University of Essex

1. University of Essex BIODEEP-WP3 Analysis of species diversity, community structures and phylogeny of microorganisms and meiofauna in the Mediterranean deep-sea hypersaline anoxic basins (DHAB) Andrea Sass , Terry McGenity

2. Community structure fingerprinting via t-RFLP of 16S rDNA Summary of achievements up to date: • Amplification with fluorescent eubacterial primers successful • Water, brine and interface samples were all investigated • Not successful: • amplification with Archaeal primers • DNA extraction from most sediment samples • amplification from Discovery brine and interface

3. Brine samples: Summary of previously presented results: • brines of each basin unique patterns • l’Atalante and Bannock brines more similar • patterns stable over time • same patterns from Urania brines 1 and 2 • number of fragments relatively small

4. new achievements: amplification from l‘Atalante and Bannock basin sediments successful

5. Bannock sediment sample and brine Sediment top RFU Brine Fragment length Amplification with eubacterial primers, digestion with AluI

6. Sediment Brine L’Atalante sediment and brine Sediment top Sediment top covered with black ooze Sediment top amplification with a-caseine Sediment top extraction with phenol-chloroform RFU Sediment bottom Cruise 2002 Cruise 2002 amplification with a-casein Brine samples taken at same location and depth? Cruise 2001 Fragment length Amplification with eubacterial primers, digestion with AluI

7. patterns from sediment samples very similar to brine samples • no unique fragments found in sediment samples Extraction methods yield DNA only from the brine fraction of the sediment? Community in the sediments widely the same as in brines? • sediments consist mainly of brine • both sediments and brine are anoxic

8. Interfaces Summary of previous presented results: Patterns from interfaces unique for each basin and different from brines and oxic seawater Unique community or accumulation of microorganisms from the water column? Bannock interface samples taken in intervals at different salinities Certain fragments occur only at certain salinities within the interface

9. Oxic water, 3000 m depth Salinities 3.8-7.6% Salinities 8.7-10.8% Salinity 12.7% Salinities 13.8-15.9% Salinity 21.5% Salinity 25% = brine Bannock interface RFU Fragment length Amplification with eubacterial primers, digestion with AluI

10. Sediment traps 2 weeks 6 months RFU 12 months Fragment length Amplification with eubacterial primers, digestion with AluI

11. new achievements: investigation of Urania basin interface with greater spatial resolution

12. Salinity 5.7 ‰ Salinity 6.3 ‰ Salinity 9.0 ‰ Salinity 13.1 ‰ Salinity 15.4 ‰ Salinity 15.6 ‰ Salinity 18.4 ‰ Salinity 21.1 ‰ Brine Urania interface RFU Fragment length Amplification with eubacterial primers, digestion with AluI

13. fingerprints change gradually from interface to brine • no succession of microbial communities within interface interface not as stable as Bannock interface?

14. Bannock and Urania upper interfaces: community fingerprints are different from oxic deep-sea water change in communcity structure starts at seawater salt concentration above the sampled interface?

15. RFU Urania interface compared with oxic water samples Urania interface oxic water 3500m depth oxic water 3000m depth Fragment length samples taken 2001 Amplification with eubacterial primers, digestion with AluI

16. Different pattern in water and interface samples due to different amount of water filtered ? Patterns from the same DNA extract with different amounts of DNA addes to PCR reaction RFU Fragment length Amplification with eubacterial primers, digestion with AluI

17. Summary of newly achieved results: • Microbial community in Bannock and l‘Atalante sediments similar to those in corresponding brines • Succession of different comminties at different salinities within the interface more pronounced in Bannock interface than in Urania interface • Upper interfaces different from sea water