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Research

Elevated atmospheric CO2 alters root symbiont


Blackwell Science Ltd

community structure in forest trees


Petra M. A. Fransson, Andrew F. S. Taylor and Roger D. Finlay
Department of Forest Mycology and Pathology, SLU, Box 7026, S-75007 Uppsala, Sweden

Summary

Author for correspondence: • Changes in below-ground ectomycorrhizal (ECM) community structure in


Petra M. A. Fransson response to elevated CO2 and balanced nutrient addition were investigated in a
Tel: +46 18 672797 37-yr-old Picea abies forest.
Fax: +46 18 309245
Email: petra.fransson@mykopat.slu.se
• Trees in whole-tree chambers were exposed to factorial combinations of ambient/
elevated CO2 (700 ppm) and fertilization (+/–). ECM fungal community structure
Received: 5 July 2001
was determined in 1997 and 2000 using a combination of morphotyping and
Accepted: 4 September 2001
molecular analyses. Samples were taken both from chambers and from reference
trees receiving the same fertilization treatments but without chambers.
• Significant effects on ECM community structure were found in response to ele-
vated CO2. Neither elevated CO2 nor fertilization altered species richness; however,
there was considerable variation among samples, which may have masked
treatment effects on individual species. After 3 yr, the effects of elevated CO2 on
community composition were of the same magnitude as those seen after 15 yr of
fertilization treatment.
• Our results show that increasing atmospheric CO2 concentrations affect the
community structure of root symbionts colonizing forest trees. The potential effects
of altered ECM community structure on allocation and turnover of carbon and
nutrients within forest ecosystems are discussed.

Key words: ectomycorrhizal fungi, community structure, elevated CO2, carbon,


ordination analysis.

© New Phytologist (2001) 152: 431–442

sequestration and emission of carbon. Boreal forest trees rely


Introduction on ectomycorrhizal (ECM) fungi for their nutrient uptake,
Within European forests there is a fine balance between the and almost 100% of forest tree short roots are colonized by
quantity of carbon (C) fixed via photosynthesis and that ECM fungi (Taylor et al., 2000). The fungi, in turn, depend
released through respiration (Valentini et al., 2000). The strongly on current assimilates from their plant hosts (Högberg
result of these two processes determines the net exchange of C et al., 2001), and up to 20% of the photosynthetically fixed
between the terrestrial biosphere and the atmosphere and carbon may be allocated by the tree to the fungus (Finlay &
whether forests act as a source or a sink for CO2. The balance Söderström, 1992). Högberg et al. (2001) found a large and
is affected by daily, seasonal and yearly variations in climatic almost immediate decrease in root associated respiration,
conditions (Hanson et al., 1993; Kirschbaum, 1995; which constituted 54% of the total soil respiration, in
Goulden et al., 1998; Lindroth et al., 1998), and by changes combination with a large decrease in ECM fruit body
in land use patterns (Schimel, 1995; Falkowski et al., 2000). production, following girdling of a pine stand illustrating the
Valentini et al. (2000) showed that total ecosystem respiration, importance of current assimilates to ECM fungi.
most notably that of roots and soil microorganisms, was the Individual ECM fungal species are likely to differ in how
main determinant of the C balance of a forest. Among soil they allocate plant derived C between growth, respiration and
microorganisms, symbiotic mycorrhizal fungi are of particular exudation. The allocation patterns can in turn be affected by
significance since they have a direct influence on both the the actual supply of C from the host plant to the fungus.

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Increased levels of CO2 in the atmosphere are known to associated with large forest trees growing under field conditions.
increase plant photosynthesis and subsequent C supply into In the present study we investigated changes in the ECM fun-
the soil (O’Neill, 1994; Rey & Jarvis, 1997; Ceulemans et al., gal community structure in a 37-yr-old Norway spruce forest
1999). Whether this increased supply of C below-ground after factorial combinations of elevated CO2 and balanced
actually leads to significant, long-term net carbon sequestra- nutrient addition.
tion in forest soils is however, questioned (Schlesinger &
Lichter, 2001). Laboratory studies of tree seedlings have
demonstrated that Suillus bovinus and Laccaria bicolor
Materials and Methods
respond differently to elevated CO2 concentrations in the
Study site
way they partition assimilates between fungal biomass and
respiration (Gorissen & Kuyper, 2000). These authors also The study was conducted at the Flakaliden field site, northern
showed that the fungi differed in their capacity to process and Sweden (64°07′ N; 19°27′ E; alt. 310 m above sea level (asl)).
transfer N to their tree hosts. In general, diversity in physio- The site was planted with Picea abies [L.] Karst. after clear-
logy and function – both among and within different fungal felling in 1963. A fertilization experiment was started in 1987
species – might be large (Cairney, 1999), but the dominance in which plots were supplied with a balanced solution of macro-
of a few species is commonly seen in investigations of below- and micronutrients on a daily basis throughout the growing
ground ECM community composition (Gardes & Bruns, season. Since the start of the experiment the fertilization
1996; Erland et al., 1999; Peter et al., 2001). Changes within treatment has included a total of 1125 kg N ha–1, added as
these dominant species thus have a potentially large influence both ammonium and nitrate. In 1996, 12 whole tree chambers
on any physiological response changing community struc- were installed around trees. Six of the chambers were placed
ture. The results of Gorissen & Kuyper (2000) also underline in an unfertilized plot and six were placed in a fertlized plot.
the importance of nutritional effects and their likely interac- The CO2-treatment started in 1998 and continued for 3 yr.
tions with C allocation. Changed ECM community structure One half of the enclosed trees on each plot received ambient
may thus contribute to changes in the C balance of boreal levels of CO2 (350 ppm), the other half received elevated levels
forest ecosystems. It is therefore important to establish how of CO2 (700 ppm). The enclosed trees are hereafter referred
individual species respond to elevated levels of CO2 under to as ambient trees and elevated trees. Three reference trees
field conditions. (without chambers) on each plot were also included in this
There are few published studies of the effects of elevated analysis. The enclosed trees were irrigated via a tubing system
CO2 on ECM mycorrhizal community structure. Rey & with amounts of water equivalent to those intercepted by the
Jarvis (1997) found indications that the mycorrhizal species same area outside the chambers. The mean distance between
composition based on morphology (morphotypes) shifted individual trees in the plots was c. 3 m. Two of the chambers
towards those species characteristic of later successional stages (ambient tree no. 2 in the unfertilized plot and ambient tree
in young birch seedlings (Betula pendula) after growing in no. 2 in the fertilized plot) were moved to new trees in 1997 and
open-top chambers under elevated levels of CO2 for 4.5 yr. 1998 as the trees died as a result of aphid and fungal attacks.
They interpreted this as an acceleration of tree ontogeny,
which may lead to the trees supporting ECM fungal species
Sampling and identification of ectomycorrhizal fungi
with a higher carbon demand. Godbold et al. (1997) trans-
ferred paper birch (Betula papyrifera) and Eastern white pine In late August in 1997, before the CO2-treatment started, and
(Pinus strobus) saplings from a forest to growth chambers and at the same time in 2000, before the final harvest of the
observed, after 24 wk of elevated CO2 exposure, an increase chamber trees, five soil cores (2.8 cm diameter) were taken
in the proportion of morphotypes that produced rhizo- from the organic soil layer beneath each of the chamber trees
morphs and large amounts of extraradical mycelium. Rygie- (6 + 6) and the reference trees (3 + 3). Forty-five soil cores
wicz et al. (2000) planted 2-yr-old Douglas fir (Pseudotsuga were thus taken from each plot, giving a total of 90 soil cores.
menziesii ) seedlings in chambers, and after 4 yr of elevated Soil samples were placed in separate plastic bags and
CO2 exposure they found small overall effects on ECM fungal maintained at 4°C, until examined. The depth of the organic
diversity, based on gross morphological traits. The authors did layer in each core was measured before soaking the sample
find an increase in ECM root tip proliferation during sum- in water for 30 min. Live root tips were then extracted by wet
mer. Markkola et al. (1996) were unable to detect any signi- sieving using a combination of 1.0 mm and 500 µm sieves.
ficant changes in ECM morphotype community structure Root tips were examined and classified into mycorrhizal
growing on roots of Pinus sylvestris as a result of CO2 enrich- morphotypes, using macroscopic and microscopic features
ment. The authors used pine seedlings, which were inculated (Agerer, 1986–98). The term morphotype is used here to
with Piloderma croceum, and then grown in pots with natural designate a recognizable group of mycorrhizal root tips.
mor humus for up to 11 months. As far as we know, there Morphotypes were either identified to species or genus level,
are no studies of the effects of elevated CO2 on ECM fungi or unidentified. The unidentified morphotypes were given

