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doi:10.2204/iodp.proc.336.105.2012

Microbiology

Hard rock samples were collected for microbiological studies from every interval cored with the RCB in Hole U1383C. Roughly 12% (6.11 m total) of core recovered from Hole U1383C was taken as whole-round samples from the core splitting room and dedicated to microbiological analysis. The 79 hard rock microbiology samples span a range of lithologic units, alteration states, and presence of chilled margins, and some contain at least one vein or fracture. Table T8 provides a description of the collected samples, and appendix Figures AF1–AF79 provide photographs of the whole-round samples taken (see the “Appendix”). Additionally, a few background contamination samples were collected for shore-based DNA analysis, including any recovered plastic bags that held the fluorescent microsphere solutions in the core catcher.

In accordance with protocols described in “Microbiology” in the “Methods” chapter (Expedition 336 Scientists, 2012a), samples were selected in the core splitting room as quickly as possible after core recovery, following initial discussion with petrologists on sample representation and photographing the sample before it was removed from the core liner. When sample volume permitted, samples were preserved for shore-based DNA and RNA analysis, FISH, cell counting analysis, isotopic analysis, and fluorescent microsphere analysis as well as ship-based culturing and enrichment experiments. Microspheres were used during all coring operations to help in evaluating core contamination. Microsphere density was evaluated in all samples following iterative washing of the exterior of the whole-round samples with sterile seawater. Table T8 summarizes the results of the microsphere contamination survey, listing whether the microspheres were detected in the first or last of 3–4 total washes with sterilized seawater. Of the 79 hard rock samples collected from Hole U1383C, 30 had microspheres in the first wash but none in the final wash, 19 did not have any detectable microspheres in any wash, and 30 had microspheres in all washes.

Several enrichment and cultivation experiments were started with rock samples collected from Hole U1383C (see “Microbiology” in the “Methods” chapter [Expedition 336 Scientists, 2012a]). Carbon-fixation incubations were initiated with several glassy basalt samples: 336-U1383C-13R-2-MBIOB, 19R-1-MBIOA, 20R-1-MBIOA, and 22R-1-MBIOA. Enrichments for carbon- and nitrogen-cycling (carbon fixation, methane oxidation, and nitrate reduction) microorganisms were conducted with samples from 18 different basalt samples. Enrichments for methanogens, sulfate reducers, sulfide oxidizers, and nitrate-reducing iron-oxidizing bacteria were conducted with rocks from several horizons: Samples 336-U1383C-6R-1-MBIOA, 9R-3-MBIOD, 10R-2-MBIOC, 16R-2-MBIOB, 20R-2-MBIOB, 30R-2-MBIOB, and 32R-2-MBIOA. Enrichments for heterotrophic metabolisms will be set up with material from Samples 336-U1383C-13R-2-MBIOB, 19R-1-MBIOA, 20R-1-MBIOA, and 22R-1-MBIOA.

Forty-one samples from Hole U1383C were scanned with the Deep Exploration Biosphere Investigative portable tool (DEBI-pt) deep UV fluorescence instrument for evaluation of microbial and organic carbon, mineral composition, and the presence of fluorescent microsphere contamination controls on the surfaces of rock fragments. Laboratory tests revealed that the fluorescent microspheres (made of polystyrene) used as contamination checks during coring have a distinct fluorescence pattern (Fig. F27) when excited by deep UV light. During subsampling of rocks collected from Hole U1383C, care was taken to document the orientation (i.e., exterior and interior) of fragments used for DEBI-pt scanning to enable evaluation of whether microsphere contamination tracers were found on either the exterior or interior of the samples. Of the samples scanned, only two exterior pieces (Samples 336-U1383C-2R-2-MBIOE and 8R-2-MBIOB) showed unequivocal fluorescence patterns consistent with those of microspheres. Other samples contained spectral signatures that had some resemblance to microspheres; however, suspect regions also contained fluorescence from a variety of other sources, precluding definitive identification at that time. All other samples showed varying degrees of microbial and organic carbon abundance. Some samples contained very little microbial fluorescence (e.g., Sample 336-U1383C-6R-1-MBIOA; Fig. F28), whereas others had a higher concentration of biomass (e.g., Sample 336-U1383C-26R-1-MBIOA; Fig. F29). The majority of fluorescence in the low-biomass Sample 336-U1383C-6R-1-MBIOA (Fig. F28) had spectral characteristics consistent with calcium, potassium, and magnesium silicates. A small amount of biomass and other low molecular–weight aromatic hydrocarbons occurred in some of the areas with altered basalt, whereas very little signal was detected in the areas with ochre alteration. In contrast, Sample 336-U1383C-26R-1-MBIOA had a higher degree of organic and microbial fluorescence distributed throughout the scanned region (Fig. F29).

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