Divergence in functional traits in seven species of neotropical palms of different forest strata

Functional traits are morphological and physiological characteristics that determine growth, reproduction, and survival strategies. The leaf economics spectrum proposes two opposing life history strategies: species with an “acquisitive” strategy grow fast and exploit high-resource environments, while species with a “conservative” strategy emphasize survival and slow growth under low resource conditions. We analyzed intra and interspecific variation in nine functional traits related to biomass allocation and tissue quality in seven Neotropical palm species from understory and canopy strata. We expected that the level of resources of a stratum that a species typically exploits would determine the dominance of either the exploitative or conservative strategy, as well as degree of divergence in functional traits between species. If this is correct, then canopy species will show an acquisitive strategy emphasizing traits targeting a larger size, whereas understory species will show a conservative strategy with traits promoting efficient biomass allocation and survival in the shade. Two principal components (57.22% of the variation) separated palm species into: (a) canopy species whose traits were congruent with the acquisitive strategy and emphasized large size (i.e., diameter, height, carbon content, and leaf area), and (b) understory species whose traits were associated with efficient biomass allocation (i.e., dry mass fraction -DMF- and tissue density). As we unravel the variation in functional traits in palms, which make up a substantial proportion of the tropical flora, we gain a deeper understanding of how plants adapt to environmental gradients.


Introduction
Functional traits are morpho-physio-phenological characters that impact fitness through their effects on growth, reproduction, and survival (Violle et al. 2007).Variation in functional traits influence plant life history strategies, resource allocation for growth and defense (Coley et al. 1985), niche differentiation, and environmental filtering (Westoby et al. 2002;Poorter et al. 2008;Boukili and Chazdon 2017).Leaf economics spectrum theory has shown the existence of a universal trade-off between two opposing life history strategies (Wright et al. 2004).On one hand, species exhibiting an "acquisitive" strategy are characterized by high photosynthetic rates, thinner and low-cost leaves, with high nitrogen and phosphorus content and high specific leaf area (SLA) that exploit high-resource environments.On the other hand, species with "conservative" strategies have thicker, tougher, low SLA leaves, with low photosynthetic rates, higher construction costs, low nitrogen content, and long lifespans, and exploit low-resource environments (Westoby et al. 2002;Wright et al. 2004).These strategies are consistent across biomes demonstrating the universality of this trade-off (Díaz et al. 2004(Díaz et al. , 2016)).
Palms (Arecaceae) are one of the most diverse and widely distributed groups of plants in tropical and subtropical areas, with more than 2600 species and 181 genera (Baker and Dransfield 2016) that dominate many tropical ecosystems (Mejia and Kahn 1990;Myers 2013).Palms are "hyperdominant" elements in the Amazon lowlands (ter Steege et al. 2013), with 6 out of the 10 most abundant species being palms.Although palms have a limited contribution to carbon stocks in diverse tropical rainforests (Fauset et al. 2015), they influence forest function (Boukili and Chazdon 2017), play a crucial role in food webs, provide habitat and food to a multitude of animal species (Zona and Henderson 1989;Howard et al. 2001;Onstein et al. 2017), and are invaluable to many human groups who use them as raw materials for construction, food, drink, clothing, fuel, medicine, and fibers (Jones 1995;Henderson 2002;Dransfield et al. 2008;Sylvester et al. 2012;Sander et al. 2023).To improve our understanding of the ecological role of palms of different forest strata on food webs, forest succession, and community structure, it is essential to improve the quantification and analysis of their inter and intraspecific variation in functional traits (Westerband et al. 2021).
Palms have been excluded from most inventories of functional traits in tropical forests (DeWalt and Chave 2004;Chave et al. 2005;Lorenz and Lal 2010).As monocots, they have a different structure, allometry, and strategies of resource use relative to trees (Tomlinson 2006(Tomlinson , 2011)).With a few exceptions, palms are monopodial and lack aerial branching, have only one shoot meristem, and lack dormancy and secondary growth.In palm species where stem diameter and stem height show a significant relationship, diameter increases through sustained primary growth (i.e., through the division, lignification, and expansion of parenchyma cells, which also differentiate into fibers, Henderson 2002; Tomlinson 2011).In addition, leaf longevity and leaf construction costs are higher in palms than in dicotyledonous trees (Renninger and Phillips 2016), which have smaller leaves and could drop leaflets rather than the entire compound leaf to acclimate to new light conditions.Within the family Arecaceae there is considerable morphological variation which is reflected in niche differentiation and habitat filtering (Henderson 2002).
Our main objective is to analyze the intra and interspecific variation in nine functional traits related to biomass allocation and tissue quality (tissue density, DMF, slenderness ratio, carbon content, diameter, height, leaf area, and root:shoot ratios based on biomass and carbon content) in seven palm species from two forest strata (understory and canopy).We tested the hypothesis that the dominant light environment in the stratum (understory and canopy) that a palm species typically exploits at maturity, will determine the strategy of resource use, and thus, the degree of functional trait similarity among species of the same stratum.Therefore, palms that complete their life cycle in the understory will show a conservative resource-use strategy that emphasizes efficient biomass distribution (dominant traits in this strategy will be tissue density, slenderness ratio, root:shoot ratios, and DMF), in contrast to palms exploiting better lit environments (canopy species) that will show an acquisitive strategy, and therefore will be characterized by functional traits favoring large size and leaf area (i.e., diameter and height, total carbon content, leaf area).Exploring the variation of functional traits within palms, one of the most abundant life forms in tropical forests, could significantly expand our understanding of how tropical plants in general adapt to environmental gradients.

