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Keywords:

  • arbuscular mycorrhizal fungi;
  • colonization rate;
  • colonization extent;
  • functional groups;
  • intraradicle mycelium;
  • extra-radicle mycelium

Summary

  1. Top of page
  2. Summary
  3. Introduction
  4. Methods and Materials
  5. Results
  6. Discussion
  7. Acknowledgements
  8. References
  •  Arbuscular mycorrhizal fungi (AMF) are important components of terrestrial communities but the basic ecology of individual AMF, including their colonization strategy, remains unclear. The colonizing behaviours of 21 AMF isolates from three families (Acaulosporaceae, Gigasporaceae and Glomaceae) were compared to test for a relationship between AMF taxonomy and colonization strategy.
  •  Both the rate and extent of colonization were considered by measuring percentage root colonization, root fungal biomass, soil hyphal length and soil fungal biomass over 12 wk.
  •  Most Glomaceae isolates colonized roots before Acaulosporaceae and Gigasporaceae isolates. The fastest colonizers were also often the most extensive. Taxonomic differences were apparent in the amount and proportion of fungal biomass found in roots vs in soil. Glomaceae isolates had high root colonization but low soil colonization, Gigasporaceae isolates showed the opposite trend whereas Acaulosporaceae isolates had low root and soil colonization. These results were similar for four different host plants.
  •  The results indicate that the colonizing strategies of AM fungi differ considerably and that this variation is taxonomically based at the family level. Arbuscular mycorrhizal fungal taxonomy therefore has a functional basis.

Introduction

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

Arbuscular mycorrhizal fungi (AMF) (Division Zygomycetes, Order Glomales) are involved in an important mutualism with most terrestrial plants. Differences in morphological and developmental traits of AMF have been used to classify them into seven genera and five families (Morton & Redecker, 2001). It is not yet clear whether existing taxonomic groups, which are based almost exclusively on morphological traits, are useful predictors of their ecology. There is some evidence for different ecological strategies among AMF, both in terms of their function in ecosystems (Newsham et al., 1995; van der Heijden et al., 1998) and on a smaller, physiological and morphological scale. For example, AMF isolates have recently been shown to differ with respect to phosphorus metabolism in extra-radicle hyphae (Boddington & Dodd, 1999) and also in their nutrient transfer efficiencies (Smith et al., 2000). However, studies that examine an array of AMF species from different taxonomic groups are lacking and life-history strategies for AMF have yet to be described.

A life-history strategy provides an ecological description of how an organism fulfills its life-cycle requirements. A life-history strategy can be described using a wide range of species traits but usually these include colonizing ability, dispersal ability, tolerance of stress and disturbance, investment into reproduction vs vegetative growth, and mode of reproduction (Grime, 1977; Pianka, 1970).

In this paper we focus on one aspect of AMF life-history; namely, colonizing ability. Our objectives are to describe the colonizing strategies of some AMF and to determine whether there is a taxonomic basis for variation in colonization ability among AMF. Colonization ability includes both the rate and extent of colonization. These are commonly used measures of AMF activity so they make a good starting point for a description of AMF life-history strategies. Because AMF colonize plant roots and soil, we studied the rate and extent of colonization of both plant roots and soil by 21 different AMF from three families (Acaulosporaceae, Glomaceae and Gigasporaceae). Specifically, we addressed the following four questions:

Do taxonomic groups differ in colonization rate?

It is generally believed that members of Glomaceae and Acaulosporaceae have a highly infective extra-radicle mycelium whereas members of Gigasporaceae regenerate most frequently from spores (Tommerup & Abbot, 1981; Biermann & Lindermann, 1983; Morton, 1993; J. N. Klironomos & M. M. Hart, unpublished). We predicted that members of the Gigasporaceae would colonize plant roots more slowly than members of either the Acaulosporaceae or the Glomaceae (Table 1). We suspected that a spore might require specific conditions or a dormancy period for regeneration. Therefore, AMF that depend on spores for colonization are likely to be at a disadvantage compared with AMF that can colonize immediately from hyphal fragments.

