Leaf area renewal, root retention and carbohydrate reserves in a clonal tree species following above-ground disturbance

Authors

  • Simon M. Landhäusser,

    Corresponding author
    1. Centre for Enhanced Forest Management, 4–42 Earth Sciences Building, Department of Renewable Resources, University of Alberta, Edmonton, Alberta T6G 2E3, Canada
      Simon M. Landhäusser, Centre for Enhanced Forest Management, 4–42 Earth Sciences Building, Department of Renewable Resources, University of Alberta, Edmonton, Alberta T6G 2E3, Canada (Tel. +780 492 6381; fax +780 492 1767; e-mail simon.landhausser@ualberta.ca).
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  • Victor J. Lieffers

    1. Centre for Enhanced Forest Management, 4–42 Earth Sciences Building, Department of Renewable Resources, University of Alberta, Edmonton, Alberta T6G 2E3, Canada
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Simon M. Landhäusser, Centre for Enhanced Forest Management, 4–42 Earth Sciences Building, Department of Renewable Resources, University of Alberta, Edmonton, Alberta T6G 2E3, Canada (Tel. +780 492 6381; fax +780 492 1767; e-mail simon.landhausser@ualberta.ca).

Summary

  • 1Removal of the above-ground portion of clonally regenerating trees results in a massive imbalance in the ratio of root to leaf area. We investigated the resilience of clones following above-ground disturbance in terms of root carbohydrates, leaf area renewal and root retention.
  • 2In a 2 × 2 factorial experiment, 40 Populus tremuloides saplings were cut at times of high (late fall) or low (spring after leaf flush) root carbohydrate reserves. Leaf area renewal was manipulated by allowing either only one or all root suckers to re-grow.
  • 3Root starch concentrations were 10 times higher at the time of cutting in the fall compared with the spring, whereas sugar concentrations were only 10% higher. When saplings were cut in fall and all suckers were allowed to develop (FA treatment), suckers were taller, and had more biomass and leaf area and higher leaf area ratio than all other treatments. In particular, leaf area and leaf area ratio recovered within a year to near pre-treatment levels compared with at least a 50% reduction in the other treatments.
  • 4At the end of the first season after cutting FA saplings also had the greatest root mass and lowest amount of dead root mass, and root starch concentrations of these saplings had returned to pre-treatment values, compared with a 20% recovery for spring-cut saplings with a single sucker. Root mass and root starch concentrations were correlated with leaf mass in all treatments.
  • 5Adequate carbohydrate reserves at the time of disturbance and rapid redevelopment of leaf area by extensive sprouting seem critical for resilience of the clone and allow for the retention of the clonal root system and a rapid rebuilding of carbohydrate reserves. Clones with poor sucker and leaf area development showed extensive root mortality and reduced carbohydrate reserves and their prospects for growth in the following year were comparatively poor.

Introduction

Resilience to above-ground disturbance such as fire, grazing or cutting is well studied in clonal herbaceous plants (e.g. Ashmun et al. 1982; Nobel & Marshall 1983; Chapman et al. 1990, 1991; Kemball & Marshall 1995; de Kroon & van Groenendael 1997). However, clonal trees have been less studied because of their longevity, large size and woody nature (Jenik 1994; Peterson & Jones 1997). Mature trees are able to accumulate great amounts of root biomass (up to 40 Mg/ha in Populus tremuloides Michx. clones; Strong & LaRoi 1983; Ruark & Bockheim 1987). Cutting the above-ground portion of a clonal tree therefore leaves a very large root system without the support of leaf area compared with an herbaceous species. Although leaf area recovery is thought to be important for root system maintenance in clonal tree species (Tschaplinsky & Blake 1995; Reichenbacker et al. 1996), there is little experimental work on this or on the role of root carbohydrate reserves following removal of the above-ground structures.