Study species
We harvested 87 individuals of seven palm species belonging to two forest strata: the canopy and the understory (Fig. 1).Canopy species included Socratea exorrhiza, Iriartea deltoidea, and Euterpe precatoria var longevaginata.Socratea exorrhiza; (Mart.)H. Wendl.(southern Nicaragua to Brazil, 0-750 masl), is a solitary palm with a cone of separate stilt roots covered in spines, which can reach up to 4 m above the ground (Henderson et al. 1995).Iriartea deltoideia; Ruiz & Pav.(SE Nicaragua to Brazil, 0-800 masl) is also single stemmed with a cone of clustered stilt roots that can reach up to 1.5 m above the ground.Euterpe precatoria; Mart.1842 (Belize to Bolivia, 0-1150 masl), is also a solitary palm with a cone of clustered orange stilt roots that can reach more than 4 m in height in extreme cases (Avalos and Schneider 2011).Maximum canopy height varies among these species, with S. exorrhiza reaching 30 m, while both I. deltoidea and E. precatoria have a maximum height of 25 m (Grayum 2003).Understory species included Prestoea decurrens, Chamaedorea tepejilote, Geonoma interrupta and Asterogyne martiana.Prestoea decurrens; H. E. Moore (Nicaragua to Ecuador, 0-900 masl), is a clonal  species reaching 10 m in height (Grayum 2003).Chamaedorea tepejilote; Liebm.(S Mexico to Colombia, 0-1600 masl) is a dioecious species which can grow up to 5 m (Grayum 2003;Castillo-Mont et al. 1994).Geonoma interrupta; (Ruiz & Pav.) Mart.(S Mexico to Peru, 0-850 masl) has a solitary stem and may reach 6 m in height (and over 10 m in exceptional cases), being one of the tallest species in the genus (Grayum 2003).Finally, Asterogyne martiana; (H.Wendl.)H. Wendl.ex Drude (Belize to Ecuador, 0-1000 masl) is an understory species with a decumbent stem often reaching 2 m in height, and with simple, bifid leaves.We sampled 10 palms per species, except for C. tepejilote (22) and A . martiana (15), making sure to include individuals of a wide range of sizes per species.

