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Species turnover during succession in evergreen forests is associated with variation in a suite of leaf structural and functional traits. Shade-tolerant late-successional evergreen trees have leaves with a lower photosynthetic capacity, lower nutrient concentrations and a larger leaf mass per unit area (LMA) than fast-growing light-demanding species, when comparisons are made in a standardized light environment (Lusk & Warton, 2007; Sterck et al., 2006; Walters & Reich, 1999). In this sense, adaptation of evergreen leaf structure and function to shade seems to share features in common with adaptation to certain other unproductive environments (Reich et al., 2003; Westoby et al., 2002). The general correlation of these traits worldwide with leaf lifespan (Wright et al., 2004) suggests, at face value, that a large LMA is required to achieve a long leaf lifespan.
Curiously, plastic responses of LMA to shade run counter to the constitutive response evident in comparative studies of evergreens of differing shade tolerance (Björkman, 1972; Kitajima, 1994; Lusk et al., 2008). There is often a strong similarity between plastic and evolutionary responses of quantitative traits to environmental gradients (Conover & Schultz, 1995), yet shade leaves invariably have smaller LMA than sun leaves of the same species (Givnish, 1988; Walters & Reich, 1999). How can this apparent divergence of plastic and evolutionary responses of LMA to shade be explained? Furthermore, shade leaves often live longer than sun leaves of the same species, despite their lower LMA (Reich et al., 2004). If prolongation of leaf lifespan requires a large LMA, how can we explain the persistence of shade leaves?
In addressing this paradox, it is helpful to consider the different components of leaves that contribute to variation in LMA (Lusk et al., 2008). We can think of a leaf as composed of structural components (cell walls, cuticle) and symplastic components (cell contents, including proteins, electrolytes and nonstructural carbohydrates) (Roderick et al., 1999; Shipley et al., 2006). The large LMA of shade-tolerant evergreens probably reflects a large structural component, protecting the leaf against herbivores and physical stresses. By contrast, plastic responses to light are probably dominated by variation in the amount of cell contents per area. The most conspicuous anatomical difference between sun and shade leaves is in the amount of palisade mesophyll (Onoda et al., 2008), associated with differences in photosynthetic capacity (Niinemets & Tenhunen, 1997; Terashima et al., 2001). Shade leaves would therefore be expected to have a slightly larger cell wall fraction than sun leaves, and there is some empirical support for this idea (Onoda et al., 2008; Poorter et al., 2006).
It is also helpful to consider the main threats to leaf survival. Wind abrasion, tatter and other damage caused by the elements mainly affects plants in exposed, open habitats (MacKerron, 1976), as well as tree crowns in the forest canopy (Putz et al., 1984). By contrast, plants in all habitats are exposed to herbivores, which have been estimated to cause annual leaf area losses of 14–26% in humid evergreen forests (Lowman, 1984). Although plants have evolved a celebrated diversity of chemical defences, especially against insect herbivores (Coley & Barone, 1996; Rosenthal & Janzen, 1979), mechanical strength remains the most effective generalist defence of fully expanded leaves, and mechanical properties are often good predictors of the degree of herbivory (Choong, 1996; Coley, 1983; Peeters et al., 2007). Overall fracture resistance is, in turn, primarily a function of the amount of cell wall per unit area (Choong, 1996). The cell wall fraction of leaves is also likely to influence the attractiveness of plants to chewing herbivores, by determining the cost–benefit economics of feeding choices. A shade leaf with a small LMA but a large cell wall fraction might be unattractive because of the modest return of energy and nutrients from the cell contents, compared with the amount of energy required to cut, chew and digest the leaf. If herbivores are the biggest single threat to leaves, cell wall fraction may be a robust correlate of leaf lifespan, irrespective of growth light environment.
We examined structural and mechanical properties of sun and shade leaves of 13 rainforest evergreens of differing light requirements, in an attempt to reconcile plastic and constitutive responses of LMA to shade. We addressed the following hypotheses: adaptation of evergreens to light gradients involves variation in the amount of cell wall per unit area; plastic responses of LMA to light gradients mainly reflect variation in the amount of cell contents per unit area; plastic responses of leaves to light affect leaf mechanical properties less than LMA. To this end, we measured LMA and cell wall content of sun and shade leaves of each species, and conducted punch and shear tests.
