Methane Production from Glucose by a Mixed Culture of Bacteria in the Chemostat: the Role of Citrobacter

Abstract

A chemostat culture under a nitrogen atmosphere with a formate and inorganic salts medium fed at dilution rates up to 0·1 h−1 and inoculated from a sewage works anaerobic digester, proved to be an easy way of isolating a stable methanogenic bacterial culture. Formate-limited growth yielded a mixed culture containing predominantly methanogenic bacteria morphologically similar to Methanobacterium formicicum and Methanobacterium ruminantium. When glucose was substituted for formate, a facultative anaerobe capable of metabolizing glucose was selected. This species far outnumbered any other on aerobically incubated nutrient agar and was identified as a species of Citrobacter. The main products of glucose fermentation by this citrobacter were acetate, formate, ethanol and lactate. When the spent medium from the citrobacter fermentation of glucose was fed to the methanogenic culture, formate and lactate appeared to be the substrates which limited methane production. When each of the citrobacter products (acetate, ethanol and lactate) was fed as the sole carbon source to a methanogenic culture selected by growth on formate, lactate by itself could maintain methane production, but acetate and ethanol by themselves were not substrates for methane production. Lactate is thus a major intermediate in the conversion of glucose to methane by the mixed culture.