Soil organic matter distribution and below-ground competition between Calluna vulgaris and Nardus stricta

Authors


‡Author to whom correspondence should be addressed. E-mail: d.r.genney@abdn.ac.uk

Summary

  • 1We conducted an experiment to investigate the effect of organic matter distribution on below-ground competition between the dwarf shrub Calluna vulgaris (L.) Hull and the coarse upland grass Nardus stricta (L.).
  • 2The two species were grown alone, or in competition in pots where the substrate was either a sand layer overlain by a peat layer (layered), or the same volume of peat evenly mixed with sand throughout the pot (mixed).
  • 3Root length and allocation in the top, middle and bottom of each pot was measured, along with plant biomass.
  • 4Calluna allocated the greatest proportion of root length to the top of the pot, irrespective of the peat distribution. Nardus allocated a large proportion of root length to the bottom of the pot. Root allocation by Nardus to the top and middle of the pot was positively related to the distribution of peat.
  • 5Nardus grown alone attained greater shoot mass in the layered than in the mixed substrate, but this was not the case when Calluna was present. Calluna was the superior competitor in both layered and mixed-substrate pots, but more so in the latter.
  • 6This was attributed to (i) the ability of Calluna to exclude Nardus roots from the upper organic material in the layered substrate, and (ii) the greater plasticity of root allocation of Nardus, enabling it to avoid competition with Calluna.

Introduction

Plant roots are able to exploit below-ground resource heterogeneity in several ways. An increase in root length allocation within nutrient-rich patches is one of the most striking and frequently demonstrated (Hutchings & de Kroon 1994; Robinson 1994). However, species differ in root system morphology (Fitter 1985), and in the scale and precision of their response to patches of different size and quality (Einsmann et al. 1999; Farley & Fitter 1999). Such differences may affect the outcome of interspecific competition, although there is relatively little experimental evidence to support or refute these suggestions (but see Cahill & Casper 1999; Fitter 1982; Robinson et al. 1999). The ability of species to proliferate roots within a nitrogen-rich, organic patch can increase nitrogen capture, and hence competitive ability (Hodge et al. 1999).

Tilman (1982) suggested that soil heterogeneity should lead to an increase in species coexistence, and that this effect should be most marked in nutrient-poor habitats. The concentration of nutrients in surface horizons, caused by organic matter accumulation and nutrient deposition at the soil surface, is a widespread and predictable source of nutrient heterogeneity in undisturbed soils. This is a particularly strong feature of heathland soils, characterized by organic upper horizons above leached, mineral, lower horizons (Gimingham 1972). Grass roots are found at greater depth when co-occurring with ericaceous dwarf shrubs (Gimingham 1972), and this has been suggested as a mechanism that reduces resource competition in heathland communities (Smith & Read 1997).

The ericaceous dwarf shrub Calluna vulgaris (L.) Hull and the coarse grass Nardus stricta (L.) are naturally co-occurring species of Scottish upland heaths. In recent decades, large areas of Calluna heath have been replaced by Nardus-dominated grassland. Preferential grazing of Calluna over Nardus is though to be the main mechanism of this replacement, particularly at high nutrient levels (Hartley 1997).

Calluna and Nardus have contrasting root systems (Genney et al. 2000), in common with other ericaceous dwarf shrubs and grasses (Aerts 1993; Read 1996). Calluna produces an abundance of fine hair-roots (≈ 0·1 mm diameter) that proliferate in the upper organic soil horizons (Aerts 1993; Gimingham 1960). These roots are normally heavily colonized by ericoid mycorrhizal (ErM) fungi. This is of functional significance because ErM fungi can access N and P from decaying plant and fungal matter within the litter and humic horizons, and consequently increase host-plant nutrition (Kerley & Read 1998). In contrast, Nardus is a relatively coarse-rooted grass that produces numerous, thick, unbranched roots (1–2 mm diameter) that penetrate to depths of 50 cm or more, along with shallower, fine lateral roots (<0·5 mm diameter) (Chadwick 1960). Nardus roots may be colonized by arbuscular mycorrhizal (AM) fungi (Ali 1969), which improve uptake of immobile nutrients such as phosphorus (Smith & Read 1997) and have recently been shown to increase decomposition of, and nitrogen capture from, patches of organic matter (Hodge et al. 2001).

