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Despite the significant selective pressure that herbivores and pathogens exert on plants, marked, genetically based, quantitative variations in chemical defenses (genetic polymorphisms) persist in plant populations (Simms, 1992; Karban & Baldwin, 1997; Mauricio, 2000). Defense costs, in the form of tradeoffs between growth, reproduction and defense, provide the most widely proposed explanation for why defensive traits remain variable (Simms & Rausher, 1987; Herms & Mattson, 1992).
Several prominent theories of plant defense against herbivores, such as optimal defense (McKey, 1974; Rhoades, 1979) and growth–differentiation balance (Herms & Mattson, 1992), suggest that optimization of costs and benefits leads to genetically variable levels of defense production within plant populations. In essence, these theories suggest that when herbivores are present and benefits outweigh costs, plant defenses increase fitness. If, however, defenses are costly and herbivores are absent, then high levels of defense decrease fitness. Defense costs are generally defined as a decrease in growth or reproduction in defended individuals in the absence of herbivores (Simms & Rausher, 1992).
According to several reviews, the number of studies that have identified costs associated with allocation to defense roughly equals the number that failed to do so (Simms, 1992; Bergelson & Purrington, 1996; Koricheva, 2002; Strauss et al., 2002). Although these equivocal results have led to some doubt about the importance of costs, it is more likely that they reflect an incomplete understanding of the mechanisms by which costs are realized, and the importance of ecological factors in mediating costs (Berenbaum, 1995; Purrington, 2000; Strauss et al., 2002). Costs of defense are probably manifest via a plant's biotic and abiotic interactions (e.g. competition, herbivore complexes, abiotic stress: Koricheva, 2002; Strauss et al., 2002). As noted by Purrington (2000), hundreds of studies have identified costs, and recent literature is focusing not on whether defense is costly, but rather under what conditions and by what mechanisms costs are or are not realized.
Resource availability may have a marked influence on the realization of defense costs. For example, high resource availability may diminish allocation costs, thereby allowing for both growth and defense (Siemens et al., 2002; Osier & Lindroth, 2006). Correspondingly, environmental stress may significantly increase the magnitude of defense costs (Weis & Hochberg, 2000; Marak et al., 2003; Siemens et al., 2003; Osier & Lindroth, 2006). According to the growth–differentiation balance hypothesis (Herms & Mattson, 1992), if levels of defense remain stable, but resources such as light or nutrients decrease, then fewer resources will be available for growth or reproduction. Further, ecological stress (water, nutrients, competition) can induce plants to produce increased concentrations of secondary metabolites (Gershenzon, 1984; Inbar et al., 2001; Osier & Lindroth, 2006), thereby compounding costs. Theoretically, however, environmental stress may not always compound the costs of defense (Bergelson & Purrington, 1996). Bryant et al. (1983) predicted that nitrogen limitation constrains growth more than photosynthesis, leading to a relative increase in the fixed carbon pool. ‘Excess’ C is predicted to result in increased allocation to ‘C-based’ secondary metabolites (tannins; phenolic glycosides, PG) at little cost to growth.
Aspen (Populus tremuloides) experiences considerable intra- and interspecific competition, particularly when it is young (Landhäusser & Lieffers, 1998). Marsh reed grass (blue joint grass, Calamagrostis canadensis) is an aggressive native species in mixed boreal forests that quickly colonizes and dominates disturbed areas (Dyrness & Norum, 1983). Marsh reed grass and other herbaceous cover can suppress both sexual and asexual aspen regeneration due to shading, thermal inhibition (Landhäusser & Lieffers, 1998; Ball et al., 2002), and root competition (Scholes & Archer, 1997; Landhäusser & Lieffers, 1998; Powell & Bork, 2004). Although light limitation is typically assumed to be the most important stress, competition for water and nutrients is often equally, if not more important for plants (Wilson, 1988; Coomes & Grubb, 2000; Powell & Bork, 2004). Belowground competition is a significant stress for regenerating aspen, and competitive interactions in the first season of growth are very important for seedling survival (Powell & Bork, 2004). As suggested by Purrington (2000), plant competition may increase the magnitude of growth–defense tradeoffs.
A primary objective of this study was to assess the independent and interactive effects of plant genotype, nutrient availability and belowground competition on allocation to secondary metabolites and growth. We were interested in the extent to which environmental stress (nutrient stress and belowground competition) changes patterns of biomass allocation (e.g. root : shoot; leaf mass ratio; specific leaf area; condensed tannins; PG concentrations) in young trembling aspen. A second objective was to identify potential tradeoffs between growth and allocation to secondary metabolites and to assess the degree to which environmental stress mediates defense costs. Tradeoffs between growth and allocation to secondary metabolites were assessed via negative phenotypic correlations. Finally, we examined growth determinants (specific leaf area; photosynthesis; leaf mass ratio) to identify potential mechanisms by which costs may be realized.
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Aspen typically exhibits considerable genetic variation with regard to plant chemistry and growth (Lindroth & Hwang, 1996; Hwang & Lindroth, 1997; Osier & Lindroth, 2006). This study sought to identify potential relationships between growth and allocation to secondary metabolites as they vary among genotypes and in response to nutrient availability and belowground competition.
