Coupling of Methanothermobacter thermautotrophicus Methane Formation and Growth in Fed-Batch and Continuous Cultures under Different H₂ Gassing Regimens

In nature, H₂- and CO₂-utilizing methanogenic archaea have to couple the processes of methanogenesis and autotrophic growth under highly variable conditions with respect to the supply and concentration of their energy source, hydrogen. To study the hydrogen-dependent coupling between methanogenesis and growth, Methanothermobacter thermautotrophicus was cultured in a fed-batch fermentor and in a chemostat under different 80% H₂-20% CO₂ gassing regimens while we continuously monitored the dissolved hydrogen partial pressures (pH₂). In the fed-batch system, in which the conditions continuously changed the uptake rates by the growing biomass, the organism displayed a complex and yet defined growth behavior, comprising the consecutive lag, exponential, and linear growth phases. It was found that the in situ hydrogen concentration affected the coupling between methanogenesis and growth in at least two respects. (i) The microorganism could adopt two distinct theoretical maximal growth yields (YCH₄ max), notably approximately 3 and 7 g (dry weight) of methane formed mol⁻¹, for growth under low (pH₂ < 12 kPa)- and high-hydrogen conditions, respectively. The distinct values can be understood from a theoretical analysis of the process of methanogenesis presented in the supplemental material associated with this study. (ii) The in situ hydrogen concentration affected the "specific maintenance" requirements or, more likely, the degree of proton leakage and proton slippage processes. At low pH₂ values, the "specific maintenance" diminished and the specific growth yields approached YCH₄ max, indicating that growth and methanogenesis became fully coupled.