The desmid Oocardium stratum is sporadically and exclusively found in active meteogene travertine habitats. The restriction to these peculiar biotopes is probably linked to specific carbon requirements. We studied inorganic carbon (Ci) uptake under controlled laboratory conditions. First, we developed a method for isolating and cultivating O. stratum. Colonies visible to the naked eye (green pinhead-shaped formations) were collected from a tufa spring in Lower Austria. We then grew unialgal cultures on solidified agar prepared with Wood’s Hole culture medium, buffered with 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) to pH 7 and enriched with CaCO3. Scanning and transmission electron microscopy were applied to study the condition of cells shortly after they started in vitro growth. For Ci uptake, pH-drift experiments were performed including the external carbonic anhydrase inhibitor acetazolamide and the inhibitor of membrane-bound adenosine triphosphatase (ATPase) sodium orthovanadate. We compared the Ci affinity of Oocardium to Acutodesmus acuminatus and Chlorella sp. grown under identical conditions. Whereas A. acuminatus was common in slightly alkaline, eutrophic surface waters, Chlorella sp. was isolated from a soil sample. Owing to its peculiar habitat with abundant free CO2, we expected a clear preference for CO2, no enzymatic catalysis of Ci uptake and absence of active HCO3- uptake for Oocardium. Our experiments confirmed these hypotheses. In contrast to A. acuminatus, we found no HCO3- uptake in Chlorella sp. and O. stratum. Moreover, enzyme inhibition assays did not suggest activity of those enzymes that are commonly responsible for active Ci uptake.