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Keywords:

  • arbuscular mycorrhiza;
  • below-ground competition;
  • common mycorrhizal network (CMN);
  • defoliation;
  • seedling establishment

Summary

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. References
  9. Supporting Information
  • 1
    Competition is considered to be one of the main biotic factors shaping plant communities. Experiments on the role of mycorrhizal symbiosis in plant competition have reported conflicting results.
  • 2
    We studied the effect of arbuscular mycorrhizal symbiosis on below-ground interactions between seedlings and established adult plants in a system where plants compete for below-ground resources. In this glasshouse experiment, seedlings of Gnaphalium norvegicum were grown with and without an arbuscular mycorrhizal fungus, in the presence and absence of a large conspecific adult plant. The ability of adult plants to support the fungus was modified by defoliating 0%, 50% or 75% of its leaf area.
  • 3
    We found that mycorrhiza increased below-ground competitive intensity. The mycorrhizal benefit to the seedlings was low in the vicinity of non-defoliated adult plants, but increased with increasing defoliation intensity of the adult plant. This was mirrored by reductions in mycorrhizal benefit that adult plants gained at the highest level of defoliation.
  • 4
    These results emphasize the importance of below-ground competition during seedling establishment and show that competition for mycorrhiza-mediated resources may be an important factor underlying seedling establishment in nutrient-poor systems. Defoliation of neighbours can increase the beneficial effect of mycorrhizae to seedlings establishing in the vicinity of larger plants, suggesting that grazing or mowing may improve seedling establishment by decreasing below-ground competition.

Introduction

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. References
  9. Supporting Information

Recent research has highlighted the importance of below-ground competition in seedling establishment and plant coexistence (Casper & Jackson 1997). Below-ground competition is usually considered to be size-symmetric (Schwinning & Weiner 1998; Cahill & Casper 2000), and it has been shown able both to reduce plant performance more than above-ground competition (Wilson 1988) and to affect seedling establishment negatively (Aguilera & Lauenroth 1993). However, there is also evidence of facilitative interactions, particularly in harsh environments, where the presence of an adult plant may provide shelter and suitable microsites for seedling establishment (Choler et al. 2001). Arbuscular mycorrhizas (AM) have been shown to mediate below-ground competition between plants (Hart et al. 2003; van der Heijden et al. 2003). Arbuscular mycorrhizal symbioses are globally distributed and most vascular plants house the Glomeromycotan fungi that form AM symbioses in their roots (Brundrett 2002). AM fungi are obligate symbionts that depend solely on plant carbon; indeed, they may consume up to 20% of total net photosynthetic carbon (Jakobsen & Rosendahl 1990). In return, the fungi improve plant nutrient and water acquisition (Marschner & Dell 1994; Augé 2001) and defend plant roots against pathogens (Mosse et al. 1981; Newsham et al. 1995). The mycorrhizal symbiosis can be very beneficial to the seedlings of herbaceous plant species when growing singly in pot cultures (Moora & Zobel 1998; Kytöviita et al. 2003). However, several factors are known to affect the mycorrhizal benefit (i.e. host plant response to mycorrhizal infection) including soil nutrient availability, light environment and identity of the AM fungal partner (van der Heijden et al. 1998; Gehring 2003; Schroeder & Janos 2004). Competition is considered to be one of the main biotic factors shaping those late-successional plant communities that are typically intensively mycorrhizal (Allen & Allen 1990). However, the role of mycorrhizal symbiosis in plant competition is not clear in systems where most plants are highly mycotrophic and depend on mycorrhiza-mediated nutrient and water acquisition and where competition for resources is intense. It has been shown that increasing plant density reduces the mycorrhizal benefit to plants (Koide 1991; Allsopp & Stock 1992), suggesting that mycorrhizal symbiosis may be of less importance in highly competitive systems where plant density is high. Furthermore, there is evidence that seedlings gain little mycorrhizal benefit when they grow in the vicinity of an adult plant (Moora & Zobel 1998; Kytöviita et al. 2003), indicating that mycorrhizas may mediate size-asymmetric competition. In nature, seedlings can become mycorrhizal as early as the cotyledon stage (Read et al. 1976) and it has also been suggested that some facilitative interactions may exist between seedlings and adult plants through AM symbiosis (Gange et al. 1993). Simard & Durall (2004) postulated that seedlings growing in the vicinity of an established plant are colonized faster by AM fungi than seedlings growing alone. Furthermore, when seeds germinate in vegetated sites, the resulting seedlings are very likely to form a symbiosis with mycelium that is already in symbiosis with established vegetation. Thus, seedlings growing in the vicinity of the adult plants may access nutrients using the pre-existing fungal hyphal network supported by the established plant and, as such, may gain the nutritional benefits from AM symbiosis earlier than seedlings growing alone.

