intersection method. The dry litter 1 g was milled and homogenised in water 100 ml for 5 min. Subsequently,
an aliquot of this homogenate was diluted to obtain four samples 1 mg litter per ml of water. Four membrane filters
were prepared according to Sundman and Sivela¨ 1978. The values are reported as mg dry fungal biomass g
21
of dry litter on the basis of the average 9.3 mm
2
cross section of the hyphae, a density of 1.1 g ml
21
and a dry mass of 15 Berg and So¨derstro¨m, 1979. The measures were made on
the three samples, each with three replicates. 2.8. pH and soluble substance measurements
The pH of the litter was determined by shaking litter in distilled water for 10 min 0.5 g dry litter in 10 ml water.
The suspension was left to stand for 10 min and the super- natant used to measure the pH with an electronic pH meter
HI 8424, HANNA Instruments, Sarmeola di Rubano PD, Italy.
Water soluble substances were determined by soaking the undecomposed leaf litter in distilled water 1 g of dry litter
in 70 ml of water at 20–22 8C for 24 h Berg and Wessen,
1984, modified. The samples underwent a two-fold sonica- tion for 2 h at the start and at the end of leaching period in a
sonicator bath filled with water and ice. To prevent micro- bial growth, 2–3 drops of sodium hypochlorite were added.
The dry weight of these substances was obtained by calcu- lating the difference between the dry weight of the litter
before and after leaching. Measurements on each sample were made in triplicate.
2.9. Statistics The mass loss over time was fitted to a simple exponential
curve Olson, 1963 ln
x
t
= x
2kt
where x is the original mass of leaf litter, x
t
the amount of litter remaining after time t, t is time year and k the decom-
position rate yr
21
. The half-life for decomposition t
12
, that is the time necessary to reach 50 mass loss, was
calculated
t
1=2
0:691=k:
The significance of differences among the litters was tested by two-way analysis of variance ANOVA followed
by Tukey test. Correlations were determined using the simple Pearson correlation coefficient.
3. Results
3.1. Initial chemical composition Among the macronutrients, the initial content of the C, N,
P, K and Ca was higher in the Cistus than in the Myrtus litter Table 2. In contrast, the initial content of Mg was higher in
the Myrtus litter while that of Na was similar in both litters. Among the micronutrients, particular attention was given to
A. Fioretto et al. Soil Biology Biochemistry 32 2000 1847–1855 1850
Table 2
Initial pH,
nutrient and
water solubl
e substanc
es contained
in Cistu
s an
d M
yrtus
Litter pH
Water sol
uble substa
nces C
mg g
2 1
dry wt.
N mg
g
2 1
dry w
t. P
mg g
2 1
dry wt.
K mg
g
2 1
dry wt.
Mg mg
g
2 1
dry wt.
Ca mg
g
2 1
dr y
w t.
Na mg
g
2 1
dry wt.
Mn mg
g
2 1
dry wt.
Cistus 5.3
10.8 414
10.8 0.68
5.84 1.22
11.7 1.22
0.09
Myrtus 5.4
16.8 375
9.8 0.55
3.86 2.62
10.7 1.35
0.07
Mn because it is essential for lignin-degrading enzymes Perez and Jeffrey, 1992; Archibald and Roy, 1992. Its
initial content was higher in the Cistus than in the Myrtus litter Table 2.
The water soluble substances were about 35 more abun- dant in the undecomposed leaf litter of Myrtus than of the
Cistus. The pH values were similar Table 2.
3.2. Litter decomposition Fig. 1 shows the decomposition dynamics over 15 months
of Cistus leaf litter incubated under Cistus and under Myrtus Myrtus replaced Cistus in the succession as well as Myrtus
litter incubated under Myrtus. The average decomposition rates were similar for all the
litter types during the first eight months of incubation. Subsequently, they were higher for litters under Myrtus
than under Cistus shrubs Fig. 1. After 15 months, litter under Cistus had lost 30 of the initial mass and those
under Myrtus had lost 40. Significant differences
P
, 0:001 were found between the mass loss patterns of Myrtus,
as well as of the Cistus under Myrtus, and that of Cistus under Cistus.
The data conformed to a first order exponential decay curve. The decay constants, determined until 240 d when
decomposition was independent from incubation microsites, ranged from 0.30 to 0.33 yr
21
Table 3. Subsequently, they increased for litters under Myrtus reaching the value k
0:48 and 0.57 yr
21
for Myrtus and Cistus, respectively. Over the experimental period the half-lives of Cistus and
Myrtus litters incubated under Myrtus were similar
t
1=2
1:5 yr and lower in Cistus under Cistus
t
1=2
2:1 yr:
Both litters incubated under Myrtus showed reduced decomposition rate during the dry summer. In the same
period the decomposition rate of Cistus litter incubated under Cistus, did not change.
3.3. Litter respiration Fig. 2 shows respiration rates as well as water contents for
litters at each sampling. The respiration rates showed seaso- nal variations: the highest value occurring in the wet
seasons, and the lowest in summer. In addition, they were correlated with the water content of the litter that, however,
had higher values in autumn than in spring. Only Cistus incubated under Myrtus showed a respiration pattern signif-
icantly different from the others
P
, 0:001: 3.4. Enzyme activities
During the decay process, extractable cellulase and
A. Fioretto et al. Soil Biology Biochemistry 32 2000 1847–1855 1851
Fig. 1. Residual mass of the initial of decomposing leaf litter of Cistus B—B and Myrtus V—V incubated under the relative shrubs. The
residual mass of Cistus litter incubated under Myrtus O– –-O is also reported. The values are means SE of 20 measurements.