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numbers as they appeared in the processing. The total were transformed (log10) before analysis, and the analyses
abundance (total number of mycorrhizal root tips per core) were performed with CANOCO 4.0 (ter Braak & Smilauer,
and relative abundance (number of each morphotype/total 1998). Monte Carlo permutation tests (n = 199) were
no. of living mycorrhizal root tips) of each mycorrhizal type performed to test the significance of the relationship between
were recorded. Nonmycorrhizal root tips were also recorded. morphotype data and treatments. In the CCA ordination
Subsamples of morphotypes were kept in the freezer for diagram vectors are included to make the presentation clearer,
internal transcribed spacer-restriction fragment length poly- although these are not strictly applicable since the environ-
morphism (ITS-RFLP) analysis as a supplement to the mental variables are not continuous.
morphotyping to confirm consistency within and between Possible treatment effects of elevated CO2 and chambers
single morphotypes. DNA was extracted from the mycorrhizal on ECM morphotype richness, total root tip numbers and
root tips according to Gardes & Bruns (1993), excluding the individual ECM morphotype abundance (total) were ana-
initial freeze-thawing step. The ITS region of the rDNA was lysed for each plot using one-way ANOVA. Morphotype
amplified by PCR following the modified protocol by richness was tested using both the numbers of identified
Henrion et al. (1994). PCR was performed using High Fidelity morphotypes, that is taxa identified to species or genus level,
DNA polymerase and High Fidelity buffer (Roche). The and the total numbers of morphotypes, that is both identified
reaction mix had final concentrations of each nucleotide at and unidentified taxa. The data from 2000 were tested for
0.2 mM, each primer at 0.3 µM, polymerase at 0.026 µ µl–1, both the elevated CO2 and the chamber factors, while the
Mg2Cl at 3.1 mM. DNA template was added as 25% of the data from 1997 were only tested for the chamber factor.
final reaction volume. Cycling parameters were modified to To test whether the effects of CO2 and chamber treatments
1 cycle at 94°C for 3 min followed by 35 cycles of 30 s at were the same for both fertlized and unfertilized plots, that is
94°C, 45 s at 50°C, 1 min at 72°C ending with 1 cycle of 7 min whether the possible effects of CO2/chamber and fertilization
at 72°C. The primers used were ITS1-ITS4 as described by treatments upon individual components of the community
White et al. (1990). PCR products were separated on a 1% structure interact, a two-way ANOVA was performed. Since
agarose gel in 0.5 X TBE-buffer at 4.7 Vcm–1, stained with the fertilization treatment was not replicated, the significance
ethidium bromide and visualized by UV-light. The amplified levels for the possible effects of the fertilization treatment
DNA was digested using the restriction enzymes Hinf 1, Mbo1 alone upon individual components cannot be used. It is however,
and Taq1 (Promega Corporation, Madison, WI, USA), for 2.5 h statistically correct to test the interaction and this part of the
in 37°C (Hinf 1, Mbo1) and 65°C (Taq1). Digestion products results from the two-way ANOVA is reported (B. Vergerfors-
were separated on a 2.3% Metaphor gel in 0.5 X TBE-buffer Persson, pers. comm.). The significance levels for the possible
at 4.7 Vcm–1, stained and visualized as previously. Fragment effects of CO2/chamber treatment upon individual com-
lengths were estimated and band patterns analysed with ponents are not reported since they were already tested
Taxotron® software system (Institute Pasteur, Paris, France). through one-way ANOVA as described in the above section.
Mean values ± standard errors for total numbers of morpho-
types, number of identified and unidentified morphotypes
Statistical analysis
and levels of colonization are reported.
To investigate community structure and possible treatment
effects, Detrended correspondence analysis (DCA) and
Canonical correspondence analysis (CCA) were carried out
Results
on the total numbers of mycorrhizal root tips in each
Ordination analyses of community structure
morphotype (ter Braak & Smilauer, 1998). DCA represents
an indirect gradient analysis, based on the morphotype The Canonical correspondence analysis showed significant
abundance data, that examines the total variation in the effects of elevated CO2 on ECM fungal community structure
community samples. CCA represents a direct gradient in the year 2000 (Fig. 1a). 20.9% of the variation in the
analysis, where the morphotype abundance data is explained species data set can be attributed to the fertilization-,
by environmental variables and part of the total variation chamber- and CO2-treatments. The eigenvalues for CCA axes
in the community samples is thus examined. In this case 1 and 2 were 0.371 and 0.303. The first two CCA axes
the environmental variables are the treatments, that is no together display c. 15% of the total variance in the species
fertilization – fertilization, no chamber – chamber and data set, and this part of the variation is shown in Fig. 1(a).
ambient CO2 – elevated CO2. An interyearly comparison of The Monte Carlo permutation tests showed that there were
community structures was made by DCA analysis. The two statistically significant (P = 0.005) relationships between
ambient trees where the chambers were moved between the ECM morphotype composition and environmental variables.
two samplings were excluded from the analysis. Arrows are When the test was repeated for one environmental variable
used in the DCA ordination graph to indicate the shift in at a time, the results were also significant for fertilization
fungal community structure between 1997 and 2000. Data and CO2 effects, but not for the chamber effect.

