Next-generation sequencing to inventory taxonomic diversity in eukaryotic communities: a test for freshwater diatoms

The recent emergence of barcoding approaches coupled to those of Next Generation Sequencing (NGS) have raised new perspectives for studying environmental communities. In this framework, we tested the possibility to derive accurate inventories of diatom communities from pyrosequencing outputs with an available DNA reference library. We used three molecular markers targeting the nuclear, chloroplast and mitochondrial genomes (SSU rDNA, rbcL, and cox1), and three samples of a mock community composed of 30 known diatom strains belonging to 21 species. In the goal to detect methodological biases, one sample was constituted directly from pooled cultures, whereas the others consisted of pooled PCR products. The NGS reads obtained by pyrosequencing (Roche 454) were compared first to a DNA reference library including the sequences of all the species used to constitute the mock community, and secondly to a complete DNA reference library with a larger taxonomic coverage. A stringent taxonomic assi...