Table 3 Decay constant k yr
21
SE of leaf litter of M. communis and C. incanus incubated in situ calculated at different incubation times. The coefficient of
determination r
2
of the exponential decay interpolation of weight loss is also reported. Significance for P
, 0:05; P , 0:01; P , 0:001 0–242 days
242–460 days 0–460 days
Cistus 0:32 0:02
0:30 0:03 0:33 0:02
Under Cistus r
2
0:98
r
2
0:94
r
2
0:99
Cistus 0:29 0:03
0:57 0:15 0:46 0:04
Under Myrtus r
2
0:97
r
2
0:88
r
2
0:95
Myrtus 0:33 0:03
0:48 0:16 0:45 0:04
Under Myrtus r
2
0:98
r
2
0:82
r
2
0:96
Fig. 2. Respiration rates of decomposing litters at the field moisture and water content of litter at sampling. Values are means SD of three
measurements with three replications. Cistus B—B and Myrtus V— V litter incubated under respective shrubs; O– –-O Cistus litter incu-
bated under Myrtus shrubs.
xylanase activities showed seasonal variations with a maxi- mum in autumn and a minimum in late spring and summer
Fig. 3. These patterns were similar to those of microbial respiration Fig. 2 and a correlation between these activities
and respiration was found Table 4. The highest activities occurred in Myrtus litter
P
, 0:001: No significant differ- ences were found in the Cistus in relation to the two incuba-
tion sites. The a-amylase activity in both litters was high at the start
of incubation and declined rapidly as decomposition progressed Fig. 3. In the Cistus litters the activity fell to
almost zero after two months. Consequently, no seasonal variation was observed. Between the litters, Myrtus had
the highest average a-amylase activity although the initial value was the lowest one. However, there was no significant
difference between them.
The b-amylase activity was similar in both kinds of litter but showed an irregular pattern Fig. 3. The highest activ-
ities of a- and b-amylase were less than 5–10 of those of cellulase and xylanase.
Laccase activity remained at low initial levels during the first 8 months Fig. 4, subsequently increasing rapidly as
A. Fioretto et al. Soil Biology Biochemistry 32 2000 1847–1855 1852
Fig. 3. Cellulase, xylanase, a- and b-amylase activities of litters of Cistus incubated under Cistus B—B and of Myrtus V—V and Cistus incubated under Myrtus O– –-O during their decomposition. Values are means SD of three measurements with three replicates of each.
Table 4 Coefficient of determination r
2
between properties examined during litter decomposition of Cistus and Myrtus. Significance for P , 0:05;, P , 0:01;
P , 0:001
Cistus under Cistus Cistus under Myrtus
Myrtus under Myrtus Water content — respiration
r
2
0:56
r
2
0:36
r
2
0:79
Water content — cellulase activity
r
2
0:80
r
2
0:80
r
2
0:50
Water content — xylanase activity
r
2
0:70
r
2
0:67
r
2
0:59
Water content — peroxidase activity
ND ND
r
2
0:60
Respiration — cellulase activity r
2
0:50
r
2
0:69
r
2
0:51
Respiration — xylanase activity r
2
0:60
r
2
0:73
r
2
0:74
Fungal biomass — laccase activity
r
2
0:88
r
2
0:85
r
2
0:94
Fungal biomass — mass loss r
2
0:50
r
2
0:94
r
2
0:77
decomposition progressed. The highest activities occurred in Myrtus litter while the lowest were found in Cistus litter
under Cistus. Differences, although more evident in the last phase of incubation, were significant
P
, 0:001 between Myrtus and Cistus, apart from the incubation site.
Peroxidase activity was found only in Myrtus litter Fig. 4. It showed a seasonal pattern, with a maximum in autumn
and a minimum in late spring and summer. In Cistus, peroxidase was below the detection limit of the assay for
almost the entire incubation period. However, a low activity was measured after a year of decay. Significant differences
were found between Myrtus and Cistus litters
P
, 0:05: 3.5. Fungal biomass
During the first eight months of incubation, the fungal biomass of decomposing litter under Myrtus slowly
increased in the wet period and decreased in the dry summer, while under Cistus it also increased in summer.
Subsequently, biomass rapidly increased in autumn Fig. 5, mainly in the litters under Myrtus. At the end of the
incubation period, the fungal biomass was 4–5 fold higher than the initial value for all the litters.
No correlation was found between the overall fungal biomass of Cistus and Myrtus litters and the xylanase and
cellulase enzyme complexes. Positive and significant corre- lation were found between fungal biomass and laccase
activity Table 4 and between fungal biomass and mass loss Table 4.
3.6. Changes in pH during decomposition Although litters of Myrtus and Cistus had similar pH
values at the start Fig. 6, they showed significantly differ- ent changes during decomposition
P
, 0:001: The pH in Myrtus litter stayed at 5.0 for about a year and then
increased to 6.5, whilst Cistus increased to 6–6.5 after just two months of incubation and stayed at this value for
the remaining incubation period. The Cistus litter incubated under Cistus and Myrtus shrubs showed a similar pattern.
4. Discussion
