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LETTER
Genetic variability in a population of arbuscular
mycorrhizal fungi causes variation in plant growth
Alexander M. Koch, Daniel Croll
and Ian R. Sanders*
Department of Ecology
and Evolution, University of
Lausanne, Lausanne,
Switzerland
*Correspondence: E-mail:
ian.sanders@unil.ch
Abstract
Different species of arbuscular mycorrhizal fungi (AMF) alter plant growth and affect
plant coexistence and diversity. Effects of within-AMF species or within-population
variation on plant growth have received less attention. High genetic variation exists
within AMF populations. However, it is unknown whether genetic variation contributes
to differences in plant growth. In our study, a population of AMF was cultivated under
identical conditions for several generations prior to the experiments thus avoiding
environmental maternal effects. We show that genetically different Glomus intraradices
isolates from one AMF population significantly alter plant growth in an axenic system
and in greenhouse experiments. Isolates increased or reduced plant growth meaning that
plants potentially receive benefits or are subject to costs by forming associations with
different individuals in the AMF population. This shows that genetic variability in AMF
populations could affect host-plant fitness and should be considered in future research
to understand these important soil organisms.
Keywords
Arbuscular mycorrhizal fungi, benefits, costs, functional variability, genetic diversity,
Glomus intraradices, intraspecific variation, plant growth, population, symbiosis.
Ecology Letters (2006) 9: 103–110
INTRODUCTION
Arbuscular mycorrhizal fungi (AMF) are abundant in soils
of most terrestrial ecosystems where they form symbioses
with roots of most plants (Smith & Read 1997). Arbuscular
mycorrhizal fungi (phylum Glomeromycota) are obligate
biotrophs and supply plants with phosphorous in exchange
for carbohydrates (Harrison 1997). The fungi are an ancient
group of presumed asexual organisms and only c. 160
morphospecies are known (Redecker et al. 2000; Kuhn et al.
2001). Arbuscular mycorrhizal fungi grow clonally and are
coenocytic, harbouring many nuclei in a common cyto-
plasm. The symbiosis with AMF can improve plant nutrient
acquisition, enhance stress and pathogen tolerance and
increase plant diversity (Grime et al. 1987; Newsham et al.
1995; Smith & Read 1997).
Inoculation with different AMF species differentially
alters the growth and coexistence of different plant species
(Streitwolf-Engel et al. 1997; van der Heijden et al. 1998a,
2003) and increasing species richness of AMF increases
plant diversity and productivity (van der Heijden et al.
1998b). However, these studies used single individuals as
representatives of each AMF species where each culture was
initiated from only one spore. These studies, therefore, did
not address whether intraspecific variation could account
for some of the observed differences among taxa. The
question of inter- and intraspecific variation in AMF and its
effects on plant growth has been partially addressed.
Variation in plant growth was higher among plants
inoculated with different species than those inoculated with
different isolates of the same species (Hart & Klironomos
2002). However, this does not mean that within isolate
variation is not ecologically important. More recently
considerable within AMF species variation has been
observed. Large differences in plant growth and phosphor-
ous uptake were reported within AMF species, thus,
showing the potential ecological importance of within
AMF species variation (Munkvold et al. 2004). The AMF
isolates compared in the study by Munkvold et al. (2004)
evolved in different environments and differed in their
culturing histories. The isolates were probably also genetic-
ally different. Therefore, these large intraspecific differences
could have been caused by environmental differences or
genotypic differences or both. Different isolates of an AMF
species from one geographic location have also been shown
to cause variation in the amount of biologically fixed
Ecology Letters, (2006) 9: 103–110 doi: 10.1111/j.1461-0248.2005.00853.x
2005 Blackwell Publishing Ltd/CNRS
nitrogen in clover plants (Gamper et al. 2005). Recently, a
population of G. intraradices was shown to be comprises of
individuals that were genetically different from each other
(Koch et al. 2004). These individuals were isolated from the
same field and maintained for successive generations in
exactly the same environment. These AMF isolates showed
considerable differences in their amplified fragment length
polymorphism (AFLP) banding patterns and their pheno-
types, especially in extraradical hyphal density, a trait that is
thought to play an important role in phosphorous acquisi-
tion. Variation in these traits likely has a genetic basis
because culturing under identical conditions over several
generations eliminated possible environmental effects.
