Above and belowground community analysis
Statistical analysis were performed in R using RStudio (RStudio Team,
2015). The community composition of plant species recorded in all plots
along transects 1 and 4 was visualized by non-metric Multidimensional
Scaling (nMDS) using Bray-Curtis dissimilarities. Both plant and
belowground analyses were made with the vegan software package (version
2.5-7; Oksanen et al., 2019). The two-sample I-test was used to test for
significant difference in mean gravimetric soil moisture between the two
soil moisture regimes (wet vs. mesic-dry). The three OTU occurrence
tables were transformed to relative abundances for each sample and used
for community analysis. nMDS ordination plots were generated using the
‘metaMDS´function in vegan. To down weight the importance of common
taxa, the analysis was repeated using square root transformation of
relative abundance data prior to calculation of the Bray-Curtis
dissimilarity. Marginal and individual PERMANOVAs were conducted on all
three datasets and two standardizations described above, using the
‘adonis’ function in the vegan to test for the marginal and overall
effect of soil moisture regime (wet vs. mesic-dry) and presence/absence
of F. meleagris on shaping belowground microeukaryotic
communities at the study site.
After taxonomic assignment as described above, the three OTU occurrence
tables were divided into two separate datasets for fungal and protist
communities separately (ie. non-fungal microeukaryotes). The ordination
and PERMANOVA tests described above were repeated for these
taxonomically distinct communities.
Distribution and abundance of OTU_Ss were visualized across the
contrasting soil conditions wet vs. mesic-dry soil conditions and
presence/absence of F. meleagris using a Venn-diagram (Heberle et
al., 2015). The relative abundance of unique and shared fungal and
protist OTU_Ss were calculated across samples for the two contrasting
conditions. To identify differentially abundant taxa in contrasting soil
moisture regimes (wet vs. mesic-dry) and presence/absence of F.
meleagris , the ‘phyloseq-to-deseq´ function in the phyloseq package (v
1.34.0; (McMurdie & Holmes, 2013) was applied to OTU_S occurrence
tables separately. The generated phyloseq object was analysed using the
DESeq2 tool (DESeq package version 1.30.1; Love et al., 2014).
Identified taxa and their differential abundance were illustrated using
the ggplot2 R package (Wickham, 2016).