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Fig. 1 Ordination diagram based on


Canonical correspondence analysis (CCA) of
the ectomycorrhizal (ECM) community in a
37-yr-old Norway spruce stand subject to
elevated CO2 (700 ppm) and optimal
fertilization for 3 and 15 year, respectively.
The diagram shows the part of the variation
within the communities (21%) that can be
attributed to the treatments. Arrow length
indicates the relative importance of
environmental variables. (a) Open symbols,
unfertilized trees; closed symbols, fertilized
trees. Ambient CO2 trees with (open
diamonds) and without (open circles)
chamber and elevated CO2 with chamber
(open diamond within square). (b) Species
scores for ECM taxa that appeared on three
or more of the sampled trees. Abbreviation:
A. byssoides, Amphinema byssoides;
C. geophilum, Cenococcum geophilum;
P. gelatinosa, Piceirhiza gelatinosa;
P. rosa-nigrescens, Piceirhiza rosa-nigrescens;
P. byssinum, Piloderma byssinum;
P. croceum, Piloderma croceum; Russula
spp.; tomentelloid group 1, clamped
tomentelloids; tomentelloid group 2,
unclamped tomentelloids; T. fibrillosa,
Tylospora fibrillosa; Unknown, unknown
basidiomycete.

The positioning of the samples relative to the end point of are grouped near the origin and between the two plot treat-
the vectors in the CCA ordination diagram relates individual ment clusters, and can thus not be related to any of the treat-
trees to different treatments (Fig. 1a). The ordination diagram ments. Piloderma byssinum seemed to be negatively affected
shows the elevated CO2 trees clustered together in the upper by the fertilization treatment, and the unknown basidio-
left part. For the unfertilized plot, elevated CO2 trees are mycete appeared to be favoured by the CO2 treatment.
clearly separated from the ambient CO2 trees and reference In 1997, before the CO2 treatment started, the CCA ana-
trees. The fertilized plot shows a similar pattern, although lysis showed a clear separation between the unfertilized and
the fungal communities on trees receiving elevated CO2 fertilized plots, and 14.4% of the total variance of the species
levels were not so clearly separated from ambient and refer- could be explained by the fertilization and chamber treat-
ence trees. ments (figure not shown). The significance of the treatments
Eleven taxa occurred on three or more of the trees exam- was tested as previously and the ECM morphotype distribu-
ined and the CCA species scores are plotted together with the tion was significantly (P = 0.01) related to fertilization and
environmental variables in Fig. 1(b). Of these, Amphinema chamber treatments. Variation in community structure in the
byssoides, Cenococuim geophilum, tomentelloid group 1 and unfertilized plot was larger compared to the variation in the
Tylospora fibrillosa, all seemed to be favoured by the fertiliza- fertilized plot. Eigenvalues for CCA axes 1 and 2 were 0.50
tion treatment. Piloderma croceum and tomentelloid group 2 and 0.38, respectively. Seven taxa in 1997 occurred on three

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Fig. 2 Ordination diagram based on


Detrended correspondence analysis (DCA) of
the ectomycorrhizal (ECM) community in a
37-yr-old Norway spruce stand, showing the
interyearly variation between 1997 and
2000. The Norway spruce stand was subject
to elevated CO2 and optimal fertilization
treatments. The direction of arrows in the
ordination diagram indicates the shift in
community structure for individual trees
between the two years, and long arrows
indicates larger differences than small arrows.
(a) All trees in the unfertilized plot remained
separated from the trees in the fertilized plot,
with one exception. The fungal community
structures of elevated trees, that are marked
in red, seem to become more similar to
those of the fertilized plot (b). The division
between the unfertilized plot and fertilized
plot is indicated by the dashed line. Red/blue,
elevated trees in unfertilized/fertilized
plot; orange/turqoise, ambient trees in
unfertilized/fertilized plot; black,
reference trees.

or more of the trees. Cenococcum geophilum, T. fibrillosa and treatment in the fertilized plot. All trees in the fertilized plot
tomentelloid group 1 were favoured by the fertilization treat- are grouped together in the DCA diagram (Fig. 2b). Despite
ment, while Piloderma species and Inocybe spp. were mainly the variation in ECM community structure between years,
associated with the trees in the unfertilized plot. especially for the reference trees in the unfertilized plot,
ambient trees and reference trees in the unfertilized plot
remain separated from the fertilized plot. In general the
Inter-yearly variation in community composition
variation both within and between years was considerable.
A comparison of the ECM community structure between
years was made using DCA analysis. The results showed that
Effects of elevated CO2 and fertilization treatment on
the composition of fungi colonizing tree roots in the
individual components of the community structure
unfertilized plot remained different from that in the fertilized
plot (Fig. 2a), with an exception. The elevated CO2 treatment In 1997, a total of 58 ECM morphotypes was found, 14
seemed to drive the community structure in the unfertilized which were identified to species or genus level and 44 that
plot towards that in the fertilized plot (Fig. 2a). This is were unknown (Table 1). In 2000, a total of 42 morphotypes
explained by changes in the relative abundance of some taxa, was found, 14 were identified and 27 were unknown types
but the specific changes, that is which of the fungal taxa (Table 2). For all investigated trees in 2000, the number of
are responsible for the shift in community structure, are not identified ECM morphotypes per tree (5.3 ± 0.5) exceeded
easily distinguishable. In this analysis no apparent shift the number of unknown morphotypes (1.7 ± 0.4). In 1997,
in community structure was seen as a result of the CO2 the number of unidentified morphotypes was higher

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Table 1 Relative abundance of ectomycorrhizal (ECM) morphotypes, sampled in 1997 from Norway spruce plots receiving no treatment or fertilization. Individual trees were either covered with
whole tree chambers, not recieving CO2 treatment in 1997, or free-standing reference trees receiving no additional treatment

