Construction Costs of Carnivorous and Non-Carnivorous Plants at Harvard Forest 2006-2008

doi:10.6073/pasta/533b5dda2244faf3d27eea6d02d781c5 Construction Costs of Carnivorous and Non-Carnivorous Plants at Harvard Forest 2006-2008 Jim Demetrios Karagatzides Aaron Ellison https://orcid.org/0000-0003-4151-6081 2023 English

Leaf traits, including photosynthetic rates, leaf mass area, and leaf nutrient content covary in a coordinated way for a wide range of plant taxa. This covariation results from trade-offs between the costs of constructing plant tissues and the benefits accrued from photosynthesis. Carnivorous plants have been found to be outliers in the "universal spectrum of leaf traits" because they have very low photosynthetic rates for the amount of nitrogen in their leaves and traps. But no studies have measured simultaneously the actual construction costs of carnivorous traps and rates of photosynthesis to determine the amortization required to recover the investment (the "payback time") and thereby calculate the "marginal gain" of "investing" in carnivorous structures. The objective of this study was to measure construction costs (CCmass, grams of glucose required to build 1g of ash-free dry mass of tissue) and photosynthesis (Amass, nmol CO2 g-1 s-1) for traps, leaves, roots, and rhizomes of 15 carnivorous plant species with differing mechanisms of prey capture and consumption (pitfall traps, snap-traps, sticky pads) grown under greenhouse conditions. Payback time (h) was calculated as the quotient of CCmass and Amass after conversion to nmol of carbon per gram of ash-free dry mass. There were highly significant differences amongst species for CCmass of traps but there were no significant differences for CCmass amongst traps, roots and rhizomes. Mean (+- SD) CCmass for traps (1.14 +- 0.24 g glucose g-1) was significantly lower than the mean CCmass of leaves of 267 non-carnivorous plant species (1.47 +- 0.17 g glucose g-1). However, all 15 carnivorous plants examined in this study had low Amass and thus, the marginal gain of carnivory is small with a long payback time (524-1641 h). Our results of low CCmass for carnivorous traps is contrary to the oft-stated expectation of a high cost to construct elaborate carnivorous traps. Payback time integrates traits used to assess leaf scaling relationships with construction costs, and locates carnivorous plants at the "slow and tough" end of the universal spectrum of leaf traits.

carnivorous plants leaves photosynthesis plant physiology LTER controlled vocabulary populations inorganic nutrients LTER core area Harvard Forest HFR LTER USA HFR default

This dataset is released to the public under Creative Commons CC0 1.0 (No Rights Reserved). Please keep the dataset creators informed of any plans to use the dataset. Consultation with the original investigators is strongly encouraged. Publications and data products that make use of the dataset should include proper acknowledgement.

Creative Commons Zero v1.0 Universal https://spdx.org/licenses/CC0-1.0.html CC0-1.0 https://harvardforest.fas.harvard.edu/exist/apps/datasets/showData.html?id=hf112 Harvard Forest Greenhouse. Coordinates based on WGS84 datum. -72.19 -72.19 +42.53 +42.53 330 330 meter 2006 2008 genus Dionaea species sp. genus Drosera species sp. genus Nepenthes species sp. genus Sarracenia species sp. complete Information Manager Harvard Forest

324 North Main Street Petersham MA 01366 USA

(978) 724-3302 hf-im@lists.fas.harvard.edu Harvard Forest

324 North Main Street Petersham MA 01366 USA

(978) 724-3302 (978) 724-3595 https://harvardforest.fas.harvard.edu

Overview We examined construction costs of 15 carnivorous plant species in three plant families and two orders (Ericales: Sarraceniaceae; Caryophyllales: Nepenthaceae and Droseraceae) grown in a climate-controlled glasshouse at Harvard Forest. These included eleven species of North American pitcher plants (Sarracenia alabamensis Case and Case, S. alata (Wood) Wood, S. flava L., S. jonesii Wherry, S. leucophylla Raf., S. minor Walt., S. oreophila (Kearney) Wherry, S. purpurea L., S. rosea Naczi, Case and Case, S. rubra Walt., and Darlingtonia californica Torr. (Sarraceniaceae)); two Asian pitcher plants (Nepenthes x coccinea (a Victorian-era hybrid of [N. rafflesiana Jack x N. ampullaria Jack] x N. mirabilis (Lour.) Druce) and N. x miranda (a modern hybrid of N. maxima Reinw. ex Nees x [N. northiana Hook f. x N. maxima]) x (Nepenthaceae)); the sundew Drosera filiformis Raf. (Droseraceae), and the Venus fly-trap Dionaea muscipula Ellis (Droseraceae). Sarracenia, Darlingtonia, Drosera filiformis, and Dionaea are native to North America, whilst the Nepenthes species are hybrids of species native to southeast Asian tropical lowlands. The modified leaves of Dionaea, Drosera and North American Sarracenia and Darlingtonia both photosynthesize and trap prey. Individuals of Nepenthes, by contrast, have a flat lamina (perhaps a modified petiole) and an attached cylindrical trap that is modified from a leaf or leaflet (Arber 1941); thus, we measured construction costs of both laminae and pitchers of Nepenthes. All plants used in our study had reached reproductive maturity; excluding juvenile plants minimized the potential confounding effects of non-functional traps that are too small to capture prey (common in juvenile plants) and heterophylly relative to adult plants (Franck 1976).