Table 1.  Predicted differences in the rate, extent and location of colonization by three arbuscular mycorrhizal fungi (AMF) families
Colonization traitAcaulosporaceaeGigasporaceaeGlomaceae
RateFastSlowFast
ExtentLimitedExtensiveLimited
LocationRootSoilRoot

Do taxonomic groups differ in colonization extent?

Taxonomic differences among AMF in hyphal structure and mycelial architecture are well known. Members of the Glomaceae and Acaulosporaceae tend to have very delicate, diffuse hyphae whereas members of the Gigasporaceae tend to have robust, densely aggregated hyphae (Jakobsen et al., 1992; Smith et al., 2000). Such differences could be the result of differences in hyphal longevity. Delicate, diffuse hyphae may simply be shorter-lived than robust, densely aggregated forms. This is based on the premise that large-bodied organisms have slower rates of turn-over than small-bodied organisms. We predicted that AMF with delicate, diffuse hyphae (i.e. Glomaceae and Acaulosporaceae) would be limited in the extent of their colonization. Conversely, AMF with robust, densely aggregated hyphae (i.e. Gigasporaceae) would be better able to create a large mycelium (Table 1).

Do taxonomic groups differ in the extent of root vs soil colonization?

AMF may have two very different colonization patterns: mycelia primarily within roots vs mycelia primarily within soil. Based on our personal observations, we predicted that members of Glomaceae and Acaulosporaceae should colonize roots more profusely than members of Gigasporaceae. Conversely, members of the Gigasporaceae should colonize soil more than members of the Acaulosporaceae or Glomaceae (Table 1). This prediction is based on the assumption that AMF cannot invest heavily in both intra- and extra-radicle mycelia simultaneously. We reasoned that it is unlikely that a plant host could sustain a symbiosis involving extensive carbon drain.

Do taxonomic groups differ in both rate and extent of colonization?

Based on the arguments made above, we predicted that AMF differ in both their rate and extent of colonization. It is important to establish the relationship between colonization and extent for AMF because ecological theory dictates that there is a necessary trade off between the two. That is, an organism cannot be, at once, a quick and extensive colonizer (Grime, 1977; Pianka, 1970).

Methods and Materials

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

AMF isolate

We chose 21 isolates from collections of AMF maintained at the University of Guelph and at Premier Tech (Riviere du Loup, Que. Canada). All isolates were isolated from the Long-term Mycorrhizal Research Station, at the University of Guelph, Ontario, Canada, unless otherwise indicated. Isolates were chosen to represent three AMF families: Acaulosporaceae, Glomaceae and Gigasporaceae. More than just one isolate was chosen per family to provide replication. More isolates were chosen from the Glomaceae than from the other two families because Glomaceae is a larger family (Morton & Redecker, 2001).

Family Acaulosporaceae

Acaulospora morrowaie Spain and Schenck

Acaulospora spinosa 1 Walker and Trappe

Acaulospora spinosa 2

Entrophospora columbiana Spain and Schenck

Family Gigasporaceae

Gigaspora gigantea (Nicol. & Gerd.) Gerdemann & Trappe

Gigaspora margarita (Becker & Hall)

Scutellospora calospora (Nicol. & Gerd.) Walker & Sanders

Scutellospora heterogama (Nicol. & Gerd.) Walker & Sanders

Scutellospora pellucida (Nicol. & Schenck) Walker & Sanders

Family Glomaceae

Glomus aggregatum Schenk & Smith emend. Koske

Glomus claroideum Schenck & Smith

Glomus constrictum Trappe

Glomus etunicatum Becker & Gerdemann

Glomus geosporum (Nicol. & Gerd.) Walker

Glomus intraradices Quebec1 (Q1) Schenck & Smith*

Glomus intraradices Quebec 2 (Q2)*

Glomus intraradices Kansas (K)*

Glomus intraradices Isreal (I)*

Glomus intraradices France (F)*

Glomus intraradices Japan (J)*

Glomus mosseae (Nicol. & Gerd.) Gerdemann & Trappe

*indicates that isolates do not originate from the LTMRS.

