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Soil Biology & Biochemistry: C. Le Guillou, D.A. Angers, P.A. Maron, P. Leterme, S. Menasseri-Aubry
Soil Biology & Biochemistry: C. Le Guillou, D.A. Angers, P.A. Maron, P. Leterme, S. Menasseri-Aubry
INRA, UMR 1069 Sol Agro et hydrosystme Spatialisation, F-35000 Rennes, France
Agrocampus Ouest, UMR 1069 Sol Agro et hydrosystme Spatialisation, F-35000 Rennes, France
c
Universit europenne de Bretagne, France
d
Agriculture et Agroalimentaire Canada, Centre de Recherche sur les Sols et les Grandes Cultures, 2560 Boulevard Hochelaga, Qubec, Qubec, Canada G1V 2J3
e
INRA, UMR 1229 Microbiologie du Sol et de lEnvironnement, F-21065 Dijon, France
b
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 21 December 2011
Received in revised form
7 March 2012
Accepted 9 March 2012
Available online 29 March 2012
The dynamics of soil water-stable aggregation (WSA) following organic matter (OM) addition are
controlled by microbial activity, which in turn is inuenced by carbon substrate quality and mineral N
availability. However, the role of microbial communities in determining WSA at different stages of OM
decomposition remains largely unknown. This study aimed at evaluating the role of microbial
communities in WSA during OM decomposition as affected by mineral N. In a 35-day incubation
experiment, we studied the decomposition of two high-C/N crop residues (miscanthus, C/N 311.3; and
wheat, C/N 125.6) applied at 4 g C kg1 dry soil with or without mineral N addition (120 mg N kg1 dry
soil). Microbial characteristics were measured at day 0, 7, and 35 of the experiment, and related to
previous results of WSA. Early increase in WSA (at 7 days) was related to an overall increase of the
microbial biomass (MBC) with wheat residues showing higher values in MBC and WSA than miscanthus.
In the intermediate stage of decomposition (from day 7 to 35), the dynamics of WSA were more associated with the dynamics of microbial polysaccharides and greatly inuenced by mineral N addition.
Mineral N addition resulted in a decrease or leveling off of WSA whereas it increased in its absence. We
suggest that opportunistic bacterial populations stimulated by N addition may have consumed binding
agents which decreased WSA or prevented its increase. To the contrary, microbial polysaccharide
production was high when no mineral N was added which led to the higher WSA in the late stage of
decomposition in this treatment. The late stage of decomposition was associated with a particular fungal
community also inuenced by the mineral N treatment. We suggest that WSA dynamics in the late stage
of decomposition can be considered as a narrow process3 where the structure of the microbial
community plays a greater role than during the initial stages.
2012 Elsevier Ltd. All rights reserved.
Keywords:
Bacteria
Fungi
Organic matter quality
N
Decomposition
Aggregate stability
1. Introduction
Water-stable aggregation (WSA) is an important characteristic
of soil functioning (Carter, 2002). It is a determinant factor of soil
susceptibility to erosion (Le Bissonnais, 1996), soils ability to
sustain plant germination and early growth (Angers and Caron,
1998) and is a mechanism of organic matter (OM) protection in
soils (Balesdent et al., 2000). Among factors affecting WSA, soil
organic matter is a major one (Tisdall and Oades, 1982) that can be
* Corresponding author. INRA, UMR Sol Agro et hydrosystme Spatialisation, 65
Route de Saint-Brieuc, CS 84215-35042 Rennes Cedex, France. Tel.: 33 2 23 48 54 73;
fax: 33 2 23 48 54 80.
E-mail address: Safya.Menasseri@agrocampus-ouest.fr (S. Menasseri-Aubry).
0038-0717/$ e see front matter 2012 Elsevier Ltd. All rights reserved.
doi:10.1016/j.soilbio.2012.03.009
127
Table 1
Biochemical characteristics of the crop residues.
Crop residue
C/N
Lignin/N
Hemicellulose
Cellulose
Lignin
Wheat
Miscanthus
125.6
311.3
21.2
88.1
13.2
6.3
34.0
28.1
45.6
52.4
7.2
13.2
MWD
wi *xi
Table 2
Absolute values of measured variables on unamended soil samples at day 0, 7 and 35
within N treatments (with (N) or without (N) mineral N supply). Values represent
the mean of three replicates. Values between brackets represent the standard deviation of the means.
