Cell suspensions of Thiovulum sp., collected from enrichment cultures, were grown, maintained, and harvested for periods up to 7 months. In open-flow cultures run with aerated seawater, a continuous supply of hydrogen sulfide was provided by diffusion through a semipermeable membrane from either a live culture of Desulfovibrio esturaii, neutralized sodium sulfide, or a N2-H2S gas mixture. Attempts to grow Thiovulum in pure culture failed despite variation in concentrations of dissolved oxygen and hydrogen sulfide in stratified as well as in completely mixed systems. Uptake of 14CO2 and some organic compounds by purified cell suspensions was measured, and values were corrected for the activity of heterotrophic as well as autotrophic contaminants as determined in control experiments. Cell populations exhibited maximum uptake activities during formation of the characteristic veils. Substantial uptake of CO2 in air-saturated seawater was coincident with an optimal concentration of hydrogen sulfide of about 1 mM. Glutamate and a selection of vitamins (B12M biotin, and thiamine) did not significantly affect the uptake of CO2. No substantial uptake of carbon from acetate, glutamate, mannitol, and Casamino Acids was found. Within the range of error indicated, the data are consistent with acceptance of a chemolithotrophic nature of Thiovulum.