The aim of the present study is to test whether genetic
differences among individuals of an AMF population could
significantly alter the growth of plants. In our experiments,
we used isolates from a population of Glomus intraradices that
had been maintained in the same environment (Koch et al.
2004) in order to reduce environmental effects. In a system
where the AMF are grown with transformed carrot roots on
an artificial medium, we tested whether genetically different
isolates differentially affected root growth. In a second
experiment, in the same culture system, we tested whether
altering the environment would differentially alter the effect
of a given isolate on root growth. In a greenhouse
experiment, we tested whether the growth of two plant
species, that had previously been shown to grow differently
when forming the symbiosis with different AMF species
(van der Heijden et al. 2003), was altered by the colonization
of genetically different G. intraradices individuals that all
come from one field (Koch et al. 2004). Because overall
growth of AMF inoculated plants did not differ from that of
non-mycorrhizal control plants in this first experiment, we
performed a second greenhouse experiment, where a poorer
soil mixture was used. To test whether differences in soil
moisture altered growth and AMF-dependency of host
plants, two different watering regimes were also included. In
all experiments genetically different AMF isolates signifi-
cantly altered host growth, and the effects ranged from
detrimental to beneficial depending on the environmental
conditions and the identity of the isolate. Our results show
that ecologically relevant genetic and functional diversity
exists within AMF populations revealing a further level of
complexity of AMF biology and natural ecosystems.
MATERIALS AND METHODS
All fungal material used in our experiments belonged to the
AMF species G. intraradices Schenck & Smith. The species
identification was based on spore morphology and internal
transcribed spacer (ITS) sequences (Koch et al. 2004). The
AMF single spore isolates originate from four different plots
from one agricultural field in Ta
¨nikon, Switzerland (Anken
et al. 2004; Koch et al. 2004). The agricultural field contained
two replicate plots nested within two tillage treatments
(plots A and B experienced no tillage and plots C and D
experienced tillage). The distances among the plots varied
from 29.2 m (distance from plot A to plot B) to 85.2 m
(distance from plot B to plot C). Single G. intraradices spores
were obtained from trap cultures that had been started from
mixtures of soil samples from each of the four plots. These
single spores were then placed individually onto germinating
roots of Plantago lanceolata roots to start single spore cultures.
From each of these cultures, a single healthy-looking spore
was used to colonise transformed carrot roots in an axenic
culture system. The isolates were subsequently kept under
identical environmental conditions for two generations prior
to experiment 1 and for at least seven generations before all
other experiments to reduce possible maternal effects (Koch
et al. 2004). The nomenclature of the fungal isolates used
here is the same as in Koch et al. (2004); capitalized letters
indicate the origin of the plot and the numbers indicate
different single spore lines.
Experiment 1: the effect of different isolates on host root
growth
The aim of experiment 1 was to test whether different
isolates altered the growth of host roots. Sixteen AMF
isolates (four isolates from each of the four plots) of
G. intraradices were cultured in an axenic system with a clone
of transformed carrot roots as described in Koch et al.
(2004). Root length was measured 3, 6, 9, 12 and 15 weeks
after inoculation of each plate by counting the number of
root intersects with a rectangular grid (2 cm interspace). The
measures over time allowed the calculation of maximum
root growth rate by determining the maximum slope of the
least square fit of the logistic curve
y¼aðbþexpðcweekÞÞ1
for each of the 229 replicate plates of the experiment. For
full details of experiment 1 see Koch et al. (2004).