Control plot Fertilization plot

Chamber trees Reference trees Chamber trees Reference trees


Tree treatment
Replicate tree no. 1 21 3 4 5 6 1 2 3 1 22 3 4 5 6 1 2 3

Amphinema sp. 3.8 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1.4 0 0


Cenococcum 0 0 8.8 14.3 0 <1 1.8 14.3 1.9 0 3.6 14.9 1.2 7.5 12.1 19.6 8.6 14.7
Cortinarius spp. 0 0 3.0 0 64.2 0 25.5 9.3 <1 0 0 0 0 0 0 0 1.7 0
Dermocybe spp. 0 0 0 0 0 <1 0 1.8 0 0 0 0 0 0 0 0 3.4 2.0
L. rufus 0 25.0 0 0 0 0 34.1 0 0 0 0 0 0 0 0 0 0 0
Lactarius spp. 88.5 0 0 0 0 0 <1 0 0 <1 0 40.4 0 0 11.0 0 0 0
Piceirhiza bicolorata 0 0 0 0 0 0 8.1 0 0 0 0 0 0 0 0 1.4 0 0
P. rosa-nigrescens 0 0 0 0 0 0 0 3.3 0 0 0 0 0 0 0 0 0 0
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P. byssinum 0 0 0 11.1 0 0 0 5.0 24.5 0 0 0 0 0 0 8.7 0 0


P. croceum 0 0 10.0 0 1.1 0 4.1 9.0 9.2 0 0 10.6 0 6.8 21.1 0 0 0
Russula spp. 0 75.0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Tomentelloid 0 0 7.1 0 0 0 0 0 0 85.2 0 4.2 10.8 17.3 5.3 18.1 0 7.3
Tylospora sp. 0 0 0 0 10.5 0 0 0 3.2 4.9 88.0 6.4 71.1 48.1 0 34.1 67.4 43.3
Unknown tips 7.7 0 14.0 12.7 21.1 68.9 11.6 51.0 49.6 0 6.0 6.4 0 10.5 45.8 16.7 18.9 10.7
Nonmyco. tips 0 0 57.1 61.9 3.1 29.9 14.3 6.3 11.1 9.6 2.4 17.1 16.9 9.8 4.7 0 0 22.0
Total no. tips 78 56 170 63 95 241 440 335 216 312 167 47 83 133 190 138 175 150
Types id. 2 2 5 2 3 2 5 6 6 3 2 4 3 4 4 6 4 4
Types unid. 1 0 1 3 2 2 5 1 8 0 2 2 0 5 4 3 3 6
Colonization (%) 100 100 42.9 38.1 96.9 70.1 85.7 93.7 88.9 90.4 97.6 82.9 83.1 90.2 95.3 100 100 78.0

Abundances are expressed as percentage of root tips examined. 1,2The chamber was moved to a new tree in 1997 and 1998, respectively.
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Table 2 Relative abundance of ectomycorrhizal (ECM) morphotypes, sampled in 2000 from Norway spruce plots receiving no treatment or fertilization. Individual trees were treated with ambient
or elevated levels of CO2 in whole tree chambers, or were free-standing reference trees recieving no additional treatment

Control plot Fertilization plot

Ambient Elevated Reference trees Ambient Elevated Reference trees


Tree treatment
Replicate tree no. 1 21 3 1 2 3 1 2 3 1 22 3 1 2 3 1 2 3

Amphinema sp. 0 0 0 0 0 4.5 0 6.0 19.9 26.9 1.4 0 0 8.1 0 1.0 6.5 8.7
Cenococcum 0 0 11.1 0 3.7 10.5 3.7 13.1 3.7 20.1 0 18.0 35.0 18.3 10.2 31.4 8.8 21.4
Cortinarius spp. 69.2 0 10.2 0 0 10.9 5.6 6.1 25.0 1.5 0 0 0 0 0 1.0 0 0
Dermocybe cf. 0 0 0 0 0 0 0 6.6 0 0 0 0 0 0 0 0 0 <1
cinnamomeus
Dermocybe spp. 0 0 0 0 0 1.4 <1 0 0 0 0 0 0 0 0 0 0 0
L. deterrimus 0 0 0 0 0 0 0 0 3.2 0 0 0 0 0 0 0 0 0
Lactarius spp. 0 0 0 0 0 25.0 0 0 0 0 0 0 0 0 1.3 0 0 0
Piceirhiza gelatinosa 0 0 0 0 7.4 1.4 0 0 0 0 0 14.6 10.0 0 31.5 6.9 <1 2.4
P. rosa-nigrescens 0 0 6.5 0 0 0 0 1.6 2.8 0 21.4 0 0 0 0 0 0 0
P. byssinum 0 80.0 0 0 5.1 0 10.0 0 3.7 0 0 0 0 8.6 0 0 0 0
P. croceum 11.6 0 0 20.5 4.4 0 20.7 0 0 0 0 12.6 0 0 43.4 0 0 21.0
Russula spp. 0 0 0 0 0 <1 <1 0 5.1 0 0 0 0 0 0 0 0 0
Tomentelloid 0 0 20.4 56.8 0 0 0 3.8 0 0 6.9 <1 0 1.1 1.3 0 <1 1.6
Tylospora sp. 15.4 0 50.0 0 4.4 4.1 51.8 12.0 25.9 42.3 60.7 45.1 40.0 40.9 8.8 20.5 51.8 37.7
Unknown 3.8 20.0 1.8 9.1 74.3 34.5 7.6 49.7 10.7 9.2 9.6 8.7 15.0 21.5 3.5 38.2 17.6 6.8
Non-myco. tips 0 0 0 13.6 <1 7.3 0 1.1 0 0 0 <1 0 1.5 0 1.0 14.1 0
Totals (no. tips) 26 5 108 44 136 220 301 183 216 130 145 206 20 186 226 102 170 252
Types is 3 1 6 3 5 8 7 8 8 4 4 6 3 5 7 5 5 8
Types unid 1 0 1 1 5 6 0 2 2 2 2 2 0 3 1 1 1 1
Colonization (%) 100 100 100 86.4 100 92.7 100 98.9 100 100 100 100 100 98.5 100 99.0 85.9 100

Abundances expressed as percentage of root tips examined. CO2 concentrations: ambient (350 p.p.m) and elevated ( 700 p.p.m).
1,2
The chamber was moved to a new tree in 1997 and 1998, respectively.