Following disturbances that remove the above-ground portion of trees there is a massive imbalance in the root to leaf area ratio of the clone. We hypothesize that rapid root and stump sprouting is necessary to rebuild leaf area quickly in order to sustain the respiratory demands of the large root system. The resilience of the clone, in terms of renewal of above-ground stems and leaves, initially relies on reserves mobilized from below-ground organs that are protected from disturbance (Bowen & Pate 1993; Iwasa & Kubo 1997; Sakai et al. 1997). Generally, in the years immediately following disturbance, clonal plants are able to rebuild part of the above-ground structure while either partially or fully retaining the original root system (Koop 1987; Peterson & Jones 1997), but the extent of root retention in woody clonal species is not well understood. There are seasonal changes in carbohydrate reserves of roots in woody shrubs, usually with highest levels in the late fall, after leaf senescence, and lowest levels in the late spring after leaf flush (Zasada et al. 1994; Landhäusser & Lieffers 1997). If the same pattern applies to large trees, clones are likely to have higher root reserves for re-sprouting when disturbed in the autumn than in late spring. We hypothesize that a large root system that is depleted of reserves at the time of disturbance will be a burden (carbohydrate sink) for the clone, as these roots will have a large respiratory demand but few reserves available to rebuild leaf area. Until a more appropriate balance between leaves and roots is developed, above-ground growth will be slow, as a large share of the fixed carbon will go to the maintenance of the large root system. Alternatively, a portion of the root system might die back in order to correct the imbalance of the root to leaf area ratio.

Aspen (Populus tremuloides Michx.) is a widespread tree species that suckers profusely from adventitious buds on shallow surface roots following disturbance such as fire or cutting (Schier 1973a), provided that soils are warm (Maini 1967; Schier & Campbell 1978). However, low numbers of suckers have been observed on some sites (Shepperd 1993), perhaps due to thick organic layers that insulate the soil and prevent spring and summer warming (Zasada & Schier 1973; Hogg & Lieffers 1991) or to intensive browsing by ungulates (Smith et al. 2000). Under these unfavourable conditions, even a large and healthy clone may produce relatively few suckers, resulting in low clonal leaf area.

This experiment examines the resilience of Populus tremuloides clones following disturbance at high and low root carbohydrate reserves. Specifically it tests the impact of different levels of leaf area development in the first year on the maintenance of the original clonal root system and on the rebuilding of carbohydrate reserves in the clone.

Methods

PLANT MATERIAL, GROWTH CONDITIONS AND INITIAL MEASUREMENTS

During the spring of 1997, 61-year-old Populus tremuloides seedlings obtained from a commercial nursery were grown for 2 months in pots (25-cm diameter) in a growth chamber. Actively growing seedlings were then transplanted to an open field at the University of Alberta experimental station near Ellerslie, in June 1997. Seedlings averaged 80 cm in height at planting and each was planted into a 1 × 1 by 0.5 m deep plot, which had been excavated, lined with a thick plastic sheet (with bottom holes for drainage) and refilled with a 3 : 1 sand-peat mixture. The plots were arranged 2 m apart in a rectangular pattern with 10 rows containing six plots each. The seedlings were staked and grown for two additional growing seasons (1998 and 1999), during which all plots were kept free of weeds, and the trees were watered and fertilized.

Saplings were randomly allocated to six groups of 10 saplings, such that each group had similar numbers from the edge of the plantation. In late September 1999, after leaves had turned yellow but just before abscission, all leaves were collected separately from each of the 60 saplings and total leaf mass and area were determined. In late October, when soil temperatures were below 5 °C and when root reserves were thought to be high, saplings in three of the six groups were cut at the soil surface. One of these groups was immediately excavated to determine root mass and carbohydrate reserves. As in subsequent collections, roots were grouped into three diameter sizes, < 1.5 mm, 1.5–8 mm and > 8 mm, and three subsamples were collected for each root size and sapling. Height, stem diameter at root collar and above-ground biomass were determined for each of the 30 cut saplings. Twigs (1-year-old branches) and older branch samples (> 2 years) were collected for carbohydrate analysis. The remaining 30 saplings were cut in late May 2000, immediately after bud flush, when root systems were thought to be low in carbohydrate reserves, and one group was excavated to determine root mass and carbohydrate reserves. Total above-ground biomass was determined and twig samples and the newly flushed leaves were collected from all saplings. Thus, by spring 2000 all saplings had been cut and two groups remained from the fall and two from the spring cuts.