Fungal inoculum

Because AMF may regenerate from spores as well as from hyphal fragments, we used an inoculum consisting of hyphal fragments, spores and root fragments. The starting amounts of isolates were equalized in terms of fungal biomass (ergosterol). Samples of inoculum from each isolate were homogenized and ergosterol was extracted with hexane by HPLC following the method of Grant & West (1986). Commercial ergosterol (5,7,22-ergostatrien-3b-ol) was used as the standard. Once we determined the concentration of ergosterol in whole inoculum, each isolate was diluted to achieve a common ergosterol density. We chose to equalize our samples based on the biomass of propagules instead of the number of propagules for two reasons. First, it would be very impractical to count propagule numbers for AMF because they are able to propegate from hyphal fragements as well as spores, and it is difficult to say how many hyphal fragments constitute a propagule. Second, biomass standardization is superior for our purposes, since we are attempting to reveal differences among AMF based on a standardized starting point. Thus, for our purposes, it is counter-productive to standardize based on the final extent of colonization.

Soil pretreatment

Isolates were cultured in soil before the application of experimental treatments by growing them with a surrogate host. We chose leek (Allium porrum L.) as the surrogate host because it is commonly used in trap cultures and is known to host a wide variety of AMF. Surrogates were grown in Conetainers (4 cm in diameter × 20.5 cm deep) (Stuewe and Sons Inc., Corvallis, OR, USA) which were two-thirds filled with a 1 : 1 mixture of sterile, low P potting soil and silica sand. This mixture was first sterilized by heating it to 120°C for 30 min, left to cool overnight, then heated again at 120°C for 30 min. A week later, AMF inoculum was added to each container and covered with additional soil-sand mixture. Three Allium seeds were added per Conetainer and allowed to grow for 30 d, at which time shoots were harvested and removed. The ‘cultured’ soil in the Conetainers was then subjected to experimental treatments.

Experimental treatments

Three experimental treatments were applied to Conetainers using a completely randomized factorial design. The three experimental treatments were (1) 21 AMF isolates, (2) six harvest dates and (3) four host plants, for a total of 504 different treatments. These were replicated five times for a total of 2520 experimental units.

(1) AMF isolate

We used 21 different AMF isolates as described above.

(2) Harvest date

Conetainers were harvested six times over a 12-wk period: at 5, 12, 19, 26, 54 and 70 d. These dates were chosen to measure differences in the rate of colonization among AMF and to ensure full colonization for testing for differences in colonization extent among AMF. Data from sequential harvests were used to calculate the rate of colonization and data from the last harvest at 70 d were used to calculate the extent of colonization. At each harvest, five randomly selected replicate Conetainers were sampled from each treatment.

(3) Host plant

We used four host plants representing two growth habits: grasses and forbs. We chose two species from each growth habit for the purposes of replication. These were: English plantain (Plantago lanceolata L.), common plantain (Plantago major L.), Kentucky-blue grass (Poa pratensis L.) and annual blue grass (Poa annua L.). These species were chosen as highly mycotrophic plants, which are commonly found in old fields and meadows, the origin of most of the AMF isolates. Three seedlings of each species (c. 1 cm radicle) were added to each Conetainer, at which point 50 ml of soil filtrate (mesh size 35 µm) from each isolate was added to each Conetainer to control for differences in other soil organisms among Conetainers. Locations of plants in each treatment were randomized on 14 greenhouse benches (1.5 m by 8 m) at Premier Tech., Riviere-du-Loup, Quebec, Canada. Each Conetainer was subjected to 14 h of light (12.2 Watts/m2 over a 24-h period) for 16 wk between July and October 2000. Conetainers were watered and fertilized as needed with a low P fertilizer.