Palm harvesting, morphological measurements, and estimation of carbon sequestration
Detailed harvesting methods are described in Avalos et al. (2022).In summary, harvesting took place from Sept 2013 to May 2015, with the goal of obtaining a representative sample with a wide range of size per species (Fig. 1).For canopy palms, this sample did not include individuals close to the maximum heights reported for the species due to the difficulty of obtaining permission to harvest very tall individuals (Fig. 2F).Stem diameter was measured at 1.3 m above the ground, at half the stem length in palms less than 1.3 m tall, or immediately above the stilt roots in palms with a root cone that surpassed 1.3 m in height.The harvested palms were separated into modules (stems, roots, and leaves) and the total fresh biomass of each module was measured in the field with a Pesola ® Macro-Line Spring Balance (30 ± 0.25 kg).Then, these samples were dried in an oven at 65 °C for 48 h or until constant weight.Carbon content was measured with an automated TruSpec CN analyzer, LECO Corporation, at the Laboratory of the Department of Systematic Botany of the University of Ulm, Germany, and an automatic analyzer for nitrogen and elemental carbon, VarioMacrocube, at the University of Costa Rica.The average carbon fraction for the palms analyzed here was 43.9% ± 1.28 (Cambronero et al. 2018), but we used the average carbon fraction obtained for each species multiplied by the estimated total biomass per individual and module (root, shoots, leaves).Dry mass fraction or tissue moisture (DMF) was measured as the ratio of total dry over total fresh biomass per individual.
We measured the total length of the stem, or stem height, from the point of connection with the roots to the base of the petiole of the youngest leaf.This included the underground stem in A. martiana.Slenderness ratio was calculated as the ratio of stem height in m to diameter in cm (Niklas 1994;Niklas et al. 2006).
To measure stem tissue density (specific gravity, ρ, g/mL) we used with a Haglof 2-wire incremental borer following the methods of Chave et al. (2005).We selected an entry point for the borer near the base of the stem, in the middle, and near the base of the crown.This tissue sample was placed in a test tube, sealed, and transferred to the laboratory for estimation of tissue density by volume displacement.
To determine the total leaf area, we followed the methods of Avalos and Sylvester (2010).We collected three leaves (one young, one intermediate, and one mature), cleaned them with a dry cloth, and measured leaf area with a LICOR LI-3100 C leaf area meter (LICOR, Lincoln, NE, USA).From these measurements we estimated the total leaf area per individual palm by averaging the leaf area of these three leaves and multiplying it by the total number of fronds.

Root:shoot ratios
We calculated root:shoot ratios as the dry biomass of roots over the dry biomass of above-ground parts (stems and leaves).This corresponded to the root:shoot ratio of dry biomass.In addition, we obtained the root:shoot ratio of the carbon fraction after calculating the carbon fraction of the above-ground and below-ground biomass (this was the root:shoot ratio based on the carbon fraction).

Analysis of the correlation structure of morphological traits of understory vs canopy palms
We used a principal component analysis (PCA) based on the correlation matrix of the ln-transformed values of nine morphological traits related to biomass allocation and tissue quality: the total amount of carbon content in kg per palm, diameter in cm (diam), total stem height from the base of the stem to base of the petiole of the youngest leaf, DMF, stem tissue density, leaf area, slenderness ratio and root:shoot ratios (calculated based on the dry biomass, and carbon content levels).The ln-transformed variables were standardized before running the analysis.We used the scores of the first two components to inspect the distribution of palm species across forest strata in the multidimensional space defined by correlation structure of morphological traits.To measure differences in trait structure between strata and among canopy and understory species we used the two first principal component to summarize the variation in morphological traits following this nested ANOVA model: where Y ijk = is the kth observation of the scores of the component associated to the stratum-i of the species-j, α i = effect of the stratum-i, (αβ) ij = effect of the species-j nested within the stratum-i, and ε ijk = error term.We applied this model to the first and second principal components.R statistical software was used for all analyses (packages corrplot, RColor-Brewer, PerformanceAnalytics, factoextra and FactoMineR, R Core Team 2022).

Results
Palms showed a wide range of variation in morphological traits with biomass increasing from understory to canopy species (Fig. 2).The understory species Prestoea decurrens and G. interrupta included very large individuals and overlapped in height, leaf area, and diameter with canopy species (Fig. 2E-G).These two species also showed overlap in the amount of sequestered carbon with canopy species (Fig. 2I).Canopy species showed less variation than understory species in DMF and tissue density (Fig. 2C, D).The maximum heights obtained for canopy species were 8.5 m (I.deltoidea), 12 m (S. exorrhiza), and 10.2 m (E.precatoria), all mature palms.

Correlation matrix of morphological traits
The highest correlations were found between diameter and total carbon (Pearson correlation coefficient = 0.89), diameter and total leaf area (0.79), height and leaf area, and tissue density and DMF (both with 0.74), and leaf area and total carbon (0.69, Fig. 3A).Root:shoot ratios showed little association with the rest of the variables.The correlogram distinguished two groups of traits with positive correlations.The first group included traits related to palm size (i.e., height, carbon content and leaf area), whereas the second group included traits associated to biomass distribution (i.e., DMF and tissue density, Fig. 3B).These two groups were negatively associated with each other.