All four groups suckered in the summer of 2000. All suckers were allowed to grow unimpeded for one group from each of the fall and the spring-cut treatments (FA and SA, respectively). In the remaining two groups (FS and SS), the dominant sucker was selected within the first 10 days of suckering and had its lower leaves removed regularly so that only 50% of the total sucker height had leaves, while all other emerging suckers were removed biweekly. In late September 2000, just before leaves abscised, all leaves were collected from all suckers to determine total leaf area for each treatment combination. All clones were excavated at the end of October, when soil temperatures were below 5 °C. Above-ground biomass, maximum sucker height, leaf area and dry mass of live and dead roots were determined and leaf area ratio (leaf area over total plant mass) calculated. Due to rapid decomposition of dead fine roots over the growing season only the amount of dead medium and coarse roots could be reliably measured. Dead and live roots were visually separated by their colouration and the separation of xylem and phloem. In addition, samples of suckers and live roots of the three sizes mentioned above were collected for carbohydrate analysis.

CARBOHYDRATE ANALYSIS

The three subsamples of each tissue were combined and oven dried at 68 °C. Samples were ground coarsely with a coffee grinder and then with a Wiley Mill (40-mesh). After the removal of interfering substances from tissue samples with chloroform, sugars were extracted three times with hot ethanol (85%). Sugar concentrations were then determined colourimetrically using phenolsulphuric acid (Smith et al. 1964). Remaining starch was subsequently solubilized by sodium hydroxide and hydrolysed to glucose by an enzyme mixture of α-amylase (ICN Biomedicals Inc., Montreal, Canada, Cat. no. 190151, from Bacillus licheniformis) and amyloglucosidase (Sigma, Aldrich Corp., St. Louis Missouri, USA, A3514, from Aspergillus niger) for 41 h, then measured colourimetrically using glucose oxidase/peroxidase-o-dianisidine solution (Sigma Glucose Diagnostic Kit 510 A).

DATA ANALYSIS

Pre-treatment growth and carbohydrate data were analysed as a randomised design with two levels of a single treatment (fall-cut and spring-cut). Post-treatment growth and carbohydrate data were analysed as a randomized 2 × 2 factorial design with two levels of season (fall and spring cut) and two levels of sprouting (single and all suckers) as fixed main effects. After log transformation of leaf area and leaf area ratio, all response variables met the assumptions of normal distribution and homogeneity of variances. t-Tests were used to analyse pre-treatment variables, and analysis of variance procedures and least significant difference multiple comparisons were used for the post-treatment variables. Both were performed with general linear models available in SAS 6.11 (SAS Institute Inc., Cary, North Carolina, USA). Relationships between leaf mass and root mass and leaf mass and root carbohydrate reserves were explored with linear regression. Slopes and intercepts of the regression lines were compared by testing for coincidental regression (Sokal & Rohlf 1981).

Results

PRE-TREATMENT CONDITIONS

There were no differences in height (anova, effect of season: P = 0.341), above-ground (P = 0.447) and root dry mass (P = 0.385), basal stem diameter (P = 0.29) and leaf area (P = 0.239) between the two seasons of cutting prior to treatment application. Mean values (± SD) of P. tremuloides saplings prior to treatment were 271 ± 42 cm for height, 42.1 ± 7.4 mm for basal stem diameter and 1.74 ± 0.6 m2 for total leaf area. At the time of cut, starch concentrations in roots were approximately 10 times greater in the fall cut saplings than in those cut after the flush of foliage (Table 1; anova, effect of season: P < 0.0001). Overall, fine- and medium-sized roots had higher starch concentrations than large roots. Although the starch concentrations of the stem tissues were less than one-twentieth of those in roots, they also showed a marked decline between the fall and spring samples (Table 1, P = 0.0007), but there were no treatment differences in leaves. Sugar concentrations in shoots and roots tended to be slightly lower in the spring than in the fall in five of the six fractions but were about 50% lower in newly expanding leaves collected in the spring than in the fall-collected yellow leaves (Table 1, P < 0.0001).