In a previous study (Genney et al. 2000) we demonstrated that, when grown alone in a layered organic/sand substrate, Nardus allocated the greatest proportion of its root length to the organic layer. However, Calluna was the superior competitor when the species were grown together (competition with Calluna reduced Nardus mass more than competition with other Nardus), and this was attributed to the ability of Calluna partially to exclude Nardus roots from the surface organic layer. It was not, however, possible to determine whether this was caused by the superior ability of Calluna to respond selectively to the nutrient-rich layer by plastic root allocation, or because it had predetermined root allocation. The study reported here was designed to distinguish between these two possibilities. Our objectives were to measure (i) the vertical root length allocation of Calluna and Nardus when grown alone (monocultures) in a layered or in a mixed peat/sand substrate; and (ii) the comparable root length allocation patterns when the species are grown together (mixtures), and also to determine how root allocation patterns affect Calluna/Nardus competition in the layered and mixed substrate.

Materials and Methods

Preparation of Materials

Even-sized Calluna cuttings (6 cm long) were collected in September 1999 from a large number of plants at Grandhome Moss near Aberdeen, north-east Scotland (grid reference NJ907124). After removing the leaves from the bottom 2 cm of each cutting, they were planted in an unheated propagator in a mixture (1 : 1, v : v) of washed sand and Terragreen in a greenhouse until roots were produced. Nardus seeds were collected in late August 1997 from Glen Derry, Aberdeenshire (grid reference NO040958). After 5 min surface sterilization with 1·0% sodium hypochlorite, the seeds were germinated on autoclaved sand.

In order to produce mycorrhizal Calluna, the ErM endophyte Hymenoscyphus ericae (Read) Korf & Kernam (Strain He 101) was grown on malt extract agar. Fresh mycelium from the margins of actively growing colonies was grown in shaking modified Melin–Norkans liquid medium (pH 5·8) (Brundrett et al. 1996) at room temperature for 14 days. The mycelium was filtered and washed five times with distilled water, before being mixed with distilled water and macerated in a blender to produce a slurry containing 7·8 mg ml−1 dry mycelium.

Pot Preparation and Growth Conditions

For the layered substrate, plant pots (120 × 120 × 120 mm) were prepared containing 80 mm sterile, acid-washed medium sand (mean particle size = 0·1 mm, Hepworth Minerals and Chemicals Ltd, Cheshire, UK; pH adjusted to 3·8 with HCl) overlain with a 40-mm-deep organic layer of mixed Irish moss peat (sieved to 2 mm) and sand (1 : 1, v : v). In order to avoid a sharp boundary between substrate layers, the bottom 5 mm of each organic layer was blended into the top 5 mm of the underlying sand. For the mixed-substrate pots an equal volume of peat was distributed evenly throughout the pot (Fig. 1b). All pots were flushed daily for 2 weeks with distilled water (pH adjusted to 3·8 with HCl) to reduce available mineral nutrient levels.

Figure 1.

(a) Planting arrangement of Nardus seedlings (N) and Calluna cuttings (C) grown alone and together. The position of plants from which sequential shoot measurements were made are underscored. (b) Peat distribution (shading) in layered and mixed-substrate pots. The vertical divisions represent the top (T), middle (M) and bottom (B) slices from which roots were analysed at the destructive harvest.

