Utilizing ITS1 and ITS2 to study environmental fungal diversity using pyrosequencing.

Area of Science:

• Mycology
• Environmental Microbiology
• Molecular Ecology

Background:

• High-throughput sequencing generates shorter reads, prompting a focus on either the ITS1 or ITS2 sublocus for fungal diversity analysis.
• The choice of internal transcribed spacer (ITS) sublocus can potentially influence the interpretation of environmental fungal community structure.

Purpose of the Study:

• To investigate the impact of analyzing either the ITS1 or ITS2 sublocus on fungal diversity datasets.
• To compare the fungal diversity profiles obtained from ITS1 and ITS2 amplicon sequencing.

Main Methods:

• DNA extraction from environmental samples (water, sediments, soil).
• Amplification of the total internal transcribed spacer (ITS) locus.
• 454-sequencing targeting both ITS1 and ITS2 regions.

Main Results:

• No significant differences in sequence counts, operational taxonomic units (OTUs), or dominant OTUs between ITS1 and ITS2 datasets.
• Lower fungal diversity observed in the ITS2 dataset compared to ITS1.
• Differential fungal taxonomic identification in soil samples, with ITS1 showing more Basidiomycota and ITS2 showing more Ascomycota.
• Only one-third of OTUs were common to both datasets, potentially due to sequence removal, primer selectivity, or BLASTn query differences.

Conclusions:

• While ITS1 and ITS2 datasets offer similar fungal community-level insights, they provide distinct information.
• Analyzing both ITS1 and ITS2 regions is recommended for in-depth fungal diversity analysis due to their complementary nature.