Table 1.  Pre-treatment sugar and starch concentrations in leaves, stems and roots of saplings at the time of fall and spring cuts (mean ± SE; n = 10). For each variable (sugar and starch concentration) different letters indicate differences between treatments within a particular tissue (t-test; P < 0.05)
TissueSugar (% dry weight)Starch (% dry weight)
Fall-cutSpring-cutFall-cutSpring-cut
Leaf19.6 ± 0.99a10.2 ± 1.42b0.09 ± 0.005x0.08 ± 0.01x
Small stem14.3 ± 0.49a13.0 ± 0.64a0.16 ± 0.02x0.05 ± 0.01y
Large stem7.5 ± 0.45a6.1 ± 0.34b0.20 ± 0.03x0.04 ± 0.01y
Small root16.2 ± 0.34a15.4 ± 0.87a13.23 ± 1.09x1.64 ± 0.36y
Medium root14.6 ± 0.54a12.1 ± 0.91b8.48 ± 1.09x0.83 ± 0.25y
Large root12.1 ± 0.89a8.6 ± 0.81b5.68 ± 0.96x0.49 ± 0.25y

MEASUREMENTS AT THE END OF THE EXPERIMENT

Overall, saplings cut in the fall developed taller suckers than those cut in spring (Table 2; anova, effect of season P < 0.0001). For fall-cut saplings, mean height was 158.8 cm when all suckers were allowed to grow compared with 109.5 cm in the single sucker treatment. For spring-cut saplings, however, sucker height was not different between both sprouting treatments (Table 2), resulting in a season × sprouting interaction (P = 0.008). Above-ground dry mass for the SA treatment was 24% of that for the FA treatment (Table 2). Both leaf area and leaf area ratio (LAR) were greater in fall-cut saplings than in spring-cut saplings (Table 2; anova, effect of season: both P < 0.0001). When all suckers were allowed to sprout, leaf area in fall-cut saplings was three times higher than in spring-cut saplings and was similar to values measured before the treatments (Table 2).

Table 2.  Clonal growth characteristics after 1 year of suckering, compared with pre-treatment values. Cutting was done either in the fall or spring and either all suckers or only a single sucker was allowed to develop (mean ± SE; n = 10). Different letters indicate differences among post-treatment means (LSD; P < 0.05)
 All suckersOne suckerPre-treatment
Fall-cut (FA)Spring-cut (SA)Fall-cut (FS)Spring-cut (SS)
Total above-ground mass (g)343.1 ± 31.2a83.9 ± 23.2b37.5 ± 9.0c18.7 ± 1.7d 849 ± 44
Maximum sucker height (cm)158.8 ± 7.8a81.1 ± 10.1c109.5 ± 11.6b84.6 ± 7.4cN/A
Total live root mass (g) 662 ± 68a 473 ± 77b 317 ± 50b 363 ± 52b 556 ± 25
Dead coarse root mass (g)4.9 ± 3.9b 30 ± 14.1a4.9 ± 2.6b15.9 ± 4.7ab0
Total leaf area (m2)1.49 ± 0.17a0.51 ± 0.13b0.09 ± 0.01c0.05 ± 0.01d1.74 ± 0.2
Leaf area ratio (cm2 g−1)14.8 ± 0.89a8.45 ± 1.04b2.42 ± 0.3c1.26 ± 0.19d11.7 ± 0.6

LEAF AREA VS. ROOT MASS RELATIONSHIP

Over the course of the experiment dry mass of the root system increased in the FA treatment and was significantly higher than in the other three treatments, where it was 48%, 40% and 28% less than the initial value (FS, SS and SA, respectively) (Table 2). Overall, there were more dead medium to large roots in the spring-cut treatments (Table 2; anova, effect of season: P = 0.027).

