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Synergistic eects of water temperature and dissolved
nutrients on litter decomposition and associated fungi
Veronica Ferreira, Eric Chauvet
To cite this version:
Veronica Ferreira, Eric Chauvet. Synergistic eects of water temperature and dissolved nutrients on
litter decomposition and associated fungi. Global Change Biology, Wiley, 2011, vol. 17, pp. 551-564.
�10.1111/j.1365-2486.2010.02185.x�. �hal-00942851�
To link to this article : DOI: 10.1111/j.1365-2486.2010.02185.x
http://doi.wiley.com/10.1111/j.1365-2486.2010.02185.x
To cite this version: Ferreira, Veronica and Chauvet, Eric Synergistic effects
of water temperature and dissolved nutrients on litter decomposition and
associated fungi. (2011) Global Change Biology, vol. 17 (n° 1). pp. 551-564.
ISSN 1354-1013
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Synergistic effects of water temperature and dissolved
nutrients on litter decomposition and associated fungi
V E R O
´
N I C A F E R R E I R A
*
and E R I C C H A U V E T w z
*
IMAR-CMA, Department of Life Sciences, University of Coimbra, PO Box 3046, 3001-401 Coimbra, Portugal, wEcoLab
(Laboratoire d’e
´
cologie fonctionnelle), Universite
´
de Toulouse; UPS, INPT, 29 rue Jeanne Marvig, F-31055 Toulouse, France,
zCNRS, EcoLab, F-31055 Toulouse, France
Abstract
In woodland streams, the decomposition of allochthonous organic matter constitutes a fundamental
ecosystem process,
where aquatic hyphomycetes play a pivotal role. It is therefore greatly affected by water temperature and nutrient
concentrations. The individual effects of these factors on the decomposition of litter have been studied previously.
However, in the climate warming scenario predicted for this century, water temperature and nutrient concentrations are
expected to increase simultaneously, and their combined effects on litter decomposition and associated biological
activity remains unevaluated. In this study, we addressed the individual and combined effects of water temperature
(three levels) and nutrient concentrations (two levels) on the decomposition of alder leaves and associated aquatic
hyphomycetes in microcosms. Decomposition rates across treatments varied between 0.0041 day
1
at 5 1C and low
nutrient level and 0.0100 day
1
at 15 1C and high nutrient level. The stimulation of biological variables at high nutrients
and temperatures indicates that nutrient enrichment of streams might have a higher stimulatory effect on fungal
performance and decomposition rates under a warming scenario than at present. The stimulation of fungal biomass and
sporulation with increasing temperature at both nutrient levels shows that increases in water temperature might
enhance fungal growth and reproduction in both oligotrophic and eutrophic streams. The stimulation of fungal
respiration and litter decomposition with increasing temperature at high nutrients indicates that stimulation of carbon
mineralization will probably occur at eutrophied streams, while oligotrophic conditions seem to be ‘protected’ from
warming. All biological variables were stimulated when both factors increased, as a result of synergistic interactions
between factors. Increased water temperature and nutrient level also affected the structure of aquatic hyphomycete
assemblages. It is plausible that if water quality of presently eutrophied streams is improved, the potential stimulatory
effects of future increases in water temperature on aquatic biota and processes might be mitigated.
Keywords: aquatic hyphomycetes, ecosystem functioning, global change, interactions, litter decomposition, nutrient enrichment,
streams, temperature
Introduction
Earth is presently going through a warming period, and
s
imulations considering a doubling in atmospheric CO
2
predict a 1.1–6.4 1C increase in air temperature by the year
2100 (IPCC, 2007). Water temperature of streams and rivers
is expected to mirror this increase (Stefan & Sinokrot, 1993;
Eaton & Scheller, 1996), which may lead to altered com-
munity structure (Hogg & Williams, 1996; Mouthon &
Daufresne, 2006), species distribution (Winterbourn, 1969;
Eaton & Scheller, 1996; Castella et al., 2001), interspecific
relationships (Webster et al., 1976; Beisner et al., 1997;
Mouritsen et al., 2005; Jiang & Morin, 2007), biodiversity
(Petchey et al., 1999; Castella et al., 2001) and ecological
processes (Petchey et al., 1999; Baulch et al., 2005).
Small, moderate to high latitude/altitude forest streams,
where water temperature is generally low, are particularly
sensitive to temperature increases (Stefan & Sinokrot,
1993). In these streams, the primary source of carbon
and energy for aquatic food webs is terrestrially derived
organic matter supplied by the riparian vegetation, whose
shade also limits primary production (Vannote et al., 1980).