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(2.7 ± 0.5), and the number of identified morphotypes elevated CO2 on ECM community structure is important
lower (3.7 ± 0.3). Between 1 and 14 morphotypes were found to establish since individual ECM fungal species may differ
per tree with a mean value of 6.4 ± 0.8 morphotypes per greatly in both physiology and function (Cairney, 1999). In
tree in 1997 and 7.1 ± 3.0 in 2000. ITS-RFLP analyses our study the ECM community structure of Norway spruce
showed satisfactory consistancy between and among tested trees changed in response to elevated levels of CO2. Canonical
morphotypes. correspondence analysis was performed in order to compare
The level of colonization was lower and more variable in the response of the fungal community as well as single ECM
1997, with a mean of 85.2% ± 4.3, compared to 2000 when taxa to elevated CO2 and fertilization. Almost 21% of the
the mean colonization level was 97.9% ± 1.1. ECM morpho- total variation in the species data set could be attributed to
type richness and total root tip numbers were generally not the CO2 and fertilization treatments, reflecting the fact that
affected by the treatments, with a few exceptions (see below). the fungal community composition was affected by other
In Tables 1 and 2, the relative abundances of Norway spruce environmental variables than the present treatments. Using
root tips colonized by each ECM morphotype are listed for CCA analysis, this is as large a part of the total variation one
each tree. Unknown morphotypes were assigned numbers as can normally expect to explain (ter Braak & Verdonschot,
they occurred during processing, and the category called 1995). High variablility among individual samples was
tomentelloid fungi includes both Tomentella and Pseudoto- obvious here and is a general feature of ECM community data
mentella species, that is both clamped and unclamped tomen- (Horton & Bruns, 2001), but despite this a significant effect
telloids. No individual ECM morphotype, except P. croceum of the CO2 treatment could be detected. Samples from
and C. geophilum (see below), showed a significant response to individual trees and chambers may have contained tree roots
the different treatments in either of the years. This apparent from other, nearby trees, but we consider the samples to have
lack of response by individual ECM morphotypes to treat- been representative of the conditions experienced by the trees.
ments was mainly due to the high variability within the data. Changes in ECM community structure due to elevated CO2
Significantly (F1,7 = 6.27, P = 0.04) more nonmycorrhizal levels have been reported previously for seedlings and saplings
root tips and unidentified ECM morphotypes (F1,7 = 7.00, (Godbold et al., 1997; Rey & Jarvis, 1997; Rygiewicz et al.,
P = 0.03) were found on the elevated CO2 trees compared 2000), but not for large forest trees grown under field
to ambient trees and reference trees in the unfertilized plot in conditions as far as we are aware.
2000. No significant effects of elevated CO2 were detected for The shift in community structure in the present study was
the fertilized plot. mainly due to a change in abundance of a few common spe-
Significantly fewer root tips were found in the chambers in cies. Similar responses were reported by Godbold et al. (1997)
the unfertilized plot both in 1997 (F1,7 = 12.25, P = 0.01) and for paper birch saplings exposed to elevated levels of CO2,
in 2000 (F1,7 = 7.17, P = 0.03). Significantly fewer identified where the frequencies of dominant morphotypes changed
ECM morphotypes (F1,7 = 15.75, P = 0.005) and fewer Pilo- significantly. Responses to elevated CO2 in the present study
derma croceum root tips (F1,7 = 16.99, P = 0.004) were found were stronger in the unfertilized plot, compared with the
in the chambers compared to reference trees in the unferti- fertilized plot, as shown in Fig. 1. This was possibly due to the
lized plot in 1997. Significantly fewer Cenococcum geophilum same ECM morphotypes responding in a similar way both to
root tips (F1,7 = 6.08, P = 0.04) were found in the chambers elevated CO2 and fertilization, thus making the actual effect
in the fertilized plot in 1997. of CO2 on community composition in the fertilized plot less
pronounced. Fifteen years of fertilization may already have
shifted the community in the fertilized plot so that the CO2
Interactions between elevated CO2 and fertilization
treatment had little effect once started. The DCA ordination
treatments
diagram of interyearly differences in ECM community structure
In 1997, significant interactions were found between the showed that morphotype composition for two out of three
fertilization treatment and the chamber treatment for the elevated trees in the unfertilisated plot became more similar to
total number of root tips (F3,14 = 6.69, P = 0.02) and for root those found in the fertilized plot, indicating such a response.
tips colonized by P. croceum (F3,14 = 10.91, P = 0.005). No In the present study the morphotype richness was high and
significant interactions were found between elevated CO2 this is in accordance with previous reports on below-ground
treatment and fertilization treatment in 2000. ECM diversity (Erland et al., 1999; Rygiewicz et al., 2000;
Peter et al., 2001). Richness was not affected by elevated CO2
in accordance with the studies by Rygiewicz et al. (2000) and
Discussion Godbold et al. (1997). Rey & Jarvis (1997) did not report on
Increased levels of CO2 in the atmosphere are known to affect the total numbers of taxa found, or the abundance of these,
both host plants (Saxe et al., 1998; Ceulemans et al., 1999; in their study of CO2 effects on ECM fungi. In the present
Norby et al., 1999) and mycorrhizal fungi (Hodge, 1996; study, balanced fertilization also had no effect on morphotype
Fitter et al., 2000; Treseder & Allen, 2000). The effects of richness. Different fertilization treatments has earlier been