Principal component analysis
We identified four principal components with an eigenvalue > 1 that explained 87.48% of the variation (Table 1).The first component (40.32%) showed a similar weight for variables related to palm size (e.g., diameter, total carbon content, leaf area, and height).The other three components had a similar weight.The second component (16.89%) had a similar contribution of tissue density and DMF.The third component (16.35%) had a dominant contribution of root:shoot ratio based on total dry biomass.Slenderness ratio and root:shoot ratio based on total carbon content dominated the fourth component (13.91%).
The distribution of palms in the multidimensional space defined by the first two components showed segregation according to forest strata and demonstrated that canopy species were more associated to palm size traits (diameter, stem height, leaf area, total carbon content, Fig. 4), while understory species were more related to biomass distribution traits (tissue density and DMF), which confirms our hypothesis of functional trait segregation according to forest strata.Understory species showed higher variation in functional traits.In the first principal component there were significant differences between strata (nested ANOVA, r 2 = 0.66, F 1,80 = 105.85,P < 0.0001, Fig. 5A) and species within a stratum (F 5,80 = 14.06,P < 0.0001, Fig. 5B).Here,  the tallest individuals of the understory species, P. decurrens and G. interrupta, overlapped with the shortest individuals of the canopy species.This overlap was also evident in the PCA score plot (Fig. 4).In the case of the second principal component, there were significant differences among strata (nested ANOVA, r 2 = 0.22, F 1,80 = 24.39,P = 0.0001, Fig. 5C) but not for species within a stratum (F 5,80 = 1.92,P = 0.09, Fig. 5D).On average, understory species showed higher scores than canopy species along the DMF (Fig. 2C) and tissue density axis (Figs.2D, 4), but differences among species were not significant.

Discussion
Our results demonstrate that palm species adapt to the light gradient following a continuum of "conservative" and "acquisitive" strategies along the leaf economic spectrum (Westoby et al. 2002;Wright et al. 2004).The classification of palms in forest strata revealed a segregation in the resource use strategy between understory and canopy species, as deduced from the principal components analysis.This segregation was not clearcut as not all traits separated canopy from understory species.There was overlap between the tallest individuals of the understory species and the shortest individuals of the canopy species in the space defined by the principal components, but not for all traits.The differentiation between canopy and understory species was more pronounced along PC1 relative to PC2.PC1 mainly reflected traits associated with size, such as stem  diameter, height, leaf area, and total sequestered carbon.Canopy palms invest more in these traits, while understory species target traits related to biomass distribution and packing (PC2), such as DMF and tissue density.These differences in allocation strategies demonstrate general trends, either maximizing survival in the shade while increasing growth in height to reach the canopy, or reaching reproductive maturity in the shade, and were consistent with our hypothesis that the dominant light environment in the stratum that a palm reaches at maturity will determine resource use strategy and, thus, will influence niche segregation.Contrary to our expectations, some functional traits were not effective in separating understory versus canopy species.For instance, shoot:root and slenderness ratios overlapped among forest strata.In the canopy species S. exorrhiza and E. precatoria slenderness ratio increases with stem height (Avalos et al. 2019), which is related to a greater investment in stature once sufficient mechanical support is achieved at the base of the stem.This in turn facilitates a greater biomass allocation to a larger crown formed by fronds with long lifespans.Slenderness ratio showed greater variation in understory species, which could reflect the greater light heterogeneity of this stratum relative to the canopy (Montgomery and Chazdon 2001).Root:shoot ratios did not show a clear separation between palms of different strata.This could have been affected by the error involved in estimating root biomass, since roots are difficult to extract and measure (Hairiah et al. 2001).In palms, there is a dearth of comparative data for root biomass, and consequently, root:shoot ratios (but see Goodman et al. 2013;Avalos et al. 2022).