Decomposition of this organic matter is carried out mainly
by aquatic hyphomycetes and shredding invertebrates
(Hieber & Gessner, 2002; Pascoal & Ca
´
ssio, 2004), through
mineralization, incorporation into biomass and conversion
into fine particulate organic matter (Gonza
´
lez & Grac¸a,
2003; Gulis & Suberkropp, 2003b, c; Pascoal & Ca
´
ssio,
2004). Litter decomposition, being primarily a biological
process, is expected to be affected by increased water
temperature. Correlative studies have demonstrated a
positive relationship between water temperature and de-
composition rates of litter incubated along altitudinal or
latitudinal gradients (Irons et al., 1994; Fabre & Chauvet,
Correspondence: V. Ferreira, tel. 1 351
239 855 760, fax 1 351 239
855 789, e-mail: veronica@ci.uc.pt
1998), probably mediated by temperature effects on the
biota (Fabre & Chauvet, 1998). Simulations also predict
higher decomposition rates with increased water tempera-
ture, through enhancement of invertebrate and microbial
activity (Buzby & Perry, 2000). Higher water temperatures
have been shown to affect litter decomposition in labora-
tory experiments, both directly, by promoting leaching of
soluble compounds (Chergui & Pattee, 1990), and indir-
ectly, by stimulating fragmentation and consumption by
selected invertebrate shredders (Gonza
´
lez & Grac¸a, 2003;
Azevedo-Pereira et al., 2006) and by enhancing microbial
activity (Carpenter & Adams, 1979). High temperatures
also stimulate production of fungal assemblages asso-
ciated with leaves (Suberkropp & Weyers, 1996), and
growth and sporulation by some species of aquatic hy-
phomycetes cultivated individually (Koske & Duncan,
1974; Grac¸a & Ferreira, 1995; Chauvet & Suberkropp,
1998; Rajashekar & Kaveriappa, 2000; Dang et al., 2009),
which can result in accelerated decomposition rates. How-
ever, species have a temperature tolerance range below
and above which their activity is reduced or suppressed.
In face of the predicted increases in water temperature
many species of invertebrates (Quinn et al., 1994) and
fungi (Koske & Duncan, 1974; Rajashekar & Kaveriappa,
2000) may have their activity inhibited, making it difficult
to predict the response of the decomposition process in
streams under a global warming scenario. In addition, the
influence of modified interspecific interactions is largely
unknown (Webster et al., 1976).
Additionally, there are several factors that are asso-
ciated with increased temperature, which might modu-
late its effects on stream biota and processes. One such
factor is increased evapotran spiration, and consequently
increased pollutant and nutrient concentrations in water
bodies (Murdoch et al., 2000). This, associated with in-
creasing needs for water and production of wastewater
by a growing human population, will result in decreased
ecological status of freshwaters, i.e. decreased ability to
provide ecosystem services as water purification. The
effects of dissolved nutrients on the decomposition of
submerged litter and associated biological activity have
been well studied. Nutrient enrichment generally stimu-
lates litter decomposition and associated biota (Elwood
et al., 1981; Suberkropp & Chauvet, 1995; Rosemond et al.,
2002; Gulis & Suberkropp, 2003a; Niyogi et al., 2003;
Pascoal et al., 2003; Gulis et al., 2004, 2006; Ferreira et al.,
2006b). In streams where a given inorganic nutrient (i.e. N
or P) is not limiting, further increases in its concentration
in water may however not enhance litter decomposition
or activity of associated microbes (Grattan & Suberkropp,
2001; Abelho & Grac¸a, 2006; Baldy et al., 2007). Also, when
increases in dissolved nutrients occur simultaneously
with variation in other factors (e.g. increases in sedimen-
tation, decreases in dissolved O
2
) their stimulatory effect
might be offset (Pascoal & Ca
´
ssio, 2004). Inhibition
of decomposition rates might occur when increases in
dissolved nutrients reach toxic levels (Lecerf et al., 2006).
The effect of interactions between increased water
temperature and nutrient concentrations on litter de-
composition and associated biological activity remains
to be assessed. Nonetheless, evidence from other sys-
tems suggests that the effects of factors associated with
global changes acting in combination might not be
predictable from the effects of factors considered indi-
vidually (Rozema et al., 1997; Hoffman et al., 2003;
Przeslawski et al., 2005). In this study, we addressed
the effects of water temperature (three levels) and
nutrient concentrations (two levels) on the decomposi-
tion of alder leaves, and associated aquatic hyphomy-
cete biomass, activity and assemblage structure in
simulated stream microcosms, in a complete factorial
design. As the rate of biological processes is dependent
on temperature, since they are basically enzyme driven
(Brown et al., 2004), and fungi can retrieve nutrients
from both the substrate and the water (Suberkropp,
1998), we predict that fungal biomass and activity, and
consequently decomposition rates, will increase with
temperature and nutrient concentrations. Given that
aquatic hyphomycetes exhibit temperature and nutri-
ents optima (Gulis & Suberkropp, 2003a; Pascoal et al.,
2005a; Ferreira et al., 2006b; Artigas et al., 2008), changes
in the structure of the assemblages are anticipated.
