Abstract Enrichment cultures for H2-CO2, methanol- or acetate-utilizing methanogens were prepared from two rice field soil samples. All the cultures except one acetate enrichment showed significant methane production. Pure cultures of Methanobacterium- and Methanosarcina-like organisms were isolated from H2-CO2 and methanol enrichment cultures, respectively, and were characterized for various nutritional and growth conditions. The organisms had an optimal pH range of 6.4–6.6 and a temperature optimum of 37°C. The Methanobacterium isolates were able to utilize H2-CO2 but no other substrates as sole energy source, while the Methanosarcina isolates were able to utilize methanol, methylamines or H2-CO2 as sole energy sources. Both Methanobacterium isolates and one isolate of Methanosarcina were able to use dinitrogen as the sole source of nitrogen for growth. The isolates used several sulfur compounds as sole sources of sulfur.