Glycerol-supplemented medium promotes transition of Cupriavidus necator from heterotrophic to lithoautotrophic growth

H₂-driven biotransformations in the chemolithoautotrophic bacterium, Cupriavidus necator, are gaining attention to obtain valuable industrial molecules. However, cultivation variability and long lag phases have been reported in C. necator chemolithoautotrophic cultures. Few studies have focused on the management of these reported long lag phases. This article proposes a cultivation strategy based on serial precultures with glycerol to reduce metabolic transition. Two preculture media with and without glycerol were adapted in a seed-train to study the transition towards lithoautotrophy. Cultures were compared by monitoring chemolithoautotrophic growth in small-scale system. The presence of glycerol in fructose preculture medium completely removed the lag phase during subsequent lithoautotrophic cultivation compared to fructose only precultures (4-5 h). Glycerol can be used as carbon source to improve the transition to lithoautotrophy in C. necator, while reducing the lag phase. This preculture setup can be implemented to study the effects of any relevant biomolecule, while avoiding the random nature of culture start that makes interpretations difficult. The study provided valuable insights in the management of C. necator precultures, and lithoautotrophic cultivation systems to support the development of C. necator as a platform organism.