DOI: 10.5061/dryad.547d7wm7h
title: Data from: Invasion-induced root-fungal disruptions alter plant water and nitrogen economies

description: Despite widespread evidence that biological invasion influences both the
 biotic and abiotic soil environments, the extent to which these two
 pathways underpin the effects of invasion on plant traits and performance
 is unknown. Leveraging a long-term (14-yr) field experiment, we show that
 an allelochemical-producing invader affects plants through biotic
 mechanisms, altering the soil fungal community composition, with no
 apparent shifts in soil nutrient availability. Changes in
 belowground fungal communities resulted in high costs of nutrient uptake
 for native perennials and a shift in plant traits linked to their water
 and nutrient use efficiencies. Some plants in the invaded community
 compensate for the disruption of nutritional symbionts and reduced
 nutrient provisioning by sanctioning more nitrogen to
 photosynthesis and expending more water, which demonstrates a
 trade-off in trait investment. For the first time, we show that
 the disruption of belowground nutritional symbionts can drive plants
 toward alternative regions of their trait space in order to maintain water
 and nutrient economics.
description: Our study site is in Fox Chapel, PA (40.520237, -79.900932) in
 the Trillium Trail Nature Reserve, on steep slopes (25- 75%) with
 Gilpin-Upshur-Atkins soils of shale, sandstone, and red clay shale bedrock
 (USDA 2015). The mean annual precipitation is 36-46 inches and the mean
 annual air temperature of 41° to 62° F (USDA 2015). The experimental
 design includes five 14 x 14 m plots that were surrounded by 2.5 m tall
 wire fencing (erected in 2002) to exclude deer. Beginning in 2006, we
 weeded <i>Alliaria </i>by hand<i> </i>from the
 left side of each plot and carried the weeded material offsite. The right
 side of each plot was left at ambient <i>Alliaria</i> field
 densities, which averaged 15.2% cover (Roche<i> et al.</i>
 2020). This experiment is a split plot design with two treatments per
 plot, referred to here after as the weeded and ambient treatments. To
 minimize allelochemical leaching from the ambient treatment to the weeded
 treatment within each plot, the <i>Alliaria</i> treatments
 were positioned parallel to the slope. We also installed a 0.5 m buffer
 between the ambient and weeded treatments within each plot to prevent
 native plant roots in the weeded treatment from being exposed to
 <i>Alliaria</i> roots and allelochemicals. In the spring of
 subsequent years, seedlings that emerged from the seed bank were hand
 pulled from the weeded treatment. To prevent seed dispersal and
 re-infestation of <i>Alliaria</i>, we installed temporary
 aboveground mesh barriers between the weeded and ambient sides of the
 plots prior to <i>Alliaria</i> seed maturation that were
 removed post seed dispersal each year. Hand pulling and annual seedling
 removal was an effective method of invasion suppression, with a 0.08% mean
 abundance of <i>Alliaria </i>in the weeded plots
 (Roche<i> et al.</i> 2020).  
 <strong><i>##### Leaf-level morphological and
 physiological traits #####</i>   </strong>                    
                                                                          
