DOI: 10.5061/dryad.zw3r228bd
title: Non-structural carbohydrates predict survival in saplings of temperate trees under carbon stress

description: 1. Non-structural carbohydrates (NSCs) mediate plant survival when the
 plant’s carbon (C) balance is negative, suggesting that NSCs could predict
 plant survival under C stress. To examine this possibility, we exposed
 saplings of six temperate tree species to diverse levels of C stress
 created by the combination of two light conditions (full light
 availability and deep shade) and two defoliation levels (severe
 defoliation and non-defoliation). We then measured survival, biomass, and
 total NSCs and soluble sugar (SSs) concentrations in different organs of
 both dead and live saplings. 2. We estimated mean NSCs and SSs contents
 and concentrations per sapling and fitted logistic generalized
 mixed-effects models to determine if NSCs and SSs predict survival. Using
 inverse prediction modelling, we also determined whether there is a common
 NSCs and SS threshold across species at the time of sapling’s death. 3.
 Defoliation and shade reduced the mean sapling’s NSCs and SSs contents,
 indicating C stress. Mean sapling NSCs and SSs contents and concentrations
 predicted survival and the robustness of the models improved with the
 inclusion of species. At death, saplings of the exotic deciduous tree
 species Acer pseudoplatanus exhibited significantly lower mean NSCs and
 SSs contents than saplings of the evergreen conifer species Podocarpus
 nubigenus and lower stem NSCs and SSs concentrations than the broadleaf
 evergreen species Drimys winteri. 4. The energetic role that NSCs and SSs
 play in plants under C stress was evidenced by the capacity of these
 compounds to predict sapling survival under C stress. No common threshold
 of NSCs and SSs contents or concentrations for sapling survival amongst
 species was found, indicating that the level of these compounds may not be
 good proxies for interspecific comparisons of tolerance to C stress.
 Presumably, there are species-specific limits for the mobilization and use
 of NSCs and SSs in metabolism. 5. Our results anticipate that the
 inclusion of NSCs and SSs in modelling will improve predictions regarding
 tree responses to ongoing climate change. Nonetheless, a better
 understanding of the many roles that carbohydrates play in plant survival
 under C stress is required to scale predictions up to the community level.
description: We conducted an outdoor pot
 experiment over two growing seasons at the El Mallín nursery (Corporación
 Nacional Forestal, Conaf), located in Puerto Aysén, southern Chile (45°
 59′′ S, 71° 52′′ W, 30 m above sea level [asl]). We studied
 two winter deciduous species (Acer pseudoplatanus and Nothofagus pumilio)
 and four evergreen species (N. betuloides, N. nitida, Podocarpus nubigenus
 and Drimys winteri) of contrasting successional status and light
 requirements. In October 2013 (early spring), two-year-old individuals of
 each species were transplanted to 2 L pots, except A. pseudoplatanus
 whichas was transplanted into 4 L pots. All species’ saplings were
 randomly assigned to four blocks with two light treatments (severe shade
 and open conditions). The open light condition was represented by full
 light availability, whereas the severe shade condition was achieved by
 placing the pots on the ground beneath the sparse shade of Alnus
 glutinosa trees, and by completely covering the lateral sides and the top
 with frames of a black Raschel mesh with 80% shading. In order to provoke
 further C limitation in saplings, by late spring (December 2014), a
 complete manual defoliation was applied to half of the saplings of each
 light condition. By late March 2015 (early autumn), all saplings were
 assessed for survival and subsequently harvested. Saplings were considered
 dead when they had shed all their leaves or when their leaves were all
 brown, in addition, the stem was dead (i.e. dry and brown). A total of 130
 saplings, including 75 live and 55 dead, were harvested for biomass and
 non-structural carbohydrate (NSC) and soluble sugar (SS)
 determination. Organ-specific NSC concentrations were scaled up to the
 whole sapling using the weighted mean concentrations across all sapling
 organs, which consider the organ specific concentration (% dry matter) of
 NSCs and the organ-specific biomass. Additionally, organ-specific NSCs and
 SS contents (i.e. pool size, or mass) were calculated per sapling as the
 product of NSCs or SS concentration and the biomass of each organ.
 Organ-specific contents were then added to obtain whole-plant NSCs and SS
 contents. Finally, the NSCs and SS fraction represented by each organ was
 estimated for each sapling as the ratio between the organ-specific content
 and the whole-plant content.  All statistical analyses were conducted in R
 version 4.2.0 (R Development Core Team, 2022). to determine
 if the light and defoliation treatments provoked C stress across species,
 we fitted linear mixed-effects models (LMMs) using the
 R/NLME package. we evaluated treatment
 and species effects on the NSCs and SS contents and concentrations of each
 organ and of the whole-sapling, as well as for NSCs and SS
 fractions. The fixed
 effects of the model were light condition, defoliation level, species, and
 all of their interactions. The random effect included the blocks. Similar
 models were used to compare biomass. Sapling survival was further analyzed
 using generalized linear
 mixed-effects models (GLMMs), with a binomial family and a logit link
 function because the sapling was considered either live or dead, with
 random effects for blocks and fixed effects for light condition,
 defoliation level, species, and the interaction amongst
 them. Similar to the previous analysis, we tested the
 significance of fixed effects using likelihood-ratio tests
 (LRTs).  NSCs and SS contents and concentrations were
 compared between live and dead saplings using LMMs, where the block was
 the random factor and the survival, the species, and the interaction
 between them were the fixed factors.To determine whether the levels of C reserves
 predict sapling survival across species, we fitted mixed-effects binary
 logistic regression models, i.e. logistic GLMM using the R/LME4 package
 v1.1-27 (Bates et
 al. 2015),
 considering the NSCs or
 SS contents or concentrations for each organ or for
 the whole-sapling, with species as fixed
 factors, the block as a
 random factor, and the survival response as the binary response
 variable (Barker
 Plotkin et al. 2021). To
 determine if the means were significantly different from one another, we
 performed post-hoc comparisons with a Tukey's honest significance
 test, and a α of 0.05. For all models, we assessed model fit by examining
 standardized residuals using the R/DHARMA package (Hartig 2020). Finally, to determine if the
 NSCs and SS
 levels predicting survival
 under C stress were similar across species, we estimated the
 NSC contents and
 concentrations and their
 respective confidence intervals for 1%, 25%, 50%, 75% and 100% survival by
 inverse prediction, using the R/GGEFFECTS package (Lüdecke 2018). For NSCs, the model structure was as
 follows: Survival ~
 1+species+NSCs+(1|block) where Survival is the response variable
 expressed as either 1, live sapling, or 0, dead sapling; species and NSCs
 (or SSs) correspond to the fixed factors, whereas block is the random
 effect. These analyses
 were performed at both the organ- and whole-sapling
 level.