Abstract
The development of high-throughput sequencing (HTS) technologies has
greatly improved our capacity to identify fungi and unveil their
ecological roles across a variety of ecosystems. Here we provide an
overview about current best practices in metabarcoding analysis of
fungal communities, from experimental design through molecular and
computational analyses. By re-analysing published datasets, we find that
operational taxonomic units (OTUs) outperform amplified sequence
variants (ASVs) in recovering fungal diversity, which is particularly
evident for long markers. Additionally, analysis of the full-length ITS
region allows more accurate taxonomic placement of fungi and other
eukaryotes compared with the ITS2 subregion. We conclude that
metabarcoding analyses of fungi are especially promising for co-analyses
with the functional metagenomic or transcriptomic data, integrating
fungi in the entire microbiome, recovery of novel fungal lineages and
ancient organisms as well as barcoding of old specimens including type
material.