Characterization of Enrichment Cultures of Nitrate-Reducing Bacteria from Deep-Sea Hydrothermal Vents Caroline Toney* and Costantino Vetriani Deep-sea vents are one of the last frontiers of the unexplored world. These unique environments spew forth super- heated fluid that is laden with metals. The base of life which allow other organisms to live in this ecosystem are microorganisms which undergo chemosynthesis as a means of primary production. Introduction Figure 2. Sampling sites of deep-sea hydrothermal vents on the East Pacific Rise Objectives • To grow cultures from samples taken at vent sites at 9°N and 13°N. •To isolate anaerobic, nitrate- reducing bacteria using different techniques and media. •To compare the 16S rRNA gene sequences of these organisms to the database using BLAST (Basic Local Alignment Search Tool) to Media Carbon Source Electron Donor Electro n Accepto r Temp (°C) HB1 CO 2 H 2 NO 3 50 & 75 Sulfurovu m CO 2 N 2 , thiosulfat e NO 3 35 Sulfurimo nas CO 2 H 2 , thiosulfat NO 3 35, 50 ASW Organic N NO 3 30 Caroline Toney [email protected] (828) 429-9476 Figure 1. Illustration of deep-sea hydrothermal vent flow. (Brock et al., 2006). Site Media Temp (°C) Identity (%) Description Alvinella Tubes HB1 50 99,97 Caminibacter mediatlanticus; Nautilia profundicula, abyssi Sulfurimona s 35, 50 86 Sulfurospirillum carboxydovorans Alvinella sulfide HB1 50 97, 96 Caminibacter mediatlanticus HB1 75 98 Desulfobacterium pacificum Crab Spa Sulfurovum 35 93;96;93 Thioprofundum lithotrophica; Rhodobacter capsulatus; Marinobacter hydrocarbonoclasticus Sulfurimona s 35, 50 97 Shewanella haliotis Anaerobic ASW 30 100 Klebsiella pneumoniae Bottom Seawate r Anaerobic ASW (B) 30 99 Shewanella haliotis Samples were taken during Nautile Dive (ND) #: ND 1726 Alvinella pompejana tubes ND 1727 Alvinella pompejana encrusted sulfides ND 1729 Fluid from Crab Spa; Bottom Seawater Acknowledgements We want to thank the crew of the R/V L’Atlante and the crew and pilots of the DSV/Nautile for sample collection We would like to offer our thanks to the members of the Deep Sea Microbiology Lab at Rutgers University their expertise, as well. This work was supported by an NSF grant. Table 2. Results of sequencing and BLAST. Conclusions •The molecular characterization of the organisms cultured in the samples taken from the dives reveals that there may be new species among them. •The sequences of the 16S rRNA gene from the organisms grown show that further isolation is needed for some samples in order to have a pure culture from which further investigation of the species can be done. • Future work would include characterization of isolated organisms. Morphological observation and studies of optimal growth in temperature, salinity and pH will be executed. Metabolism will be studied through growth under different conditions (i.e. electron donors and acceptors, pressure, etc.), in organic substrates, and resistance to antibiotics will also be tested. Table 1. Characteristics of anaerobic media used. Alvinella pompejana. (noc.soton.ac.uk) Crab Spa (whoi.edu)