Experiment 2: the effect of decreased nutrient availability
on host root growth
The same model system as in experiment 1 was used to test
how the growth of the host roots and four genetically
different fungal isolates were altered when the availability of
two key nutrients, sucrose and phosphate, was decreased.
To do this, we selected four G. intraradices isolates, which
were sampled from different plots and which were
previously shown to differ in their AFLP fingerprint profiles
(Koch et al. 2004). The four AMF isolates (A4, B3, C3 and
D1) were cultivated in the standard M medium and in
medium with reduced sucrose and phosphate concentra-
104 A. M. Koch, D. Croll and I. R. Sanders
2005 Blackwell Publishing Ltd/CNRS
tions. The concentration of either sucrose or potassium
phosphate in the growth medium was reduced to 50% and
25% compared with the standard M medium respectively
(Becard & Fortin 1988). The reduced amount of potassium
was compensated by adding the equivalent amount of K in
the form of KCl to the medium. The five treatments are
abbreviated as M (standard), S50, S25, for the reduced
sucrose, and P50, P25 for the reduced phosphorous
treatments respectively. After the end of experiment 1, the
isolates and non-mycorrhizal (NM) roots were subcultured
four times on M medium over a period of 1 year before
being used for this experiment. For roots inoculated with
each of the four isolates and the NM roots, 40 parental
plates were randomized and used to inoculate 60 new plates,
i.e. 12 new replicate plates of each of the five treatments.
After 15 week of growth, root length of each plate was
measured as in experiment 1, and hyphal length and spore
density were quantified as in Koch et al. (2004). Contam-
inated plates (n¼3) and plates with no AMF hyphal
growth (n¼21) were removed from the statistical analyses,
which reduced the total number of plates from 300 to 276.
Experiments 3: the effect of different isolates on total dry
weight of two host species
The aim of this greenhouse experiment was to test whether
the growth of the two plant species Prunella vulgaris L. and
Brachypodium pinnatum (L.) P.B. was altered when inoculated
with different isolates of G. intraradices. To cover as much of
the detected genetic diversity of this population as possible,
we chose six isolates that were phenotypically and genetic-
ally the most different in a previous study (Koch et al. 2004).
The seeds of the host plants were obtained from Fenaco
(Winterthur, Switzerland). Seeds of the two host species
were surface sterilized in 6% bleach for 10 min, rinsed three
times with sterile water and put on seed trays with moist
vermiculite for germination in the greenhouse on 27 August
2002. A total of 140 pots (10.5-cm diameter ·8-cm high)
were filled with 450 mL of a 1 : 1 : 1 (vol : vol : vol)
mixture of a previously sieved (5-mm mesh width) high
nutrient brown soil from an old field on the campus of the
University of Lausanne (2684, 2.4 and 27 lmol L
)1
N,
soluble P and K respectively), washed sand and pure quartz
sand (1 mm). The soil mixture was steam autoclaved twice
at 120 C 2 weeks before planting. On 8 September 2002,
seedlings of similar size of either of the two host species
were transferred to 70 pots that had been well watered. Two
seedlings of the same species were transplanted per pot.
Each seedling was inoculated with 0.2 mL spore suspension
that contained 500 spores of one of the isolates A4, B1, B3,
C2, C3 and D1, NM plants received 0.2 mL tap water. The
spores used as inoculum were previously grown for
4 months with transformed carrot roots on split plates,
and the cultures were kept under identical conditions for
several years to eliminate maternal effects (Koch et al. 2004).
After 16 days of growth the smaller of the two seedlings was
removed. This resulted in 10 replicate pots for each isolate
and NM treatment for each plant species. Each pot was
watered every other day with 30 mL tap water. Day length
was 16 h, supplemented with artificial light when necessary.
