A metabarcoding framework for facilitated survey of endolithic phototrophs with tufA

Background: In spite of their ecological importance as primary producers and microbioeroders of marine calcium carbonate (CaCO3) substrata, endolithic phototrophs spanning both prokaryotic (the cyanobacteria) and eukaryotic algae lack established molecular resources for their facilitated survey with high throughput sequencing. Here, the development of a metabarcoding framework for the elongation factor EF-Ttu (tufA) was tested on four Illumina-sequenced marine CaCO3 microfloras for the characterization of their endolithic phototrophs, especially the abundant bioeroding Ostreobium spp. (Ulvophyceae). The framework consists of novel tufA degenerate primers and a comprehensive database enabling Operational Taxonomic Unit (OTU) identification at multiple taxonomic ranks with percent identity thresholds determined herein. Results: The newly established tufA database comprises 4057 non-redundant sequences (from 1339 eukaryotic and prokaryotic phototrophs, and 2718 prokaryotic heterotrophs) i...