In a fully crossed, factorial, replacement design, eight plants of either Nardus, Calluna or both species (four of each) were planted in layered and mixed-substrate pots (Fig. 1a). Eight replicates of each competition/substrate placement combination resulted in a total of 48 pots. Rooted Calluna cuttings were planted (along with a 0·5 ml inoculation of live H. ericae slurry) into the pots and allowed to establish for 2 weeks prior to the addition of Nardus seedlings (one blade ≈ 10 mm high) into their relative positions (Fig. 1a). ErM inoculum was added because the hair-roots of Calluna are normally highly colonized by ErM endophytes in the field. In addition, this experiment was not conducted under sterile conditions, therefore inoculation minimized the risk of uneven ErM contamination from external sources. For consistency, each Nardus seedling also received a 0·5 ml inoculation of H. ericae slurry. The pots were placed in a randomized array (re-randomized every 2 weeks) in a glasshouse (day 25 ± 0·2 °C, night 17·5 ± 0·04 °C, supplementary high-pressure sodium lamp lighting between 15 : 00 and 00 : 00 h). The peat was the only source of nutrients throughout the experiment, and the substrate was kept constantly moist by watering from above and below with distilled water (pH adjusted to 3·8 with HCl).

Non-Destructive Shoot Measurements

The number of leaf blades >5 mm long and maximum green blade length were measured for one inner and one outer Nardus plant per pot (positions shown in Fig. 1a) at monthly intervals throughout the growth period.

Harvest

After 6 months the final shoot measurements were made and all plants were harvested. Nardus and Calluna shoots were weighed after oven-drying at 80 °C for 48 h. The ability of Calluna to compete with Nardus in layered and mixed substrates was determined with an aggressivity index (Acn) (equation 1; Snaydon 1991).

image(eqn 1)

where Wcc and Wnn are the total mass per plant of c (Calluna) and n (Nardus) grown alone; and Wcn and Wnc are the total mass per plant when both species were grown together.

The substrate in the pots was cut into three 40-mm-deep, horizontal slices (Fig. 1b). All roots were extracted from each slice by washing with a water jet over a 100 µm sieve, and manually separated by species with fine tweezers. In 10 randomly selected pots, a small subsample of roots from each species was removed and the presence/absence of mycorrhizal colonization was determined visually by clearing in 10% KOH (20 min, 80 °C), acidifying in 1% HCl (1 h, room temperature), and staining with 0·05% trypan blue in acid glycerol (20 min, 80 °C). Nardus roots that had grown out of the bottom of the pots were harvested separately. Each root sample was scanned on a flat-bed scanner, and total root length was measured using image analysis software (winrhizo ver. 3·10b; Regent Instruments Inc., Canada). Where the sample was too large to be scanned (total root length over ≈ 35 m) a weighed subsample was used. After drying in an oven at 80 °C for 48 h, the root samples were weighed.

Statistical Analysis

minitab (ver. 12·21) was used to perform glm-anova to compare treatment effects of final harvest measurements. The change in shoot measurements over time was analysed with glm-anova by comparing power regression coefficients (of mean inner and outer plants per pot), between placement and competition treatments. The model included pots as a random factor nested within the main factors (competition and substrate placement type), and month of measurement as a covariate.

The effect of placement and competition type on proportional root length allocation between substrate slices was tested by incorporating ln(T/B) and ln(M/B) into a manova analysis, where T, M and B represent the proportion of total root length allocated to the top, middle and bottom slice, respectively. Univariate, post hoc Tukey tests were used to compare individual means.

Homoscedasticity of variance and normality of residuals were tested to ensure suitability for parametric statistics. Where these conditions were not met, Box–Cox analysis was used to determine the most suitable transformation function. Mean values are presented ±1 SE. Due to the labour-intensive nature of root harvesting and measurement, roots from only seven randomly selected replicates for each factor combination were harvested.