The temperatures ranged between 18 and 30 C. The
position of the individual pots was randomized weekly. The
shoots of all plants were harvested 100 and 200 days after
inoculation. This was done by cutting off all plant tissue
1 cm from the base of the shoot. This ensured re-growth by
remaining meristems. At the final harvest, after 300–
310 days growth, the shoot of each host plant was separated
from the roots and the roots were carefully washed. After
recording the fresh weight of the roots, a small root sample
was cut and stored in 50% ethanol and the remaining fresh
weight of the roots was recorded. At all harvests shoots and
roots where separately dried at 80 C for 2 days and then
weighed. The stored root samples were stained with Trypan
Blue as described in Munkvold et al. (2004) and inspected
for AMF colonization. All plants survived to the end of the
experiment and none of the NM plants showed any AMF
colonization. All plants that received inoculum were clearly
colonized by AMF.
Experiment 4: the effect of different isolates on plant
growth under two watering regimes
The aim of this experiment was to test whether the same
set of isolates used in experiment 3 also altered plant
growth in conditions where there was a main positive
effect of AMF inoculation on plant growth. In order to
achieve this, we set-up two different watering regimes to
test whether differences in soil moisture influenced AMF
dependency on plant growth. The experimental procedures
in experiment 4 were as in experiment 3, except where
indicated. The substrate mixture consisted of 3 : 2
(vol : vol) Terra Green (Oil Dry US Special, Maag Technic
SA, Crissier, Switzerland) and pure quartz sand to which
we added 4% (vol) of the same soil as in experiment 3.
Single seedlings were inoculated and transplanted on 29
February 2004, using the same six AMF isolates, a NM
treatment, and the same two 2-host species. For each host
species and each AMF isolate and the NM treatment 16
replicate pots were used in the experiment. Day length was
set at the natural day/night rhythm. Each pot was
previously filled with 325 g of dried substrate and watered
to water holding capacity (WHC) and later to 40–55% of
WHC. In addition, eight replicate pots were established
with each of the same two host plant species and each of
the isolates A4, B3, and C2 and NM plants. These plants
received an increased water regime where soil moisture was
Functional variability in AMF populations 105
2005 Blackwell Publishing Ltd/CNRS
kept between 70% and 100% of WHC. All plants were
watered with deionized water (Option 1; Lab Services AG,
Wohlen, Switzerland). After 3 months, each pot was
fertilized once with 10 mL of full-strength Hoagland
solution containing no phosphorous and the K concentra-
tion was adjusted by adding KCl. After 4 months, all
shoots were harvested. The pots were subsequently stored
in a cold room at 4 C. All the roots were washed and
weighed within the following 3 weeks. One plant died
during the experiment and in roots of four plants where
inoculum was added no fungal colonization was observed.
These five individuals were removed which left data from
283 plants for the analyses.
Statistical analyses
In experiment 1, data were analysed using a nested
ANOVA
model with the main factors tillage treatment, plot (nested in
treatment), isolate (nested in plot) and inoculation (plate
nested in isolate), as in Koch et al. (2004). In the results, only
significant main effects are given. In experiment 2, data were
analysed using a crossed two-way
ANOVA
with treatment and
isolate as main fixed factors. The analyses were carried out
separately for the different phosphorous and sucrose
treatments and the root growth data was analysed both
with and without NM roots. In experiments 1 and 2,
Pearson’s correlation coefficient was calculated for com-
parison of different growth variables. The data of experi-
ments 3 and 4 were also analysed with a crossed two-way
ANOVA
with both host species and isolate as fixed factors.
To test whether the overall growth of NM plants differed
from AMF inoculated plants, all the isolates were pooled. In
order to test for an AMF isolate by plant–species
interaction, NM plants were omitted from the analysis. In
experiment 4, data were analysed separately for the two
water treatments. Variables were transformed, if necessary,
to meet the requirements of the statistical tests (Zar 1984).
All analyses were performed with the statistical programs
JMP 5.0 (SAS Institute Inc., Cary, NC, USA) and R version
1.8.0 (R. Development Core Team, 2003, www.R-project.org).