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In agreement with our first hypothesis, the relationships between species shade tolerance and LMA of both sun and shade leaves were entirely the result of variation in CWA (Fig. 1). In both sun and shade, the four most shade-tolerant species had CWA values that were, on average, three times higher than the four most light-demanding species. This correlates closely with the biomechanical results, showing that leaves of shade-tolerant species were more resistant to fracture than those of their light-demanding associates (Fig. 1). By contrast, mass of cell contents per unit area was not related to shade tolerance or to mechanical properties (Figs 1, 2). Several other studies have reported similar patterns. Coley (1983) reported that shade-tolerant rainforest saplings had higher leaf fibre content and ‘toughness’ (more appropriately called punch strength) than species associated with treefall gaps. Similarly, successional position of tropical rainforest species has been shown to correlate with leaf lignin content (Poorter et al., 2004) and fracture resistance (Reich et al., 1995). The sum of the evidence seems to confirm that adaptation of evergreens to shade, like adaptation to nutrient-poor habitats (Reich et al., 2003), usually involves prolongation of leaf lifespan through investment in a large structural fraction.
Although leaves of most shade-tolerant species appear to be defended primarily by their mechanical properties, there may be exceptions. Laureliopsis philippiana (Looser) Shodde (Atherospermataceae), one of the most shade-tolerant trees of the temperate rainforests of South America, has long-lived leaves that nevertheless have high nitrogen concentrations (> 1.5% N: Lusk & Contreras, 1999) and quite a low dry matter content (CH Lusk, unpublished). Nitrogen concentrations > 1.5% have also been found in other shade-tolerant species of the same family, such as Atherosperma moschatum Labill. and Doryphora sassafras Endl. in southeastern Australia (Iddles et al., 2003; J. Read, pers. comm.); by contrast, many other shade-tolerant species in the temperate rainforests of the southern hemisphere seem to have nitrogen concentrations of c. 1% (Iddles et al., 2003; Lusk & Contreras, 1999). The Atherospermataceae are rich in alkaloids (Urzua & Cassels, 1978; Webb, 1949, 1952), which probably act as a chemical defence against herbivores. These observations seem to support the proposal that herbivores, rather than damage by weather or falling debris, are the main threat to leaf survival in shaded understoreys.
There was partial support for our second hypothesis, that plastic responses to light gradients mainly involve changes in the amount of cell contents per unit area. Sun leaves averaged 47% more cell contents per unit area than shade leaves, compared with 22% more cell wall mass. This is consistent with anatomical comparisons of sun and shade leaves, which often show large differences in the amount of palisade mesophyll, but less difference in structural features such as cuticle and cell wall thickness (Onoda et al., 2008; Terashima et al., 2001). As a result, the cell wall fraction of shade leaves was slightly higher than that of sun leaves (Fig. 1). This is fairly consistent with the results of a study of 22 diverse plant species (Poorter et al., 2006), which reported that shade leaves had significantly higher concentrations of structural carbohydrates than sun leaves. Similarly, Onoda et al. (2008) found that shade leaves of the herb Plantago major (Plantaginaceae) had a larger cell wall fraction than sun leaves.
Consistent with our third hypothesis, light environment had less effect on leaf biomechanics than on LMA (Tables 2, 3; Fig. 1). Shade leaves required only slightly less force-to-punch than sun leaves of the same species (Fig. 1h), and a larger specific force-to-punch (Fig. 1i). The greater strength of shade leaves at a given LMA (Fig. 2b) reflected their having a slightly larger cell wall fraction than sun leaves, and a slightly higher leaf density. Our results therefore generalize the findings of Onoda et al. (2008), who reported that, at a given LMA, shade leaves of P. major were stronger than sun leaves. Onoda et al. (2008) found that this was the result not only of a larger cell wall fraction in shade leaves, but also of a higher specific strength of cell walls. By contrast, we found that the relationship of force-to-punch with CWA did not differ much between sun and shade leaves (Fig. 2; Table 4), implying that light environment had little effect on specific cell wall strength. Sun and shade leaves differed more noticeably in work-to-shear (Fig. 1j), although not commensurately with differences in LMA. The percentage of leaf area occupied by veins tends to be higher in sun leaves than in shade leaves (Eschrich et al. 1989), and this may explain the greater work required to shear the former. By contrast, punch tests were conducted on intercostal regions of the lamina; the resulting lack of sensitivity to differences in vein area is probably one factor underlying the similarity of the results between punch tests on sun and shade leaves (Fig. 1h).