Results

Above Ground

Nardus

Competition with Calluna reduced Nardus shoot mass by 59% (F1,28 = 230, P < 0·001; Fig. 2), and there was a reduction in the rate at which new leaf-blades were produced (F1,28 = 54·5, P < 0·001; Fig. 3). There was, however, an interactive effect of competition on the response of shoot mass to peat distribution (F1,28 = 9·66, P < 0·005; Fig. 2). When grown in Nardus-only pots, shoot mass was 29% greater when the peat was layered rather than evenly distributed (Tukey: P < 0·05). However, when grown with Calluna, there was no difference in Nardus shoot mass between substrate types (Tukey: P > 0·1). Although the pattern of response was the same, the interactive effect on the rate of leaf-blade increment was not significant (F1,28 = 1·23, P > 0·1).

Figure 2.

Shoot dry mass per Nardus (shaded) and Calluna (unshaded) plants between different peat placement (layered/mixed) and competition (alone/together) pot types. Means ± SE are shown; different letters represent within-species differences at P < 0·05 (n = 8).

Figure 3.

Leaf blade production per Nardus plant throughout the growth period in layered (solid lines) and mixed (dashed lines) substrates. (•) Species grown alone; (○) species grown together. Means ± SE are shown; n = 8.

Calluna

Total Calluna mass per plant was greater when in competition with Nardus, which resulted in positive values of Acn in both substrate types. However, Acn was greater when the peat was layered (0·71) rather than evenly mixed (0·49). Calluna shoot mass was always greater when the peat was layered (F1,28 = 38·4, P < 0·001), but the difference between substrate types was greater when Calluna was grown with Nardus (46%) than when grown in monoculture (22%) (Fig. 2).

Below Ground

Specific root length of both species was unaffected by treatment [Nardus = 23·0 cm mg−1 (SE = 0·74); Calluna = 51·4 cm mg−1 (SE = 3·69)], therefore only root length data are presented.

Nardus

Irrespective of peat distribution, competition with Calluna reduced root length per Nardus plant by 66% (F1,24 = 117, P < 0·001; Fig. 4b). When grown in monoculture, there was no effect of peat distribution on root length. When grown with Calluna, however, plants in pots where peat was evenly mixed had 56% greater root length than plants in pots where peat was layered (competition × placement: F1,24 = 8·95, P < 0·01; Fig. 4b). There was an interactive effect of competition and peat distribution on the pattern of root length allocation between slices (F2,23 = 9·29, P < 0·001; Fig. 4a). When the peat was layered, Nardus allocated a greater proportion of root length to the organic top slice compared to the inorganic middle slice (Tukey: P < 0·001) (Fig. 5). However, the difference was greatest (3·5-fold) when Nardus was grown in monoculture, compared to when grown with Calluna (twofold) (Fig. 4a). When the peat was evenly mixed, an equal proportion of root length was allocated to the top and middle slices (Tukey: P > 0·1). Nardus always allocated a large proportion of root length to the bottom layer, although a greater proportion was allocated to the bottom layer when grown with Calluna (0·67) than when grown in monoculture (0·42) (Tukey: P < 0·001; Fig. 4a). In addition, a greater proportion of root length was allocated in the bottom slice when the peat was evenly distributed, rather than layered, when grown with Calluna (Fig. 4a). No AM colonization was found in any of the Nardus root samples.

Figure 4.

(a) Proportional distribution of Nardus root length between the three substrate slices; (b) root length per plant between different substrate placement (layered/mixed) and competition (Nardus alone/Nardus + Calluna) pot types. Means ± SE are shown; different letters represent differences at P < 0·05 (n = 7).

Figure 5.

Nardus root proliferation in the upper organic slice of the layered substrate pots.

