DOI: 10.5061/dryad.2280gb5tp
title: Leaf CNPK concentrations and isotopic signatures

description: Previous meta-analyses suggested that carnivorous plants – despite access
 to N, P, and K from prey – have significantly lower leaf concentrations of
 these nutrients than non-carnivores. Those studies, however, largely
 compared carnivores in nutrient-poor habitats with non-carnivores in more
 nutrient-rich sites, so that the differences reported might reflect
 habitat differences as much as differences in nutrient-capture strategy.
 Here we examine three carnivorous and 12 non-carnivorous plants in the
 same nutrient-poor bog to compare their foliar nutrient concentrations,
 assess their patterns of nutrient limitation using leaf NPK stoichiometry,
 and estimate %N derived from prey by carnivores using a mixing model for
 stable N isotopes. We hypothesized that (1) carnivore leaf nutrient
 concentrations approach or exceed those of non-carnivores in the same
 nutrient-poor habitat; (2) species in different functional groups show
 different patterns of stoichiometry and apparent nutrient limitation; and
 (3) non-carnivores might show evidence of employing other means of
 nutrient acquisition or conserv­ation to reduce nutrient limitation. At
 Fallison Bog in northern Wisconsin, carnivorous plants (Drosera
 rotundifolia, Sarracenia purpurea, Utricularia macrorhiza) showed
 significantly lower leaf % C and N:P ratio, higher δ15N, and no difference
 from non-carnivores in leaf N, P, K, and δ13C. Sedges had significantly
 lower leaf % P, % C, and N:K ratio, and higher K:P ratio than non-sedges
 restricted to the Sphagnum mat, and may tap peat N via
 aerenchyma-facilitated peat oxidation (oxipeditrophy). Evergreen
 erica­ceous shrubs exhibited significantly higher levels of % C and lower
 values of d15N than mat non-ericads. Calla palustris – growing in the
 nutrient-rich moat at the bog’s upland edge – had very high values of leaf
 N, K, δ15N, and N:P ratio, suggesting that it may obtain nutrients from
 minero­trophic flows from the adjacent uplands and/or rapidly decay­ing
 peat. Stoichiometric analyses indicated that most species are N-limited. A
 mixing model applied to δ15N values for carnivores, non-carnivores, and
 insects produced an estimate of 50% of leaf N derived from prey for
 Utricularia, 42% for Sarracenia, and 41% for Drosera.
description: Leaf samples of the above species were gathered <em>in
 situ</em> – <em>Utricularia</em> and
 <em>Nuphar</em> in the bog pond, <em>Calla</em>
 and <em>Carex pseudocyperus</em> in the moat, and all other
 species in the grounded and floating mats. We collected samples from 2-6
 scattered individuals per species (mean = 3.3 ± 1.1 SD) from three
 carnivorous and twelve non-carnivorous species (see supplementary Table
 S1), including almost all the common herbaceous and shrub species present.
 Leaf samples were transported on ice from the field to a drying oven, and
 then dried at 70°C for 72 hrs. Insect samples were captured via sweep and
 aerial netting and then oven dried. Large Odonata and other insect-eating
 forms were excluded to avoid complications due to the higher
 δ<sup>15</sup>N values expected in such predators; in most
 instances, these are massive and unlikely to be captured by carnivorous
 plants. Dried samples were ground in a Wiley mill before being sent for
 quantifi­ca­tion of δ<sup>13</sup>C,
 δ<sup>15</sup>N, and N, P, and K concentrations to the Nippert
 lab at Kansas State University. At KSU, dried leaf
 samples were further ground into a fine homogenous powder, and 2 mg
 aliquots were placed into tin capsules. These capsules were loaded into an
 Elementar Pyrocube Elemental Analyzer (EA), in which the samples were
 combusted at 950°C. The resulting gas was analyzed for %C and %N, and then
 injected into an Elementar Geovision Isotope Ratio Mass Spectrometer
 (IRMS) to measure δ<sup>15</sup>N and
 δ<sup>13</sup>C in the gas. Isotopic data were calibrated with
 lab standards to convert the raw data to δ<sup>15</sup>N (air)
 and δ<sup>13</sup>C (VPBD). The lab standards are calibrated
 annually with IAEA standards. The run precision was 0.04 per mil
 δ<sup>15</sup>N and δ<sup>13</sup>C.
 To measure P and K concentrations (% dry mass), 0.25 g of
 prepared tissue was weighed into a 50 ml Kimax digestion tube. In a fume
 hood, 2 ml of concentrated sulfuric acid was added, then 1 ml 30% hydrogen
 peroxide. After oxidation subsided, the tube was placed on a digestion
 block at 285°C for 20 min. After cooling to room temperature, another 1 ml
 of hydrogen peroxide was added before returning the tube to the digestion
 block. This was repeated until the acid in the tube was colorless. After
 cooling, the sample was diluted to 50 ml with ddH<sub>2</sub>0
 and mixed by inverting the tube twice. P and K concentrations were then
 measured using inductively coupled plasma spectrometry.