Conservative strategy of understory palms
Understory species were more associated to tissue density and DMF than canopy species.Tissue density is a leaf economics spectrum trait related to stem construction costs, plant architecture and stability, stem hydraulic conductance (Chave et al. 2006), as well as plant mortality (Kraft et al. 2010) and relative growth rate (Iida et al. 2016).Tissue density is a key predictor of carbon sequestration in the general model of Chave et al. (2014) since it is positively related to the aboveground biomass of tropical forest species.The model of Chave et al. (2014) was not significantly different from the family model for Arecaceae proposed by Avalos et al. (2022), which included only diameter and stem height.Likewise, DMF was part of the composite variable used to estimate the aerial biomass of Arecaceae in the study of Goodman et al. (2013), whose model also converged with those of Chave et al. (2014) and Avalos et al. (2022).The importance of tissue density and DMF needs to be further investigated as very few palm species have been examined to date.In our case, understory species, as a group, showed higher DMF and tissue density than canopy species, but also had higher variation, and hence differences among species were not significant.Higher DMF and tissue density point towards a resource-conservative strategy emphasizing slow growth and increased survival under low light (Poorter 2009).
Many understory palms display their leaf area with a high degree of efficiency by reducing leaf overlap and increasing light interception (Takenaka et al. 2001;Alvarez-Clare and Avalos 2007).Efficiently allocating biomass resources in the shade has a higher selective value in shade-adapted palms, as well as in palm species that start their life cycle in the understory (Westoby et al. 2002).Shade-adapted palms are positioned at the end of the resource conservative strategy and can complete their life cycle and reproduce in the understory (Chazdon 1986b;Sylvester and Avalos 2013;Avalos 2019).When light conditions improve, palms that start in the shade can opportunistically increase their leaf area (Sylvester and Avalos 2013) and augment height growth (Chazdon 1986a;Gatti et al. 2011).Canopy palms such as S. exorrhiza and E. precatoria increase their slenderness ratio with height as they escape the understory and expose their crown to better lit conditions (Avalos et al. 2019).Once these palms cross a height threshold and have more access to light, they target their allocation strategy in securing high resource acquisition by increasing their crown footprint and switching to the resource acquisitive strategy of canopy species.It is likely that understory species, such as P. decurrens and G. interrupta, which reach maximum heights of 10 m, may follow a similar strategy.The ample morphological variation shown in the space defined by the first two principal components demonstrates these general trends and illustrates the diversity of allometric strategies within understory and canopy species.

Acquisitive strategy of canopy palms
Canopy palms invest in increasing size, as reflected in larger height and diameter, higher slenderness ratios, and larger crown area.Frond morphology and crown architecture change during the transition from the understory to the canopy (Rich et al. 1995).While in the understory, small I. deltoidea and S. exorrhiza develop less stratified crowns whose fronds present wedge-shaped leaflets or fins with a similar angle orientation.At about 7-8 m in height, these palms produce fronds with narrow, longitudinal leaflets in between the fins and generate a multilayer crown as they reach better-lit environments.According to Rich et al. (1995), I. deltoidea and S. exorrhiza palms > 11-17 m produce more heterogeneous fronds with a variety of morphologically diverse leaflets resulting in a multilayered crown.In addition to changing crown morphology and increasing frond lifespan, slenderness ratios also increase, which demonstrates a clear niche shift from shade tolerant to a more light-demanding strategy (Avalos 2023).The patterns of biomass allocation observed here are consistent with this acquisitive resource strategy as canopy palms transition from the understory to the canopy, showing a phenotype dominated by functional traits associated with achieving larger size consistently with trait growth theory (Westoby et al. 2022).