Photosynthesis Among the pitcher plants, we measured plants with at least three mature, fully expanded pitchers. There were six replicate plants for each species with the exception of N. x coccinea (N = 4) and Darlingtonia californica, N. x miranda, and S. rosea (N = 2 each). The Venus' fly-trap Dionaea muscipula and the sundew Drosera filiformis were flowering at the time of measurement, but the other species were not. Maximum photosynthetic rate (Aarea, umoles CO2 m-2 s-1) of one trap (and lamina for Nepenthes) on each plant was determined using a LI-COR 6400 infrared gas analysis system (LI-COR, Lincoln, NB, USA) fitted with a 3-cm x 2-cm cuvette. We also measured phyllodia (flat, non-trapping leaves) that were produced by three of the species (S. flava, S. leucophylla, S. oreophila) during our study. All measurements were taken between 0900 and 1400 hours at a photosynthetic photon flux density (PPFD) of 1200 umol m-2 s-1 during 23-25 July 2006. Photosynthetic rate for Dionaea was the mass-based average of the trap and its petiole. Several Drosera leaves were placed in the cuvette to maximize surface area of the cuvette occupied. In this and in those few instances when the sample did not cover the entire surface area of the cuvette (e.g., Dionaea, N. x coccinea), photosynthetic rates were adjusted for the proportion of the cuvette covered by leaf tissue. In those cases where only two plants were available, 2-4 pitchers were sampled from a plant and averaged for that individual.

Harvest Plants were harvested immediately after photosynthetic rates were measured. Pitchers were cut longitudinally with a stainless steel razor and washed with tap water to remove any prey, detritus and extra-floral nectar. Pitchers were subsequently rinsed with distilled-deionized water, patted dry with paper towel and spread on the conveyer belt of a Li-Cor 3000 to measure leaf area (+- 1 mm2). Roots and rhizomes were washed separately with tap water and rinsed with distilled-deionized water. Plant tissue was dried at 70 deg C to constant mass, weighed (+- 0.001 g) and ground to a fine powder with a stainless steel capsule and ball bearing in a Wig-L-Bug grinder (Bratt Technologies, LCC, East Orange, NJ). For aboveground tissue, the entire pitcher or phyllode used for Aarea measurements was ground to a fine powder; any other aboveground tissue was oven-dried and weighed to obtain total dry mass of each structure. Root and rhizome masses were measured separately, and each was ground to a fine powder. Leaf areas and associated masses were used to calculate LMA (g m-2) and to re-express Aarea as Amass (nmoles CO2 g-1 s-1).