Control for background fungi

It is possible that some nonGlomalean fungi may be counted in ergosterol measurements of root and fungal soil biomass. To estimate the amount of fungi other than AMF in our treatments, we included 20 control Conetainers in the experiment. These controls were similar in every way to treated Conetainers except they lacked AMF inoculum.

Colonization rate and extent

The four dependent variables in this study were (1) the rate of initial root contact measured in terms of earliest date of intraradicle colonization (2) the extent of root colonization measured by percentage intraradicle colonization and intraradicle fungal biomass (3) the extent of soil colonization measured by soil hyphal length and soil fungal biomass and (4) the extent of root vs soil colonization. Two different measures of the extent of colonization were included because hyphal length and percentage colonization are measures of AMF density in two dimensions (length and width) while fungal biomass is a volumetric measure of AMF density.

Rate of initial root contact

The rate of initial root contact was measured at the earliest date of root colonization evident in any of the five replicates. At each harvest date, the root system of the host plant was cleaned by first shaking off excess soil and then washing the roots with water. Next, roots were sonicated for 15 s to remove any residual matter, including external mycelium. The clean root system was then cut into 2 cm fragments from which eight fragments were randomly selected, stained with Chlorazol Black (Brundrett, 1991) and mounted on glass slides to determine percentage colonization. The presence (or absence) of fungal material (AMF hyphae or spores or vesicles) was recorded using a gridline intersect method (McGonigle et al., 1990) and the percentage of 100 intersections ‘colonized’ was calculated. The rate of soil colonization was not considered because AMF inoculum was already present in the soil at the start of the experiment.

Extent of root colonization

The extent of root colonization was determined by measuring both percentage colonization and root fungal biomass. Per cent colonization was assessed using the method described above. Root fungal biomass was measured indirectly as the concentration of ergosterol in roots. Eight 2 cm fragments of clean roots were randomly selected, homogenized and their ergosterol concentration was determined as described above.

Extent of soil colonization

The extent of soil colonization was determined by measuring soil hyphal length and soil fungal biomass. Soil hyphal length was determined as follows. A 10-g portion of soil was taken from each Conetainer and suspended in 250 ml of water. Sodium hexametaphosphate (3.6% w/v) was added and left for 16–18 h to break up soil aggregates. The soil suspension was then agitated in a blender at high speed for 2 mins and stirred with an electronic stir bar. One 6 ml aliquot per sample was removed from halfway between the beaker edge and the vortex and this aliquot was added to 250 ml of distilled water plus 30 ml of sodium hexametaphosphate solution. This mixture was stirred to resuspend hyphae and 10 ml aliquots were transferred to 50 ml centrifuge tubes where they were centrifuged five times at 1000 × g. Pellets were resuspended in glycerol and centrifuged again at 75 × g for 30 s. The supernatant was filtered onto a 20-µm polyester filter which was then stained with Chlorazol Black and decanted over 1.2 µm nitrocellulose filter paper. These filters were mounted on glass slides, dried and made transparent by mounting in immersion oil. The presence (or absence) of hyphae was recorded at 140 intersections per sample and hyphal length per gram dry soil was then calculated as described by Newman (1966).

Soil fungal biomass was determined indirectly by measuring the concentration of ergosterol in 20 ml of root-free soil.

Extent of root vs soil colonization

To determine whether isolates differed in their colonization of roots vs soil, the ratio of fungal biomass in roots to fungal biomass in soil was calculated using the measurements of fungal biomass determined above.

Background fungi

Soil and root ergosterol measurements did not distinguish between AMF and other fungi. However, the amount of nonAMF fungi in soil was unlikely a confounding factor for several reasons. First, soil was sterilized before use and second, ergosterol values in control Conetainers (no AMF added) were low. Third, ergosterol values in treated containers (AMF added) were highly correlated with percentage colonization values for AMF.

Statistical analysis

The following statistical procedures were applied to test for significant (P < 0.05) differences among taxonomic groups of AMF in their rate and (or) extent of colonization. SPSS 7.0 (SPSS, 1995) was used for all analyses.

