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.

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 Alliaria by hand from the left side of each plot and carried the weeded material offsite. The right side of each plot was left at ambient Alliaria field densities, which averaged 15.2% cover (Roche et al. 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 Alliaria 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 Alliaria 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 Alliaria, we installed temporary aboveground mesh barriers between the weeded and ambient sides of the plots prior to Alliaria 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 Alliaria in the weeded plots (Roche et al. 2020).

##### Leaf-level morphological and physiological traits #####                                                                                                                                                                                                       

To characterize the effect of the Alliaria 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_sat), and stomatal conductance (g_s) using a Li-COR LI-6400XTR portable photosynthesis system, equipped with a CO₂ 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^-2s^-1, ambient temperature and humidity, and a reference chamber CO₂ concentration of 400 μmol mol^-1, following Heberling et al. (2017). A saturating light level of 800 μmol m^-2s^-1 was based on detailed light response curves for these understory species at our field site (Heberling et al. 2017; Heberling et al. 2019a). To minimize within-plant variability for Maianthemum, we restricted our sampling to either the 2^nd or 3^rd leaf from the terminal end of the stem following Cornelissen et al. (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 Alliaria treatments x 3 species x 2 life stages). To measure SLA, we collected two to three 5.32 cm² leaf tissue samples from each individual and calculated the mean leaf surface area per g dry mass (cm² g^-1). 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_sat /g_s; µmol CO₂ mmol H₂O^-1) and photosynthetic nitrogen use efficiency (PNUE = A_sat / N_area; μmol CO₂ gN^-1 s^-1) (Field et al. 1983; Wright et al. 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 plant_traits_performance_EcoLetters.

Before doing the physiology analyses, we subset the raw data to exclude Ci values <60. 

The treatment column in the CSV file refers to the two garlic mustard treatmetns (i.e., 'Ambient' and 'Weeded')

The Sp column in the 'plant_traits_performance_EcoLetters' 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.

##### fungal community composition #####                                                                                                                                                                                                                                     

The associated CSV file for the fungal community composition results is named Soil_AM_Fungal_communites_EcoLetters. 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.

##### root conolozation rates #####                                                                                                                                                                                                                                                 

The associated CSV file for the root colonization results is named AM_fungal_root_colonization_EcoLetters.

Refer to the methods section of the published paper for details on how the root sampls were processed. 

The treatment column in associated files refers to the two garlic mustard treatmetns (i.e., 'Ambient' and 'Weeded')

The sp 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.

##### aboitic soil proporties #####                                                                                                                                                                                                                                                   

The associated CSV file for the abiotic soil proporties results is named soil_properties_EcoLetters.