Shade leaves might be relatively unattractive to chewing herbivores because feeding on them involves a lower ratio of benefits to costs than feeding on sun leaves (Fig. 1i). Analyses often link leaf longevity to LMA (Wright et al., 2004), which seems reasonable on the basis of LMA being well correlated with measures of leaf fracture resistance in large datasets (Read et al., 2005). However, if herbivores are the biggest threat to leaf survival, the cost–benefit economics of herbivory may also be highly relevant to both inter- and intraspecific variation in leaf lifespan. Although a wide range of chewing herbivores may be capable of cutting through the laminas of shade leaves, feeding on them may be a relatively unattractive option for some herbivores because they offer smaller benefits per unit area (energy and nutrients in cell contents) per unit force required for ingestion (Figs 1, 2). The small amount of mesophyll in shade leaves might also make them unattractive to leaf miners. Cafeteria experiments have shown that generalist insect herbivores usually prefer sun leaves to shade leaves (Maiorana, 1981), although field studies comparing actual rates of herbivory on sun and shade leaves have yielded varying results (Louda et al., 1987; Lowman, 1985; Maiorana, 1981). The longer lifetimes of shade leaves compared with sun leaves (Reich et al., 2004) reflect more gradual self-shading in low-light environments where plants grow slowly (Ackerly & Bazzaz, 1995). However, the persistence of shade leaves also implies that they are at least as well protected as sun leaves; we suggest that at least part of this protection takes the form of economic unattractiveness to herbivores.
We found no evidence that plasticity of leaf structural or mechanical parameters was related to shade tolerance. In no case did the slopes of trait relationships with species’ light requirements differ significantly between sun and shade leaves (Table 3; Fig. 1). It has been hypothesized that leaf traits of light-demanding species will respond more plastically to light environment than those of their shade-tolerant associates (Bazzaz, 1979). Some of the strongest evidence to date in support of Bazzaz’s hypothesis comes from a comparative study of 16 sympatric Psychotria species, which reported that a wide range of physiological and morphological traits were more plastic in light-demanding species (Valladares et al., 2000). A review of seedling data gathered from diverse studies found that, of several traits analysed, only specific leaf area (1/LMA) showed evidence that amplitude of plasticity was related to light requirements, and even that evidence was not especially strong (Walters & Reich, 1999; Table 2). Experimental treatments often confound ontogenetic drift with plastic responses to treatments, because of growth rate differences between species at high resource levels (Coleman et al., 1994). Observational field studies, when they involve adequate quantification of light environments and comparisons of similar-sized plants, provide good opportunities to minimize ontogenetic effects on apparent plasticity. Studies such as these have not lent much support to the hypothesis of greater plasticity in light-demanding species (Chazdon & Kaufmann, 1993; Lusk, 2002; Lusk & Reich, 2000).
Our results point to a means of reconciling the puzzling divergence between inter- and intraspecific relationships of LMA with leaf lifespan. Long leaf lifespans are characteristic of both constitutive and plastic response of evergreens to shade; both these responses also involve increased cell wall fraction of leaves, despite divergent trends in LMA. The persistence of shade leaves, despite their low LMA, may thus reflect the economics of herbivory, as shade affects the amount of cell contents per unit area more than leaf strength. Our results also show that cell wall fraction and punch strength are more robust correlates of species light requirements or successional position than LMA, which is strongly responsive to light environment (Tables 2, 3). Although LMA is useful as an indicator of biomass partitioning (Lambers & Poorter, 1992; Westoby et al., 2002), neutral detergent fibre and punch tests might be preferable in studies aiming to index functional diversity of plant species in environments subject to wide variation in light intensity, such as forests.