Complementary hypotheses to the leaf economics spectrum
Explaining the variation in niche partitioning requires the analysis of more functional traits, especially those that integrate plant-wide strategies in response to environmental gradients (Kraft et al. 2008).Key traits of the leaf economic spectrum, such as leaf area, SLA, maximum diameter, maximum height, leaf nitrogen content, and tissue density (Chave et al. 2006) have great potential to integrate changes through ontogeny in response to environmental gradients (Westoby et al. 2002;Wright et al. 2010), as well as to predict demographic and population processes such as density dependence (Umaña et al. 2018).There is a paucity of longitudinal studies examining functional traits in long-lived species (but see Wright et al. 2010;Boukili and Chazdon 2017), especially in linking functional traits to plant demography (Poorter et al. 2008;Worthy and Swenson 2019).The current and historical effects of temperature and precipitation seasonality should also be considered since climatic factors have a strong effect on the distribution of functional traits such as SLA and maximum height (Göldel et al. 2015).
The metabolic scaling theory (Castorena et al. 2022) is an integrative attempt to scale individual and species variation in functional traits to predict impacts in ecosystem functions.This theory has focused on body size as a driver of ecological, ecosystem, and evolutionary processes.Implicit in this assumption is the importance of ontogenetic niche shifts, which are mediated by changes in functional traits associated with ontogenetic size or stage (Westoby et al. 2022).These shifts are interpreted from an optimization approach, which is consistent with the economic spectrum of the leaf and with the theories of optimization of the useful life of the leaf (Kikuzawa 1995).Such efforts still require the integration of interspecific and intraspecific variation in functional traits, demography, and community and ecosystem ecology (Enquist et al. 2007(Enquist et al. , 2017)).
There are significant obstacles to achieving a synthesis that would integrate the metabolic scaling theory, the leaf economics spectrum, and other explanations to scale the variation in functional traits to community and ecosystem ecology.Monitoring the physiological performance of species of large size and long-life spans in highly diverse forests, as well as increasing the knowledge of functional trait variation in poorly known life forms, such as epiphytes, ferns, and palms, is still difficult.Functional ecology has the potential to identify resource use strategies using traits of universal importance that separate species according to their resource use strategy (Visser et al. 2016).This research becomes increasingly urgent as fragmentation and changes in land use intensify the response of ecological systems to climate change.

Conclusions
Our classification of palm species into different forest strata reflected different strategies of segregation in resource use strategy among understory and canopy species.Tissue density and DMF showed that understory species followed a "conservative" resource use strategy, while canopy palm species were associated to traits that reflected palm size and that conformed to the "acquisitive" strategy of resource use within the leaf economics spectrum (Westoby et al. 2002;Wright et al. 2004).These findings show general trends in resource partitioning to maximize survival in the shade or invest in height growth to reach the canopy.The functional overlap between the shortest individuals of canopy species and the tallest individuals of understory species reflects the continuity of the light gradient of the forest profile and the plasticity to adapt to it.Our results demonstrate the need to include more palm species and to carry out longitudinal studies that would consider the species´ ontogeny and population dynamics to understand the role of functional traits influencing plant responses to environmental gradients.

Fig. 1
Fig. 1 Palm species included in this study.Canopy species: A Crown of Socratea exorrhiza, B Juvenile stage of S. exorrhiza of 5 m in height, C Stilt root cone of S. exorrhiza showing separated roots covered by small thorns, D Crown of Iriartea deltoidea, E A 6 m long frond of a 25 m tall I. deltoidea palm, F Stilt root cone of I. deltoidea showing clustered, blackish roots, G Habit of Euterpe precatoria, H Top of crown of E. precatoria I, Stilt root cone of E. precatoria showing clustered, orangish roots.Understory species: J Habit of Prestoea decurrens, K detail of leaflet insertion in P. decurrens, L Base of the stem of P. decurrens showing adventitious roots, M Habit of Chamaedorea tepejilote, N Immature fruits of C. tepejilote, O Adventitious roots in C. tepejilote, P Habit of Geonoma interrupta, Q Habit of Asterogyne martiana showing flowers and immature inflorescences, R Reproductive individual of A. martiana, S Seedling of A. martiana, the coin has a diameter of 3 cm

Fig. 2
Fig. 2 Variation in nine morphological traits in seven species of neotropical palms of different forest strata.Palm species are referred to by stratum (U = understory, C = canopy) followed by the initial of the genus and species name (PD = Prestoea decurrens, GI = Geon-

Fig. 3 A
Fig. 3 A Correlation matrix of nine morphological traits in seven species of neotropical palms of different forest strata.Values indicate the magnitude of the Pearson Correlation Coefficient of the Ln-transformed values of morphological traits, and asterisks indicate the level of significance (p values = 0.0001, 0.001, 0.01, correspond to ***, **,

Fig. 4
Fig. 4 Spatial distribution of canopy and understory palms in the space defined by the first two principal components of nine morphological traits.Trait abbreviations follow Table 1

Fig. 5
Fig. 5 Segregation of canopy and understory species in terms of the scores of the first and second principal components summarizing 57.22% of the variation in functional traits for stratum, and species within a stratum for PC1 (panels A and B) and for PC2 (panels C and D).Palm species are referred to by stratum (U = understory,

Table 1
Summary