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shown to either decrease ECM richness (Alexander & Fairley, This has been demonstrated both from different types of pot
1983; Rühling & Tyler, 1991; Peter et al., 2001) or not to studies (O’Neill et al., 1987; Godbold & Berntson, 1997)
affect richness at all (Kårén & Nylund, 1997; Jonsson et al., and from studies conducted in field soil (Tingey et al., 1997),
1999). although Lewis et al. (1994) and Walker et al. (1997) found
Godbold et al. (1997) suggested that birch saplings grown no effect. The degree of ECM root tip colonization in boreal
under elevated CO2 levels could support mycorrhizal species forests, however, is often close to 100% (Taylor et al., 2000)
with a higher C-demand. Twelve morphotypes were found by and in those types of systems elevated CO2 will have no effect
Godbold et al. (1997) on paper birch and Eastern white pine, on colonization levels. The response of seedlings germinated
with an average of six morphotypes per individual birch and grown according to nursery practices or other seminatural
sapling. They observed an increase in the number of mor- growth conditions might be expected to differ from those seen
photypes forming extensive extraradical mycelia and rhizo- with rooted trees in a more undisturbed environment. What
morphs. Increased production of mycelium due to elevated may happen in the field is that the production of fine roots
CO2 has been reported for several different ECM species increases as a result of elevated CO2, fine roots being more
(Ineichen et al., 1995; Rouhier & Read, 1998a) as well as for responsive to CO2 compared to the rest of the root system,
arbuscular mycorrhizal (AM) fungi (Rouhier & Read, 1998b; but it is not clear if this response would persist (Norby et al.,
Sanders et al., 1998). The increase in fungal biomass is sus- 1999). Norby et al. (1999) did not find any support for an
tained by the extra C fixed by and supplied from the host increased root : shoot ratio in their analysis of open-top
plant under enriched CO2 regimes (Godbold et al., 1997). chamber field experiments. In the present study the degree of
However, the turnover of mycelium is likely to be faster com- colonization in 1997 before the CO2 treatment started was
pared to roots, and an increase in the amount of mycelium variable, due to irrigation problems in some of the chambers.
could lead to a faster turnover of C within the plant-fungus Increases in the numbers of nonmycorrhizal root tips can be
system (Fitter et al., 2000). In the present study it was observed after dry periods and subsequent addition of water
impossible to say what happened with the extraradical myce- when the root tips start to proliferate. In the sampling in 2000
lial production. all levels of colonization were high as expected, and in an ear-
Rey & Jarvis (1997) found indications that the mycorrhizal lier study at the same field site high levels of colonization were
species composition in birch shifted towards later successional also observed (Fransson et al., 2000).
stages after CO2 treatment, and they interpreted this as an Elevated CO2 affected few individual components of the
acceleration of tree ontogeny. This may lead to the trees sup- ECM community, only nonmycorrhizal root tips and uniden-
porting ECM fungal species with a higher carbon demand, in tified ECM morphtypes in the unfertilized plot increased sig-
accordance with the conclusions of Godbold et al. (1997). nificantly with CO2 treatment. The high variablility among
Rey & Jarvis (1997) also reported that trees grown in elevated samples may have masked significant effects of CO2 on indi-
CO2 invested more C into fine roots, and did not experience vidual taxa. The below-ground ECM community is often
any decline in nutrient concentrations in plant tissues. The dominated by a few common species (Gardes & Bruns, 1996;
authors interpret the increased fine root density as likely to Erland et al., 1999; Peter et al., 2001) with a long tail of rare
have met the nutrient demands of the trees. In addition, the species in the distribution curve. Although there is still little
above mentioned increased production of mycelia under known about differences in carbon use efficiency between
elevated CO2 may also contribute to more efficient nutrient ECM fungal species, there may be differences in their parti-
acquisition. tioning of C, and in their response to elevated CO2 concen-
Due to the dry conditions in the chambers in our study, the trations (Dosskey et al., 1990; Rouhier & Read, 1998a;
number of root tips decreased compared to reference trees, Gorissen & Kuyper, 2000). Changes in abundance of domi-
although attempts were made to irrigate. This also affected the nating fungi may thus affect the C-allocation pattern of the
colonization levels to some extent (see below), but we may ECM community.
consider that the irrigation problem should not have affected Variation in ECM community composition both within
the overall response of the ECM community structure to seasons and among years has been reported (Malajczuk &
elevated CO2 because in the CCA ordination diagram, the Hingston, 1981; Wu et al., 1993), and was also evident in the
chamber effect was small and not significant. Irrigation has present study. Rygiewicz et al. (2000) found seasonal patterns
previously been shown not to affect the community compo- in the number of ECM root tips and in two of the dominant
sition at the Flakaliden field site (Fransson et al., 2000), since morphotypes (the Rhizopogon group and C. geophilum) to
the site is not water limited. Water limitation may influence respond differentially to CO2. Effects of elevated CO2 on
the response of host plants and associated mycorrhizal fungi ECM fungi have also been shown to vary over time by O’Neill
to increasing CO2, as reported for longleaf pine (Runion et al. (1987). Samples in our study were taken at the end of the
et al., 1997). growth season at both sampling occasions to avoid some of
An increase in colonization of ECM root tips due to ele- the variation attributed to seasonal changes in community
vated levels of CO2 has often been reported in the literature. structure. Despite the considerable interyearly variation in

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community composition a clear pattern was revealed in the of starch and succrose) or environmental limitations (root
DCA analysis. The ECM community structure of all fertilized restrictions due to pot size, nutrient availability). For trees in
trees was clearly separated from that of the unfertilized trees, the field, the situation seems to be quite different, and down-
with the exception of the elevated trees in the unfertilized regulation has usually not been seen in longer-term experi-
plot. These became more similar to the fertilized plot after ments (Saxe et al., 1998; Curtis & Wang, 1998; Norby et al.,
three years of CO2 treatment. 1999). But although down-regulation in the field may not
After 3 yr, the effects of CO2 on community composition generally be considered to be as pronounced as for short-term
were of the same magnitude as those seen after 15 yr of ferti- pot experiments, it cannot be dismissed as a potentially
lization treatment (indicated in Fig. 1 by the similar length of important factor in the over-all C-cycling. In the Flakaliden
CO2 and fertilization vectors). Trees are seen to respond experiment, down-regulaton of net photosynthesis was
within hours-days to elevated CO2, and the response of the observed (G. Wallin, pers. comm.).
below-ground symbiont will be mediated through the host Fitter et al. (2000) suggested that the response of mycor-
plant. The dependence of ECM fungi on current assimilates rhizal fungi could counteract down-regulation by providing
have been shown both in laboratory and field experi- the plants with more nutrients, but only up to a certain point.
ments (Söderström & Read, 1987; Lamhamedi et al., 1994; Eventually the whole system will come to a new equilibrium
Högberg et al., 2001). Physiological responses by ECM fungi since some factor will always be limiting in ecosystem pro-
to changes in C-supply are therefore likely to take place shortly cesses. In conclusion, our results show that elevated CO2 can
after the plant responds. Shifts in community composition, have an impact on the community structure of ECM fungi
however, may take a longer time to be induced. We do not in mature forests, and potentially alter carbon and nutrient
know if the induced change in ECM community structure allocation and turnover within forest ecosystems. Since soil
observed in the present study is transient in the present fertility has been shown to limit C sequestration by forest
study, but evidence from long-term studies on the effects of ecosystems in an enriched CO2 environment (Oren et al.,
N-fertilization upon ECM communities suggests that once 2001) the role of ECM fungi could be even more important
initiated, changes in community structure may be more than earlier believed.
enduring compared to physiological responses.
C- and N-nutrition within ECM fungi are intimately
linked. The response by the fungus to elevated CO2 levels is
Acknowledgements
most likely to be affected by availability, uptake and allocation This work was supported by the Swedish National Energy
of N. We found no detectable interactions between elevated Administration (STEM) and the Knut and Alice Wallenberg
CO2 and fertilization for individual components of the com- foundation. We wish to thank Jörg Brunet for valuable help
munity, as might have been expected. Interactions between with the ordination analysis, Birgitta Vegerfors-Persson for
elevated CO2 and N treatments were reported by Runion evaluating the statistical analysis and Björn Lindahl for
et al. (1997), with increased colonization, fine root lengths discussing the ordination results.
and numbers of ECM root tips under CO2 enrichment only
under high N conditions. The authors suggested that sink/
source relationships were a major factor regulating ECM References
fungal responses to elevated CO2. Agerer R. 1986 –98. Colour atlas of ectomycorrhizae. Münich: Germany:
Klironomos et al. (1997) found increased growth of extra- Einhorn-Verlag.
matrical mycelium of AM fungi colonizing Populus tremuloides Alexander IJ, Fairley RI. 1983. Effects of N fertilisation on populations of
under low N-availability/high CO2 treatment, and a decrease fine roots and mycorrhizas in spruce humus. Plant and Soil 71: 49 –53.
under high N-availability. Körner et al. (1997) suggested that ter Braak CJF, Smilauer P. 1998. Canoco reference manual and user’s guide to
Canoco for Windows: software for canonical community ordination, Version 4.
microbial activity of a late successional alpine grassland eco- Ithaca, NY, USA: Microcomputer Power.
system was colimited by C supply and N availability. We still ter Braak CJF, Verdonschot PFM. 1995. Canonical correspondence analysis
know very little about individual species responses. and related multivariate methods in aquatic ecology. Aquatic Science 57:
The issue of whether or not trees will experience a down- 255–289.
regulation or acclimation of photosynthesis as a response to Cairney JWG. 1999. Intraspecific physiological variation: implications for
understanding functional diversity in ectomycorrhizal fungi. Mycorrhiza 9:
long-term exposure to elevated CO2 has been debated over 125–135.
the years. If a down-regulation occurs in the field it might Ceulemans R, Janssens IA, Jach ME. 1999. Effects of CO2 enrichment on
have large effects on the capacity of forests to sequester trees and forests: lessons to be learned in view of future ecosystem studies.
CO2 and act as the predicted sink for atmospheric C. Results Annals of Botany 84: 577–590.
from short-term pot studies have shown considerable down- Curtis PS, Wang XZ. 1998. A meta-analysis of elevated CO2 effects on
woody plant mass, form, and physiology. Oecologia 113: 299 – 313.
regulation of photosynthetic rates within days-weeks (Curtis Dosskey MG, Linderman RG, Boersma L. 1990. Carbon-sink stimulation
& Wang, 1998; Ward & Strain, 1999). This down-regulation of photosynthesis in Douglas-Fir seedlings by some ectomycorrhizas. New
seems to be a result either of sink-limitations (accumulation Phytologist 115: 269–274.

