Stoeck Thorsten

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Stoeck
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Thorsten
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  • Article
    Massively parallel tag sequencing reveals the complexity of anaerobic marine protistan communities
    (BioMed Central, 2009-11-03) Stoeck, Thorsten ; Behnke, Anke ; Christen, Richard ; Amaral-Zettler, Linda A. ; Rodriguez-Mora, Maria J. ; Chistoserdov, Andrei Y. ; Orsi, William D. ; Edgcomb, Virginia P.
    Recent advances in sequencing strategies make possible unprecedented depth and scale of sampling for molecular detection of microbial diversity. Two major paradigm-shifting discoveries include the detection of bacterial diversity that is one to two orders of magnitude greater than previous estimates, and the discovery of an exciting 'rare biosphere' of molecular signatures ('species') of poorly understood ecological significance. We applied a high-throughput parallel tag sequencing (454 sequencing) protocol adopted for eukaryotes to investigate protistan community complexity in two contrasting anoxic marine ecosystems (Framvaren Fjord, Norway; Cariaco deep-sea basin, Venezuela). Both sampling sites have previously been scrutinized for protistan diversity by traditional clone library construction and Sanger sequencing. By comparing these clone library data with 454 amplicon library data, we assess the efficiency of high-throughput tag sequencing strategies. We here present a novel, highly conservative bioinformatic analysis pipeline for the processing of large tag sequence data sets.The analyses of ca. 250,000 sequence reads revealed that the number of detected Operational Taxonomic Units (OTUs) far exceeded previous richness estimates from the same sites based on clone libraries and Sanger sequencing. More than 90% of this diversity was represented by OTUs with less than 10 sequence tags. We detected a substantial number of taxonomic groups like Apusozoa, Chrysomerophytes, Centroheliozoa, Eustigmatophytes, hyphochytriomycetes, Ichthyosporea, Oikomonads, Phaeothamniophytes, and rhodophytes which remained undetected by previous clone library-based diversity surveys of the sampling sites. The most important innovations in our newly developed bioinformatics pipeline employ (i) BLASTN with query parameters adjusted for highly variable domains and a complete database of public ribosomal RNA (rRNA) gene sequences for taxonomic assignments of tags; (ii) a clustering of tags at k differences (Levenshtein distance) with a newly developed algorithm enabling very fast OTU clustering for large tag sequence data sets; and (iii) a novel parsing procedure to combine the data from individual analyses. Our data highlight the magnitude of the under-sampled 'protistan gap' in the eukaryotic tree of life. This study illustrates that our current understanding of the ecological complexity of protist communities, and of the global species richness and genome diversity of protists, is severely limited. Even though 454 pyrosequencing is not a panacea, it allows for more comprehensive insights into the diversity of protistan communities, and combined with appropriate statistical tools, enables improved ecological interpretations of the data and projections of global diversity.
  • Preprint
    Protistan community patterns within the brine and halocline of deep hypersaline anoxic basins in the eastern Mediterranean Sea
    ( 2008-10-21) Edgcomb, Virginia P. ; Orsi, William D. ; Leslin, Chesley ; Epstein, Slava S. ; Bunge, John ; Jeon, Sunok ; Yakimov, Michail M. ; Behnke, Anke ; Stoeck, Thorsten
    Environmental factors restrict the distribution of microbial eukaryotes but the exact boundaries for eukaryotic life are not known. Here we examine protistan communities at the extremes of salinity and osmotic pressure, and report rich assemblages inhabiting Bannock and Discovery, two deep-sea superhaline anoxic basins in the Mediterranean. Using a rRNA-based approach, we detected 1538 protistan rRNA gene sequences from water samples with total salinity ranging from 39 g/kg to 280 g/Kg, and obtained evidence that this DNA was endogenous to the extreme habitats sampled. Statistical analyses indicate that the discovered phylotypes represent only a fraction of species actually inhabiting both the brine and the brine-seawater interface, with as much as 82% of the actual richness missed by our survey. Jaccard indices (e.g., for a comparison of community membership) suggest that the brine/interface protistan communities are unique to Bannock and Discovery basins, and share little (0.8-2.8%) in species composition with overlying waters with typical marine salinity and oxygen tension. The protistan communities from the basins’ brine and brine/seawater interface appear to be particularly enriched with dinoflagellates, ciliates and other alveolates, as well as fungi, and are conspicuously poor in stramenopiles. The uniqueness and diversity of brine and brine-interface protistan communities make them promising targets for protistan discovery.
  • Article
    Evidence for isolated evolution of deep-sea ciliate communities through geological separation and environmental selection
    (BioMed Central, 2013-07-08) Stock, Alexandra ; Edgcomb, Virginia P. ; Orsi, William D. ; Filker, Sabine ; Breiner, Hans-Werner ; Yakimov, Michail M. ; Stoeck, Thorsten
    Deep hypersaline anoxic basins (DHABs) are isolated habitats at the bottom of the eastern Mediterranean Sea, which originate from the ancient dissolution of Messinian evaporites. The different basins have recruited their original biota from the same source, but their geological evolution eventually constituted sharp environmental barriers, restricting genetic exchange between the individual basins. Therefore, DHABs are unique model systems to assess the effect of geological events and environmental conditions on the evolution and diversification of protistan plankton. Here, we examine evidence for isolated evolution of unicellular eukaryote protistan plankton communities driven by geological separation and environmental selection. We specifically focused on ciliated protists as a major component of protistan DHAB plankton by pyrosequencing the hypervariable V4 fragment of the small subunit ribosomal RNA. Geospatial distributions and responses of marine ciliates to differential hydrochemistries suggest strong physical and chemical barriers to dispersal that influence the evolution of this plankton group. Ciliate communities in the brines of four investigated DHABs are distinctively different from ciliate communities in the interfaces (haloclines) immediately above the brines. While the interface ciliate communities from different sites are relatively similar to each other, the brine ciliate communities are significantly different between sites. We found no distance-decay relationship, and canonical correspondence analyses identified oxygen and sodium as most important hydrochemical parameters explaining the partitioning of diversity between interface and brine ciliate communities. However, none of the analyzed hydrochemical parameters explained the significant differences between brine ciliate communities in different basins. Our data indicate a frequent genetic exchange in the deep-sea water above the brines. The “isolated island character” of the different brines, that resulted from geological events and contemporary environmental conditions, create selective pressures driving evolutionary processes, and with time, lead to speciation and shape protistan community composition. We conclude that community assembly in DHABs is a mixture of isolated evolution (as evidenced by small changes in V4 primary structure in some taxa) and species sorting (as indicated by the regional absence/presence of individual taxon groups on high levels in taxonomic hierarchy).