Marine heterotrophic alveolates: a genomic and morphometric single cell approach.

• Applicant: Dr. Uwe John
• Subject Area: Evolution and Systematics of Plants and Fungi ; Systematics and Morphology (Zoology)
• Term: from 2017 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 351812489

Microbial eukaryotes (protists) apparently account for the majority of eukaryotic species, but comparably few taxa have been taxonomically investigated from both morphological and molecular criteria, as there are only limited sequence data. The gap between known and unknown microbial eukaryotes is even higher for heterotrophs, because most descriptions and nucleic acid based investigations depend upon laboratory cultures and heterotrophs tend to be more difficult to cultivate than autotrophs. Newly developed single cell methods, such as single cell PCR, enable classification without cultivation, but most nucleic acid based studies still rely almost entirely on published sequences, rendering unknown species invisible as poorly or completely unassigned sequences. Furthermore, data about heterotrophs is almost completely lacking from the new field of metatranscriptomics. The need for an extensive annotation of new sequences and correct taxonomic classification of the respective heterotrophic species is therefore evident. In this study, we will focus on marine heterotrophic protists, i.e. alveolates, since at times they represent the major component of planktonic biomass, transferring biological matter to higher trophic levels, and may play key functional roles in marine ecosystems. We will conduct a winter and summer field study in Greenland and a cruise around Spitzbergen to study the successional processes of heterotrophic communities and the differences in their biodiversity during annual shifts. The goal of collecting comprehensive data for completing databases will be reached with the development of a toolbox integrating microscopy, single cell multiplex PCR, single cell transcriptomics and meta-analysis of the collected environmental samples. With single cell PCR and sequencing of ribosomal DNA (SSU, LSU and ITS), we will obtain sequences for classification or identification of species and will identify cryptic species. Higher sequence resolution will be achieved with ITS2 rRNA secondary structure modeling and compensatory base change (CBC). Light microscopic analysis will serve to find established and new morphological classification features congruent with sequence data. Additionally, the organisms will be photographed and/or filmed and fixed for archiving. For selected understudied species, single cell transcriptomics will be utilized to develop new classification markers and to fill major gaps in metatranscriptomic databases. Meta-analyses of abiotic and biotic environmental data will complete the dataset of the species to offer additional biogeographical information. Existing databases will be supplied with the collected comprehensive data after thorough revision and nomenclatural analysis. The new toolbox will facilitate future identification and the data obtained will substantially enrich existing databases, thereby yielding a more comprehensive view of diversity of microbial eukaryotes and their functional role in marine ecosystems.

• DFG Programme: Priority Programmes
• Subproject of: SPP 1991: Taxon-Omics: New approaches for discovering and naming biodiversity
• Co-Investigator: Dr. Sylke Wohlrab