Calluna

Irrespective of peat distribution, plants grown with Nardus had 28% longer roots than those grown in monoculture (F1,24 = 28·0, P < 0·001; Fig. 6b). In comparison to when peat was layered, root length per plant was ≈ 20% greater when the peat was evenly distributed, irrespective of competition type (F1,24 = 15·4, P = 0·001; Fig. 6b). The proportion of root length allocated to each slice decreased with depth in all treatment combinations, and allocation between the slices was not affected by competition with Nardus (F2,23 = 0·58, P > 0·1; Fig. 6a). There was, however, an overall effect of peat distribution on the proportional root length allocation between slices (F2,23 = 17·87, P < 0·001). This was caused by an increase in allocation of root length from the top slice (0·65–0·55) to the middle (0·22–0·35) when peat was evenly distributed (Fig. 6a). All root samples were colonized by ErM fungi, and the level of colonization appeared to be comparable to that observed in field-grown plants.

Figure 6.

(a) Proportional distribution of Calluna root length between the three substrate slices; (b) root length per plant between different substrate placement (layered/mixed) and competition (Calluna alone/Calluna + Nardus) pot types. Means ± SE are shown; different letters represent differences at P < 0·05 (n = 7).

Discussion

These results demonstrate that differences in root allocation and plasticity of two species with contrasting life forms can result in different competitive outcomes, depending on the vertical distribution of nutrients.

Growth in Monocultures

The root length allocation strategy of the two species varied considerably. Calluna roots rarely reached the bottom of the pot, whereas Nardus always allocated the largest proportion of its root length to the bottom slice. This is consistent with field observations, where ericaceous dwarf shrubs form a dense hair-root mat in the litter and humic horizons (Gimingham 1960; Kerley & Read 1998) and, in common with other heathland grasses (Aerts 1993), Nardus allocates roots to a greater depth (Chadwick 1960).

When the peat was layered, both species allocated a greater proportion of root length to the top (organic) slice than the underlying (inorganic) middle slice. When the peat was evenly distributed, Nardus allocated an equal proportion of root length to the top and middle slices. However, Calluna allocated most root to the top slice, irrespective of peat distribution. Nardus exhibits plastic root allocation, a strategy that has been demonstrated for many species (Robinson 1994). On the other hand, Calluna root allocation is (at least in the establishment phase) predetermined by development, rather than peat distribution.

Differential responses of species to non-uniform supplies of nutrients have been demonstrated elsewhere (Einsmann et al. 1999; Farley & Fitter 1999), and a lack of root allocation response is not uncommon. It seems probable that the lack of response to peat placement exhibited by Calluna is due to a fundamental requirement for ErM roots to associate with a predictable source of organic matter. Where organic matter is predictably distributed at the soil surface, Calluna does not (like Nardus) potentially waste resources by allocating roots to nutrient-poor soil horizons. However, the same trait may prevent Calluna responding to situations where organic matter is uniformly distributed with depth, and may contribute to Calluna's susceptibility to drought (Gordon et al. 1999).

Both species had bigger shoots in the layered substrate compared to the mixed substrate. Increases in growth have been reported in other studies, where nutrients are concentrated within a patch, compared to the same quantity of nutrients distributed evenly (e.g. Cahill & Casper 1999; see also Casper & Jackson 1997; Robinson (1994). Possibly a smaller proportion of the immobile ions become bound to the solid phase when nutrients are concentrated in a small fraction of the soil volume, and more are available for plant uptake (Casper & Jackson 1997). It may be that mineralization is greater if a more active microbial community develops in the peat-rich slice. It is also possible that nutrients in peat are accessed more quickly by root growth into the layered compared to the mixed substrate.

Growth in Mixtures

Calluna was the superior competitor in both layered and mixed substrates. Nardus plants grown with Calluna were smaller, and produced fewer leaf blades and less root, than when grown alone. In contrast, Calluna grown with Nardus had bigger roots and shoots than when grown alone. As the only source of nutrients was the peat, it is likely that, as in our previous experimental system (Genney et al. 2000), plant growth was limited by nutrient availability. These results are consistent with other studies, where ericaceous shrubs have had a competitive advantage over grasses at low nutrient supply (Hartley & Amos 1999). It is possible that using Calluna cuttings (rather than seedlings) in competition with Nardus seedlings influenced competition. Using cuttings helped to overcome the difference in growth rate between the two species. In addition, Nardus requires gaps in the Calluna canopy for successful establishment (Alonso & Hartley 1998). Seed dispersal of Nardus into gaps created by fire, grazing or senescence of old Calluna plants is therefore an important mechanism by which Nardus invades Calluna heath. This is a further reason why Calluna cuttings were used in this study.