Estimation of construction cost Tissue construction costs (CCmass; g glucose g-1 ash-free dry mass [AFDM]) were estimated for roots, rhizomes, and the leaf tissue on which Aarea had been measured using the heat-of-combustion method (Williams et al. 1987): CC = [(0.06968 d Hc - 0.065) x (1 - Ash) + (kN)] x (1/ Eg) where d Hc is the heat of combustion (energy as kJ g-1 AFDM); Ash is the ash content (g ash g-1 dry mass); k is the oxidation state of the nitrogen substrate (nitrate = +5, ammonium = -3); N is the organic nitrogen content (g N g-1 dry mass); and Eg is the growth efficiency (the proportion of energy used to produce reductant that is consumed during the formation of tissue but not contained within the biomass). An Eg = 0.87 incorporates cost of transport and gives a good fit against the detailed biochemical analysis used as the standard (Griffin 1994). Heat of combustion was determined using a micro-bomb calorimeter (construction details available online at: http://harvardforest.fas.harvard.edu/personnel/web/aellison/research/stoichiometry/calorimetry/Micro-bomb%20Home%20Page.htm) calibrated with benzoic acid pellets of known calorific values. The calibration line was verified against a spinach reference standard (NIST 1570a) with a non-certified calorific value of 3500 calories g-1. Analysis of N = 35 spinach pellets during our assay yielded an average calorific value of 3536 cal g-1 (i.e., +1% of the expected value). In almost all cases, sample tissue was sufficient to allow each sample to be analyzed in triplicate as 2-12 mg pellets pressed from the ground sample. The Hc values obtained for the triplicate pellets of each sample were then averaged. Due to the large number of analyses (greater than 1000) we used Ni-Cr ignition wire (which contributes a small amount of heat during the reaction) rather than the more expensive platinum wire that does not give off heat from combustion. Therefore, 5 samples of Ni-Cr wire and no sample pellet were combusted to obtain the heat given off by the Ni-Cr wire and to determine the intercept of the calibration line. Total nitrogen was substituted for organic N (Nagel et al. 2005) and measured on a Carlo-Erba Model 2500CN elemental analyzer. Nagel et al. (2005) found that the substitution of total N for organic N overestimated CC by only 0.03-0.06%. Ash content (g g-1) was determined by combusting a 10-100 mg sub-sample of the powdered plant tissue in a muffle furnace at 550 deg C for 6 h. Construction costs were calculated using both k = +5 and -3 and the average value reported on an ash-free dry mass basis. Payback time represents the time to recover the carbon investment in an entire trap and was estimated as the quotient of Amass / CCmass after conversion of Amass from nmol CO2 g-1 dry mass s-1 to nmol C g-1 AFDM h-1 and conversion of CCmass from g glucose g-1 AFDM to nmol C g-1 AFDM. Calculations for payback times of pitcher construction for Nepenthes were also made using Amass of the attached lamina. We compared CCmass of traps against leaves of 267 non-carnivorous species compiled from a search of the published literature (Griffin 1994; Isagi 1994; Mitchell et al. 1995; Baruch and Gomez 1996; Dai and Wiegert 1996; Isagi et al. 1997; Marquis et al. 1997; Niinemets 1997; Spencer et al. 1997; Wullschleger et al. 1997; Baruch and Goldstein 1999; Eamus et al. 1999; Baruch et al. 2000; Nagel and Griffin 2001; Villar and Merino 2001; George et al. 2003; Suarez 2003; Nagel et al. 2004; Oikawa et al. 2004; Osunkoya et al. 2004; Barthod and Epron 2005; Nagel et al. 2005; Suarez 2005; Brunt et al. 2006; Oikawa et al. 2006; Osunkoya et al. 2007).