We used a glasshouse experiment to study the effect of AM symbiosis on below-ground interactions between seedlings and established adult plants in a system where plants compete for below-ground resources. We grew seedlings with and without an AM fungus in the presence and absence of a large adult plant. The adult plants were allocated to different defoliation treatments to manipulate their ability to support the AM fungus. We assumed that the competitive effect of seedlings on adults was negligible and focused on the effects of neighbouring adult plants on seedlings. We aimed to answer the following questions:

  • 1
    Do AM fungi affect the competitive balance between seedlings and adults? In non-mycorrhizal systems defoliation may potentially affect only the size of the adult plant and thus root competition, whereas in AM systems defoliation of the adult plant may also affect its ability to support AM fungi.
  • 2
    Does defoliation affect the competitive balance in mycorrhizal and non-mycorrhizal systems?

Materials and methods

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. References
  9. Supporting Information

study organisms and pregrowing conditions

Our study species, Gnaphalium norvegicum Gunn., is a rosette-forming perennial herb which typically grows in alpine meadows of the northern hemisphere (Hultén & Fries 1986). It has poor capabilities for vegetative reproduction and thus reproduces mainly generatively (Söyrinki 1938). In natural ecosystems it is colonized by AM fungi as early as the seedling stage (Pietikäinen et al. 2005). The seeds of G. norvegicum were collected from a low arctic meadow in September 1999, in Kilpisjärvi, north-west Finland (69°03′N, 20°50′E). To obtain mature adult plants, the seeds were first stratified at 4 °C for 2 months and then sown in sterile sand in March 2001. The seedlings were transplanted into individual pots and grown in a glasshouse for 6 months. In September 2001 the plants were placed in a climate chamber set to +8 °C where the plants were kept for 6 months until transplanting into experimental pots in March 2002. At the time of transplanting, the plants were non-mycorrhizal, about 1 year old and weighed 145 ± 11 mg (dry weight; mean ± 1 SE, n = 10).

The AM fungus Glomus claroideum (V225) was used to inoculate the plants. This fungus was originally isolated from a location situated further east than the seed material but at a similar latitude (69°08′N, 27°6′E). Under experimental conditions, this fungus has been shown to be beneficial to many low arctic herb seedlings (Kytöviita et al. 2003). The spores of the fungi were produced when in symbiosis with Sibbaldia procumbens L., another arctic herb. The spores were washed from the substrate, collected on a 50-µm mesh sieve and diluted in water. The non-mycorrhizal inoculum was prepared in a similar manner but from non-mycorrhizal plants.

experimental set-up

We performed a glasshouse experiment in which seedlings were grown in the presence and in the absence of a conspecific adult plant, with and without the AM fungus. Each adult plant was assigned to one of the three defoliation treatments: no defoliation, 50% leaf area removed or 75% leaf area removed (Fig. 1). In total we had eight different treatment combinations with 14 replicates in each.

image

Figure 1. Experimental set-up. Five seedlings of Gnaphalium norvegicum were grown in the presence and in the absence of a conspecific adult plant, with and without the arbuscular mycorrhizal (AM) fungus. Each adult plant was assigned to one of the three defoliation treatments: no defoliation, 50% leaf area removed or 75% leaf area removed (14 replicates for each treatment).

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First, plastic pots (diameter 14.5 cm, height 13 cm) were filled with 1.2 dm3 of artificial soil. The soil was prepared by mixing autoclaved (20 min at 121 °C) sand with non-sterile, unfertilized peat in a ratio of 5 : 1. To this mixture were added 1.5 g bone meal and 5 g dolomitic lime per litre of sand. Bone meal is a slow-release organic fertilizer whereas peat addition increases the organic matter content and introduces natural soil microbes into the pots, but does not contain AM propagules. The initial nutrient contents were 182 µg P and 119 µg N g−1 soil.