Day 0
N
N
Day 7
N
N
Day 35
N
N
WSA (mm)
Microbial C
(mg kg1
dry soil)
Ergosterol
(mg kg1
dry soil)
Hot-water
extractable
polysaccharide
(mg kg1
dry soil)
0.5 (0.0)
0.5 (0.0)
60.8 (6.1)
56.6 (8.7)
0.7 (0.1)
0.7 (0.1)
0.9 (0.0)
0.9 (0.0)
0.4 (0.0)
0.4 (0.0)
70.2 (9.2)
58.9 (5.7)
1.0 (0.2)
0.8 (0.2)
0.7 (0.1)
0.6 (0.1)
0.3 (0.0)
0.4 (0.1)
60.5 (8.3)
55.8 (3.6)
0.5 (0.1)
0.6 (0.1)
0.5 (0.1)
0.9 (0.3)
-1
fresh soil samples (24 g fresh soil). Microbial C was estimated as the
organic C extracted in the fumigated samples minus the organic C
extracted in the non-fumigated samples. Extractable C was
measured using a Shimadzu TOC 5050A total carbon analyzer .
Ergosterol content was determined according to the method of
Djajakirana et al. (1996) and Gong et al. (2001). Extraction from
3.5 g soil sample was realized with 120 mL ethanol and 4 g of glass
beads by shaking for 30 min on a rotative agitator. The extracts
were ltered through glass ber lters (Whatman GF/C, UK) and
evaporated under vacuum on a rotary evaporator at 40 C. The
residues were dissolved in 10 mL ethanol. Determination of
ergosterol content was performed by HPLC as described in Annabi
et al. (2007).
Hot-water extractable polysaccharide content was determined
according to the method of Puget et al. (1999). The samples (1-g
dried and ground soil sample) were extracted using 20 mL demineralized water at 80 C for 24 h. The supernatant was collected after
centrifugation (20 000 g) and the polysaccharide content measured
by a colorimetric method at 490 nm according to the phenol-H2SO4
method (Dubois et al., 1956).
Microbial DNA was extracted according to the method described
by Ranjard et al. (2003). The genetic structure of the microbial
communities was determined by using the DNA ngerprinting
technique, the automated ribosomal intergenic spacer analysis
(ARISA). The bacterial and fungal ribosomal IGS were, respectively,
amplied with two primers sets: S-D-Bact-1522-b-S-20/L-D-Bact132-a-A-18 and ITS1F/3126T, PCR conditions being described by
Ranjard et al. (2003). A uorescent-labelled primer was used for the
LiCor DNA sequencer (ScienceTec, Les Ulis, France) in B-ARISA and
F-ARISA, namely the IRD800 dye uorochrome (MWG SA Biotech,
Ebersberg, Germany). ARISA fragments were resolved on 3.7%
polyacrylamide gels and run under denaturing conditions for 15 h at
3000 V/60 W on a LiCor DNA sequencer (ScienceTec). The data were
analysed using the 1D-Scan software (ScienceTec). This software
converted uorescence data into electrophoregrams where peaks
represented PCR fragments. The height of the peaks was calculated
in conjunction with the median lter option and the Gaussian
integration in 1D-Scan, and represented the relative proportion of
the fragments in the total products.
128
120
100
80
60
40
Wh -N
Wh +N
20
Mc -N
Mc +N
0
0
10
15
20
25
30
35
40
Time (day)
Fig. 1. Microbial C during the incubation (residue-treated minus control within each N
treatment). Wheat (Wh) and Miscanthus (Mc) residues combined with 0 (N) and 120
(N) mg N kg1 dry soil addition rates. Error bars represent the standard error of the
means.
-1
3.5
Wh -N
Wh +N
Mc -N
Mc +N
2.5
2
1.5
1
0.5
0
0
10
15
20
25
30
35
40
-1
129
1
Wh -N
Wh +N
0.8
Mc -N
Mc +N
0.6
0.4
0.2
0
-0.2
-0.4
0
10
15
25
20
30
35
40
Time (day)
Fig. 3. Hot-water extractable polysaccharide content during the incubation (residuetreated minus control within each N treatment). Wheat (Wh) and Miscanthus (Mc)
residues combined with 0 (N) and 120 (N) mg N kg1 dry soil addition rates. Error
bars represent the standard error of the means.
3.2. Ergosterol
1.6
Wh -N
Wh +N
1.4
1.2
Mc -N
Mc +N
1
0.8
0.6
0.4
0.2
0
0
10
15
20
25
30
35
40
Time (day)
Time (day)
Fig. 2. Ergosterol during the incubation (residue-treated minus control within each N
treatment). Wheat (Wh) and Miscanthus (Mc) residues combined with 0 (N) and 120
(N) mg N kg1 dry soil addition rates. Error bars represent the standard error of the
means.
