DOI: 10.20350/digitalcsic/15325
title: Raw data on the effect of resource availability and herbivory on the defence-growth-reproduction trade-off of a masting Mediterranean pine

description: [General methods] The full collection of six csv files contains all the raw data used in the paper “Resource availability and herbivory alter defence-growth-reproduction trade-offs in a masting Mediterranean pine”, which is now in its pre-print version. In this article we report the results of two parallel experiments we carried out to independently asses the effect of nutrient availability and different defence induction treatments on the three-way trade-off. Thirty six ramets were allocated to the fertilization experiment, with 12 ramets of each of three genotypes being randomly assigned to one of three blocks and one of two treatments (fertilization and control). Within each block, ramets were spatially grouped in two treatment plots. Each treatment by genotype combination comprises six ramets, two per block. The remaining 45 ramets were included in the defence induction experiment and randomly assigned to one of three blocks and one of five induction treatments. Within each block, ramets were spatially grouped in five treatment plots. Each treatment by genotype combination had three ramets, one per block. In the fertilization experiment we had only two treatments (fertilized vs control). Fertilization was accomplished by applying Fertimon Rojo (Soaga S.L. 2-10-16 NPK + 2.8% Mg) and a liquid 100-20-70 NPK fertilizer applied to the substrate at the base of the trunk of each tree (see Sampedro et al. 2011 for details on this fertilizer solution). The foliar fertilization continued until the end of the experiment. In january 2010, 125 additional grams of Osmocote (KB®, Scotts, NPK 17-09-11 + 2% MgO) were also added to each tree. Fertilization was maintained throughout the whole monitoring period (2010-2015). Five treatments were assigned to the herbivory simulation experiment (applied during the first two years, 2010-2011): • Needle clipping (repeated cutting of needles tips, around 1-2 cm). • Methyl jasmonate application (brushing a 25 mM solution on debarked areas of the trunk; see details in Vázquez-González et al. 2021). • Low intensity mechanical wounding of the trunk involved making three 5 mm circular holes in the bark and phloem. • High intensity mechanical wounding of the trunk, throug six 5 mm circular hole in the bark and phloem Ramets were monitored from 2010 to 2015. Below, we provide specific details on the methods associated to its data-set and the variables it includes.
description: [growth.csv] Ramet growth was monitored for six years, beginning in 2010. Growth monitoring involved measuring maximum tree height and stem diameter at one meter height and two orthogonal directions. Tree height was measured to the nearest dm using a telescopic measuring rod, while tree callipers were used to measure stem diameter to the nearest cm.
description: [nutrients.csv] Nutrient content was only assessed for the fertilization experiment and for the needles produced in 2010. Needle samples were taken in 14 October 2010. Nitrogen concentration in needles was determined in a LECO elemental analyzer (model CHN 628) while phosphorus was analyzed by inductively coupled plasma optical emission spectroscopy (ICP-OES) using a Perkin-Elmer Optima 4300DV (Massachusetts, USA) at the central laboratory facilities of Universidade de Vigo – CACTI (www.uvigo.es/webs/cactiweb/). Nitrogen and P concentration were expressed in mg g-1 tissue on a dry weight basis.
description: [phenols.csv] See “resin.csv” below, for details on the sampling protocol for defensive compounds estimation. Polyphenolics were extracted from a subsample of 30 mg of needles or phloem, by immersion in a metanol-water solution (1:1 volume) in an ultrasonic bath of 15 min, followed by centrifugation and extract dilution. Colorimetric measurements were taken with a Biorad 650 microplate reader (Bio-Rad Laboratories, Philadelphia, PA,USA) a 740 nm.
description: [pissodes.csv] During the summer of 2011 both experiments suffered an outbreak of Pissodes castaneus. We assessed the abundance of the damage caused by this curculionid in each ramet on 16 September 2011, by visually inspecting the scars produced in the main stem and assigning them to a four category ordinal scale (none, rare, abundant and very abundant).
description: [reproduction.csv] Monitoring of ramet reproduction comprised measures of reproductive effort (number of female strobili and abundance of male strobili) from 2010 to 2013, and measures of reproductive output (number of ripe cones, total number of seeds produced and number and dry weight of healthy seeds) from 2012 to 2015. Strobili cohorts blooming in 2010 to 2013 were hence monitored to ripeness (in 2012 to 2015). All female flowering strobili were visually counted and ripe cones collected, while male strobili shoots were counted and their length visually estimated to the nearest 5 mm to get an estimate of male strobili abundance. Pine cones where air-dried for several weeks and then oven-dried for seven days to induce opening (35ºC). Empty and healthy seeds were discriminated by decanting them on cold water, where empty seeds remain floating on the surface. After separating healthy and empty seeds for each cone, they were oven-dried again at 35ºC before weighing them to the nearest 1 mg with a precision scale.
description: [resin.csv] On 14 October 2010 we collected needle samples from both experiments, to assess defensive compound (non-volatile resin and total polyphenolics) and nutrient content (nitrogen and phosphorus concentration). Both needles produced in 2009 and 2010 were collected (and analysed) separately in each ramet. Nutrient content was only assessed for the fertilization experiment and for the needles produced in 2010. Defensive compounds were also determined in phloem samples collected from all ramets on 16 april 2011 but due to logistical reasons, resin content data for phloem samples in 2010 were lost. Needle and phloem sampling was hence repeated at the end of the experiment, in 2013, in order to estimate non-volatile resin concentration. Total non-volatile resin content in phloem and needle tissue was determined by gravimetric analysis. Needle and phloem fresh samples (aprox 2-5 g) were immediately covered with hexane within pre-weighed test tubes, and sonicated for 20 min at 20ºC. Resin extraction continued for 24 h at room temperature under the fume hood and then the content of the tube was filtered with GFF filters (Whatman GF⁄ D, Whatman Int. Ltd, Maidstone, Kent, UK) into a second pre-weighed tube. The whole extraction procedure was repeated with each sample and the liquid fraction from both extractions pooled before letting them dry under the fume hood. Completely dried tubes were weighed with a precision scale to the nearest 0.0001 g. The solid fraction was oven-dried at 65ºC until constant weight. Total non-volatile resin content was estimated as mg of dried extracted resin per g of dry sample. Needle and phloem samples were analysed to estimate phenol content by the Folin-Ciocalteu method (Scalbert 1992), a colorimetric protocol using the Folin-Ciocalteu reagent and tannic acid as standard. Fresh samples were immediately oven-dried at 45ºC and manually ground in a mortar with liquid nitrogen.
description: growth.csv, nutrients.csv, phenols.csv, pissode.csv, reproduction.csv, resin.csv