Pots with an adult plant were prepared by transplanting one non-mycorrhizal G. norvegicum plant into the centre of each pot. These plants, referred to hereafter as adults, were allowed to become established for 5 weeks before sowing the seeds. In April 2002 approximately 25 seeds (stratified for 5 months at +5 °C) of G. norvegicum were sown in each pot at 5 cm distance from the centre of the pot. Most of the seeds germinated within a week. The seedlings were counted daily and marked with coloured pins as they emerged. After the first week, a cohort of five seedlings evenly spaced and of the same age or as close as possible in age was chosen in each pot. All the other seedlings were removed. The adult plant was growing in the centre of the pot and the seedlings grew around it in such a way that there was no competition for light.

The pots were placed in a glasshouse where natural light was supplemented with Osram HQI lamps to give a photoperiod of 20 h day/4 h night. The pots were watered daily or every other day with tap water. Full nutrient solution without phosphorus was applied twice during the experiment. The two fertilizations added in total 10.1 mg N and 6.5 mg K and trace amounts of Ca, Mg, Fe, Mn, Zn, Cu and Na to each pot.

mycorrhizal inoculation

The pots with an adult plant were assigned to either AM or non-mycorrhizal treatment at the time of transplanting the adult plants. Each pot received 5 mL of Gl. claroideum (V225) inoculum (containing approximately 700 spores) or the same amount of non-mycorrhizal inoculum, applied in the centre of the pot directly on the roots of the adult plant. Therefore, the colonization of the seedlings germinating 5 weeks later in the mycorrhizal pots was likely to result from contact with hyphae growing from the roots of the adult plant. The pots without an adult plant were assigned to be AM or non-mycorrhizal at the time of sowing the seeds and received either 1.5 mL Gl. claroideum (V225) inoculum containing approximately 500 spores or the same amount of non-mycorrhizal inoculum applied directly on the seeds sown.

defoliation treatments

The adult plants were defoliated twice during the experiment: just after sowing the seeds and 3 weeks later. On both occasions 0%, 50% or 75% of the leaf area was removed and these defoliation treatments are referred to as D0, D50 and D75, respectively. In total, the two defoliation events removed 0.08 ± 0.01 g and 0.13 ± 0.008 g of leaf biomass in the D50 treatment and 0.12 ± 0.02 g and 0.17 ± 0.01 g of leaf biomass in the D75 treatment in non-mycorrhizal and AM pots, respectively (mean ± 1 SE).

final harvest and measurements

The final harvest was conducted 5 weeks after sowing the seeds when the seedlings were approximately 4 weeks old. The total biomass, shoot : root ratio, AM colonization in the roots and the shoot nitrogen (N) concentration were measured in the seedlings and the adult plants. Shoot phosphorus (P) concentration was measured only from the shoots of the adult plants as there was not enough seedling mass for P analysis. The seedling material from each pot was pooled and an average per pot was used in the later calculations.

Roots were separated from the soil by washing. The harvested shoots and the washed roots were dried at 70 °C. Samples for mycorrhizal measurements were taken before drying the washed roots. Two and 10 2.5-cm root segments were taken from each of the seedling and adult plant root systems, respectively, and stored in 50% ethanol. In the adult plants, the biomass of removed root segments was assumed to be insignificant (< 1%) in relation to the total root biomass and no measures were taken to estimate the removed root biomass. To correct the seedling root biomass for the amount lost due to sampling, 25 root segments from the AM and non-mycorrhizal seedlings were dried and weighed, respectively, and the data used to calculate the total seedling root biomass for each pot.

To measure mycorrhizal colonization the root samples were stained with trypan blue as described by Phillips & Hayman (1970). Root colonization by the AM fungus was measured by calculating 10 intersections per 2.5-cm root segment using the gridline intersection method of McGonigle et al. (1990). Thus, the frequency of AM structures in the roots of the adult plant and in the roots of the seedlings was based on 100 intersections (10 intersections in two root segments in each of the five seedlings in the pot; 10 intersections in 10 root segments of the adult plant in the pot). At each intersection the presence of AM-hyphae, vesicles and arbuscules was noted.

