Phylogenetic Analysis

To visualize how the armillarioid species clusters were related, we utilized a phylogenetic approach. One exemplar sequence from each species cluster was chosen to include in the multiple sequence alignment. We utilized the full-length sequence, when possible, and not just the hypervariable regions. Species clusters for the ITS1 region and ITS2 region were analyzed separately. This was because in no study where we identified armillarioid taxa did researchers employ both ITS1 and ITS2 markers from a single sampling location such that we could absolutely ensure that both markers came from the same genetic individual or genet. As ITS1 and ITS2 sequences could not be ascribed to a single genet or individual, we did not attempt to statistically assess the phylogenetic tree correlation of the ITS1 and ITS2 sequences – this would be better suited for pangenomic studies of the armillarioid fungi using multiple genes. The public sequences we identified as having homology to our armillarioid nexus file were aligned using MAFFT version 7 (Katoh and Standley, 2013) with refinements to the alignment performed manually. RAxML-NG (Kozlov et al., 2019) was used to reconstruct this phylogeny. The two known species of Guyanagaster, sister to Armillaria and Desarmillaria species (Koch et al., 2017), were used to root this phylogeny. We defined well supported branches as having 70% or greater bootstrap support. The location metadata associated with each species cluster was mapped onto each phylogenetic tree using the ‘ggtree’ package (Yu et al., 2018) and the ‘UpSetR’ package (Conway et al., 2017) within the R 4.1.0 environment (R Core Team, 2013).