RESULTS
Experiment 1: the effect of different isolates on host root
growth
After 15-week growth, the length of transformed carrot
roots differed significantly among isolates, with up to 15%
difference in root length among different AMF isolate
treatments (
ANOVA
,F
12,46
¼3.28, P< 0.01, Fig. 1). The
maximal rate of root growth showed a similar pattern and
was positively correlated with final root length (r¼0.76,
P< 0.0001, data not shown). The maximal hyphal growth
rate was negatively correlated with both the maximal rate of
growth of the roots (r¼)0.13, P< 0.05) and the growth
rate of the roots at the time of maximal hyphal growth (r¼
)0.19, P< 0.01).
Experiment 2: the effect of decreased nutrient availability
on host root growth
Growth of transformed carrot roots was significantly altered
by both the reduced phosphorous concentration and the
different AMF isolates (F
2,162
¼5.23, P< 0.01 and
F
4,162
¼9.07, P< 0.0001 respectively, Fig. 2a). There was
no significant P-concentration by isolate interaction, indi-
cating that root growth in the different phosphorous
treatments responded in the same way with each of the
different AMF isolates. The results were qualitatively the
same when the NM roots were excluded from the analysis.
Final hyphal length and spore density also differed among
AMF isolates (F
3,129
¼65.67, P< 0.0001 and F
3,129
¼
45.85, P< 0.0001 respectively, Fig. 2b, c) and among P-
concentrations (F
2,129
¼5.64, P< 0.01 and F
2,129
¼7.68,
P< 0.001 for the two variables respectively) with no
significant interaction terms.
Growth of transformed carrot roots decreased consider-
ably with the reduced sucrose concentrations (Fig. 2a,
F
2,142
¼247.35, P< 0.0001 and F
2,109
¼209.07,
P< 0.0001, with and without NM roots respectively). Root
growth also differed among isolates (F
4,142
¼7.00,
P< 0.0001 and F
3,109
¼8.87, P< 0.0001, with and with-
out NM roots respectively, Fig. 2a). The growth of NM
roots was lower than inoculated roots in M medium.
However, the growth of NM roots was higher than some
AMF inoculated roots in medium with lowered sucrose
concentration, which resulted in a significant sucrose
concentration by AMF isolate interaction (F
8,142
¼2.79,
P< 0.01, Fig. 2a). The interaction term was not significant
when the NM roots were omitted from the analysis. Hyphal
Figure 1 Mean final length of transformed D. carota roots after
15 weeks of growth in M medium and inoculated with 16 different
single spore isolates of the AMF Glomus intraradices. Error bars
represent + 1 SE.
106 A. M. Koch, D. Croll and I. R. Sanders
2005 Blackwell Publishing Ltd/CNRS
length and spore density differed among isolates (F
3,109
¼
17.44, P< 0.0001 and F
3,109
¼11.08, P< 0.0001 respec-
tively, Fig. 2b, c). Hyphal length and spore density also
differed greatly in the different sucrose concentrations
(F
2,109
¼207.41, P< 0.0001 and F
2,109
¼230.73, P<
0.0001 respectively) with growth being much lower at low
sucrose concentrations compared with the M medium.
There was no significant AMF isolate by sucrose
concentration interaction for either variable.
When all isolates were pooled, hyphal length and spore
density were positively correlated in each of the five
treatments of different nutrient availability (P< 0.0001, for
all five treatments). However, there were significant negative
correlations between hyphal and root length on M medium
(r¼)0.37, P< 0.05) and between spore density and root
length on M medium (r¼)0.34, P< 0.05) and in the S50
treatment (r¼)0.32, P< 0.05).