The N concentration was analysed using the dynamic flash combustion technique (CE Instruments EA 1110 Elemental Analysers). In some seedling samples there was too little plant shoot material to measure the N concentration reliably. Therefore, the number of replicates was reduced; the number of replicates for the shoot N concentration and N content in the seedling data was between 4 and 14. Analysis of P concentration in plant shoots was modified from the procedure described by John (1970). Dried and milled plant tissue was acid-digested using the Paar001H program in the Paar Physica multiwave sample preparation system (Perkin Elmer). Phosphorus concentration was measured as absorbance at 882 nm (UV-160A; Shimadzu). Reference plant material gave identical P concentrations when determined as above and when determined by HCl extraction of ashed samples followed by plasma emission spectroscopy (Novalab Oy, Finland).

calculations and statistical analysis

Mycorrhizal benefit was defined as the mass or nutrient content ratio of mycorrhizal (AM) to non-mycorrhizal (NM) plants. This ratio has been used in previous studies as a measure of mycorhizal benefit (Kytöviita et al. 2003; Ruotsalainen & Kytöviita 2004). In the seedlings, the mycorrhizal benefit was calculated as the ratio of AM to non-mycorrhizal in terms of both total biomass and shoot N content. In the adult plants, the mycorrhizal growth benefit was calculated as the AM to non-mycorrhizal ratio of the sum of total biomass at final harvest and biomass removed in the two defoliation occasions. Below-ground competitive intensity (BCI; Cahill & Casper 2000) was calculated for the seedlings in each pot. First, the mean biomass of seedlings growing alone was calculated separately for AM and non-mycorrhizal seedlings. This was then used for calculating the BCI:

  • BCI = (mean biomass of seedlings growing alone – seedling biomass with adult plant)/(mean biomass of seedlings growing alone).

Statistical analyses were conducted using SPSS 8.0 for Windows (SPSS Inc.). Treatment effects were analysed with a two-factor anova. When testing effects on seedling variables (biomass, root : shoot ratio, shoot N concentration and N content), the two factors were neighbouring adult plant with four levels (seedlings alone, with D0, D50 and D75 adult plant) and mycorrhiza with two levels (AM, NM). When testing effects on the adult plant variables (biomass, root : shoot ratio, shoot N and P concentration), the factors were defoliation with three levels (D0, D50, D75) and mycorrhiza with two levels (AM, NM). When testing effects on BCI, the two factors were defoliation intensity of the adult plant with three levels (D0, D50, D75) and the mycorrhiza with two levels (NM, AM). The homogeneity of variances was tested with Levene's test. To satisfy the assumptions of anova, i.e. normal distribution and homogeneity of variance, dependent variables were transformed when necessary. In the case of the total biomass data of the adult plant, the assumption of homogeneity of variance was slightly violated (Levene's test, P = 0.029) even after transformations, but this was taken into consideration when interpreting the results. Only when a significant interaction between two treatments was found was one-factor anova (not shown) followed by the Student–Newman–Keuls multiple comparisons test performed comparing the eight (in the case of the seedling data) or six (in the case of the adult plant data) treatment combination means. The interpretation of the interaction was based on these comparisons.

A plant's size is likely have direct effects on its competitive potential, and as the mycorrhizal adult plants tended to be larger than the non-mycorrhizal adult plants, seedlings could potentially respond mainly to the adult plant size and not to the shared mycorrhizal symbiont. In order to elucidate the effects of adult plant size irrespective of the treatment on the seedling biomass, we conducted regression analyses on total seedling and total adult plant biomass separately for all treatments. Furthermore, we tested if the AM fungal hyphal colonization rate in seedlings’ own roots or in the adult plant roots explains the seedling total biomass separately for each defoliation treatment using linear regression analysis.

The treatment effects on the mycorrhizal colonization rate in AM seedlings and in AM adult plants were tested with one-factor anova. In these one-factor anova models, the treatment factor had four levels (AM seedlings alone, AM D0, AM D50, AM D75) for variables presenting seedling mycorrhizal colonization, and three levels (AM D0, AM D50, AM D75) for variables presenting mycorrhizal colonization of the adult plant. The effects of defoliation of the adult plant on mycorrhizal benefit in the seedlings were tested with one-factor anova followed by the Student–Newman–Keuls test.

