Long- and short-read metabarcoding technologies reveal similar
spatio-temporal structures in fungal communities
Abstract
Fungi form diverse communities and play essential roles in many
terrestrial ecosystems, yet there are methodological challenges in
taxonomic and phylogenetic placement of fungi from environmental
sequences. To address such challenges we investigated spatio-temporal
structure of a fungal community using soil metabarcoding with four
different sequencing strategies: short amplicon sequencing of the ITS2
region (300–400\ bp) with Illumina MiSeq, Ion Torrent
Ion S5, and PacBio RS II, all from the same PCR library, as well as long
amplicon sequencing of the full ITS and partial LSU regions
(1200–1600\ bp) with PacBio RS II. Resulting community
structure and diversity depended more on statistical method than
sequencing technology. The use of long-amplicon sequencing enables
construction of a phylogenetic tree from metabarcoding reads, which
facilitates taxonomic identification of sequences. However, long reads
present issues for denoising algorithms in diverse communities. We
present a solution that splits the reads into shorter homologous regions
prior to denoising, and then reconstructs the full denoised reads. In
the choice between short and long amplicons, we suggest a hybrid
approach using short amplicons for sampling breadth and depth, and long
amplicons to characterize the local species pool for improved
identification and phylogenetic analyses.