Experiment 3: the effect of different isolates on total dry
weight of two host species
After 10 months of growth in the green house, there was no
significant effect of mycorrhizal inoculation on the total dry
weight of either plant species when comparing AMF
inoculated with NM plants (Fig. 3). However, inoculation
with different AMF isolates significantly altered the total dry
weight of the plants (F
5,108
¼4.50, P< 0.0001, Fig. 3). The
total dry weight of B. pinnatum plants was greater than that of
P. vulgaris (F
1,108
¼44.82, P< 0.0001) and there was no
significant isolate by host interaction. All plants that received
AMF inoculum were clearly colonized and no AMF
colonization was observed in any of the NM plants. The
P. vulgaris plants inoculated with isolate B1 tended to be
smaller than plants colonized with other G. intraradices isolates
as well as non-mycorrhizal plants at all three harvests. In
contrast, at all three harvests B. pinnatum plants colonized
with isolate B3 were larger than plants colonized with the
other isolates and NM plants (data not shown). The total dry
weight of the roots was strongly correlated with the total dry
weight of the shoots for both host species (r¼0.84,
P< 0.0001 and r¼0.82, P< 0.0001 for B. pinnatum and
P. vulgaris respectively).
Figure 2 (a) Mean final length of transformed D. carota roots, (b) mean hyphal length and (c) mean spore density of four Glomus intraradices
isolates. Transformed D. carota roots were each colonized with one of four genetically different arbuscular mycorrhizal fungi (AMF) isolates
of G. intraradices (solid circle A4, black triangle B3, open circle C3, cross D1) or non-mycorrhizal (open squares). The roots were grown on
five different media; standard M medium (M), M medium with phosphate reduced to 50% (P50), phosphate reduced to 25% (P25), sucrose
reduced to 50% (S50) and sucrose reduced to 25% (S25). Although data from phosphorous and sucrose treatments are graphed together, the
black-shaded triangles below each graph indicate that
ANOVA
s were performed separately for the different phosphate and sucrose
concentrations and both analyses included the standard M medium. Different letters left of symbols of the same medium composition
indicate a significant difference (P< 0.05) according to the Tukey–Kramer honest significant test (HSD, P< 0.05).
Figure 3 Mean total dry weight of plants that were inoculated with
six genetically different Glomus intraradices isolates (A4, B1, B3, C2,
C3 and D1) or left uninoculated [non-mycorrhizal (NM)] after a
growth period of 10 months. Two host species, Brachypodium
pinnatum (black shading) and Prunella vulgaris (unshaded), were used
in the experiment. Error bars represent + 1 SE and different letters
above bars of the same plant species indicate a significant
difference (P< 0.05) according to the Tukey–Kramer honest
significant test (HSD) test.
Functional variability in AMF populations 107
2005 Blackwell Publishing Ltd/CNRS
Experiment 4: the effect of different isolates on plant
growth under two watering regimes
At the end of experiment 4, the roots of all plants that had
received AMF inoculum were colonized with AMF and
none of the NM plants were colonized with AMF. In
contrast to experiment 3, AMF colonized plants grew
significantly larger than NM plants after a growth period of
4 months under dry conditions (F
1,215
¼146.42,
P< 0.0001, Fig. 4a). The AMF growth enhancement was
greater for P. vulgaris than B. pinnatum. Plants colonized with
isolate C2 were larger than plants colonized with isolate C3
and overall the AMF isolates significantly altered final total
plant dry weight (F
5,175
¼3.01, P< 0.05) with no
significant host effect and isolate by host interaction.
Mycorrhizal B. pinnatum plants had a significantly higher
root to shoot ratio than P. vulgaris (F
1,175
¼185.98,
P< 0.0001, Fig. 4b) and there was no significant isolate
effect or isolate by host interaction. The absence of a
significant isolate by host interaction indicates that the
growth response of the two plant species did not differ
according to which isolate they had been inoculated with.
Under wetter growth conditions the plants grew larger than
under dry growth conditions (Fig. 4a, c). Plants inoculated
with AMF were colonized and tended to be larger than the
NM plants, which were not colonized with AMF (F
1,60
¼
2.80, P< 0.1, Fig. 4c), but there were no significant isolate,
host or interaction effects.