Results

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. References
  9. Supporting Information

adult plant biomass, n and p concentrations and am colonization

The adult plants grew well during the experiment; non-mycorrhizal plants increased their biomass sixfold, and mycorrhizal plants increased their biomass ninefold. The total biomass and shoot N concentration in the adult plants were affected both by the defoliation and by the mycorrhizal treatments (Table 1). The shoot P concentration and root : shoot ratio were only affected by mycorrhiza. The total biomass of adult plants in the 50% defoliation treatment was decreased by approximately 36% in comparison with non-defoliated adult plants, and the 75% defoliation treatment decreased total biomass by approximately 60% (Table 2). Defoliation increased the shoot N concentration in the AM treatment. The mycorrhizal symbiosis was functional: the AM adult plants had a lower root : shoot ratio, a higher total biomass and about three times higher shoot P concentration than the non-mycorrhizal adult plants. The mycorrhizal growth benefit was 1.40 ± 0.08, 1.42 ± 0.07 and 1.28 ± 0.07 in the 0%, 50% and 75% defoliation treatments, respectively (mean ± 1 SE, n = 14).

Table 1. anova statistics for main and interaction effects of defoliation (def) and mycorrhiza (myc) on total biomass, root : shoot ratio, shoot nitrogen [N] and phosphorus [P] concentration in adult plants
Source of varianceTotal biomassRoot : shoot[N][P]
d.f. error 78 MS error 21.927d.f. error 78 MS error 0.029d.f. error 78 MS error 0.043d.f. error 78 MS error 0.009
FPFPFPFP
def48.790.000 1.700.19013.410.000  2.300.107
myc21.350.00037.010.00033.080.000600.730.000
def × myc 1.030.363 0.0010.999 4.440.015  2.360.102
Table 2.  Treatment effects on adult plant total biomass (excluding the defoliated mass), root : shoot ratio, shoot nitrogen [N] and phosphorus [P] concentration. Values are mean ± 1 SE, n = 14. Treatments are mycorrhizal (AM), non-mycorrhizal (NM), and 0%, 50% and 75% of leaf area defoliated (D0, D50 and D75). Letters indicate significant differences between means (multiple comparisons Student–Newman–Keuls test, only performed when a significant interaction between factors was found in two-factor anova, see Table 1)
TreatmentTotal biomass (mg)Root : shoot[N] (mg g−1)[P] (mg g−1)
NM D0 978 ± 1375.0 ± 0.416.7 ± 0.8 ab1.04 ± 0.07
NM D50 622 ± 585.9 ± 0.616.1 ± 0.6 a1.11 ± 0.11
NM D75 412 ± 535.3 ± 0.717.7 ± 0.5 ab1.00 ± 0.04
AM D01369 ± 782.9 ± 0.117.6 ± 0.4 ab3.07 ± 0.16
AM D50 875 ± 453.3 ± 0.218.8 ± 0.4 b3.66 ± 0.20
AM D75 519 ± 292.8 ± 0.221.9 ± 0.5 c3.92 ± 0.20

No AM structures were observed in the non-mycorrhizal treatments and the colonization rate in these plants was zero. All adult plants assigned to mycorrhizal treatment were colonized by AM fungi: frequency of hyphae was 23 ± 2% and frequency of arbuscules 10 ± 1% of root length (mean ± 1 SE, n = 56). Frequency of vesicles was very low (less than 1% of root length). The defoliation treatments did not affect either the frequency of AM hyphae (F2,39 = 0.111, P = 0.895) or the frequency of arbuscules (F2,39 = 0.495, P = 0.613).

biomass, n concentration and am colonization in seedlings

In the seedlings, both the mycorrhiza and the neighbouring adult plant with different defoliation treatments significantly affected the biomass, root : shoot ratio and shoot N concentrations (Table 3). Seedlings grown in the absence of an adult plant were larger than those grown in the presence of an adult plant (Table 4). In general, AM seedlings grew larger than their non-mycorrhizal controls. However, both the AM and the non-mycorrhizal seedlings were of the same size when grown in the presence of a non-defoliated adult plant. Defoliation of the adult plant affected the total biomass of the seedlings only in the AM treatment (Table 4). The total biomass of AM seedlings was higher the greater the extent of the defoliation of the adult plant. The regression analyses on seedling and adult plant biomasses showed that whereas seedling biomass was not systematically affected by the adult plant biomass (see supplementary Fig. S1), hyphal colonization rate in adult plant roots did have a significant effect on seedling total biomass in non-defoliated systems (R2 = 0.478, P = 0.006). When the adult plant was defoliated, no significant relationship between the hyphal colonization rate in adult plant roots and seedling total biomass was found. In addition, the hyphal colonization rates in seedlings’ own roots did not explain the seedling total biomass.