The 20 saplings excavated immediately after cutting showed a strong relationship between leaf and root mass (Fig. 1). The higher root weight of fall-cut saplings resulted in a difference in the intercepts of the pre-treatment regression lines (Fig. 1; test of coincidental regression: P = 0.0181) but not in the slopes (P = 0.241) (y = 2.638 × x + 55.1; R2 = 0.88 and y = 2.051 × x + 82.4; R2 = 0.88 for fall and spring, respectively). After 1 year, both the slopes and intercepts were different for the spring and fall treatments (Fig. 1; test of coincidental regression: both P < 0.0001; y = 4.807 × x + 309.6; R2 = 0.40 and y = 3.403 × x + 283.8; R2 = 0.64, for spring and fall, respectively).

Figure 1.

Relationship between leaf mass at time of leaf senescence and root mass of saplings before cutting (for both fall (□) and spring (▵) cutting treatments, n = 10) and 1 year after cutting in the fall (▪) and spring (▴) (n = 20 sprouting treatments combined). Regression lines show significant linear relationships.

NON-STRUCTURAL CARBOHYDRATES

Sugar concentration in both the small (anova, effect of sprouting: P = 0.0132) and medium (P = 0.017) diameter roots was lower in the single sucker treatments (FS and SS) compared with the all sucker treatments (FA and SA) (Fig. 2). The season and sprouting treatments had no effect on sugar concentrations in large roots, stems and leaves (Fig. 2).

Figure 2.

Root, stem and leaf sugar and starch concentration before treatments (fall values shown) and after 1 year of suckering. Treatments were cutting in fall or spring with either a single sucker or all suckers allowed to develop (mean ± SE; n = 10). Different letters indicate differences among post-treatment means (LSD; P < 0.05). Note different scales for starch concentrations for roots and aerial tissues.

Starch concentrations in stems and leaves of the suckers did not differ between season or sprouting treatments but increased during the treatments (Fig. 2). Starch concentrations were higher in the roots than in above-ground parts (Table 1). In roots, starch concentrations were affected by season of cutting and by the sprouting treatments. The much higher starch concentrations in FA roots than in any of the other three treatments were similar to those measured in the fall pre-treatment (Fig. 2). Averaged across all root sizes, starch concentrations in the SS treatment reached only 20% of the level determined for the previous fall.

The mean starch concentration in roots of the saplings after 1 year of suckering increased linearly with leaf mass (y = 0.061 × x + 2.05; R2 = 0.58; Fig. 3a), although there was no relationship in pre-treatment saplings. Total starch content in the root system (estimated from total root mass) was positively correlated with leaf mass both before and after treatments (y = 0.241 × x + 4.52; R2 = 0.70 and y = 0.516 × x + 2.23; R2 = 0.86, respectively; Fig. 3b). There was, however, a sharper increase in total starch content with increasing leaf mass for roots following suckering than in the pre-treatment saplings (Fig. 3b; test of coincidental regression: P < 0.0001), suggesting a faster accumulation of starch in the roots with recent suckers.

Figure 3.

Root starch concentration (a) and total starch content (b) in relation to leaf mass. Each graph depicts pre-treatment (□) data from the fall-cut treatment (n = 10) and combined data from all season and sprouting treatments (•) (n = 40). Lines show significant linear relationships.

Discussion

The capacity for sucker initiation and subsequent leaf area development appears to determine the resilience of P. tremuloides clones to above-ground disturbance. Suckers were taller, developed more leaf area and the clone retained more of its root system when the saplings were cut in the late fall, at the time of high root starch reserves, than in spring. By the end of the first season following suckering, the starch concentrations in the roots of the fall-cut saplings with all suckers growing (FA) had reached pre-treatment levels. For the spring-cut saplings with all suckers (SA), however, starch concentrations were only half that of pre-treatment saplings, and for the treatments with single suckers, concentrations were less than 25% of pre-treatment. This suggests that development of leaf area on suckers is critical for re-building root starch reserves for the next growing season. The leaf mass of suckers in the SA treatment had to support the respiratory demands of proportionately more non-assimilating biomass than in the FA treatment (see the differences in LAR in Table 2). If suckers do not establish a large leaf area, they may not be able to rebuild reserves or supply the carbon necessary to maintain respiration of the whole root system. Saplings cut when they have low root carbohydrate concentrations are likely to show suppressed growth in the next growing season.

