Genomic DNA extraction PCR amplification of 16S rDNA Separate mixture of 16S rDNAs by cloning into plasmid vector and transformation of host cells ATTCGATCCG... B. subtilis B. cereus M. luteus P. aeruginosa P. stutzeri Sequence individual cloned genes, compare sequence to database, and identify original bacteria Bacteria in sample Laboratory Methods 10 AY |Psychrobacter.alimentarius F282 CGO 1 F238 CGO 1 F227 CGO 1 F103 CGO 1RC AB |Psychrobacter.pacificensis AA29 BRR 1RC F213 CGO 1RC AB002657| 16S Mariana trench mud AJ |Beer bottling plant biofilm L |Thyasira.flexuosa AA04 CGO 1RC A196 NIS 1 AA53 BRR 1 AA33 BRR 1 AA61 BRR 1 AF384206|Ectothiorhodospira.sp AJ404972|Alcalilimnicola.halodurans A092 NIS 1RC A182 NIS 1 A021 NIS 1RC AA31 NIS 1 L |Marine.aggregate gi gb gi gb gi gb gi gb AF321239|Pseudomonas.sp. F104 CGO 1 PFU492830|Pseudomonas.fulgida AF388206|Pseudomonas.sp AF Pseudomonas.putida.MT PSEIAM13|Pseudomonas.fragi ABE308319|Azotobacter.beijerin AA77 NIS 1RC AB |Pseudomonas.pachastrellae AA92 NIS 1 AJ |Marinobacter.sedimentalis DA30 CGO 1 DA16 CGO 1 DA42 CGO 1 DA55 CGO 1 DA72 CGO 1 DA79 CGO 1RC DA22 CGO 1RC DA38 CGO 1 DA49 CGO 1 DA37 CGO 1 DA50 CGO 1 F019 CGO 1RC F046 CGO 1RC F180 CGO 1 F044 CGO 1 F078 CGO 1 F030 CGO 1RC F184 CGO 1 F124 CGO 1RC AY |Marine.oligotrophic.water AA05 NIS 1RC AB |deep sea sediments DA73 CGO 1 U |Continental shelf Cape hateras AF |Continental shelf off Oregon CA36 CGO 1 gi gb gi gb gi gb F145 CGO 1 gi gb gi gb gi gb AA62 NIS 1 AA87 BRR Aquifex.aeolicus.VF5 16S rRNA sequences identified as within the Class Pseudomonadales (2002 data) Colors indicate what location and type of sample from which the sequences were obtained  Bamboo coral from Murray Seamount  Black coral from Murray Seamount  Bamboo coral Isidella from Warwick Seamount  Water collected at Murray Seamount  Caribbean coral (Rohwer et al., 2002)  Closest BLASTN match in Genbank  Biofilm from rock at Murray Seamount  Bamboo coral from Chirikof Seamount Specific to Coral Life as we know it can be divided into three broad groups, the Eucarya, Archaea, and Bacteria. The only group visible to us with the naked eye is the Eucarya, containing animals, plants, and fungi. The Bacteria and Archaea are single-celled microbes and are only visible with use of a microscope. However, despite their small size, they have important roles in many spheres, and interactions with many larger organisms. These interactions can be either detrimental or beneficial to the eukaryote host. What is TIGR? TIGR (The Institute for Genomic Research) is a private, not-for-profit research institute located in Rockville, Maryland. TIGR principally conducts large-scale genome sequencing projects; sequencing targets include many microbial species, but also plant, animal, and fungal organisms. The DNA sequence data obtained from these projects is used to derive genome-based insights into the biology of the target organism. TIGR has been participating in research cruises aboard R/V Atlantis, at the invitation of WHOI and NOAA OE, since In Search of Bacterial Communities Inhabiting Deep-Sea Corals on Gulf of Alaska Seamounts Kevin Penn and Naomi Ward, TIGR Acknowledgements We thank the Captain and crew of the R/V Atlantis, as well as members of the Alvin group, for all their help during both the 2002 and 2004 expeditions. Special thanks to Randy Keller, Brad Stevens, Tom Shirley, Tom Guilderson, Amy Baco- Taylor, and Peter Etnoyer for taking us on board. Thanks also to WHOI and NOAA’s Ocean Explorer program (especially Catalina Martinez) for supporting our participation. Acknowledgements We thank the Captain and crew of the R/V Atlantis, as well as members of the Alvin group, for all their help during both the 2002 and 2004 expeditions. Special thanks to Randy Keller, Brad Stevens, Tom Shirley, Tom Guilderson, Amy Baco- Taylor, and Peter Etnoyer for taking us on board. Thanks also to WHOI and NOAA’s Ocean Explorer program (especially Catalina Martinez) for supporting our participation. Bamboo coral observed during Alvin dive We know that bacterial populations are associated with these corals, but their function (parasitic, or beneficial to the coral) remains unclear. Collection of bamboo coral specimen using Alvin’s manipulators Bamboo coral specimen successfully transferred to the surface in Alvin’s basket Collection of exopolysaccharide (mucus) produced by bamboo coral. Coral mucus may protect the coral from desiccation, predators, and disease. We know that the mucus contains many different species of bacteria. The mucus presumably provides shelter and nutrition to the bacteria, but is not yet known whether the bacteria provide a benefit to their coral host. We use 16S rDNA cloning and sequencing to determine community composition, without the need to cultivate bacteria in the lab. This approach has been shown to recover bacterial diversity much more extensively than cultivation-based methods. 16S rDNA sequences can be used to draw a family tree of relationships between different bacteria. These data suggest that different coral species harbor specific groups of bacteria. Seamounts visited during the 2004 expedition to the Gulf of Alaska