Reactor-scale cultivation of the hyperthermophilic methanarchaeon Methanococcus jannaschii to high cell densities

For the hyperthermophilic and barophilic methanarchaeon Methanococcus janna schii, we have developed a medium and protocols for reactor-scale cultivati on that improved the final cell yield per liter from similar to 0.5 to simi lar to 7.5 g of packed wet cells (similar to 1.8 g dry cell mass) under aut otrophic growth conditions and to similar to 8.5 g of packed wet cells (sim ilar to 2 g dry cell mass) with yeast extract (2 g liter(-1)) and tryptone (2 g liter(-1)) as medium supplements. For growth in a sealed bottle it was necessary to add Se to the medium, and a level of 2 mu M for added Se gave the highest final cell yield. In a reactor M. jannaschii grew without adde d Se in the medium; it is plausible that the cells received Se as a contami nant from the reactor vessel and the H2S supply. But, for the optimal perfo rmance of a reactor culture, an addition of Se to a final concentration of 50 to 100 mu M was needed, Also, cell growth in a reactor culture was inhib ited at much higher Se concentrations. These observations and the data from previous work with methanogen cell extracts (B. C. McBride and R. S. Wolfe , Biochemistry 10: 4312-4317, 1971) suggested that from a continuously spar ged reactor culture Se was lost in the exhaust gas as volatile selenides, a nd this loss raised the apparent required level of and tolerance for Se. In spite of having a proteinaceous cell wall, M,jannaschii withstood an impel ler tip speed of 235.5 cms(-1), which was optimal for achieving high cell d ensity and also was the higher limit for the tolerated shear rate. The orga nism secreted one or more acidic compounds, which lowered pH in cultures wi thout pH control; this secretion continued even after cessation of growth.