www.newphytologist.com © New Phytologist (2001) 152: 431– 442


NPH_276.fm Page 441 Friday, November 2, 2001 10:02 AM

Research 441

Erland S, Jonsson T, Mahmood S, Finlay RD. 1999. Below-ground Körner C, Diemer M, Schappi B, Niklaus P, Arnone J. 1997. The responses
ectomycorrhizal community structure in two Picea abies forests in of alpine grassland to four seasons of CO2 enrichment: a synthesis. Acta
southern Sweden. Scandinavian Journal of Forest Research 14: 209–217. Oecologica-International Journal of Ecologica 18: 165 –175.
Falkowski P, Scholes RJ, Boyle E, Canadell J, Canfield D, Elser J, Gruber N, Lamhamedi MS, Godbout C, Fortin JA. 1994. Dependence of Laccaria
Hibbard K, Högberg P, Linder S, Mackenzie FT, Moore B, Pedersen T, bicolor basidiome development on current photosynthesis of Pinus strobus
Rosenthal Y, Seitzinger S, Smetacek V, Steffen W. 2000. The global seedlings. Canadian Journal of Forest Research 14: 412 – 415.
carbon cycle: a test of our knowledge of earth as a system. Science 290: Lewis JD, Thomas RB, Strain BR. 1994. Effects of elevated CO2 on
291– 296. mycorrhizal colonization of Loblolly Pine (Pinus Taeda L.) seedlings.
Finlay RD, Söderström B. 1992. Mycorrhiza and carbon flow to the soil. In: Plant and Soil 165: 81–88.
Allen M, ed. Mycorrhiza functioning. London, UK: Chapman & Hall, Lindroth A, Grelle A, Moren AS. 1998. Long-term measurements of boreal
134 –160. forest carbon balance reveal large temperature sensitivity. Global Change
Fitter AH, Heinemeyer A, Staddon PL. 2000. The impact of elevated CO2 Biology 4: 443–450.
and global climate change on arbuscular mycorrhizas: a mycocentric Malajczuk N, Hingston FJ. 1981. Ectomycorrhizae associated with Jarrah.
approach. New Phytologist 147: 179 –187. Australian Journal of Botany 29: 453–462.
Fransson PMA, Taylor AFS, Finlay RD. 2000. Effects of continuous Markkola AM, Ohtonen A, Ahonen-Jonnarth U, Ohtonen R. 1996. Scots
optimal fertilization on belowground ectomycorrhizal community pine responses to CO2 enrichment. 1. Ectomycorrhizal fungi and soil
structure in a Norway spruce forest. Tree Physiology 20: 599–606. fauna. Environmental Pollution 94: 309–316.
Gardes M, Bruns T. 1993. ITS primers with enhanced specificty for Norby RJ, Wullschleger SD, Gunderson CA, Johnson DW, Ceulemans R.
basidiomycetes – application to the identification of mycorrhizae and 1999. Tree responses to rising CO2 in field experiments: implications for
rusts. Molecular Ecology 2: 113 –118. the future forest. Plant, Cell & Environment 22: 683 – 714.
Gardes M, Bruns TD. 1996. Community structure of ectomycorrhizal fungi O’Neill EG. 1994. Responses of soil biota to elevated atmospheric carbon-
in a Pinus muricata forest: above- and below-ground views. Canadian dioxide. Plant and Soil 165: 55–65.
Journal of Botany 74: 1572 –1583. O’Neill EG, Luxmoore RJ, Norby RJ. 1987. Increases in mycorrhizal
Godbold DL, Berntson GM, Bazzaz FA. 1997. Growth and mycorrhizal colonization and seedling growth in Pinus Echinata and Quercus Alba in
colonization of three North American tree species under elevated an enriched CO2 atmosphere. Canadian Journal of Forest Research 17:
atmospheric CO2. New Phytologist 137: 433–440. 878–883.
Gorissen A, Kuyper TW. 2000. Fungal species-specific responses of Oren R, Ellsworth DS, Johnsen KH, Phillips N, Ewers BE, Maier C,
ectomycorrhizal Scots pine (Pinus sylvestris) to elevated [CO2]. New Schäfer KVR, McCarthy H, Hendrey G, McNulty SG, Katul GG. 2001.
Phytologist 146: 163 –168. Soil fertility limits carbon sequestration by forest ecosystems in a
Goulden ML, Wofsy SC, Harden JW, Trumbore SE, Crill PM, Gower ST, CO2-enriched atmosphere. Nature 411: 469 – 472.
Fries T, Daube BC, Fan SM, Sutton DJ, Bazzaz A, Munger JW. 1998. Peter M, Ayer F, Egli S. 2001. Nitrogen addition in a Norway spruce
Sensitivity of boreal forest carbon balance to soil thaw. Science 279: stand altered macromycete sporocarp production and below-ground
214 –217. ectomycorrhizal species composition. New Phytologist 149: 311– 325.
Hanson PJ, Wullschleger SD, Bohlman SA, Todd DE. 1993. Seasonal and Rey A, Jarvis PG. 1997. Growth response of young birch trees (Betula
topographic patterns of forest floor CO2 efflux from an upland oak forest. pendula Roth.) after four and a half years of CO2 exposure. Annals of
Tree Physiology 13: 1–15. Botany 80: 809–816.
Henrion B, Chevalier G, Martin F. 1994. Typing truffle species by PCR Rouhier H, Read DJ. 1998a. Plant and fungal responses to elevated
amplification of the ribosomal DNA spacers. Mycological Research 98: atmospheric carbon dioxide in mycorrhizal seedlings of Pinus sylvestris.
37– 43. Environmental and Experimental Botany 40: 237– 246.
Hodge A. 1996. Impact of elevated CO2 on mycorrhizal associations and Rouhier H, Read DJ. 1998b. The role of mycorrhiza in determining the
implications for plant growth. Biology and Fertility of Soils 23: 388–398. response of Plantago lanceolata to CO2 enrichment. New Phytologist 139:
Högberg P, Nordgren A, Buchmann N, Taylor AFS, Ekblad A, Högberg 367–373.
MN, Nyberg G, Ottosson-Löfvenius M, Read DJ. 2001. Large-scale Rühling Å, Tyler G. 1991. Effects of simulated nitrogen deposition to
forest girdling shows that current photosynthesis drives soil respiration. the forest floor on the macrofungal flora of a beech forest. Ambio 20:
Nature 411: 789 – 792. 261–263.
Horton TR, Bruns TD. 2001. The molecular revolution in ectomycorrhizal Runion GB, Mitchell RJ, Rogers HH, Prior SA, Counts TK. 1997. Effects
ecology: peeking into the black-box. Molecular Ecology 10: 1855–1871. of nitrogen and water limitation and elevated atmospheric CO2 on
Ineichen K, Wiemken V, Wiemken A. 1995. Shoots, roots and ectomycorrhiza of longleaf pine. New Phytologist 137: 681– 689.
ectomycorrhiza formation of Pine-seedlings at elevated atmospheric Rygiewicz PT, Martin KJ, Tuininga AR. 2000. Morphotype community
carbon-dioxide. Plant, Cell & Environment 18: 703–707. structure of ectomycorrhizas on Douglas fir (Pseudotsuga menziesii Mirb.
Jonsson L, Dahlberg A, Brandrud T-E. 1999. Spatiotemporal distribution Franco) seedlings grown under elevated atmospheric CO2 and temperature.
of an ectomycorrhizal community in an oligotrophic Swedish Picea abies Oecologia 124: 299–308.
forest subject to experimental nitrogen addition: above- and below-ground Sanders IR, Streitwolf-Engel R, van der Heijden MGA, Boller T, Wiemken A.
views. Forest Ecology and Management 132: 143–156. 1998. Increased allocation to external hyphae of arbuscular mycorrhizal
Kårén O, Nylund J-E. 1997. Effects of ammonium sulphate on the c fungi under CO2 enrichment. Oecologia 117: 496 – 503.
ommunity structure and biomass of ectomycorrhizal fungi in a Norway Saxe H, Ellsorth DS, Heath J. 1998. Tree and forest functioning in an
spruce stand in southwestern Sweden. Canadian Journal of Botany 75: enriched CO2 atmosphere. New Phytologist 139: 395 – 436.
1628 –1642. Schimel DS. 1995. Terrestrial ecosystems and the carbon-cycle. Global
Kirschbaum MUF. 1995. The temperature-dependence of soil organic- Change Biology 1: 77–91.
matter decomposition, and the effect of global warming on soil organic-C Schlesinger WH, Lichter J. 2001. Limited carbon storage in soil and litter
storage. Soil Biology and Biochemistry 27: 753–760. of experimental forest plots under increased atmospheric CO2. Nature
Klironomos JN, Rillig MC, Allen MF, Zak DR, Kubiske M, Pregitzer KS. 411: 466–469.
1997. Soil fungal-arthropod responses to Populus tremuloides grown under Söderström B, Read DJ. 1987. Respiratory activity of intact and excised
enriched atmospheric CO2 under field conditions. Global Change Biology ectomycorrhizal mycelial systems growing in unsterilised soil. Soil Biology
3: 473 – 478. and Biochemistry 19: 231–236.

