of inorganic carbon (primarily C02 + HCC>3" ) in the incubation 

 media. To determine the activity of inorganic carbon, replicate 

 50 fl aliquots were placed in scintillation vials containing 0.1 

 ml of hyamine hydroxide. Ten ml of Fluorosol (National 

 Diagnostics) was added to each sample, and the disintigrations 

 per minute determined by scintillation counting in a Beckman LS 

 6800 liquid scintillation counter and corrections made for 

 background and counting efficiency (~ 92%). The total inorganic 

 carbon pool in the seawater or saline incubation media was 

 determined by gas chromatography (Childress, Arp, and Fisher 

 1984). Rates of total inorganic carbon fixation were calculated 

 from these measurements (Strickland and Parsons 1972). 



Elemental Sulfur Determination 



For the analysis of elemental sulfur, pieces of tissue (0.1 - 

 4.0 g wet weight) were dried for 18 hours in a 60xC drying oven 

 and then extracted for 24 hours with cyclohexane in a micro- 

 soxhlet apparatus. The extract was cleaned up by passage through 

 a fluorosil column and concentrated by allowing most of the 

 solvent to evaporate. Sulfur in the extract was quantified by 

 gas chromatography according to the method of Richard et al. 

 (1977). Ten fl of cyclohexane was injected onto the column. The 

 injector temperature was 240xC and the initial column temperature 

 was 150xC programmed to 220xC over the course of the separation. 

 A six foot glass column with a 2 mm bore, packed with 5% SP2401 

 on 100/120 mesh Supelcoport was used to separate the sulfur. 

 The sulfur was detected and quantified using a thermal 

 conductivity detector. The detection limit for elemental sulfur 

 was ca. 0.001% of the dry weight of the sample (depending 

 somewhat on sample size). The identity of the separated sulfur 

 was confirmed by the distinctive smell of sulfur vapor coming out 

 of the gas chromatograph. 



Metabolism Measurements 



The net fluxes of C0„, ? , CH., and other gases were measured 

 for pieces of gill, trophosome preparations and whole animals to 

 test for methane consumption. The gases were analyzed by gas 

 chromatography of 0.5ml aliquots. The gill and trophosome 

 incubations were prepared as described previously and were 

 carried out in glass syringes. The whole animal incubations were 

 carried out in a flowing water respirometer system (Anderson, 

 Childress, and Favuzzi 1987). 



RESULTS 



The bulk of the results are presented in Table 1. The 

 undescribed hydrocarbon seep mytilid which has been shown to 

 contain methanotrophic symbionts (Childress et al . 1986) has a 

 very different pattern of properties than do the other species 

 tested. It is the only species tested which has endosymbionts 



180 



