To investigate the methanogenic population dynamics during the start-up under gradually increasing free ammonia levels, five mesophilic UASB reactors seeded with different sludges were operated for 140 days. Changes in the methanogenic community investigated by using 16S rDNA/rRNA based molecular methods such as denaturing gradient gel electrophoresis (DGGE), fluorescent in-situ hybridization (FISH), cloning and DNA sequencing. Free ammonia nitrogen (FAN) in the reactors was gradually increased from 50 to 130 mg l(-1) by increasing total ammonia nitrogen (TAN) from 1000 to 2500 mg l(-1) in the feed and keeping reactor pH at 7.7 in the active zone. Even at highest FAN level, COD and VFA removal efficiencies above 90 and 98% were obtained, respectively. However, Methanosaeta-related species mainly prevailing in seed sludges were substituted for Methanosarcina as the abundant methanogens although reactor performances were almost comparable and constant.