Five hundred tons of antibiotics are consumed yearly in the world. In this study, the biodegradation characteristics of tetracycline (TET) under nitrate-reducing, sulfate-reducing, and methanogenic conditions were determined by batch tests. Also, effects of TET on mixed microbial cultures were revealed by microbiological analysis. In this scope, gas generation and composition, dissolved organic carbon, and electron acceptor concentrations were monitored during 120 days. Additionally, changes on quantities of specific microbial groups were determined by Q-PCR. TET showed non-biodegradable behavior under nitrate-and sulfate-reducing conditions, whereas slightly biodegradable behavior under methanogenic conditions approximately 46 % degradation. The effects of TET on the abundance of mixed culture varied according to taxonomic units. Sulfate-reducing bacteria were inhibited by TET, while archaeal, bacterial, and methanogenic populations were not affected significantly.