Do taxonomic groups differ in colonization rate?

Differences among AMF families in their date of first colonization was tested using a Kruskal–Wallis test. Results for all four plant hosts were pooled to maximize sample size.

Do taxonomic groups differ in colonization extent?

Differences in percentage colonization (and fungal biomass) among AMF families were tested using a one-way analysis of variance (ANOVA). Data met ANOVA assumptions of normality and equality of variance. Root colonization data for the four plant hosts were pooled because the Family*Host interaction term in ANOVA was not significant. By contrast, soil colonization data for the four plant hosts were analyzed separately because the Family*Host term in ANOVA was significant. For simplicity, however, we have presented all graphs with hosts pooled, since the differences among host for soil colonization were quantitative in nature, thus trends were similar in all hosts.

Do taxonomic groups differ in root vs soil colonization?

Differences in root vs soil colonization among AMF families were tested using a one-way ANOVA with the ratio of root to soil fungal biomass as the test variable. Data met ANOVA assumptions of normality and equality of variance. Data for the four plant hosts were analyzed separately because the Family*Host interaction term in ANOVA was significant.

Do taxonomic groups differ in both rate and extent of colonization?

Discriminant analysis was used to test for a significant difference among AMF families in both the rate and extent of colonization. Discriminant analysis attempts to find canonical functions from a pool of predictor variables (in this case percentage colonization, root fungal biomass, soil hyphal length, root fungal biomass) to reliably recognize differences among predefined groups (in this case AMF families). The canonical functions are linear equations of the predictor variables that best distinguish the groups. Predictor variables were entered into the analysis step-wise, minimizing Wilk’s lambda at each step.

Results

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

Do taxonomic groups differ in colonization rate?

Mycorrhizal fungi of some isolates were first evident in roots during the first week of the experiment and all 21 isolates had colonized host roots by week eight (Fig. 1). Colonization rates differed significantly among AMF families (χ2df=2 = 16.1, P < 0.0001). Between 92% and 100% of 12 Glomaceae isolates had colonized by week 4. By contrast, some isolates of Gigasporaceae and Acaulosporaceae colonized as late as weeks 6–8.

image

Figure 1. Harvest date on which each of the 21 Arbuscular mycorrhizal fungi (AMF) isolates had first colonized roots of a plant host. Host species have been pooled. Acaulosporaceae, grey columns; Gigasporaceae, open columns; Glomaceae, closed columns.

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Do taxonomic groups differ in colonization extent?

Root colonization (%)

The extent of root colonization by the 21 AMF isolates ranged from a mean of 3.2–84.8% (Fig. 2). Percent colonization differed significantly among AMF families (F2,83 = 8.5, P < 0.0001). Gigasporaceae and Acaulosporaceae isolates generally had lower mean percentage colonization, ranging from 3.2 to 73.4%. Isolates of Glomaceae usually had higher mean percentage colonization, ranging from 20.6 to 84.8%

image

Figure 2. Mean (±1 SE) extent of percentage root colonization for 21 Arbuscular mycorrhizal fungi (AMF) isolates. Host species have been pooled. Acaulosporaceae, grey columns; Gigasporaceae, open columns; Glomaceae, closed columns.

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Root fungal biomass

For the 21 AMF isolates, fungal biomass within plant roots ranged from a mean of 0.82–4.64 µg g−1 root d. wt (Fig. 3). Fungal biomass differed significantly among AMF families (F2,83 = 165.7, P < 0.0001). Gigasporaceae and Acaulosporaceae isolates generally had the least fungal root biomass, ranging from a mean of 0.82–1.41 µg g−1 root d. wt. Glomaceae isolates tended to have the most fungal biomass, ranging from a mean of 1.74–4.64 µg g−1 root d. wt.

image

Figure 3. Mean (±1 SE) root fungal biomass (ergosterol) for 21 arbuscular mycorrhizal fungi (AMF) isolates. Host species have been pooled. Acaulosporaceae, grey columns; Gigasporaceae, open columns; Glomaceae, closed columns.