Table 3. anova statistics for main and interaction effects of neighbouring adult plant (adult) and mycorrhiza (myc) on seedling total biomass, root : shoot ratio and shoot nitrogen concentration [N] and content
Source of varianceTotal biomassRoot : shoot[N]N content
d.f. error 104 MS error 0.039d.f. error 104 MS error 0.019d.f. error 73 MS error 0.067d.f. error 73 MS error 0.052
FPFPFPFP
adult 52.210.000  6.730.00018.940.000 38.450.000
myc141.190.000321.640.00039.360.000161.370.000
adult × myc  9.580.000  3.450.019 0.900.448  6.5190.001
Table 4.  Seedling total biomass, root : shoot ratio and shoot nitrogen [N] concentration and N content when grown in different mycorrhiza and neighbouring adult plant treatments. Values are mean ± 1 SE, n = 14 for total biomass and root : shoot ratio, n = between 4 and 14 for N concentration and N content. Treatments are arbuscular mycorrhizal (AM), non-mycorrhizal (NM), seedlings alone (alone), and seedlings with 0%, 50% and 75% defoliated adult plant (D0, D50 and D75). Letters indicate significant differences between means (multiple comparisons Student–Newman–Keuls test, only performed when a significant interaction between factors was found in two-factor anova, see Table 3)
TreatmentTotal biomass (mg)Root : shoot[N] (mg g−1)N content (µg)
NM alone 3.32 ± 0.50 b1.75 ± 0.18 D19.9 ± 0.9 32.2 ± 8.8 x
NM D0 1.50 ± 0.21 a1.86 ± 0.09 D15.4 ± 0.8 10.2 ± 1.9 w
NM D50 1.26 ± 0.11 a1.87 ± 0.19 D16.2 ± 1.1 10.6 ± 1.5 w
NM D75 1.66 ± 0.20 a1.93 ± 0.14 D17.8 ± 0.5 12.2 ± 1.7 w
AM alone14.63 ± 1.31 d0.44 ± 0.02 A25.2 ± 1.1263.8 ± 33.5 z
AM D0 2.03 ± 0.21 a0.84 ± 0.09 C18.1 ± 0.4 23.8 ± 2.4 x
AM D50 3.77 ± 0.78 b0.76 ± 0.10 BC19.8 ± 0.6 51.5 ± 12.4 x
AM D75 6.29 ± 0.98 c0.59 ± 0.02 B21.9 ± 0.5 92.0 ± 15.4 y

As in the adult plants, the root : shoot ratio in the seedlings was lower in the AM than in the non-mycorrhizal treatment (Table 4). However, in the AM seedlings the root : shoot ratio was decreased by increasing defoliation intensity of the adult plant.

The seedling shoot N concentration and N content was higher in the AM seedlings than in their non-mycorrhizal controls (Tables 3 & 4). In the presence of a neighbouring adult plant, the shoot N concentration and N content of the seedlings were lower than in the absence of an adult plant. When the neighbouring adult plant was defoliated, in both AM and non-mycorrhizal systems, the N concentration of seedling shoots was higher than in the respective non-defoliated systems. However, only in the AM systems was the shoot N content of the seedlings grown in the presence of a defoliated adult plant higher than that of the seedlings grown with a non-defoliated adult plant. As a result, the mycorrhizal benefit, in terms of both biomass and N content, was highest in seedlings grown without an adult plant, and lowest in those seedlings grown in the presence of a non-defoliated adult plant (Fig. 2). In addition, defoliation of the adult plant increased the mycorrhizal benefit to the seedlings.

image

Figure 2. Effects of neighbouring conspecific adult plants on mean mycorrhizal benefit in seedlings measured as the ratio of mycorrhizal to non-mycorrhizal in terms of biomass and shoot N content. The adult plants were subjected to three levels of defoliation (0%, 50% and 75% of leaf area removed). Values are means ± 1 SE, n = 14 for biomass benefit and n = between 4 and 14 for shoot N content benefit. Letters indicate significant differences between means (multiple comparisons Student–Newman–Keuls test).

