Simulated heap bioleaching of low-grade high pyrite-bearing chalcocite ore was conducted at 40 °C with aeration of CO 2 and N 2 . Ore samples were collected at day 43, 64, 85, 106 and subjected to microbial community analysis by 16S rRNA gene clone library. Phylogenetic analyses of 16S rDNA fragments revealed that the retrieved sequences are mainly related to genus Acidithiobacillus, Leptospirillum and Sulfobacillus. Aeration of CO 2 and N 2 significantly impacted the microbial community composition. When CO 2 was aerated, the proportion of genus Acidithiobacillus considerably increased, whereas the proportion of genus Leptospirillum and genus Sulfobacillus declined. However, with the aeration of N 2 , the proportion of genus Acidithiobacillus and Leptospirillum increased, but genus Sulfobacillus decreased. When there was no aeration, the microbial community was similar to the inocula with the proportion of genus Leptospirillum mounted. These results indicated that the limitation of oxygen could change the bioleaching microbial community and the aeration of CO 2 and N 2 was favourable for the growth of sulfur-oxidizer (At. caldus) and iron-oxidizer (L. ferriphilum) respectively, which could be used for the regulation of microorganisms' role in mineral bioleaching.