The apparent sulfur oxidation activities of four pure thermophilic archaea, Acidianus brierleyi (JCM 8954), Metallosphaera sedula (YN 23), Acidianus manzaensis (YN 25) and Sulfolobus metallicus (YN 24) and their mixture in bioleaching chalcopyrite were compared, which were characterized indirectly by the evolution of the cells concentration, pH value and sulfate ions concentration in solution. The results show that the mixed culture contributed significantly to the raising of leaching rate, which suggests that the mixed culture had a higher sulfur oxidation activity than the pure culture. Meanwhile, the results also indicate that the changes of parameters characterizing the sulfur oxidation activity of thermophilic archaea are often influenced by many factors, so it is hard to reflect accurately the specific sulfur oxidation activities among the different sulfur-oxidizing microbes when bioleaching chalcopyrite at different conditions. Accordingly, an efficient method to characterize microbial sulfur oxidation activity appears to be desirable.