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The BCI experienced by the seedlings was affected both by mycorrhiza (F1,78 = 16.03, P < 0.001) and the defoliation of the neighbouring adult plant (F2,78 = 6.40, P = 0.003). The BCI was higher in the AM treatment, but the defoliation of the neighbouring adult plant decreased the BCI both in AM and non-mycorrhizal treatments (Fig. 3).

image

Figure 3. Below-ground competitive intensity (BCI) experienced by the seedlings as affected by arbuscular mycorrhiza and a neighbouring conspecific adult plant that was defoliated by 0%, 50% and 75%. Values are means ± 1 SE, n = 14.

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The frequency of AM hyphae in the roots of the AM seedlings averaged about 21 ± 1% of root length, and the average frequency of arbuscules was approximately 14 ± 1% (mean ± 1 SE, n = 56). The frequency of vesicles was less than 1% of root length. The frequency of hyphae (F3,52 = 2.016, P = 0.123) and arbuscules (F3,52 = 2.607, P = 0.061) were unaffected by the neighbouring adult plant with different defoliation treatments. No AM structures were observed in the non-mycorrhizal treatments and the colonization rate in these seedlings was zero.

Discussion

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. References
  9. Supporting Information

The results of this experiment emphasize the importance of competition in below-ground interactions between adult plants and seedlings. The AM symbiosis was very beneficial to seedling growth in the absence of an adult plant; the biomass of the mycorrhizal seedlings growing alone was approximately five times greater, and their shoot N content was approximately eight times higher than the non-mycorrhizal controls. The large benefit of mycorrhizae to seedlings grown in the absence of an adult plant shows that the seedlings were capable of allocating carbon to the fungus and forming a functional relationship with the AM fungus. However, the mycorrhizal benefit to seedlings grown in the presence of a non-defoliated adult plant was low. These results are similar to the findings of Moora & Zobel (1998) and Kytöviita et al. (2003) under laboratory conditions. Furthermore, they are in line with field observations that seedlings do not receive significant benefits when associated with adult plants (Gehring & Connell 2006) and the benefit of mycorrhizal inoculum is greatest in the absence of neighbouring vegetation (Scherff et al. 1994).

In this experiment we excluded light competition and focused on the below-ground interactions between seedlings and an adult plant. Competition for below-ground resources is composed of direct root competition (resources taken up by plant roots without mycorrhizal mediation) and competition for mycorrhiza-mediated resources. Thus, in the non-mycorrhizal systems, the effects of a neighbouring adult plant can be attributed solely to root competition. In the mycorrhizal experimental systems, it was not possible to separate resource acquisition by the mycorrhizal and non-mycorrhizal pathways in the roots of inoculated plants. However, there is evidence that mycorrhizal symbiosis inactivates the plants’ own P uptake pathway, and up to 100% of plant total phosphorus gain can be through arbuscular mycorrhizal fungi, while only a minor fraction may be directly acquired through plant roots (Smith et al. 2004). Therefore, we suggest that in the mycorrhizal systems the seedlings and adult plant were competing for limited nutrient resources provided by their AM fungal symbiont and that the competition for mycorrhiza-mediated resources was more important than root competition. Furthermore, we found no significant systematic relationship between adult plant root biomass and seedling biomass. Thus, root competition alone cannot explain the pattern of seedling growth.