Harvard Forest Long-Term Ecological Research Harvard Forest

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(978) 724-3302 (978) 724-3595 https://harvardforest.fas.harvard.edu https://ror.org/059cpzx98 pointOfContact The Harvard Forest Long-Term Ecological Research (LTER) program examines ecological dynamics in the New England region resulting from natural disturbances, environmental change, and human impacts. National Science Foundation LTER grants: DEB-8811764, DEB-9411975, DEB-0080592, DEB-0620443, DEB-1237491, DEB-1832210. hf112-01-measure.csv measurements hf112-01-measure.csv 33712 f5825141c86d61db1b3aa8280fc8d780 1 \r\n column , https://harvardforest.fas.harvard.edu/data/p11/hf112/hf112-01-measure.csv genus all genera and species self-explanatory, but note: Nepenthes x coccinea is a Victorian-era hybrid of [N. rafflesiana Jack x N. ampullaria Jack] x N. mirabilis (Lour.) Druce). N. x miranda is a modern hybrid of N. maxima Reinw. ex Nees x [N. northiana Hook f. × N. maxima]. genus species all genera and species self-explanatory, but note: Nepenthes x coccinea is a Victorian-era hybrid of [N. rafflesiana Jack x N. ampullaria Jack] x N. mirabilis (Lour.) Druce). N. x miranda is a modern hybrid of N. maxima Reinw. ex Nees x [N. northiana Hook f. × N. maxima]. species struct plant structure Trap pitchers (includes tendril, if present), sticky pad, snap-traps (trap and its petiole) RE reproductive effort and includes the flower head and its stalk Phyll phyllodia which are flat leaves not modified into pitchers LfwP lamina with conjoined carnivorous trap LfwoP lamina without a conjoined carnivorous trap Roots roots Rhizome rhizome envt environment where plants were harvested GH greenhouse TS Tom Swamp JB James Bay (Fort Albany, Ontario, Canada; LAT. N52 11 43.7, LONG. W81 47 47.0 20.4 [deg. min. sec.]) year year of sampling YYYY re reproductive status at time of harvest 0 no flower 1 flower present replic replicate number. Sum indicates that multiple plants were pooled into one sample (usually because of small sample mass of individuals). Sum# indicates multiple parts from the same plant pooled (where # indicates the replicate) replicate number cc.mass the average tissue construction cost (grams glucose per gram ash-free dry mass [AFDM]) using the heat of combustion method and based on plant uptake of nitrate and ammonium in equal proportions. Williams, K., Percival, F., Merino, J., & Mooney, H.A. (1987) Estimation of tissue construction cost from heat of combustion and organic nitrogen content. Plant, Cell and Environment, 10, 725-734. gramsPerGram .01 real NA missing value cc.nitr tissue construction cost (grams glucose per gram ash-free dry mass [AFDM]) using the heat of combustion method and plant uptake of nitrate gramsPerGram .01 real NA missing value cc.amm tissue construction cost (grams glucose per gram ash-free dry mass [AFDM]) using the heat of combustion method and plant uptake of ammonium gramsPerGram .01 real NA missing value amass mass-based photosynthetic rate (nmol CO2 g-1 s-1) measured at photosynthetic photon flux density = 1200 umol m-2 s-1 nanomolesPerGramPerSecond 1 real NA missing value amass.in denotes data used in analyses involving Amass. Excluded values had suspect (low) Amass. Only traps and Nepenthes laminae were included in the analysis. 1 included in analysis 0 excluded from analysis NA NA a.area leaf-area based photosynthetic rate (µmol CO2 m-2 s-1) at photosynthetic photon flux density = 1200 umol m-2 s-1 micromolePerMeterSquaredPerSecond .01 real NA missing value lma leaf mass area. The quotient of leaf mass and leaf area (g m-2). Precision = 0.1. gramsPerSquareMeter 0.1 real NA missing value payback payback time (hours) represents the time to recover the carbon investment in a structure. It was estimated as the quotient of CCmass / Amass after conversion of CCmass from g glucose g-1 AFDM to nmol C g-1 AFDM and of Amass from nmol CO2 g-1 dry mass s-1 to nmol C g-1 AFDM h-1. Note that calculations are included elsewhere in the spreadsheet for payback times of pitcher construction for Nepenthes were also made using Amass of the attached lamina.. hour 10 whole NA missing value pnue.nitr photosynthetic nutrient-use efficiency for nitrogen (nmol CO2 mol-1 N s-1) calculated as total photosynthesis for a leaf divided by total nitrogen content in the leaf. nanomolePerMolePerSecond .01 real NA missing value lf.area leaf area squareCentimeters .01 real NA missing value dry.fresh ratio of dry mass to fresh mass number .001 real NA missing value ash tissue ash content as g ash per g dry mass gramsPerGram .001 real NA missing value hc heat of combustion (energy content) as kJ g-1 ash-free dry mass kilojoulePerGram .01 real NA missing value n nitrogen concentration (%) of oven-dried (70°C for 72 hours) tissue dimensionless .01 real NA missing value c carbon concentration (%) of oven-dried (70°C for 72 hours) tissue dimensionless .01 real NA missing value dm dry mass of structure analyzed (e.g., dry mass of one pitcher harvested from a plant) gram .001 real NA missing value tot.dm the total dry mass of that structure for an individual plant (e.g., dry mass of all pitchers on a plant) gram 0.001 real NA missing value lab.no unique lab number assigned to a sample. Field samples from 2004 preceded with “F”. unique lab number 297 hf112-02-review.csv literature review hf112-02-review.csv 19350 964551f497f2e558db400e18974dab98 1 \r\n column , https://harvardforest.fas.harvard.edu/data/p11/hf112/hf112-02-review.csv genus genus genus species species species type plant type CP carnivorous plant NCP non-carnivorous plant traps construction cost of carnivorous plant traps (including tendrils when present). Snaptraps (e.g., Dionaea muscipula) includes trap and its petiolein grams glucose per gram tissue. gramsPerGram .01 real NA missing value leaves construction costs of non-carnivorous plant leaves, or carnivorous plant phyllodia, in grams glucose per gram tissue gramsPerGram .01 real NA missing value roots construction cost of roots in grams glucose per gram tissue gramsPerGram .01 real NA missing value rhizomes construction costs of rhizomes in grams glucose per gram tissue gramsPerGram .01 real NA missing value reference paper from which data were derived. We followed Griffin (1994) and compiled construction cost data only from studies using the Williams et al. (1987) heat of combustion method. paper from which data were derived 291