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Soil hyphal length

For the 21 AMF isolates, soil hyphal lengths ranged from a mean of 1.32–8.43 m cm−3 (Fig. 4). Hyphal length differed significantly among AMF families for each of the four plant hosts (F2,20 = 364.7, 285.8, 521.9 and 499.1 for four hosts, and P < 0.0001 for all hosts). Glomaceae and Acaulosporaceae isolates had mean hyphal lengths ranging between 1 and 2 m cm−3, whereas Gigasporaceae isolates had mean hyphal lengths ranging between 6 and 9 m cm−3.

image

Figure 4. Mean (±1 standard error) soil hyphal length for 21 arbuscular mycorrhizal fungi (AMF) isolates. Host species have been pooled. Acaulosporaceae, grey columns; Gigasporaceae, open columns; Glomaceae, closed columns.

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Soil fungal biomass

For the 21 AMF isolates, soil fungal biomass ranged from a mean of 0.38–3.30 µg g−1 root d. wt (Fig. 5). Fungal biomass differed significantly among AMF families for each of the four plant hosts (F2,20 = 283.9, 222.7, 351.9 and 292.1 for four hosts, and P < 0.0001 for all hosts). Glomaceae and Acaulosporaceae isolates generally had the least fungal biomass with mean values consistently less than 0.71 µg g−1 root d. wt, whereas Gigasporaceae and Acaulosporaceae isolates had the most fungal biomass, with means ranging from 2.0 to 3.3 µg g−1 root d. wt.

image

Figure 5. Mean (±1 SE) soil fungal biomass (ergosterol) for 21 arbuscular mycorrhizal fungi (AMF) isolates. Host species have been pooled. Acaulosporaceae, grey columns; Gigasporaceae, open columns; Glomaceae, closed columns.

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Do taxonomic groups differ in root vs soil colonization?

AMF families differed significantly in the ratio of root to soil fungal biomass (F2,419 = 1252.180, P < 0.000). The mean root to soil ratio was 2.4 for Acaulosporaceae, 0.4 for Gigasporaceae and 5.3 Glomaceae (Fig. 6).

image

Figure 6. Relationship between root and soil fungal biomass for arbuscular mycorrhizal fungi (AMF) isolates classified by family. Acaulosporaceae, grey circles; Gigasporaceae, black circles; Glomaceae, asterisks.

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Do taxonomic groups differ in both rate and extent of colonization?

Discriminant analysis indicated that AMF families differed significantly in both the rate and extent of colonization (χfunction 12 = 3818.7, P < 0.0001, χfunction 22 = 1008.8, P < 0.0001) (Table 2). At step one of the discriminant analysis external fungal biomass was included, followed by internal root ergosterol, % whole colonization and soil hyphal length. These steps created two canonical discriminant functions. Function 1 represented soil colonization and function 2 represented root colonization (Table 3). An orthogonal plot of these two functions showed that the three AMF families were well differentiated (Fig. 7). Glomaceae occurred high along the root colonization axis (Function 2) but was low on the soil colonization axis (Function 1). Gigasporaceae occurred high on Function 1 but lower than Glomaceae on Function 2. Finally, Acaulosporaceae occurred much lower than both other groups on the axis for Function 2 and was equally low along the axis for Function 1.

Table 2.  Canonical discriminant functions (F1, F2) for four predictor values
FunctionEWilk’s λχ2dfP-value
F127.87350.0103523818.780.000
F2 2.34560.2989021008.98430.000
Table 3.  Correlation between the four predictor variables and the two canonical discriminant functions (F1, F2) for each of the four plant hosts (Plantago spp., Poa spp.)
 Plantago lanceolataPlantago majorPoa pratensisPoa annua
  1. * Stronger correlation with F1 vs F2 for a predictor variable.