DISCUSSION
Our results (experiments 1–4) show that genetically differ-
ent individuals of G. intraradices from a population origin-
ating from one field alter plant growth. Using an artificial
study system where AMF isolates grow with transformed
carrot roots we found, in both experiments 1 and 2, that
genetically different AMF isolates significantly influenced
root length of transformed Daucus carota. The results from
experiment 2 additionally show that altering the environ-
ment can result in AMF colonization that is costly for the
plant. We also conducted two experiments in the green-
house where, in both cases, genetically different AMF
isolates significantly altered plant growth. In experiment 3,
although there was no overall mycorrhizal benefit of the
plants (comparison of NM with AMF inoculated plants)
genetically different AMF isolates resulted in differential
plant growth. Costs of AMF colonization were also seen
for P. vulgaris plants colonized with one isolate. In experi-
ment 4, where all AMF isolates enhanced plant growth
compared with NM plants, different isolates also altered
plant growth.
In this study, we have shown that significant functional
variability exists in an AMF population. The study by
Munkvold et al. (2004) showed large differences in plant
phosphorous uptake, which were caused by isolates from
different geographic origins. Compared with our study, the
AMF isolate effects observed by Munkvold et al. (2004)
appear larger and this points to the fact that the origin of an
isolate also plays an important role. It is also possible that the
genetic differences among AMF individuals used by Munkv-
old et al. (2004) from different regions were larger than the
genetic differences among individuals within the population
used in our study. Koch et al. (2004) have already shown
Figure 4 (a) Mean total dry weight and (b) root to shoot ratio of
plants that were inoculated with six genetically different Glomus
intraradices isolates (A4, B1, B3, C2, C3 and D1) or left uninoculated
[non-mycorrhizal (NM)]. Values were at the final harvest 4 months
after inoculation and under dry growth conditions (40–55% of soil
water holding capacity, n¼219). (c) Mean total dry weight of NM
plants and plants inoculated with three arbuscular mycorrhizal
fungi (AMF) isolates (A4, B3 and C2) and 4 months after
inoculation, under wet growth conditions (70–100% of WHC,
n¼64). Two host species, Brachypodium pinnatum (black shading)
and Prunella vulgaris (unshaded), were used in the experiment. Error
bars represent + 1 SE and different letters above bars of the same
plant species indicate a significant difference (P< 0.05) according
to the Tukey–Kramer honest significant test (HSD) test.
108 A. M. Koch, D. Croll and I. R. Sanders
2005 Blackwell Publishing Ltd/CNRS
spatial genetic structure in an AMF population. Combined
with our results here, and the findings of Munkvold et al.
(2004), this suggests that both genetic and functional
diversity in AMF species is spatially structured both within
and among populations. A sampling scheme of AMF
populations from a local to a regional or even global scale
would be required to address the question of how functional
and genetic diversity is related to different spatial scales. Only
by sampling several individuals, of several species and genera
from one community would it be possible to address the
question at what taxonomic level most functional AMF
diversity is explained. To our knowledge, no such experiment
has yet been performed, but the results from Hart &
Klironomos (2002) indicate that variation between AMF
genera is higher than within the genus Glomus.
In experiment 2, reduction of two key nutrients
(phosphate and sucrose) in the growth medium decreased
root and fungal growth, but the effects were much stronger
when sugar was limiting. Under such limiting conditions of
reduced sucrose availability AMF colonized roots had a
reduced root length compared with NM roots. This
indicates that there are costs of the symbiosis for the plant
roots under these environmental conditions. This is also
supported by negative correlations between root and fungal
growth in both experiments 1 and 2. In experiment 2,
however, this was only the case on the M medium (highest
nutrient availability) and in the S50 treatment (50% reduced
sucrose availability). This indicates that nutrient availability
influences costs and benefits as well as potential conflicts
between the growth potential of host and fungal symbionts.