© New Phytologist (2001) 152: 431– 442 www.newphytologist.com


NPH_276.fm Page 442 Friday, November 2, 2001 10:02 AM

442 Research

Taylor AFS, Martin F, Read DJ. 2000. Fungal diversity in ectomycorrhizal Montagnani L, Minerbi S, Jarvis PG. 2000. Respiration as the main
communities of Norway spruce (Picea abies [L.] Karst.) and beech (Fagus determinant of carbon balance in European forests. Nature 404: 861– 865.
sylvatica L.) along north-south transects in Europe. In: Schulze E-D, ed. Walker RF, Geisinger DR, Johnson DW, Ball JT. 1997. Elevated
Carbon and nitrogen cycling in European forest ecosystems. Berlin, Germany: atmospheric CO2 and soil N fertility effects on growth, mycorrhizal
Springer Verlag, 343 –365. colonization, and xylem water potential of juvenile ponderosa pine in
Tingey DT, Phillips DL, Johnson MG, Storm MJ, Ball JT. 1997. Effects of a field soil. Plant and Soil 195: 25–36.
elevated CO2 and N fertilization on fine root dynamics and fungal growth Ward JK, Strain BR. 1999. Elevated CO2 studies: past, present and future.
in seedling Pinus ponderosa. Environmental and Experimental Botany 37: Tree Physiology 19: 211–220.
73 – 83. White TJ, Bruns T, Lee S, Taylor J. 1990. Amplification and direct
Treseder KK, Allen MF. 2000. Mycorrhizal fungi have a potential role in sequencing of fungal ribosomal RNA genes for phylogenetics. In:
soil carbon storage under elevated CO2 and nitrogen deposition. Innis MA, Gelfand DH, Sninsky JJ, hite, TJ, eds. PCR protocols:
New Phytologist 147: 189 – 200. a guide to methods and applications. New York, USA: Academic Press,
Valentini R, Matteucci G, Dolman AJ, Schulze ED, Rebmann C, Moors EJ, 315–322.
Granier A, Gross P, Jensen NO, Pilegaard K, Lindroth A, Grelle A, Wu Y, Gale MR, Cattelino PJ, Richter DL, Bruhn JN. 1993. Temporal
Bernhofer C, Grunwald T, Aubinet M, Ceulemans R, Kowalski AS, dynamics of ectomycorrhizal populations and seedling characteristics
Vesala T, Rannik U, Berbigier P, Loustau D, Guomundsson J, on red pine (Pinus resinosa). Canadian Journal of Forest Reasearch 23:
Thorgeirsson H, Ibrom A, Morgenstern K, Clement R, Moncrieff J, 810–815.

www.newphytologist.com © New Phytologist (2001) 152: 431– 442