Materials and methods
Fungal
species assemblage
An assemblage of six species of aquatic hyphomycetes was
used,
as representative of fungal diversity found on a single
leaf decomposing in natural streams (Ba
¨
rlocher, 1992). Strains
were isolated from single conidia trapped in naturally occur-
ring foam, or released from leaf accumulations, collected from
a lowland stream in central Portugal [Ribeira do Bota
˜
o;
40118
0
22
00
N, 8123
0
37
00
W; Articulospora tetracladia Ingold
(ARTE)], a Mediterranean stream in the French Pyrenees
[Maureillas; 42128
0
18
00
N, 2 147
0
57
00
E; Clavariopsis aquatica de
Wildeman (CLAQ), Flagellospora curvula Ingold (FLCU) and
Tetracladium marchalianum de Wildeman (TEMA)] and a tem-
perate mountain stream in the Massif Central, SW France
[Oreval; 43126
0
19
00
N, 215
0
41
00
E; Heliscus lugdunensis Saccardo
& The
´
rry (HELU) and Tumularia aquatica (Ingold) Descals &
Marvanova
´
(TUAQ)]. Growing colonies were kept at 15 1C, in
9 cm diameter Petri dishes with 10 mL of growth medium
(10 g malt and 20 g agar per liter of sterile distilled water), until
they were used to induce conidial production. Conidial inocu-
lations (o1-day-old) were produced at 15 1C, by incubation of
agar plugs taken from the leading edge of 7- to 14-day-old
colonies (either grown from an individual agar plug or from
conidia spread over a Petri dish) in 25 mL of nutrient solution
(75.5 mg CaCl
2
, 10 mg MgSO
4
7H
2
O, 0.5 g 3-morpholinopro-
panesulfonic acid (MOPS), 0.55 mg K
2
HPO
4
and 100 mg KNO
3
per liter of sterile distilled water), on an orbital shaker
(100 rpm). An aliquot of each specific conidial suspension,
based on conidial numbers, was used to make a combined
conidial suspension to inoculate each microcosm.
Microcosms, medium and experimental setup
Alder [Alnus
glutinosa (L.) Gaertner] leaf discs were incubated
in laboratory microcosms designed to simulate stream condi-
tions (Suberkropp, 1991). Each microcosm consisted of a 50 mL
glass chamber aerated from the bottom by a continuous air
flow (80–100 mL min
1
), which creates turbulence and keeps
the leaf discs in permanent agitation. A tap at the bottom
allowed for the aseptic drainage of the chamber and recovery
of the conidial suspension. Fresh medium (40 mL) was added
to microcosms through the open top which was otherwise
closed with a glass cap. Microcosms were incubated in the
dark at three temperatures (5, 10 and 15 1C), for the duration of
the experiment.
Half of the microcosms at each temperature were filled with a
low nutrient concentrations solution (low NP level treatment)
and half with a high nutrient concentrations solution (high NP
level treatment). The low nutrient solution was composed of
75.5 mg CaCl
2
, 10 mg MgSO
4
7H
2
O, 0.5 g MOPS, 0.055 mg
K
2
HPO
4
and 10 mg KNO
3
per liter of sterile distilled water
(5 0.01 mg PO
4
-P L
1
and 1.39mg NO
3
-N L
1
), while the high
nutrient solution was amended with 0.55 mg K
2
HPO
4
and
100 mg KNO
3
(5 0.10 mg PO
4
–P L
1
and 13.86 mg NO
3
–N L
1
)
(Dang et al., 2005). The phosphorus limitation in the solutions
(N : P 5 141) was intended to mimic the trophic conditions of
most streams (Grattan & Suberkropp, 2001), although there are
streams in which nitrogen naturally is the limiting nutrient
(Ferreira et al., 2006b). There were 12 replicate microcosms for
each of the six temperature–nutrient treatments.