Predictor variablesF1F2F1F2F1F2F1F2
Soil fungal biomass 0.71*−0.26 0.71*0.02 0.74*0.05 0.82*0.12
Soil hyphae length 0.68* 0.06 0.64*0.08 0.67*0.07 0.61*0.12
Root fungal biomass−0.21 0.66*−0.18 0.72*−0.21 0.65*−0.200.63*
Root colonization−0.04 0.15*−0.05 0.20*−0.04 0.13*−0.040.15*
image

Figure 7. Positions of arbuscular mycorrhizal fungi (AMF) isolates (classified by family) on an orthogonal plot of the two canonical discriminant functions. Acaulosporaceae, grey squares; Gigasporaceae, black squares; Glomaceae, asterisks.

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Discussion

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

We found evidence for distinct AMF colonization strategies that are related to taxonomic differences at the family level. These different strategies were evident regardless of the host plant involved.

AMF isolates showed large differences in the rate at which they initially colonize plant roots. Some isolates in our study colonized roots within 1 wk while other isolates required an additional 7 wk to colonize roots. The biological basis for this variation requires further study but the pattern of colonization we observed was consistent with our prediction that AMF regenerating primarily from spores (i.e. members of the Gigasporaceae) would be the slowest colonizers. Spore dormancy and specific environmental requirements for spore germination presumably slow the rate at which spore-regenerating AMF can colonize plant roots. Our prediction that AMF regenerating primarily from hyphal fragments (i.e. members of the Glomaceae and Acaulosporaceae) would initially colonize roots faster than spore-regenerating AMF was only partly correct. As predicted, Glomaceae isolates were generally the fastest colonizers of plant roots. Members of the Acaulosporaceae are generally assumed to have equally infective mycelia as Glomaceae (Morton, 1993). However, we found that Acaulosporaceae isolates generally colonized roots more slowly, at a rate more similar to spore-regenerating Gigasporaceae. Our results are consistent with those of Tommerup & Abbott (1981) who found that an Acaulospora spp. was unable to form a new infection from a root fragment containing hyphae whereas a Glomus spp. successfully colonized from root fragments.

The differential rate at which AMF isolates intercept a root may help to explain why some AMF have ‘parasitic’ effects on host plants in pot experiments (Francis & Read, 1995; Klironomos, 2000). ‘Parasitic’ AMF may simply be slow-colonizers and thus more demanding of plant resources for a longer time than faster colonizers.

The differential rate of colonization by AMF also has implications for methods of isolating AMF in the field. Current practices involve successful colonization of ‘trap plants’ by indigenous AMF and their subsequent sporulation. Based on our results, trap cultures that run less than 8 wk may be biased in favour of fast colonizing AMF. To represent all AMF in a community, it is important to allow adequate time for colonization of slow and fast colonizing isolates.

AMF isolates also showed large differences in the extent to which they colonize plant roots and soil. While there is some variation within families in the amount of root colonization, overall there are strong, family based trends. Based on the sum of root plus soil fungal biomass (ergosterol), Gigasporaceae isolates had the highest mean (±1 SE) extent of colonization (2.62 ± 0.04 µg g−1 root plus soil d. wt), followed by Glomaceae isolates (0.58 ± 0.09 µg g−1 root plus soil d. wt) and Acaulosporaceae isolates (0.55 ± 0.09 µg g−1 root plus soil d. wt). This pattern is consistent with our prediction that Gigasporaceae isolates would have the most extensive mycelium based on its traits (i.e. robust, densely aggregated hyphae). AMF families with more delicate, diffuse hyphae were predicted to have less extensive colonization. Acaulosporaceae isolates were consistent with this prediction but a surprising number of Glomaceae isolates showed more extensive colonization than Acaulosporaceae isolates. The reason for this difference is not clear. Our results are consistent with a study that examined the growth of extra-radicle mycelia in three AMF (Glomus spp., Acaulospora laevis, Scutellospora calospora) (Jakobsen et al., 1992). They, too, found that Scutellospora had the most extra-radicle mycelium and that it was concentrated closer to the root system compared with the more diffuse mycelia of the Glomus spp. and A. laevis.

