We use Pacbio sequencing of full-length fungal nuclear ribosomal internal transcribed spacer (ITS) region to characterize the deadwood mycobiome of Castanopsis carlesii. Pacbio sequencing for full-length fungal nuclear ribosomal internal transcribed spacer (ITS) region to characterize the deadwood mycobiome of Castanopsis carlesii

Short-read Next Generation Sequencing (NGS) platforms can easily generate thousands to hundreds of thousands of sequences per sample in a short time; however the limited length of the obtained sequences may cause a problem to the fungal taxonomic identification. Here we aimed to validate the use of long-read sequencing NGS method, Pacbio sequencing to characterize the fungal community (mycobiome) associated with deadwood of Castanopsis carlesii. Specifically, we test (i) how diverse of the deadwood mycobiome? (ii) What fungal taxonomic and functional groups are associated with this deadwood mycobiomes? (iii) Is Pacbio sequencing suitable for mycobiome study in moderately complex fungal community? and (iv) the suitability to use phylogenetic analysis based on the full-length ITS obtained by Pacbio sequencing to confirm the taxonomic information at genus and species levels. In this work, we successfully use the Pacbio sequencing to generate the long-read sequences of the full-length fungal ITS (ranged from 500 – 780 bp) of the Castanopsis carlesii mycobiome. Our results showed that the studied deadwood mycobiome are taxonomically (average 85 fungal OTUs) and functionally (five functional groups: animal endosymbiont, endophyte, mycoparasite, plant pathogen and saprotroph) diverse. Based on relative abundance data, Basidiomycota are the most frequently detected phyla (50% of total sequences), followed by unidentified phyla and Ascomycota. However, based on presence/absence data, the OTU-rich phyla were Ascomycota (55% of total OTUs, 72 OTUs) followed by Basidiomycota and unidentified phyla. The majority of fungal OTUs were identified as saprotroph (70% of successfully function assigned OTUs) followed by plant pathogen. Finally, we successfully used phylogenetic analysis based on the full-length ITS obtained by Pacbio sequencing to confirm the species identification of 14/36 OTUs with high bootstrap support (99 – 100%). Based on the sequence reads obtained per sample which are ranged from 3,047 to 13, 463, we conclude that there is a high potential to use Pacbio sequencing to characterize moderately or even more complex fungal community.