Methanogenesis from ethanol by defined mixed continuous cultures.

Methanogenesis from ethanol by defined mixed continuous cultures was studied. Under sulfate-free conditions, a Desulfovibrio strain was used as the ethanol-degrading species producing acetic acid and hydrogen. In a two-membered mutualistic coculture, the hydrogen was converted to methane by a Methanobacterium sp. and pH was maintained at neutrality by the addition of alkali. Introduction of a third species, the acetate-utilizing Methanosarcina mazei, obviated the need for external pH control. Methanogenesis by the co-and triculture was studied at various dilution rates in the steady state. The mutualistic coculture performed like a composite single species, as predicted from the theory of mutualistic interactions. Coupling between the mutualistic coculture and the acetate-utilizing methanogen was less tight. Increasing the dilution rate destabilized the triculture; at low dilution rates, instability was soon recovered, but at higher dilution rates imbalance between the rates of production and removal of acetic acid led to a drop in pH. Flocs formed in the triculture. An annulus of the Methanobacterium sp. and Desulfovibrio sp. was retained around the Methanosarcina sp. by strands of material probably derived from the Methanosarcina sp.