The defoliation treatments aimed to manipulate the ability of the adult plant to support (i.e. provide carbon to) its mycorrhizal symbiont and thus decrease their ability to compete for mycorrhiza-mediated resources. Defoliation did decrease mycorrhizal benefit in the adult plants and a similar response has also been shown in experiments with herbaceous plants by Grime et al. (1987). Furthermore, defoliation of the adult plants induced a shift in mycorrhizal benefit in the seedlings. With increasing defoliation intensity of the adult plants the mycorrhizal benefit in the seedlings increased in terms of both shoot nitrogen content and total biomass. Similar results are reported by Jakobsen (2004) with tomato seedlings competing with established cucumber plants. As defoliation of the adult plant resulted in increased seedling size only in the mycorrhizal systems, it seems that defoliation significantly affected competition for mycorrhiza-mediated resources only, and did not affect the root competition the seedlings experienced. This view is further supported by the association of AM fungal colonization rate in adult plant roots with the seedling biomass. In the non-defoliated systems there was a significant relationship between AM hyphal colonization rate in adult plant roots and the seedling biomass: the higher the colonization rate, the smaller the seedlings. When the neighbouring adult plant was defoliated no such relationship between seedling biomass and adult plant colonization rate was found. According to Eissenstat & Newman (1990) a larger Plantago lanceolata individual can affect the mycorrhizal colonization intensity in a smaller conspecific. These results indicate that plants may affect the intensity and functioning of the mycorrhizal symbiosis in their conspecific neighbours.

As AM fungi grow in the soil from root to root they can connect plants of both the same (Hirrel & Gerdemann 1979) and different species (Heap & Newman 1980; Chiariello et al. 1982; Francis & Read 1984), forming a common mycorrhizal network. The functioning of this network and its role in seedling establishment is an area of active discussion (e.g. Leake et al. 2004; Simard & Durall 2004; Selosse et al. 2006; Taylor 2006). The presence of a common mycorrhizal network was not directly demonstrated in the present experiment, but the mycorrhizal inoculation was undetaken in such a way that colonization of the seedling was most likely to result from contact with hyphae growing from the roots of the adult plant. Thus, the fact that all the seedlings in the mycorrhizal treatment were colonized by AM fungus provides indirect evidence for the presence of hyphal connections between the adult plant and the seedlings (i.e. a common mycorrhizal network). On the basis of the results of this experiment, the shift in mycorrhizal benefit induced by defoliation, and the relationship between hyphal colonization in adult plant roots and seedling growth, we hypothesize that resource flow in a common mycorrhizal network could be regulated by the carbon source strength of the plant, which may be defined as the ability of the plant to supply carbohydrates per root length per time unit. The exact physiological mechanisms regulating resource exchange in AM symbioses are currently unknown, but it has recently been shown using axenic carrot root cultures that transfer of P by an AM fungus was affected by carbon availability to the root (Bücking & Shachar-Hill 2005). This hypothesis is in line with biological market models, where an individual's decision regarding with whom to co-operate is based on a comparison of the potential benefits offered by potential partners attempting to outbid each other (Hoeksema & Bruna 2000). Applied to mycorrhizal systems with multiple plant partners, the resources provided by the AM fungus would be directed towards the plant partner providing the most carbohydrate.

In summary, the results from this experiment suggest that below-ground competition for mycorrhiza-mediated resources is an important factor underlying seedling success in nutrient-poor systems. The shared mycorrhizal symbiont increased the below-ground competition intensity experienced by the seedlings, supporting the view that mycorrhizal symbiosis does not attenuate the intraspecific competition but rather amplifies it (Allsopp & Stock 1992; Hartnett et al. 1993; Moora & Zobel 1996; Watkinson & Freckleton 1997; Facelli et al. 1999; but see Eissenstat & Newman 1990). The competition for mycorrhiza-mediated resources may hinder seedling establishment near conspecific adult plants and thus affect the plant community composition. Seedlings and adult plants are reported to be colonized by different fungal communities (Husband et al. 2002), which seems a possible way to minimize negative effects of common mycorrhizal networks on seedlings. Furthermore, our experiment shows that defoliation of vegetation can increase the benefit of mycorrhizae to those seedlings establishing in the vicinity of larger plants, suggesting that grazing or mowing may improve seedling establishment by decreasing below-ground competition.

Acknowledgements

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. References
  9. Supporting Information

We thank Tuulikki Pakonen for assistance in laboratory work. This study was financially supported by the Finnish cultural foundation, the Kone foundation and the Academy of Finland (1206981).

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  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. References
  9. Supporting Information
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Supporting Information

  1. Top of page
  2. Summary
  3. Introduction
  4. Materials and methods
  5. Results
  6. Discussion
  7. Acknowledgements
  8. References
  9. Supporting Information

Figure S1 Linear regression of total seedling biomass against total adult biomass.

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