PlutoF—a Web Based Workbench for Ecological and Taxonomic Research, with an Online Implementation for Fungal ITS Sequences

Abstract

DNA sequences accumulating in the International Nucleotide Sequence Databases (INSD) form a rich source of information for taxonomic and ecological meta-analyses. However, these databases include many erroneous entries, and the data itself is poorly annotated with metadata, making it difficult to target and extract entries of interest with any degree of precision. Here we describe the web-based workbench PlutoF, which is designed to bridge the gap between the needs of contemporary research in biology and the existing software resources and databases. Built on a relational database, PlutoF allows remote-access rapid submission, retrieval, and analysis of study, specimen, and sequence data in INSD as well as for private datasets though web-based thin clients. In contrast to INSD, PlutoF supports internationally standardized terminology to allow very specific annotation and linking of interacting specimens and species. The sequence analysis module is optimized for identification and analysis of environmental ITS sequences of fungi, but it can be modified to operate on any genetic marker and group of organisms. The workbench is available at http://plutof.ut.ee.

Ectomycorrhizal fungi contribute to soil organic matter cycling in sub-boreal forests

“…The extracted ITS1 sequences were clustered at 98% (Ryberg et al, 2008) similarity (global alignment) using CD-Hit-Est (Huang et al, 2010). Representative sequences from all clusters (operational taxonomic unit, OTU) were used for BLAST searches against fully identified entries in INSD (Benson et al, 2011) and UNITE (Abarenkov et al, 2010a), using the PlutoF online server (Abarenkov et al, 2010b). A minimum of 97% similarity across 97% of the sequence length was required for positive taxonomic identification at any level.…”

Five simple guidelines for establishing basic authenticity and reliability of newly generated fungal ITS sequences

“…Several studies have reported on the various shortcomings of the public DNA sequence corpus (e.g., Gilks et al 2002;Harris 2003;Bidartondo et al 2008), but none have succeeded in halting the continual submission of substandard entries to the databases. On the contrary, there are indications that the proportion of several classes of compromised sequences -such as chimeras and reverse complementary sequences -increases over time (Abarenkov et al 2010b). While very experienced users may perhaps be able to look through such broken data, many others may not be in a position to do so, particularly not since a growing number of people from outside mycology -even outside the academia -now use fungal sequence data as a part of their work.…”

“…One recent example is to add an UNVERIFIED keyword to highly problematic sequences and exclude the sequence from BLAST, although the sequence will still be archived in INSD. Finally, it is possible to use the third-party sequence annotation feature of UNITE/PlutoF (Abarenkov et al 2010b) to simply replace the incorrect species name with the correct one, or to mark the entry as chimeric, or to take whatever other action appropriate. Third-party annotations of sequences via PlutoF are visible to users in the European Nucleotide Archive of the INSD through a link-out function.…”

Tree diversity and species identity effects on soil fungi, protists and animals are context dependent

“…The longest sequence of each remaining 9614 clusters was selected as a representative for BLASTn sequence similarity search (word size = 7; penalties: gap = − 1; gap extension = − 2; match = 1) against the INSDC and UNITE (Abarenkov et al, 2010a) databases. In addition, we ran BLASTn searches against reference sequences of fungi in 99.0% similarity species hypotheses that include third-party taxonomic and metadata updates as implemented in the PlutoF workbench (Abarenkov et al, 2010b). For each query, we considered 10 bestmatching references to annotate taxa as accurately as possible.…”