A field pilot-scale passive treatment system was developed for in-situ bioremediation of acid mine drainage (AMD). The microbial community and its variation were analyzed. The data proved that 93.7% of total soluble Fe and 99% of soluble Fe(II) could be removed by the system. Principal coordinates analysis (PCoA) showed that a low pH and an elevated Fe concentration within the system created a unique microbial community that was dominated by acidophilic iron-oxidizing bacteria and iron-reducing bacteria. Canonical correlation analysis (CCA) indicated that the pH, iron content and total sulfur jointly determined the composition of the microbial communities. Species of Ferrovum, Delftia, Acinetobacter, Metallibacterium, Acidibacter and Acidiphilium were highly enriched, which promoted the removal of iron. Furthermore, the results revealed important data for the biogeochemical coupling of microbial communities and environmental parameters. These findings are beneficial for further application of in-situ field bioreactors to remediate AMD.