Fig. 4. WSA expressed as the mean weight diameter (MWD) during the incubation
(residue-treated minus control within each N treatment). Wheat (Wh) and Miscanthus
(Mc) residues combined with 0 (N) and 120 (N) mg N kg1 dry soil addition rates.
Error bars represent the standard error of the means. Adapted from Le Guillou et al.
(2011).
130
PC2 12%
Wh -N D7
PC2 14%
Mc -N D0
Mc +N D7
Wh +N D7
Wh +N D35
Wh -N D35
Mc -N D35
Mc +N D35
Mc -N D7
Wh -N D0
Wh +N D0
Mc +N D0
PC1 49%
PC2 14%
PC1 44%
PC2 16%
Un +N
Mc +N
Un -N
Wh -N
Mc +N
Mc -N
Wh +N
Wh +N
Un -N
Un +N
Mc -N
Wh -N
PC1 46%
PC1 26%
Fig. 5. Bacterial genetic structure analysis. Principal component analysis ordination (PC1 and PC2) of the bacterial genetic structure of the (a) wheat and (b) miscanthus treatments
from each sampling date and of the (c) day 7 and (d) day 35 considering every treatment. Statistical ellipses drawn over the plot replicates represent 90% condence. The percentage
of explained variations for the rst two axes is indicated within the gures. Unamended (Un), Wheat (Wh) and Miscanthus (Mc) residues treatments combined with 0 (N) (white
ellipses) and 120 (N) (grey ellipses) mg N kg1 dry soil addition rates at day 0 (D0), 7 (D7) and 35 (D35).
PC2 16%
131
PC2 13.1%
Mc -N D0
Wh +N D35
Mc -N D35
Mc +N D0
Mc +N D35
Wh +N D7
Mc -N D7
Wh -N D35
Wh +N D0
Wh -N D7
Mc +N D7
Wh -N D0
PC1 38%
PC2 18%
c
Mc +N
PC1 29%
Mc -N
PC2 21%
Un +N
Un -N
Un -N
Wh -N
Wh +N
Wh -N
Mc +N
Un +N
Mc -N
Wh +N
PC1 25%
PC1 37%
Fig. 6. Fungal genetic structure analysis. Principal component analysis ordination (PC1 and PC2) of the fungal genetic structure of the (a) wheat and (b) miscanthus treatments from
each sampling date and of the (c) day 7 and (d) day 35 considering every treatment. Statistical ellipses drawn over the plot replicates represent 90% condence. The percentage of
explained variations for the rst two axes is indicated within the gures. Unamended (Un), Wheat (Wh) and Miscanthus (Mc) residues treatments combined with 0 (N) (white
ellipses) and 120 (N) (grey ellipses) mg N kg1 dry soil addition rates at day 0 (D0), 7 (D7) and 35 (D35).
Table 3
Correlation coefcients (r) between the mean weight diameter (MWD) and measured microbial variables at day 7 and 35 (data are expressed as the difference between the
residue-treated and control soils within each N treatment). Bacterial and fungal genetic structure are expressed as the Euclidian distance between treated and control soils
within each N treatment for each incubation time. CeCO2 rate data are from Le Guillou et al. (2011). (ns non signicant, yP < 0.1, *P < 0.05, **P < 0.01).
Day 7
Day 35
CeCO2 rate
Microbial C
Ergosterol
Hot-water
extractable
polysaccharide
Bacterial
genetic
structure
Fungal
genetic
structure
ns
ns
0.76**
ns
ns
ns
ns
0.67*
ns
0.79**
ns
0.53y
132
Water-stable aggregate
II
Time
week
C quality
OM input
Total microbial
month
Water-stable aggregate
r = 0.76 **
N availability
biomass
Microbial
community
II
r = 0.67 *
Polysaccharide
structure
Fig. 7. This conceptual diagram depicts our understanding of the microbial contribution to the transient soil water-stable aggregation dynamics following organic matter
(OM) inputs. ( ) denotes the controls on the level of water-stable aggregates at
different stages of OM decomposition. In phaseI (week(s)) (continued line), the early
increase in water-stable aggregation is controlled by OM inputs biochemical characteristics (C-substrate quality) and related to the increase in the total microbial biomass.
In phaseII (dotted line) (month(s)), the level of water-stable aggregates is controlled by
N availability and related to the level of microbial polysaccharides. The amount of
polysaccharides would be determined by the balance between production and
consumption which are inuenced by the fungal and bacterial populations stimulated
(microbial community structure).
133