Publication
A new oomycete metabarcoding method using the rps10 gene
Publisher:
Cold Spring Harbor Laboratory
Date:
22-09-2021
DOI:
10.1101/2021.09.22.460084
Abstract: Oomycetes are a group of eukaryotes related to brown algae and diatoms, many of which cause diseases in plants and animals. Improved methods are needed for rapid and accurate characterization of oomycete communities using DNA metabarcoding. We have identified the mitochondrial 40S ribosomal protein S10 gene ( rps10 ) as a locus useful for oomycete metabarcoding and provide primers predicted to lify all oomycetes based on available reference sequences from a wide range of taxa. We evaluated its utility relative to a popular barcode, the internal transcribed spacer 1 (ITS1), by sequencing environmental s les and a mock community using Illumina MiSeq. Amplified sequence variants (ASVs) and operational taxonomic units (OTUs) were identified per community. Both the sequence and predicted taxonomy of ASVs and OTUs were compared to the known composition of the mock community. Both rps10 and ITS yielded ASVs with sequences matching 21 of the 24 species in the mock community and matching all 24 when allowing for a 1 bp difference. Taxonomic classifications of ASVs included 23 members of the mock community for rps10 and 17 for ITS1. Sequencing results for the environmental s les suggest the proposed rps10 locus results in substantially less lification of non-target organisms than the ITS1 method. The lified rps10 region also has higher taxonomic resolution than ITS1, allowing for greater discrimination of closely related species. We present a new website with a searchable rps10 reference database for species identification and all protocols needed for oomycete metabarcoding. The rps10 barcode and methods described herein provide an effective tool for metabarcoding oomycetes using short-read sequencing. Oomycetes are a group of eukaryotes related to brown algae and diatoms, many of which cause diseases in plants and animals. Improved methods are needed to rapidly characterize the ersity of oomycete species found in environmental s les. We have identified the mitochondrial 40S ribosomal protein S10 gene ( rps10 ) as being useful for oomycete community sequencing. We evaluated its utility relative to a popular barcode, the internal transcribed spacer 1 (ITS1), by sequencing environmental s les and a community we synthesized in the laboratory. The lified rps10 region is predicted to have a higher taxonomic resolution than ITS1, allowing for greater discrimination of closely related species. We present a new website with a searchable rps10 reference database for species identification and all protocols needed for oomycete community sequencing. The rps10 barcode and methods described herein provide an effective tool for characterizing oomycetes using environmental DNA sequencing.