polymerized at 65°C for 5 days. Ultrathin sections were cut 

 using an LKB Nova ultramicrotome and collected on formvar and 

 carbon-coated 200 mesh copper grids. Sections, both stained 

 (uranyl acetate and lead acetate) and unstained, were viewed and 

 photographed on a Hitachi scanning/transmission electron 

 microscope (Model H-600). 



For scanning electron microscopy (SEM) of the bacterial mat 

 samples, a portion of the glutaraldehyde-f ixed particulate 

 material was desalted by consecutive immersion in a graded series 

 of seawater/distilled water solutions (to 0% seawater) 

 immediately followed by sample dehydration by consecutive 

 immersion in a graded series of distilled water/ethanol 

 solutions (to 100% ethanol ) . After dehydration, the ethanol was 

 exchanged with liquid carbon dioxide during the critical point 

 drying procedure. Prior to examination the samples were mounted 

 onto aluminum pedestals and coated, by vacuum evaporation, with a 

 layer of either gold (in preparation for high resolution viewing) 

 or carbon (in preparation for x-ray elemental analysis). The 

 samples were viewed at 100-20, OOOx magnification using an ISI 

 scanning electron microscope (Model SS-40), and images were 

 recorded on Polaroid P/N55 type film. Energy dispersive x-ray 

 fluorescence spectroscopy was performed using a Princeton Gamma 

 Tech spectrograph. 



Methane oxidation experiments 



Bacterial methane oxidation was assessed during both the 

 Alvin and Pisces V dive programs. Preliminary experiments were 

 performed to measure the potential for methane oxidation using 

 14 C-labeled methane as a tracer. Water samples collected on 

 Alvin dives #1799 and #1800, a bacterial mat sample and a deep- 

 sea water (1000 m) control were used for these experiments. 

 Twenty-five ml of each sample was placed into a glass serum 

 bottle which was stoppered with a gas-tight rubber septum. One 

 ml (50 y Ci) of 14 C-CH 4 (5 mCi mmol" 1 , Cat. #17339, ICN 

 Radiochemicals) was injected into each serum bottle and the 

 samples were inverted (to eliminate potential loss of -^C-CH^ and 

 l^C-CC^ ) and incubated at 25°C. Following incubation periods of 

 7.5 and 18 hours, exactly 10 ml of each sample was transferred, 

 by syringe, to a second serum bottle containing a gas tight 

 serum stopper, a plastic well and a piece of fluted filter paper 

 as described by Hobbie and Crawford (1969). After the addition 

 of NaOH hydroxide (100 y l of 1 M NaOH ) to the filter paper, one 

 ml of 5 M HC1 was added to each sample. Following a 6 hour 

 passive distillation period, the serum bottles were opened and 

 the filter paper wicks (now containing the -^CG^ released from 

 the acidified solutions) were removed, placed into glass vials 

 containing 10 ml of Aquasol-2 (New England Nuclear) and their 

 radioactivities measured using a Packard Model 4640 liquid 

 scintillation counter. The acid-insoluble particulate materials 

 were collected onto Whatman glass fiber filters (GF/F) and 



196 