Although growth varied modestly among genotypes, a significant negative phenotypic correlation between RGR and levels of PGs was nevertheless observed. Similarly to the results of Osier & Lindroth (2006), this tradeoff was evident only when resources were limiting. An examination of the relationship between PG concentration and other growth determinants (photosynthesis, leaf mass ratio, specific leaf area) did not reveal a mechanism for this putative defense cost. For example, a relative increase in allocation to nonstructural leaf mass (e.g. secondary chemicals, nonstructural carbohydrates) can lead to decreased specific leaf area and therefore a dilution of N and decreased assimilation efficiency (Roumet et al., 1999). Although specific leaf area trended downward as levels of PGs increased (not shown) in this study, the relationship was not statistically significant. There is evidence that PG maintenance costs can be high in aspen (Kleiner et al., 1999), although research with willow has shown that these compounds are surprisingly stable and turn over relatively slowly (Ruuhola & Julkunen-Tiitto, 2000). The fact that the tradeoff occurred only when resources were limiting suggests that allocation or maintenance costs, as predicted by the growth–differentiation balance hypothesis (Herms & Mattson, 1992) or protein-competition model (Jones & Hartley, 1999), may explain a portion of the patterns observed.
The effects of nutrient availability and belowground competition appear to be additive for the majority of traits measured in this study. Most variables either increased or decreased in a linear fashion with increasing stress (decreased fertility and increased competition). Although nutrient uptake was not measured, the lower leaf [N] in competition treatments, observed with both high and low fertility, indicated that growth losses resulting from the competition treatment were probably related to decreases in either nutrient availability (e.g. caused by nutrient usurpation by grass) and/or aspen nutrient uptake. There was some evidence implicating the latter. Soil testing at harvest indicated that, in the low-fertility treatment, soil NO3− levels were actually slightly higher with competition than without (data not shown). Grass could have interfered with aspen's nutrient acquisition in any of several ways (Aerts & Chapin, 2000). For example, competition-mediated decreases in tree transpiration might have led to declines in transport of mobile ions, such as NO3− to the root via mass flow (McDonald et al., 2002).
Plant competition may affect plant growth and allocation indirectly via its influence on secondary metabolite production. Although Agrawal (2004) found that competition has no effect on levels of cardenolides or latex in milkweed, results of other studies indicate that competition decreases allocation to defensive compounds in Brassica napus (Cipollini & Bergelson, 2001) and tomato (Stamp et al., 2004). The effect of competition may be dependent on the class of compounds produced in the plant. For example, Marak et al. (2003) found that competition may affect allocation to C-based compounds differently from allocation to non-C-based compounds. In their study of Plantago lanceolata, competition significantly increased allocation to iridoid glycosides. They suggest that plant carbon–nutrient balance may explain such increases. The carbon–nutrient balance hypothesis (CNB; Bryant et al. 1983) predicts that nutrient stress results in ‘excess’ fixed C and therefore increased allocation to C-based chemical defenses. The theory has been debated intensely (Hamilton et al., 2001), but many plants, including aspen, appear to follow its predictions (Hemming & Lindroth, 1999; Lerdau & Coley, 2002). For example, in the present experiment CT concentrations increased with decreasing fertility and with competition.
We note, however, that a key exception to the predictions of CNB occurred in this study. Increased production of CTs under limited resource availability appeared to exact a cost for growth in aspen (increased CT was not solely a product of ‘excess’ C). Relative to that in high fertility, RGR in low fertility was generally less than would be expected for a given rate of photosynthesis per unit plant mass. This discrepancy was so large that we examined an additional index of growth potential, N mass ratio (calculated in the same manner as CT and PG mass ratios), to ascertain whether photosynthesis per unit plant mass underestimated the direct effect of N limitation on growth.
Nitrogen mass ratio was very similar to photosynthesis per unit plant mass in terms of its role as a predictor of RGR (regression not shown). Thus neither predictor explained all the growth loss exhibited by stressed trees. Rather, some correlate of N limitation constrained growth. We acknowledge that, because N, P and K were manipulated together in our soil fertility treatment, we cannot rule out the influence of P and K in explaining some of the growth differences among treatments. However, allocation and maintenance costs of CTs are second only to lipids and lignin in their relative cost (mg C g−1) to plants (Chapin, 1989), and the more than twofold increase in levels of tannins in low-fertility trees was correlated with the difference between realized growth rates in low vs high nutrient trees. Still, the difference in RGR was greater than is likely based on CT construction costs alone (Chapin, 1989; Poorter, 1994), and other factors may have played a role (e.g. P, K, water availability). Although we are unable to explain fully the magnitude of such a reduction in realized growth, resource limitation and (to a lesser extent) plant competition appear to change allocation patterns in aspen, and may result in compounded costs of chemical defense.
Herbivory and plant competition are among the primary selective agents operating in plant communities. The growth–differentiation balance hypothesis (Herms & Mattson, 1992) provides a conceptual model that describes how these two selective agents interact to maintain genetic variation in plant chemical defense in plant populations. Physiological constraints within the plant lead to a tradeoff between competitive ability (high growth rates) and resistance to herbivory (high levels of defense). If the relative importance of plant competition and herbivory vary in time and space (as they may), then natural selection will maintain variation in a population (Herms & Mattson, 1992). Aspen experiences significant selection pressure via competitive interactions (Barnes, 1966; Landhäusser & Lieffers, 1998; Powell & Bork, 2004) and from herbivores (Lindroth & Hwang, 1996). Genetically variable allocation to chemical defense in aspen may, in part, be maintained by environmental heterogeneity in the relative importance of these two opposing selective pressures.
Results from this study support the hypothesis that there is a tradeoff between growth and defense in establishing (seedling) aspen, and that the tradeoff is mediated by resource availability. When resources are limiting, not only is the cost of chemical defense realized, but changes in allocation (increased condensed tannin concentrations) compound costs by further limiting growth. The effects of this tradeoff are likely to be particularly important during aspen establishment, in both their immediate effects and their long-term influence on aspen populations (Powell & Bork, 2004).