The relative suppression of Nardus was greatest when peat was concentrated at the surface (Calluna had a more positive aggressivity index in the layered substrate). This difference in competition intensity is explained by the contrasting root allocation of the two species. Calluna root length in the top slice, in both mixed and layered substrate, was unaffected by competition with Nardus. However, Nardus was less able to colonize the top slice in the layered substrate than when grown alone. The ability of one species to reduce that of another to exploit a zone of nutrient enrichment has been demonstrated in other studies (Caldwell et al. 1991; Fitter 1976). Because the peat was evenly distributed in the mixed-substrate pots, nutrient uptake by roots in the bottom slice is a likely explanation why Nardus growth was less affected by Calluna in this homogeneous substrate.

These results support the hypothesis that resource competition between ericaceous dwarf shrubs and grasses is avoided by vertical displacement of grass roots (Smith & Read 1997). However, where organic matter is concentrated at the soil surface, Calluna is a more aggressive competitor, so Nardus gains little advantage from the ability to allocate roots to greater depth. In the field, Calluna occurs most frequently on podsolized soils (Gimingham 1960), with a nutrient-leached horizon below litter and humic horizons (Fitzpatrick 1999). The superior competitive ability of Calluna in the layered substrates may, to some extent, contribute to the dominance of Calluna on such soils. Where deeper histosols develop, or podsolic profiles are disrupted (e.g. by increased nutrient addition or cultivation for forestry), Calluna may be more susceptible to invasion by grasses such as Nardus. Similar hypotheses have been suggested to explain coexistence between species of different rooting depth in a hayfield community (Berendse 1983). Many years ago, Fitter (1982) demonstrated reduced dominance by one or two species in a mixture of grassland species, when grown in a layered fertile/infertile substrate, compared to an evenly mixed substrate.

In the field, Nardus roots are normally colonized by AM fungi (Ali 1969; Genney et al. 2001). In common with previous workers, we had difficulty in obtaining colonization in experimental plants (e.g. Heijne et al. 1994), and this study was conducted with non-mycorrhizal Nardus. In a pot study of seven woodland species, Farley & Fitter (1999) found that AM colonization had no effect on within-nutrient patch root length, and improved the phosphorus uptake of only one species. However, Hodge et al. (2000) demonstrated that AM colonization increased root length within organic patches and AM hyphae stimulated organic matter decomposition and nitrogen capture from such patches (Hodge et al. 2001). Given that AM colonization may influence the response of plants to nutrient patches, the absence of AM colonization in this study should be considered when extrapolating these results to the field.

In summary, Calluna consistently concentrated the greatest proportion of root length in the surface slice, whereas Nardus always allocated a large proportion of root length to the bottom slice and exhibited a plastic response to peat distribution in the upper two slices. These contrasting strategies resulted in the greatest inhibition of Nardus growth by Calluna when peat was concentrated in the upper slice, mainly due to the exclusion of Nardus roots from a source of peat. When peat was distributed evenly throughout the slices, Calluna was less competitive. This was due to inflexibility in Calluna's root allocation leading to an inability to exploit peat in the lower slices, whereas Nardus avoided competition with Calluna by exploiting peat in the bottom slice.

Organic matter distribution can determine the intensity of competition between Calluna and Nardus. This is due to differential advantages and disadvantages of a plastic and developmentally predetermined root foraging strategy.

Acknowledgements

We would like to thank Dave Elston (BIOSS) for statistical advice and Professor David Read for providing the H. ericae culture. This work was funded by a NERC CASE studentship to D.R.G.

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