Table of Contents

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

A list of and metadata for sequences of ICoMM GPS (Global Protistan Survery) Vent-related samples archived in the ICoMM database

These vent-related samples include 3 from off-axis at EPR and one from vent plume water at 2495 m at EPR.
They also include 3 samples from the 2007 cruise at Guaymas: control sediment, orange mat, and near-bottom CTD.

Link to VAMPS Archive at MBL
Follow the instructions below to access the site
login as: user: guest, passwd: guest

In the left panel select: "trimmed fasta sequences"

Select project: "ICM_GPS_Ev9 Global Protist Survey"

Select: "All reads"

Select" "Eukarya"

Select: "GPS_0010" (through to GPS_0016) as per the list in the data

You will able to down load thousands of sequence - these are trimmed and Q/C'd sequences.

Collection
Samples were collected from within a deep-sea hydrothermal plume using a standard
CTD/Niskin rosette deployed from the R/V Atlantis.  Microbial mats growing in warm
(above ambient temperature) locations and cold ‘control’ sediments were sampled
directly on the seafloor via DSRV Alvin using ‘push core’ sampling devices.

Plume sampling was accomplished by ‘tow-yo’ casting of the CTD/Niskin rosette directly
over a hydrothermal vent feature to within approximately 10 m above the seafloor.
Anomalies in both temperature and beam transmission signaled a high probability that
the sampling device was within a hydrothermal plume at which point the position of
R/V Atlantis was fixed and Niskin bottles were fired within the hydrographic feature.

Plume samples were confirmed at the surface by their strong sulfurous odor indicating
the presence of reduced sulfur compounds.  Sites for core sampling were generally selected
where there were sharp gradients between warm hydrothermally-influenced sediments harboring
dense microbial mats (largely Beggiatoa) and adjacent cold sediments without overlying
microbial mats.

Filtration
Water samples were pre-screened through 200 µm Nitex mesh followed by 80 µm Nitex mesh
(to reduce microbial metazoa e.g., larval stages, copepods, etc.) directly from Niskin
sampling bottles.  Microbial biomass in the < 80 µm size-fraction was collected onto GF/F
filters at low vacuum pressure  (< 10 mm Hg) and stored with 2 ml of lysis buffer (as in
Countway et al., 2005) at -80°C until extraction.  Sediments were retrieved from the seafloor
cores, sectioned on board ship at one centimeter intervals, and stored at -80°C until processing
in our home laboratories.  Samples from microbial mats were not pre-screened prior to freezing
at -80°C.

Molecular processing
High molecular weight DNA was extracted from sediment samples using the MoBio power soil
kit (MoBio, Carlsbad, CA. USA).  DNA from water samples collected on GF/Fs was extracted
using a standard Phenol:Chloroform:Isoamyl alcohol method and precipitated with 95% ethanol. 
All extracted samples were quantified on a Nanodrop 1000 (University of Delaware) or by
fluorometry (University of Southern California) using PicoGreen (Invitrogen).  DNA samples
for eukaryote ‘pyrotag sequencing’ were sent to the MBL in Woods Hole, MA for amplification
by PCR and subsequent library preparation as described in Amaral-Zettler et al. (2009).
Preliminary taxonomic assignments were made for all pyrotag sequences using the ‘GAST’
approach described in the previous reference.

Amaral-Zettler, L.A., McCliment, E.A., Ducklow, H.W., and Huse, S.M. (2009) A Method for Studying
Protistan Diversity Using Massively Parallel Sequencing of V9 Hypervariable Regions of Small-Subunit
Ribosomal RNA Genes. PLoS ONE 4: e6372. doi:6310.1371/journal.pone.0006372.

Countway, P.D., Gast, R.J., Savai, P., and Caron, D.A. (2005) Protistan diversity estimates based
on 18S rDNA from seawater incubations in the western North Atlantic. Journal of Eukaryotic Microbiology 52: 95-106.

Protistan assemblages are essential components of food webs in the vast majority of aquatic ecosystems that have been examined to date. Despite the pivotal roles that these organisms play in carbon fixation, energy utilization and elemental cycling in marine ecosystems, little information exists on the presence, abundance and activities of these species in the deep ocean in general, and in particular at and around hydrothermal vents. The latter environments should be particularly conducive to the growth of phagotrophic protists in the deep ocean because biomass is greatly elevated at these sites and organic carbon availability in these regions is largely mediated by archaeal and bacterial assemblages. This research project will address fundamental questions regarding the species diversity, abundances of specific taxa, and trophic activities of protists within the deep ocean including hydrothermal vent areas. Specifically, the PIs will determine whether deep-sea communities harbor endemic assemblages of protists, establish the identity of the protists associated with these environments that are trophically active, and document the protistan taxa that dominate these assemblages in situ. They will examine the diversity of the protistan assemblages in the deep-sea away from the vents and in the overlying water column, and in microenvironments in and around hydrothermal vents and associated with dominant macrofauna of vents. Traditional (microscopy) and molecular biological (18S rDNA) approaches will be used to characterize abundance and diversity. Observations of the ingestion of fluorescently labeled prey will be used to establish trophically active protists in situ. They will also employ 18S rDNA clone libraries, cultures and microscopy-based observations to target specific taxa for the application of fluorescent in situ hybridization (FISH) to link morphological identity to commonly occurring (and trophically active) phylotypes. A culture collection of deep-sea protists will be established to further characterize protistan diversity and also to provide specimens for baseline physiological measurements in the laboratory. Growth rates will be examined in the laboratory using cultured protists grown under temperatures, pressures and water chemistries representative of deep-sea environments. Hydrothermal vent research is an excellent vehicle for incorporation into education at all levels ranging from elementary through graduate level. The information resulting from this research will be incorporated into undergraduate and graduate courses by the PIs, and will be featured on the Caron Lab Homepage. The PIs and the postdoctoral investigator supported by this project will participate directly in an ongoing teacher education program (Centers for Ocean Science Education Excellence; COSEE-West) that will reach middle and high school students, most of whom are Hispanic, African-American or other ethnic minorities and most of whom are economically disadvantaged. This will be accomplished through existing teacher enhancement and student enrichment activities that will incorporate this research into a learning experience that will enhance student awareness of environmental science, microbiology and the natural world. This work will be the center feature for the internationally acclaimed Extreme 2000 outreach program for secondary schools which has, over the last 5 years, directly engaged over 180,000 students throughout the US and 9 foreign countries. Contributions from this project will educate the public at every level about the deepsea, and the importance of microbes in forming and maintaining the biosphere.