Alder leaves collected just after abscission on December 30,
2006 at Gibel, Midi-Pyrenees, France (43117
0
35
00
N, 1140
0
51
00
E),
and
dried at room temperature, were moistened with distilled
water and left to rehydrate overnight. Leaf discs were cut with
a 12 mm diameter cork borer and oven dried (65 1 C) during 3
days. Batches of 20 leaf discs were frozen overnight at 20 1C,
lyophilized (20 h), weighed ( 0.1 mg) to determine initial dry
mass (DM), placed inside glass tubes with an aliquot of
distilled water and autoclaved (20 min at 121 1C). Seven
batches of 20 leaf discs were given the same treatment and
were used to create a correction factor for mass loss due to
leaching during sterilization. Sterilized microcosm (30 min at
121 1C) were filled with 40 mL of the low or high nutrient
solutions, received the corresponding leaf discs, were distrib-
uted by three incubator chambers (5, 10 and 15 1C), and
aerated for 24 h. Nutrient solutions were changed, the micro-
cosms inoculated with a total of 4800 conidia equally parti-
tioned among all species of the aquatic hyphomycete
assemblage, aerated for 10 min, left to rest for ca. 2 h, and
aerated again. The nutrient solutions were replaced after 24 h
and then every 3 days for the duration of the experiment (43
days). All manipulations of microcosms took place in a flow
cabinet.
Fungal sporulation
Each of the 14 times the nutrient solutions were changed, the
co
nidial suspensions from three microcosms (the same over
time) from each treatment were stored into 50 mL centrifuge
tubes, sample volume was adjusted to 42 mL with distilled
water and preserved with 3 mL of 35% formalin. Additionally,
after 10, 16, 28 and 43 days, three microcosms from each
treatment were sacrificed, and the conidial suspensions pre-
served as above. When preparing filters for conidial counting
and identification, 150 mL of polyethylene glycol p-(1,1,3,3-tetra-
methylbutyl)-phenyl ether (Triton X-100, 0.5%) were added to
the suspension, mixed with a magnetic stirring bar, to ensure a
uniform distribution of conidia, and an aliquot of the suspension
was filtered (Millipore SMWP, pore size 5 mm). Filters were
stained with 0.08% trypan blue in 60% lactic acid, and spores
were identified and counted under a compound microscope at
320 (Grac¸a et al., 2005). For sacrificed microcosms, sporulation
rates were expressed as number of conidia mg
1
DM day
1
.
For microcosms repeatedly sampled throughout time, results
were expressed as number of conidia microcosm
1
and as
mg conidia microcosm
1
. The total conidial mass at each sam-
pling date was calculated by multiplying the number of conidia
from each species by the average mass of individual conidia
obtained from the literature (Chauvet & Suberkropp, 1998) or
calculated from biovolume data (Ba
¨
rlocher & Schweizer, 1983)
assuming a 70% water content. Cumulative conidial mass
production over time was calculated by summing up the total
conidial mass at the preceding sampling dates.
Oxygen consumption
A subset of five leaf discs from each sacrificed microcosm was
used to determine fungal oxygen consumption rates using a
closed six-channel dissolved oxygen measuring system (Strath-
kelvin 928 System, North Lanarkshire, Scotland) connected to a
computer. The oxygen electrodes were calibrated against a 4%
sodium sulfite solution prepared immediately before use (0%
O
2
), and a 100% O
2
saturated low or high nutrient solution at the
target temperature. Leaf discs were incubated in 3 mL volume
chambers filled with 100% O
2
saturated low or high nutrient
solution, homogenized with a magnetic stirring bar, kept at the
target temperature of 5, 10 or 15 1C by circulation of water
originating from a temperature-controlled water bath. Addi-
tional chambers without leaf discs were used as controls. After
1 h trial, leaf discs were enclosed in small sterile zip lock bags
and promptly frozen at 20 1C for later DM determination and
ergosterol extraction. Oxygen consumption rates were deter-
mined by the difference in the oxygen concentration in the
sample and the control over a 20 min interval and corrected
for the chamber’s volume, time and discs mass. Results were
expressed as mg O
2
g
1
DM h
1
.
Mass loss and mycelial biomass
The remaining 15 leaf discs from each microcosm were enclosed
in small sterile zip lock bags and promptly frozen at 20 1C. All
20 leaf discs from each microcosm were combined, lyophilized,
![Table 3 Summary table for repeated measures ANOVAs performed on conidial production per microcosm [log(x1 1) transformed] and sporulating species richness, associated with alder discs incubated in microcosms at three temperatures and two NP levels for 43 days](/figures/table-3-summary-table-for-repeated-measures-anovas-performed-3sdnk94y.webp)
![Table 2 Summary table for three-way ANOVAs performed on fungal oxygen consumption, mycelial biomass [log(x1 1) transformed], and sporulation rates [log(x1 1) transformed], associated with alder discs incubated in microcosms at three temperatures and two NP levels for 43 days](/figures/table-2-summary-table-for-three-way-anovas-performed-on-1mn72ln3.webp)