DOI: 10.5061/dryad.hmgqnk9p6
title: Data from: Mycorrhizal driven feedbacks and forest resilience to reduced rainfall

description: Forests can experience negative feedbacks in the growth of tree
 populations but positive feedbacks within the two dominant mycorrhizal
 types of trees: ectomycorrhizal (EM) and arbuscular mycorrhizal (AM).
 Positive feedbacks within mycorrhizal types may provide communities with
 resistance to climate change. We tested whether each mycorrhizal type led
 to positive feedbacks on seedling survival, while statistically
 controlling for the effect of congeneric trees in ambient versus rainfall
 reduced conditions. We explored two potential drivers; the variation in
 soil fungal community structure and soil chemistry. Seedlings benefited
 from growing in stands dominated by their own mycorrhizal type, and
 simultaneously, tree seedlings performed worse in the presence of adult
 trees of their own genus, but only in rainfall reduced conditions. We
 found that the composition of the EM fungal community differed between
 plots dominated by EM versus AM trees. These results indicated that
 mycorrhizal types may create positive feedbacks in dry conditions that
 should be considered when predicting future states.
description: Methods Site description and plot
 selection We assessed the effects of mycorrhizal type
 and throughfall reduction on tree seedling success in a field experiment
 in central Illinois. We quantified feedbacks of mycorrhizal type and
 congenerics by comparing tree seedling success and performance across 12
 forest plots that varied in the basal area of EM trees and percent basal
 area of trees congeneric to our transplanted seedlings. The study site was
 in Allerton Park and Retreat, a 1500-acre natural area
 (https://allerton.illinois.edu/). Within a 64-acre area old-growth upland
 forest, 12 plots with similar topography were selected that represented a
 gradient of % EM canopy tree basal area (see Fig. S1 and Table S1).
 Species compositions of plots were determined by recording the diameter at
 breast height (DBH) and species identity of each tree over 10 cm DBH in a
 circle of 30 m diameter. Tree species were designated as EM or AM
 mycorrhizal status (See Table S2). The % basal area of canopy EM tree
 species was determined by dividing the summed basal area of EM tree stems
 (<em>Quercus</em>, <em>Carya</em>, and
 <em>Tilia</em> individuals) by total basal area of all trees.
             Experimental Design The
 effects of mycorrhizal guild feedbacks, congeneric feedbacks, and
 throughfall treatments on tree seedling success were determined using a
 split-plot design. Within each of the 12 plots we established three sets
 of paired control and rain out subplots, totaling six subplots per plot.
 Canopy openness (%) was calculated using Gap Light Analyzer imaging
 software from hemispherical photographs at each of the subplots (Frazer et
 al. 1999).. In each subplot, a mix of 11 AM and EM tree seedlings were
 planted 40 cm apart. Bare-root seedlings were sourced from two nurseries.
 Tree seedlings from Engel’s Nursery in Fennville, MI were planted May
 15-19, 2017, and all tree seedlings were one year old except for
 <em>Diospyros virginiana</em> seedlings, which were one to two
 years old. All tree seedlings from Mason State Nursery in Topeka, Il were
 planted May 25-27, 2017 and were one year old. To
 reduce natural precipitation, we constructed three throughfall reduction
 shelters and three mock shelters at each site.  A 1.2m×1.5m sheet of clear
 polycarbonate plastic was attached to the top of four PVC legs,
 approximately 1.3m in length. For the ambient treatment, mock shelters had
 80 holes (d=1.27 cm) drilled into each plastic sheet (10.2% of area
 removed) to allow precipitation through to tree seedlings.  The
 throughfall reduction shelters were left unaltered and were positioned at
 an angle to allow for drain off. Soil moisture (m³/m³ water content) and
 soil temperature on one paired subplot in each of three main plots were
 measured using a TEROS 12 Moisture sensor. The ZL6 Cloud data logger
 logged data every 12 hours from June 1 to October 16, 2019. Throughfall
 reductions shelters reduced average soil moisture by 10.4% + 0.08 SE in
 the 2019 growing season.                      The
 average precipitation for the growing season (June-Oct.) for the 30 years
 prior to the start of the study (1986-2016) was 449 mm at this site (PRISM
 2004). In 2017, the precipitation for the growing season was 321 mm,
 indicating drought conditions for the first year of the study. In both the
 2018 and 2019 growing season, precipitation was slightly above normal (546
 and 532 mm, respectively), with higher precipitation in the early growing
 season in 2018 and the later growing season in 2019.
 Each tree seedling was assessed for survival and performance
 measurements across three years (2017-2019) in early growing season and
 late growing season. We assessed tree seedling survival and measured stem
 diameter and height for living plants. At the end of the third growing
 season (2019), surviving plants were harvested, collecting all aboveground
 tissue and as much belowground tissue as possible within the subplot
 perimeters. Roots were immediately assessment of mycorrhizal colonization
 and the remainder dried for biomass measurements.
             AM fungal colonization on surviving AM seedlings was
 measured on cleared and stained roots using the grid line intersect method
 at 40X magnification (Giovannetti & Mosse 1980). Ectomycorrhizal
 colonization was assessed under a dissecting microscope using methods in
 Brundrett et al. 1996. Since most seedlings died before the end of the
 experiment, any destructive sampling of the surviving seedlings was highly
 censored. For additional details, see Supplemental Methods.
             We collected soil cores (10 cm depth) from each of
 the 72 subplots (three ambient and three throughfall reduced for each of
 12 sites) in summer 2018. Fungal communities were characterized in these
 soil samples by sequencing the ITS2 region of the fungal rRNA gene, as
 amplified by the ITS3-KYO2 (Toju et al. 2012) and ITS4 (White et al. 1990)
 primer pair. Amplicons were sequenced via paired end Illumina Miseq with
 300 cycles. Sequences were clustered into exact Amplicon Sequence Variants
 using the DADA2 program (Callahan et al. 2016) as implemented in the
 QIIME2 pipeline and identified to the lowest confident taxonomic level
 using the naïve Bayesian classifier using the UNITE database for ITS reads
 (Nilsson et al. 2019). We used the FUNGUILD database to assign ASVs (ASVs
 equivalent to 100% identity OTUs) to functional groups (Nguyen et al.
 2016). For additional details, see Supplemental Methods.
 We took a soil sample from each pair of subplots at the 12 plots,
 totaling 36 samples for soil analyses at University of Wisconsin Soil
 & Forage Analysis Lab (Marshfield, WI). Soil pH was measured using
 a 1:1 soil to water dilution, concentrations of P and K were extracted
 using a Bray method, and % organic matter was measured using the
 Loss-On-Ignition method. Total N and C were assessed using the dry
 combustion method.