AMF isolates also differed substantially in their colonization of roots vs soil. Based on previous observations, we predicted that members of the Gigasporaceae would colonize soil more extensively than plant roots and, conversely, members of the Acaulosporaceae and Glomaceae would colonize roots more extensively than soil. Data for the 21 AMF isolates studied here are largely consistent with this prediction. Such large differences among isolates in root vs soil colonization have potentially important implications for their role as a plant mutualist. A high ratio of soil to root colonization may provide the plant host with the greatest benefit (increased potential for nutrient transfer from root to soil). Conversely, a low ratio of soil to root fungal biomass may provide the host plant with little net benefit because nutrient transfer may be limited by the extent of the absorptive hyphal network. In any case, our data confirm that AMF isolates exhibit a wide range of colonization strategies with respect to root vs soil colonization.

These results suggest that community level studies that use only root measurements of AMF to characterize the AMF community may be biased against AMF, which have less mycelium in roots than in soil (i.e. Gigasporaceae spp.). Both internal and external measures of colonization are important for an accurate representation of an AMF community.

By combining measures of AMF colonization ‘rate’ and ‘extent’, we found that AMF isolates from three taxonomic families could be reliably separated. In other words, AMF taxonomy has a functional basis as well as morphological and developmental basis. The strong relationship between AMF colonizing strategy and taxonomy suggests that the taxonomic status of an AMF may be a useful predictor of its ecology, at least in terms of colonizing ability. Members of the Glomaceae usually contact roots quickly and produce extensive mycelium in roots compared with in soil. Members of the Gigasporaceae typically contact roots more slowly and establish an extensive mycelium in soil rather than in roots. Members of the Acaulosporaceae also contact roots more slowly and establish a much less extensive mycelium in either roots or soil than members of the other two families.

It remains to be seen how these functional groups of AMF differ with respect to mycelial architecture. We documented major differences in the internal and external mycelium among different AMF. It is unclear, however, exactly how they are different in terms of mycelial architecture. For example, Friese & Allen (1991) characterized an assemblage of AMF in this way, differentiating absorptive and runner hyphae from hyphal bridges, germ tubes spores and root fragments. In our study, Gigaspora and Scutellospora isolates had extensive external mycelia. Because it has been shown that their mycelia are not infective (Biermann & Lindermann, 1983; Brundrett et al., 1999, J. N. Klironomos & M. M. Hart unpublished) it may be that these genera have mycelia that are lacking in infective, runner hyphae (Friese & Allen, 1991) and form many absorptive hyphae instead. Glomus isolates, conversely, have a highly infective mycelium and thus might have a mycelium comprised of runner hyphae with little absorptive hyphae. As such, Gigasporaceae may be more ‘mutualistic’ because they provide the most nutritive benefits for their hosts due to a heavy investment into primarily absorptive hyphae.

While it is clear that AMF have distinct functional groups, it is unclear whether these functional groups have any bearing on host benefit. That is, are certain functional groups of AMF better at nutrient acquisition than others? For instance, because Gigasporaceae species have a larger soil mycelium, does that increased surface area result in more nutrients for the host? Or are the AMF with high root colonization (Glomaceae) more effective at nutrient uptake due to the higher incidence of arbuscules? It is unclear at this point whether the arbuscule or the absorptive mycelium is a more limiting factor for host nutrient acquisition. Further, such large differences in mycelium sizes might mean that some AMF are more ‘costly’ to their hosts than others. That is, AMF with large mycelia may be better at nutrient uptake but may pose a larger carbon-sink than AMF with small mycelia (i.e. Acaulosporaceae). Because AMF and their host plants are so closely linked, it will be important for future studies to address the significance of AMF functional groups in host plant response.

Acknowledgements

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

The authors would like to thank John Klironomos for his invaluable advice and assistance. We would also like to thank Peter Moutoglis and staff at Premier Tech. Inc. for resources and assistance. This work was supported by the National Science and Engineering Research Council grant to MMH.

References

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