing evidence that compost amendments, incorporated into the soil to 20 cm, can enhance earthworm abundance significantly. Few comparable data on compost effects on earthworms are available from other agro-ecosystems, but Thomson and Hoffmann (2007) recently reported increased earthworm abundance (exotic Lumbricidae) under plots surface-mulched with compost made of domestic prunings for two growing seasons compared to bare plots in an Australian vineyard. The organic matter that provides the food base for the earthworm community is vitally important in determining their distribution and abundance (Edwards and Bohlen, 1996). For instance, Hendrix et al. (1992) reported a significant correlation between earthworm abundance and soil organic carbon content over a range of sites in Georgia, USA. However, from the results of the present research, we can assume that other factors than solely the soil organic C content influence the earthworm abundance. Although the final organic C content of all organically amended plots was statistically similar, ranging from 1.09% in the CSL
ARTICLE IN PRESS 148 plots to 1.19% in the VFG plots (Table 3), clear and significant differences in earthworm abundance (number and biomass) were observed, with the highest abundances found in the FYM and CSL plots (Figure 3A and B). We hypothesize that this seemingly contradicting result of a disconnection between the soil organic carbon content on the one hand and the earthworm abundance on the other can be explained by the chemical properties of the organic amendments used. The chief biochemical process of composting is humification, which is defined as the prolonged stabilization of organic substances against biodegradation. Composts are thus chemically stabilized and biologically matured amendments (Senesi and Plaza, 2007). As a result of extended microbial degradation, composts have lower total and watersoluble organic C contents, higher relative contents of humic acid like substances and a higher degree of polycondensation and polymerization than their raw starting materials. Animal manures, by contrast, are rich in incompletely humified materials including polysaccharides and S- and N-containing groups, most notably proteinaceous materials (Senesi and Plaza, 2007). These chemical differences between organic amendments also translate into different chemical characteristics of the organic matter in soil treated with these materials. For example, a recent Canadian study reported significantly higher humification indices (ratio of organic C content in unhumified fraction to fulvic and humic acids) and higher amino sugar contents (especially glucosamine) in two soil aggregate size fractions (0.25–2.00 and 42.00 mm) of loam soil treated for 3 years with fresh paper mill sludge (40 t ha1 year1) compared to soil amended with the same but composted sludge (Bipfubusa et al., 2008). In the present experiment, four applications of relatively fresh farmyard manure or cattle slurry that had not been subjected to composting resulted in much larger earthworm abundance increases than in the plots amended with any of the three composts (Figure 3A and B). We hypothesize that the nutritional value for earthworms of both the cattle slurry and farmyard manure, and hence also of the soil organic matter in plots treated with these raw materials, were higher than that of the composts and the soil OM in the compost plots, in which the applied organic matter was more decomposed and stabilized, due to the aerobic composting for several weeks or months. Our interpretation of field population data concurs with current ecological theory that has revised the traditional understanding of the trophic ecology of earthworms (Curry and Schmidt, 2007). Experimental evidence suggests firstly that
B.L.M. Leroy et al. (endogeic) earthworms are limited by available C in soil, irrespective of the total C content of soil (Tiunov and Scheu, 2004) and secondly that earthworms do not depend on micro-organisms as a food source but consume organic matter directly (Salamon et al., 2006). Since the same amounts of organic C were added in our organic treatments, it is plausible that higher amounts of nutritionally available C in the uncomposted organic treatments supported larger earthworm populations. It is also likely that earthworm abundances were not related to microbial biomass which can sometimes be elevated in soils treated with composts. One example is the study by Canali et al. (2004) in which an orchard soil receiving poultry manure for 6 years had a smaller microbial biomass and lower microbially mineralizable C content than the same soil amended with composted manure. However, in the present experiment, soil microbial biomass was not significantly different among the five organic treatments (Leroy, 2008), suggesting that availability of organic matter, and not microbes, determined the size of lumbricid community. From the third earthworm sampling onwards (autumn 2006), earthworm abundance remained fairly stable, but biomass continued to increase in response to repeated organic matter application. This pattern is similar to Irish data (Schmidt and Curry, 2001), which suggested that earthworm populations stabilize only 2–3 years after regular organic matter inputs to cultivated soil commenced. It would, however, be necessary to continue monitoring earthworm abundance longer to establish whether the treatment differences would become more pronounced in the present experiment. Based on the sampling in spring 2006, preliminary results suggested that the mineral fertilization and the presence or absence of a crop also influenced the earthworm abundance, since differences (although not significant, p40.05) in earthworm biomass and number were recorded between the MIN N, NF+ and NF plots (Figure 2A, also Leroy et al., 2007). These differences seemed to disappear in the next samplings, except in spring 2007, when the earthworm number and biomass in the MIN N plots tended to be again slightly higher than in the unfertilized plots. This is possibly due to a better crop performance on the minerally fertilized plots compared to the NF+ plots (Leroy, 2008), and hence a higher root and crop biomass. This higher biomass results in a higher amount of above and below ground crop litter, which is an important source of organic matter on which earthworms feed (Curry, 2004). Generally, the effects of the application of moderate levels of mineral fertilizer on earthworm
ARTICLE IN PRESS Earthworms and quality of exogenous organic matter abundance are considered positive in the literature, reflecting increased litter quality and quantity (Bostro ¨m, 1988; Muldowney et al., 2003). Again, future sampling would be required to establish if this effect of mineral fertilization on earthworm abundance would persist in the long term.
Acknowledgements The authors wish to thank Franky Van Peteghem and Jean-Pierre Van Maerke for their much appreciated assistance with the field work, and Luc Deboosere, Olle Victoor, Mathieu Schatteman, Tina Coddens and Sophie Schepens for their practical assistance with the earthworm sampling and in the lab. The authors thank AgriVet for the use of its experimental fields.
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