Table of Contents

• Dataset Description
  • Methods & Sampling
  • Data Processing Description
• Data Files
• Parameters
• Instruments
• Deployments
• Project Information
• Funding

Cloned SSU rRNA sequences of kinetoplastids detected in Discovery, Urania and Kryos Basin haloclines and brines.

Related Reference:

Edgcomb, V.P., W. Orsi, H.-W. Breiner, A. Stock, S. Filker, M.M. Yakimov, and T. Stoeck. 2011, Novel kinetoplastids associated with hypersaline anoxic lakes in the Eastern Mediterranean deep-sea. Deep-Sea Research I 58(10):1040-1048.

The position of the halocline was determined during the R/V Oceanus cruise using a SBE9 CTD (Sea-Bird Electronics, USA) equipped with an SBE43 oxygen sensor (Sea-Bird Electronics, USA). Samples were collected from the halocline and brine of Discovery basin using a rosette equipped with Niskin bottles (12 L) on R/V Oceanus.

RNA was extracted as described in detail previously (Alexander et al., 2009) using bead beating of filters in an extraction buffer and E-Matrix tubes (QBiogene, MP Biomedicals, USA), followed by the RNA/DNA Allprep kit (Qiagen, Hildesheim, Germany) according to the manufacturer's instructions. Total RNA was transcribed into cDNA using the Two-Step Omniscript Reverse Transcription kit (Qiagen) according to the manufacturer's instructions. Subsequently, the small subunit ribosomal RNA gene (18S rRNA) was amplified using the kinetoplastid-specific primers 14F (5′-CTGCCAGTAGTCATATATGCTTGTTTCAAGGA-3′) and 2026R (5′-GATCCTCTGCAGGTTCACCTACAGCT-3′) (von der Heyden et al., 2004).

PCR products were cloned and Sanger sequenced (bidirectionally)

BCO-DMO Processing:
original file: EdgcombKinetoplastid.xlsx
- added conventional header with dataset name, PI name, version date
- renamed parameters to BCO-DMO standard
- converted lat/lon to decimal degrees from degrees-decimal minutes
- added cruise id's
- added html links to GenBank Popset

Protists are an essential component of microbial food webs and play a central role in global biogeochemical cycles, and thus are key players in sustaining the healthy functioning of any ecosystem. Over the past few years a rich diversity of protists has been revealed in a range of extreme environments, indicating that the frontiers of eukaryotic life are still being explored. Only recently, one of the most extreme marine environments known to science was discovered in the eastern Mediterranean Sea at a depth of ~3500m, namely deep hypersaline anoxic basins (DHABs). These basins are characterized by extremely high salt concentrations (up to saturation) that have been considered anathema to life. Instead, highly diverse communities of bacteria exist in the waters of these basins. With the exception of a preliminary study to this proposal that indicated a diverse and active assemblage of protists in the water column along the halocline and below the halocline, these DHABs remain largely unexplored regarding eukaryotic life forms. The sediments of the DHABs have not been explored for protists at all.

The investigators will collect water column and sediment samples on a short cruise to two basins with different brine chemistries. An exciting combination of molecular, cultivation-independent and culture-based approaches will be used to study the microbial communities of two basins. Investigators will use those approaches to determine adaptive strategies of marine protist communities to hypersaline, anoxic environments and the degree of their potential impact on biogeochemical cycling as a result of their predation activities, the degree to which the dominant protists maintain bacterial or archaeal symbionts, and the identity of those symbionts. The original research proposal identified Bannock and Discovery Basins as the field study areas, however the 2009 cruise collected samples at Discovery and Urania Basin. Methods to be employed include RNA-based sequence analysis of diversity based on 18S rDNA genes, statistical analyses of community composition and phylotype richness, geochemical documentation of the water column and sediments using classical and microelectrode approaches, expression profiling using 3'-UTR fragments of mRNAs, sequencing of complete gene transcripts for proteins appearing to confer adaptation to hypersalinity, analysis of the proteome signatures, FISH-SEM to characterize novel extremophiles, CARD-FISH to identify eukaryote prey and putative symbionts, and TEM to assess morphology and endobiont presence in common benthic morphotypes.

Hypersaline environments rank highly in the list of extreme systems that have attracted increasing notice in science as well as by the lay public. For example, considering predictions of increasing temperatures and drought in certain regions of our planet, the number of hypersaline habitats may increase dramatically causing this ecosystem to gain importance on a global scale. Thus, an understanding of the ecosystem in these habitats will help predict future ecosystem functioning due to global change. From a different perspective, revealing the mechanisms of adaptation to high salinity has become a major objective, both for biological science and for potential commercial exploitation of natural products associated with those adaptations.