There is a massive build-up in starch reserves in roots by late October but they are nearly depleted after bud flush in the following spring. The significance of root starch concentration in the shoot recovery of re-sprouting shrubs has been reported by Bowen & Pate (1993) and Jones & Laude (1960). While our study found large differences in starch concentrations, soluble sugar concentrations, which were more than 10% of tissue dry mass for most of the tissues, were mostly not affected by the treatments. Relatively constant levels may be needed for functions such as turgor maintenance, hardening and freezing tolerance (Larcher 1995; Lambers et al. 1998). Indeed, by the end of the experiment some live medium and large roots, which had become isolated from the rest of the clone by dead root sections, had nearly normal concentrations of sugar but were completely depleted of starch.

The development of leaf area appears to be an important factor in the maintenance of the original root system. The destabilization of the relationship between root mass and leaf mass in clones caused by cutting was nearly completely corrected by the end of the first summer in the FA treatment, where there was rapid leaf area development. This resulted in a high leaf area ratio (LAR) in the FA treatment, which was higher than in the pre-treatment saplings due to the reduced above-ground mass. High LAR is associated with fast growth rates (Lambers et al. 1998). Peterson & Jones (1997) and others suggest that the clonal growth strategy in herbaceous plants is strongly associated with reproduction, expansion and resource gathering. We believe that the primary role of clonal regeneration through suckering and rapid leaf area development in a large and long-lived clonal species such as P. tremuloides is to maintain the large below-ground biomass, and thus aid the persistence (fitness) of the clone (Cook 1985; Chazdon 1992). Many of the roots in P. tremuloides will be maintained for the lifetime of the ramets (DesRochers 2000). Also, juvenile P. tremuloides are preferred forage for a variety of wildlife species and profuse suckering and leaf development therefore allows for risk spreading and should be beneficial for the survival and maintenance of the clone (Eriksson & Jerling 1990; Chazdon 1991).

In the FA treatment, the root mass at the end of the first year of suckering was, if anything, greater than prior to cutting, suggesting that the whole root system was maintained and that new root mass was added during the growing season. The three other treatments resulted in poorer development of leaf area and showed a reduction in the overall root mass. The single sucker in the FS treatment started with the same level of root starch reserves as in FA but did not grow as tall as when there were multiple suckers. Assuming that the dominant sucker was indeed selected, the respiratory demands of the large root system appear to have higher priority for allocation of carbohydrates than sucker height growth, with spring cutting causing further suppression. An earlier study of carbon allocation in P. tremuloides seedlings suggested that the root system is a high priority carbon sink when a resource such as light is low (Landhäusser & Lieffers 2001). However, variations in auxin/cytokinin ratios have also been suggested to be the cause of seasonal differences in suckering (Schier 1973b).

The need for rapid development of leaf area to retain most of the original root system has also been suggested by DesRochers (2000), who excavated juvenile sucker stands in the field. Thus, if the clonal root system is to be maintained, it is imperative that dense suckering with full leaf area development occurs in the first year. The roots of past ramets can be an important component of the root systems of suckers for 80 or more years (Strong & La Roi 1983; DesRochers 2000) and loss of parts of the root system is therefore likely to have negative effects on the growth and performance of new sprouts in P. tremuloides.

Acknowledgements

We thank Ben Seaman, Brad Pinno, Clark Protz, Erin Fraser, Jim Cutbertson, Justine Karst, Ken Stadt, Nicki Kircher, Sarah Lieffers, Vashti Thompson and Wei Liu for their assistance in the set-up, maintenance and sample collection throughout the 4 years. We thank Pak Chow for the TNC analyses and Dick Puurveen at the Ellerslie Research Station. This study has been supported by Ainsworth Lumber Inc., Alberta Pacific Forest Industries Inc., Daishowa Marubeni International Ltd, Millar Western Industries Ltd, Slave Lake Pulp Corporation, Weyerhaeuser Canada Ltd and the Natural Sciences and Engineering Research Council of Canada (NSERC).

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