                                                                          
                             To characterize the effect
 of the <i>Alliaria</i> treatments on morphological and
 physiological traits associated with water and nutrient use efficiency for
 the focal native understory species, we quantified the instantaneous
 physiology of additional reproductive and non-reproductive plants in 2018
 whose roots were not sampled. We measured the net photosynthetic rate
 (A<sub>sat</sub>), and stomatal conductance
 (g<sub>s</sub>) using a Li-COR LI-6400XTR portable
 photosynthesis system, equipped with a CO<sub>2</sub> control
 module, 2 x 3 cm leaf cuvette and a red-blue light-emitting diode (LED)
 light source (Li-Cor, Lincoln, NE, USA). Measurements were taken at a
 saturating light level of 800 μmol
 m<sup>-2</sup>s<sup>-1</sup>, ambient temperature
 and humidity, and a reference chamber CO<sub>2</sub>
 concentration of 400 μmol mol<sup>-1</sup>, following
 Heberling<i> et al.</i> (2017). A saturating light level of
 800 μmol m<sup>-2</sup>s<sup>-1 </sup>was based on
 detailed light response curves for these understory species at our field
 site (Heberling<i> et al.</i> 2017; Heberling<i> et
 al.</i> 2019a). To minimize within-plant variability for
 <i>Maianthemum</i>, we restricted our sampling to either the
 2<sup>nd</sup> or 3<sup>rd</sup> leaf from the
 terminal end of the stem following Cornelissen<i> et al.</i>
 (2003) and only used leaves without signs of senescence or damage (e.g.
 insect herbivory). Specific leaf area (SLA) and leaf carbon and nitrogen
 concentration were measured on a subset of individuals (11-18 individuals
 x 2 <i>Alliaria</i> treatments x 3 species x 2 life stages).
 To measure SLA, we collected two to three 5.32 cm<sup>2</sup>
 leaf tissue samples from each individual and calculated the mean leaf
 surface area per g dry mass (cm<sup>2
 </sup>g<sup>-1</sup>). Leaf carbon and nitrogen
 concentration were measured on dry leaf samples using an elemental
 analyzer (Costech Inc., Valencia, CA). We used these physiological
 measurements to calculate water use efficiency (WUE =
 A<sub>sat</sub> /g<sub>s</sub>; µmol
 CO<sub>2</sub> mmol
 H<sub>2</sub>O<sup>-1</sup>) and photosynthetic
 nitrogen use efficiency (PNUE = A<sub>sat</sub> /
 N<sub>area</sub>; μmol CO<sub>2</sub>
 gN<sup>-1</sup> s<sup>-1</sup>) (Field<i> et
 al.</i> 1983; Wright<i> et al.</i> 2003). A mean
 chlorophyll index of three leaflets per plant was measured using a SPAD
 502 Chlorophyll Meter (spectrum Technologies, Inc, Aurora, IL, USA). To
 standardize our sampling, we chose to take SPAD readings from the darkest
 green section of each leaflet. The Chl-sampled individuals are a subset of
 plants for which the other functional traits were measured and the same
 plants for which whole-plant performance were measured.
 The associated CSV file for the plant physology and
 performance results is named
 <i>plant_traits_performance_EcoLetters.</i>
 Before doing the physiology analyses, we subset the raw data to
 exclude Ci values &lt;60.  The
 <i>treatment</i> column in the CSV file refers to the
 two garlic mustard treatmetns (i.e., 'Ambient' and
 'Weeded') The <i>Sp</i> column in
 the '<i>plant_traits_performance_EcoLetters</i>' CSV
 refers to the three focal native species, Mai = Maianthmum, Tri =
 Trillium, and Ari = Arisaema. The units for each measurement is reported
 in the main manuscript.  Leaf area in the CSV is cm^2. To convert leaf
 area from cm^2 to m^2 (as it is reported in the manuscript), multiply leaf
 area x 10000.   <strong>#####
 fungal community composition #####  </strong>                       
                                                                          
                                                                          
                                                        
 The associated CSV file for the fungal community
 composition results is named
 <i>Soil_AM_Fungal_communites_EcoLetters</i>. Refer to the
 methods section of the published paper for details on how the soil samples
 were taken. The associated soil fungal DNA sequences have been deposited
 with links to BioProject accession number PRJNA691885 in the DDBJ
 BioProject database.  
 <strong>##### root conolozation rates ##### 
 </strong>                                                           
                                                                          
                                                                          
                                 The associated CSV file
 for the root colonization results is named
 <i>AM_fungal_root_colonization_EcoLetters</i>.
 Refer to the methods section of the published paper for details
 on how the root sampls were processed.  The
 <i>treatment </i>column in associated files refers to the
 two garlic mustard treatmetns (i.e., 'Ambient' and
 'Weeded') The<i> sp</i> column in
 the CSV refers to the three focal native species, Mai= Maianthmum, Tri
 =Trillium, and Ari = Arisaema. FoV column is short for
 field of view.  For beta regressions, percent
 colonization rates were calcuated as Arbuscules / FoV and Aseptate/FoV.
 Percent values = 0 were changed to 0.00001 for analysis
 purposes.   <strong>#####
 aboitic soil proporties #####</strong>                             
                                                                          
                                                                          
                                                                
   The associated CSV file for the abiotic soil
 proporties results is named
 <i>soil_properties_EcoLetters</i>.