These findings also suggest that the host plant does not
strictly control the symbiosis because AMF colonization
continued under decreased resource availability despite
increased costs to the host. However, overall fungal and
root growth were reduced under decreased resource
availability, but the growth reduction on fungal growth
appear larger than that of roots in the treatment of lowest
sucrose availability. This indicates that there are thresholds
of nutrient availability allowing sustainable growth of both
symbionts. This culture system is, therefore, ideal to further
investigate the conditions and consequences of AMF and
host root co-existence in controlled experiments. The fact
that we detected no significant AMF isolate by environment
effect in experiment 2 indicates that the genetically different
isolates did not differentially effect root growth in these
treatments of reduced sucrose and phosphate availability.
However, it is still possible that other AMF isolates from
this population, that were not used in this study, could
differentially modify their growth in response different
environments.
There was also no significant host species by AMF isolate
interaction in neither experiment 3 nor experiment 4
indicating that these isolates did not have specific host
species affinities that resulted in differential plant growth.
Evidence for selection for similar phenotypes in this AMF
population (Koch et al. 2004) could explain similar function
and thus the relatively consistent performance of the
isolates.
Our results are also interesting in the light of the fact that
all AMF isolates used in this study come from an agricultural
field. There is evidence showing that AMF from nutrient-
rich environments such as fertilized soils tend to be less
efficient symbionts (Johnson 1993). While, overall there was
no AMF inoculation effect in experiment 3, one AMF
isolate (B1) significantly reduced plant growth of P. vulgaris,
but had no such drastic negative effect on the second host
species B. pinnatum. In experiment 4, where the soil mixture
was poorer in nutrients and the growth conditions drier, all
G. intraradices isolates used enhanced plant growth showing
that all these individuals have maintained functions, which
can be beneficial to their hosts. It will be interesting to
further investigate whether and how environmental differ-
ences or different host species influence, interact with or
possibly maintain the genetic diversity present in AMF
populations.
In general, however, the AMF symbiosis is thought to be
mutualistic as it can improve plant growth and nutrient
acquisition, protect hosts from pathogens, alter host–
herbivore interactions, and increase plant drought tolerance
(Newsham et al. 1995; Smith & Read 1997; Auge 2001;
Gange et al. 2003; Garmendia et al. 2005; Kula et al. 2005).
Nevertheless, plants that form the AMF symbiosis often
have no growth benefit or even a reduced growth (Johnson
et al. 1997; Jifon et al. 2002; Klironomos 2003; Kyto
¨viita
et al. 2003; Brundrett 2004). The frequent observation of
commensal or parasitic AMF–plant interaction could also be
an insurance policy for plants, as in an unpredictable and
changing environment short-term costs of plants could be
outweighed by long-term net benefits. Our results show that
genetic variability in an AMF population cause a range of
different outcomes of plant growth, which also depend on
environmental conditions. We conclude that intraspecific
variability and genetic diversity in AMF populations cause
variation in plant growth, which could also be ecologically
relevant on the ecosystem level and be important for the
development of potent AMF inocula for successful restor-
ation of plant communities.
ACKNOWLEDGEMENTS
We thank Nicoletta Rinaudo, Gerrit Kuhn and Martine
Ehinger for their help in the greenhouse and the Swiss
National Science foundation (project 3100AO-105790/1) to
which support is gratefully acknowledged. We appreciate the
constructive comments of three referees that helped to
improve this manuscript.
Functional variability in AMF populations 109
2005 Blackwell Publishing Ltd/CNRS
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Editor, Rebecca Irwin
Manuscript received 3 August 2005
First decision made 14 September 2005
Manuscript accepted 3 October 2005
110 A. M. Koch, D. Croll and I. R. Sanders
2005 Blackwell Publishing Ltd/CNRS
